Biomarkers

Abstract

The present invention relates to a method of screening for NASH, and for discriminating between non-alcoholic steatohepatitis (NASH) with and without liver fibrosis in a subject having or suspected of

Description

The present invention relates to biomarkers associated with non-alcoholic steatohepatitis (NASH). The present invention relates to methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis, and to methods of screening for NASH. NASH is a type of liver disease. NASH is characterized a build-up of fat in the liver and liver inflammation. Such inflammation can seriously harm the liver. NASH is common in patients who are overweight (e.g. obese) and / or who have Type 2 diabetes. Note that NASH is synonymous with “metabolic dysfunction-associated steatohepatitis” (MASH). Throughout this document, we refer to this liver disease as NASH. NASH may be symptomless, but in some cases patients with NASH may experience extreme tiredness, weakness and / or discomfort in the upper right side of the abdomen. There is no standard treatment for patients that have NASH, but lifestyle changes including losing weight, maintaining a healthy diet and / or addressing an underlying condition such as hypothyroidism or Type 2 diabetes have been shown to affect its progression. Vitamin E may also be useful for NASH treatment. In some cases, NASH is not accompanied by liver fibrosis (scarring). This may be referred to as NASH without fibrosis. However, as NASH progresses, persistent inflammation can cause liver fibrosis (scar tissue around the liver and nearby blood vessels). NASH that is accompanied by liver fibrosis may be referred to as NASH with fibrosis. There are thus different types of NASH; NASH without fibrosis and NASH with fibrosis. NASH with fibrosis is a more severe type of NASH than NASH without fibrosis. Extensive liver fibrosis can eventually lead to cirrhosis (which is a later stage, more severe, type of liver disease). There is an urgent need to improve the current screening and diagnostics landscape for NASH. In particular, affordable and practical tools for NASH diagnostics are needed to assist healthcare professionals, for example to assist in diagnosing NASH and / or in determining whether subjects have NASH with liver fibrosis or NASH without liver fibrosis. Being able to discriminate between subjects having NASH with liver fibrosis and subjects having NASH without liver fibrosis would be useful as the severity of the NASH could be assessed. This would help clinicians to guide the management of NASH patients. Circulating biomarkers are molecules that can be measured in accessible body fluids of individuals, e.g. urine or blood (e.g. plasma), and whose levels are useful to assist in, for example, screening for disease, diagnosis of disease, and monitoring of disease progression. What is needed in the art are new methods of screening for NASH, and for distinguishing between NASH with liver fibrosis and NASH without liver fibrosis. The identification of novel biomarkers for NASH may potentially have clinical implications for a large number of patients and would be an important clinical advancement. Here, the inventors provide methods that employ certain markers which, advantageously, are useful in screening for NASH, and for discriminating (or distinguishing) between NASH with liver fibrosis and NASH without liver fibrosis. Advantageously, such methods are non-invasive and can be performed on readily obtainable samples. Surprisingly and advantageously, the present inventors have identified that the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) is different in body fluid samples from patients having NASH with liver fibrosis in comparison to patients having NASH without liver fibrosis. The present inventors have also identified that the chemical composition of chondroitin sulfate (CS) is different in body fluid samples from patients with NASH in comparison to patients with simple steatosis. These differential chemical compositions of the GAG CS (GAG profiles) as characterized by differential levels of certain GAG properties in accordance with the present invention, can act as biomarkers useful for NASH screening and for distinguishing between patients with NASH with liver fibrosis and patients with NASH without liver fibrosis. Clearly the finding that NASH screening, and distinguishing between patients with NASH with liver fibrosis and patients with NASH without liver fibrosis, can be carried out using an accessible body fluid sample, e.g. urine or plasma, from a subject is extremely advantageous. Thus, in one aspect the present invention provides a method of discriminating between non-alcoholic steatohepatitis (NASH) with liver fibrosis and non-alcoholic steatohepatitis (NASH) without liver fibrosis in a subject having non-alcoholic steatohepatitis or in a subject suspected of having non-alcoholic steatohepatitis, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS; and (ii) 4S6S CS, wherein said sample has been obtained from said subject. In another aspect, the present invention provides method of screening for nonalcoholic steatohepatitis in a subject, said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS, and (ii) 4S6S CS, wherein said sample has been obtained from said subject. In some embodiments, methods of the present invention comprise determining the level of 4S CS. In some embodiments, methods of the present invention comprise determining the level of 4S6S CS. In some embodiments, methods of the present invention comprise determining the level of 4S CS and 4S6S CS. In some embodiments, methods of the present invention comprise determining the level of one or more GAG property selected from the group consisting of: (i) absolute concentration of 4S CS; and (ii) relative concentration of 4S6S CS. The terms “absolute concentration” and “relative concentration” are discussed elsewhere herein. Unless otherwise clear from the context, the term “one or more” includes “all”. In some embodiments, methods of the present invention comprise determining the absolute concentration of 4S CS. In some embodiments, methods of the present invention comprise determining the relative concentration of 4S6S CS. In some embodiments, methods of the present invention comprise determining the absolute concentration of 4S CS or the relative concentration of 4S6S CS. In some embodiments, methods of the present invention comprise determining the absolute concentration of 4S CS and the relative concentration of 4S6S CS. Thus, in preferred embodiments of the present invention, the level of 4S CS is the absolute concentration of 4S CS, and the level of 4S6S CS is the relative concentration of 4SCS CS. In some embodiments, the body fluid sample is blood (e.g. plasma). In some embodiments, the body fluid is urine. In some embodiments, the body fluid is blood (e.g. plasma) and the method comprises determining the level of 4S CS (preferably the absolute concentration of 4S CS). In some embodiments, the body fluid is urine the method comprises determining the level of 4S6S CS (preferably the relative concentration of 4S6S CS). In some embodiments, methods comprise determining the chemical composition of CS in accordance with the invention (and optionally also the chemical composition of HS) in more than one (e.g. two) type of body fluid sample (e.g. in a blood (e.g. plasma) sample and in a urine sample). Put another way, in some embodiments, methods may comprise determining the chemical composition of CS in accordance with the invention (and optionally also the chemical composition of HS) in different (e.g. two different) body fluid sample types (e.g. in a blood (e.g. plasma) sample and in a urine sample). Thus, in some embodiments, methods of the present invention may comprise determining the level of 4S CS (preferably the absolute concentration of 4S CS) in a blood (e.g. plasma) sample) and determining the level of 4S6S CS (preferably the relative concentration of 4S6S CS) in a urine sample. Unless otherwise clear from the context, references to one or more GAG properties (or GAG forms or GAG features or GAGome features or GAGome properties) “in accordance with the invention” or “in accordance with the present invention” (or equivalent phrases) are references to one or more of 4S CS (preferably absolute concentration of 4S CS) and 4S6S CS (preferably relative concentration of 4S6S CS). In preferred methods of the present invention, the determined level of one or more GAG property in accordance with the invention is compared to a control (e.g. control level). Thus, in preferred embodiments, there is a step of comparing one or more GAG property in accordance with the invention to a control (e.g. a control level). In preferred methods of the invention, an altered level (increased or decreased as the case may be) of one or more GAG properties in accordance with the invention (4S CS (e.g. absolute concentration of 4S CS) and / or 4S6S CS (preferably relative concentration 4S6S CS)) in comparison to a control level is indicative of NASH in a subject (e.g. NASH with fibrosis or NASH without fibrosis in said subject) (e.g. indicative of whether said subject has NASH, or whether said subject has NASH with fibrosis or NASH without fibrosis). In certain methods of discriminating in accordance with the invention (and analogous methods), an altered level (e.g. increased level or higher level) of one or more GAG properties in accordance with the invention (4S CS (e.g. absolute concentration of 4S CS) and / or 4S6S CS (e.g. relative concentration of 4S6S CS)) in comparison to a control level is indicative of NASH with fibrosis (or fibrotic NASH) in said subject (indicative of whether said subject has NASH with fibrosis (or fibrotic NASH)). In certain methods of discriminating in accordance with the invention (and analogous methods), an altered level (e.g. decreased level or lower level) of one or more GAG properties in accordance with the invention (4S CS (e.g. absolute concentration of 4S CS) and / or 4S6S CS (e.g. relative concentration of 4S6S CS)) in comparison to a control level is indicative of NASH without fibrosis (or non-fibrotic NASH) in said subject (indicative of whether said subject has NASH without fibrosis (or non-fibrotic NASH)). The skilled person would be readily able to establish appropriate control levels for use in accordance with the invention. Certain appropriate control levels and how an indication of NASH with fibrosis or NASH without fibrosis can be reached (discrimination of likely NASH with fibrosis versus likely NASH without fibrosis) are discussed elsewhere herein. In some embodiments of methods of discriminating in accordance with the invention, a level of one or more of the GAG properties in accordance with the invention (4S OS (e.g. absolute concentration of 4S CS) and / or 4S6S OS (e.g. relative concentration of 4S6S CS)) that lies within an established (or given or appropriate) reference range (or reference interval) or a level that is altered (increased or decreased as the case may be) with respect to an established (or given or appropriate) reference range (or reference interval) is indicative of either NASH with fibrosis or NASH without fibrosis in said subject (indicative of whether said subject has either NASH with fibrosis or NASH without fibrosis). The skilled person would be readily able to establish appropriate reference ranges (or reference intervals) for use in accordance with the invention. Certain appropriate reference ranges (or reference intervals) and how an indication of NASH with fibrosis or NASH without fibrosis can be reached using such reference ranges are discussed elsewhere herein. Methods of the present invention comprise determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample. In some embodiments, methods of the present invention comprise determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) and the chemical composition of the glycosaminoglycan (GAG) heparan sulfate (HS) in a body fluid sample. As described elsewhere herein, in some preferred embodiments of methods of the present invention, the method comprises determining the chemical composition of the protein-free fraction of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally also of the glycosaminoglycan (GAG) heparan sulfate (HS)) in a body fluid sample. Thus, in some embodiments, the chemical composition of the protein-free fraction of chondroitin sulfate (CS) is determined. In some embodiments, the chemical composition of the protein-free fraction of heparan sulfate (HS) is additionally determined. Thus, in some embodiments, the chemical composition of the protein-free fraction of chondroitin sulfate (CS) and heparan sulfate (HS) is determined. Although in some embodiments of methods of the invention the method comprises determining the chemical composition of the protein-free fraction of CS (and optionally also the chemical composition of HS), it is not essential in all aspects of the invention for the chemical composition specifically of the “protein-free fraction” to be determined. Glycosaminoglycans (GAGs) are sugar containing molecules which can be attached to proteins on serine residues, i.e. can form a part of a proteoglycan. They are formed from linear or unbranched chains of monosaccharides (i.e. are polysaccharides) which can be sulfated. Heparan sulfate (HS), chondroitin sulfate (CS), keratan sulfate (KS), hyaluronic acid (HA) and heparin are the common types of GAG, of which HS and CS are examples of sulfated GAGs. The different types of GAG are distinguished by different repeating disaccharide units. When linked or attached to proteins, CS and HS are GAGs that share a common biosynthetic route in the linkage to the core protein, but thereafter they differ in their polymerisation in that the CS repeating disaccharide is made up of repeating N-acetylgalactosamine (GalNAc) and glucuronic acid residues (GlcA), whilst the repeating disaccharide in HS is typically made up of repeating N-acetylglucosamine (GIcNAc) and glucuronic acid (GlcA) residues. Each monosaccharide is attached by a specific enzyme allowing for multiple levels of regulation over GAG synthesis. Although GAGs can be attached to proteins, i.e. they may be in a protein-bound or proteoglycan form, GAGs can also exist in a “free” form, i.e. they can also exist in a non protein-bound or non-proteoglycan form. Such “free” forms of GAGs are referred to herein as “protein-free GAGs”. Thus, in body fluids (or body samples) there is typically a protein-free fraction of GAGs (or protein-free pool of GAGs) and a protein-bound fraction of GAGs (or protein-bound pool of GAGs). Together (i.e. protein-free fraction plus protein-bound fraction), the two fractions may be referred to as the entire GAG fraction or entire GAG pool. In preferred methods of the invention, the chemical composition of the protein-free fraction of chondroitin sulfate (CS) (and optionally also of heparan sulfate (HS)) in a body fluid sample is determined. The protein-free fraction GAGs (or disaccharide units derived therefrom as discussed elsewhere herein) for analysis may be obtained by any suitable means. As discussed elsewhere herein, body fluid samples are typically processed prior to analysis. Such processing typically comprises subjecting the GAG(s) to a processing step to obtain disaccharide units for analysis. Such a processing step typically comprises contacting said sample (or said GAG(s) in said sample) with an enzyme (e.g. a GAG lyase such as a chondroitinase or a heparinase) which digests (or fragments) the GAG(s) into disaccharide units. Without wishing to be bound by theory, such enzymes are not able to access proteinbound GAGs but rather act on (or use as their substrate) only (or essentially only) protein-free GAGs. If proteoglycans (which are proteins with GAGs bound or attached thereto) are contacted with a proteolytic agent (e.g. a protease such as proteinase K) the protein component thereof is digested by the proteolytic agent and the protein-bound GAGs are freed or released, meaning that protein-bound GAGs are (or are converted into) into a “free" or “released" form, which would then be available for digestion (or fragmentation) by an enzyme such as a GAG lyase. As in certain preferred methods of the invention it is specifically the composition of the protein-free fraction (or naturally protein-free fraction) of the GAG CS (and optionally also of the GAG HS) that is determined, then preferred methods of the invention do not comprise contacting the sample with a proteolytic agent (e.g. a protease such as proteinase K). Omitting a proteolytic agent during processing of a sample for analysis is thus a way to obtain (or obtain only or obtain essentially only) the protein-free fraction of GAGs (or disaccharides subsequently derived therefrom) for analysis. Proteinbound GAGs (proteoglycan GAGs) that have been (or become) “freed" or “released" by the action of a proteolytic agent (e.g. a protease) are not protein-free GAGs in accordance with the present invention. Thus, protein-free GAGs (or the protein-free fraction of GAGs) are GAGs (or the fraction of GAGs) that are already (or naturally) free (i.e. not protein-bound) in the absence of (or without) the sample having been treated with a proteolytic agent (e.g. a protease). Put another way, the protein-free GAGs (or the protein-free fraction of GAGs) are GAGs (or the fraction of GAGs) that are free (i.e. not protein-bound) in an original, or initial, or unprocessed sample. For example, the protein-free GAGs (or the protein-free fraction of GAGs) can be GAGs (or the fraction of GAGs) that are present in a sample, e.g. an original or unprocessed sample, and are susceptible to, or accessible to, or available for (e.g. are a substrate for) digestion (or fragmentation) into disaccharide units as described elsewhere herein, e.g. using an enzyme such as a lyase enzyme. As indicated above, protein-free GAGs (or the protein-free fraction of GAGs) may be considered the non-protein bound (or non-protein bound fraction) or non-proteoglycan form (or non-proteoglycan fraction) of GAGs. Put another way, the protein-free GAGs (or the protein-free fraction of GAGs) are GAGs not decorating a proteoglycan in the original, or initial, or unprocessed sample. Thus, in some preferred embodiments, methods comprise determining the chemical composition of the protein-free fraction of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in accordance with the invention (and optionally also of the GAG HS), in a body fluid sample, wherein the sample is subjected to processing prior to determining the chemical composition and wherein said processing does not comprise contacting said sample with a proteolytic agent (e.g. a protease such as proteinase K), or other agent which can release protein-free GAGs from protein-bound GAGs. Alternatively viewed, in some preferred embodiments, methods comprise determining the chemical composition of the protein-free fraction of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in accordance with the invention (and optionally also of the GAG HS) in a body fluid sample, wherein the sample is subjected to processing prior to determining said composition and wherein said processing does not comprise contacting said sample with a proteolytic agent (e.g. a protease such as proteinase K), or other agent which can release GAGs (or GAG chains) from proteoglycans. Thus, in some preferred embodiments, methods comprise determining the chemical composition of the protein-free fraction of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in accordance with the invention (and optionally also of the GAG HS) in a body fluid sample, wherein said sample has been obtained from said subject and has been subjected to processing prior to determining said chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units (e.g. as described elsewhere herein); and (b) does not comprise prior to (a) contacting said sample with an agent which can release GAGs (or GAG chains) from proteoglycans. Thus, in some preferred embodiments, methods comprise determining the chemical composition of the protein-free fraction of the glycosaminoglycans (GAG) chondroitin sulfate (CS) in accordance with the invention (and optionally also of the GAG HS) in a body fluid sample, wherein said sample has been obtained from said subject and has been subjected to processing prior to determining said level and / or chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units (e.g. as described elsewhere herein); and (b) does not comprise prior to (a) contacting said sample with a proteolytic agent. As indicated above, protein-bound GAGs (or the protein-bound fraction of GAGs) may be considered proteoglycan GAGs (or the proteoglycan fraction of GAGs). Alternatively viewed, protein-bound GAGs (or the protein-bound fraction of GAGs) may be considered as GAGs that typically require the protein to which they are bound to be contacted with a proteolytic agent (e.g. a protease such as a non-specific protease) in order for them to be (or become) freed or released. In some other embodiments of methods of the invention the chemical composition in accordance with the invention of the entire fraction (or entire pool) of the GAG CS (and optionally also of the GAG HS) in a body fluid sample may be determined (i.e. protein-free GAGs plus protein-bound GAGs). In such embodiments, the sample is typically contacted with a proteolytic agent (such as a protease, e.g. a non-specific protease, e.g. proteinase K) during processing of the sample. The “level” of CS or HS as referred to herein generally refers to the total level or amount (e.g. concentration) of the CS or HS present in the sample. The total level or amount (e.g. concentration) of the CS or HS present in a sample may be referred to as “Total CS” or “Total HS”. The level of CS and / or HS in a sample can be measured or determined by any appropriate method which would be well-known and described in the art. Some methods involve electrophoresis, in particular capillary electrophoresis, e.g. capillary electrophoresis with fluorescence detection, e.g. laser-induced fluorescence detection. Other suitable methods are gel electrophoresis, e.g. agarose gel electrophoresis (e.g. FACE, fluorophore-assisted carbohydrate electrophoresis) or mass spectrometry or liquid chromatography, e.g. HPLC, optionally in combination with mass spectrometry (HPLC-MS). Preferred methods involve high performance liquid chromatography (HPLC), preferably ultra-HPLC (UHPLC), in combination with mass spectrometry, e.g. MS / MS or triple quadrupole mass spectrometry. Preferred methods comprise ultra-high-performance liquid chromatography (UHPLC) coupled with electrospray ionization triple-quadrupole mass spectrometry system. Conveniently these levels can be measured as a concentration (e.g. a real or absolute level or concentration), for example, as a number of microgram per ml (pg / ml). However, again, any appropriate measure of level may be used. In methods of the present invention which involve determining the levels of CS and / or HS (e.g. the levels of the protein-free fraction of HS and / or CS) the levels of CS and or HS are typically determined separately or individually. In other words the methods do not involve the measurement of total GAG levels in a sample or the total levels of all the GAGs present in combination (e.g. in the protein-free GAG fraction) but may involve the measurement of the levels of one or more of the individual GAGs HS or CS. In some embodiments, the level (e.g. total level, or concentration) of CS and / or HS (e.g. the level of the protein-free fraction of CS and / or HS), can be determined in body fluid samples, for example in urine or blood samples. The individual monosaccharide units making up the CS and HS can have different sulfation patterns in terms of the position of the sulfate molecules and the amount / number of sulfate molecules. For CS, sulfation may most commonly occur at one or more of position 2 of the GlcA and positions 4 and 6 of the GalNAc. For HS, sulfation may occur at one or more of position 2 of the GlcA after epimerization to IdoA (iduronic acid), positions 3 and 6 of the GIcNAc, and N-sulfation of the GIcNAc. Thus, each individual disaccharide in the GAG chain may have 0 (i.e. be unsulfated), 1, 2, 3 or 4 (only in HS) sulfation forms and this in turn gives rise to different overall chemical compositions of GAG chains in terms of sulfation levels and specific disaccharide sulfation patterns. As described elsewhere herein, methods of the invention involve the determination of the chemical composition of CS (and optionally also of HS). The term “chemical composition” as used herein can refer to both the levels of the GAGs as well as the disaccharide sulfation composition of the GAGs. In particular, this term includes a determination of one or more particular forms, e.g. sulfation forms, of the disaccharides making up the the GAG CS GAG (and optionally also the GAG HS). Put another way, the term “chemical composition” refers to the amount or level of one or more of the various sulfated and / or unsulfated forms of CS or HS disaccharides, as well as, for example, some other properties of the individual GAGs present, such as total CS or HS GAG levels, or other properties related to GAG sulfation such as CS charge or HS charge as described further elsewhere herein. Such a chemical composition which is analysed or determined in the present invention can also be referred to herein as a GAG profile, GAG forms, GAG features, GAG properties, GAGome, GAGome features. Thus, for example, the term “chemical composition” as used herein may refer to a determination or analysis of the sulfation patterns (e.g. one or more of the sulfation forms) of the disaccharides making up CS (and optionally also HS). As is evident from the above discussion, in accordance with the present invention it is essential that the level of one or more GAG property selected from the group consisting of 4S CS (preferably the absolute concentration of 4S CS) and 4S6S CS (preferably the relative concentration of 4S6S CS) is determined. Thus, in accordance with the present invention and disclosure, determining the “chemical composition” must include a determination of the level of one or more GAG property selected from the group consisting of 4S CS and 4S6S CS. The level of one or more other (additional) GAG properties (e.g. as described herein) may additionally be determined (or measured), but in accordance with the present invention and disclosure it is the level of one or more GAG property selected from the group consisting of 4S CS and 4S6S CS that allows an indication (e.g. diagnosis) of NASH, for example NASH without fibrosis or NASH with fibrosis (as appropriate) to be made (or reached). For CS, there are 8 main sulfated and unsulfated forms (sulfation patterns, disaccharide sulfation forms) which are: OS CS (also referred to as unsulfated CS or CS O unit), 2S CS (also referred to as chondroitin-2-sulfate), 4S CS (also referred to as chondroitin-4-sulfate or CS A unit), 6S CS (also referred to as chondroitin-6-sulfate or CS C unit), 2S4S CS (also referred to as chondroitin-2-4-sulfate), 2S6S CS (also referred to as chondroitin-2-6-sulfate or CS D unit), 4S6S CS (also referred to as chondroitin-4-6-sulfate or CS E unit) and Tris CS (also referred to as chondroitin-2-4-6-sulfate or trisulfated CS). Each of the above is a form of CS GAG (a CS GAG form or property or feature or GAGome feature) which may be measured in the methods of the present invention. One or more of these forms may be measured, for example up to 8, e.g. 1, 2, 3, 4, 5, 6, 7 or all 8 of these sulfation forms may be measured. In some embodiments, measurement of all 8 of these sulfation forms is preferred. In some embodiments, measurement of, or of at least, the CS sulfation forms OS CS, 6S CS, 4S CS, 2S6S CS, 2S4S CS, 4S6S CS is preferred. Another GAG property for CS which may be measured in the methods of the present invention is the total concentration of CS (which may also be referred to as CS tot or Tot CS or Total CS) or the total level of CS. This is typically measured as a concentration, e.g. an absolute concentration, e.g. in pg / ml, as described elsewhere herein. In some embodiments, the total CS is measured by summing the level of all measured CS disaccharide forms listed above (the 8 main sulfated and unsulfated forms). Thus, total CS may be the sum of the levels of all measured CS disaccharide forms listed above (the 8 main sulfated and unsulfated forms). In some embodiments, the total CS is measured by summing the level of all of the CS disaccharide forms that are measured at (or above) an analytically detectable level, or at (or above) a level that has been selected as a minimum threshold level (a minimum threshold level that is considered analytically detectable or analytically meaningful, for example, the limit of detection or the lower limit of quantification for such CS disaccharide form). For example, the total CS may be the sum of all of the CS disaccharide forms that are measured at a level at, or above, a given analytically detectable level, or at, or above, a selected minimum threshold level (e.g. at a concentration of >0.1pg / ml). The level of CS disaccharide forms below a given analytically detectable level or below a minimum threshold level would typically provide only a negligible (or de minimis) contribution to the overall CS concentration, and so such CS disaccharide forms may, if desired, be excluded from the summing of CS disaccharide forms that is done to determine or measure Total CS concentration. In some embodiments the total concentration of CS is not measured. “Charge CS” is another GAG form or property which may be measured in the present invention, e.g. as part of the GAG profile. “Charge CS” refers to the total fraction of sulfated disaccharides of CS, i.e. the fraction of sulfated disaccharides of CS present or measured or detected in a sample out of the total CS disaccharides present or measured or detected in a sample (i.e. sulfated CS disaccharides / sulfated + unsulfated CS disaccharides). In some embodiments, “Charge CS” refers to the weighted sum of the concentration of all CS disaccharides divided by the total CS, where the weight is the count of sulfo groups in that disaccharide, i.e. 0 for OS CS, 1 for 4S CS, 6S CS, and 2S CS, 2 for 2S6S CS, 4S6S CS, and 2S4S CS, and 3 for Tris CS (and this is the definition of “Charge CS” used in connection with the term “Charge CS” in the Example section herein). As the measurement of “charge CS” is dependent on the measurement of other properties, i.e. the measurement of levels of sulfated and unsulfated CS disaccharides, this property is not referred to herein as an independent GAG property or CS property. In some embodiments it is preferred to measure up to 8 (e.g. 1, 2, 3, 4, 5, 6, 7 or 8) or all 8 of the CS sulfation forms (i.e. the sulfated and unsulfated forms), together with total CS. Charge CS may additionally be measured in some embodiments. In preferred embodiments, more than one (e.g. at least 2, 3, 4, 5, 6, 7 or 8) CS sulfation form is measured. In some embodiments, one or more (or all) of the following GAG properties may be measured or determined: the relative level of 4S CS with respect to 6S CS (e.g. the ratio 4S CS / 6S CS or the inverse ratio 6S CS / 4S CS), the relative level of 6S CS with respect to OS CS (e.g. the ratio 6S CS / OS CS or the inverse ratio OS CS / 6S CS) or the relative level of 4S CS with respect to OS CS (e.g. the ratio 4S CS / OS CS or the inverse ratio OS CS / 4Ss CS). In some embodiments, the relative level of 4S CS with respect to 6S CS (e.g. the ratio 4S CS / 6S CS or the inverse ratio 6S CS / 4S CS) is not measured or determined. For HS, there are 8 main sulfated and unsulfated forms (sulfation patterns, disaccharide sulfation forms) which are: OS HS (also referred to as unsulfated HS), 2S HS (which is sulfated at the 2-position of GlcA), NS HS (which is sulfated at the N-position of the GIcNAc), 6S HS (which is sulfated at the 6-position of the GIcNAc), 2S6S HS (which is sulfated at the 2-position of GlcA and the 6-position of the GIcNAc), NS6S HS (which is sulfated at the 6-position and N-position of GIcNAc), NS2S HS (which is sulfated at the 2-position of GlcA and the N-position of GIcNAc), Tris HS (which is sulfated at the 2-position of GlcA and 6-position and N-position of GIcNAc, also referred to as trisulfated HS). Note that sulfation in position 3 of the GIcNAc is also possible but rarely observed. Each of the above is a form of HS GAG (an HS GAG form or property or feature or GAGome feature) which may be measured or determined in the methods of the present invention. However, due to its rareity, in preferred embodiments of the invention, the sulfation form with sulfation in position 3 of the GIcNAc is not measured. Thus, in the methods of the invention, one or more (or all) of these 9 (or preferably 8) forms may be measured, for example up to 9 (or preferably up to 8), e.g. 1,2, 3, 4, 5, 6, 7, 8 or all 9 of these sulfation forms may be measured. In some embodiments, measurement of all 8 of these sulfation forms (excluding the sulfation form with sulfation in position 3 of the GIcNAc) is preferred. Another GAG property for HS which may be measured in the methods of the present invention is the total concentration of HS (which may also referred to as HS tot or Tot HS or Total HS) or the total level of HS. This is typically measured as a concentration, e.g. an absolute concentration, e.g. in pg / ml, as described elsewhere herein. In some embodiments, the total HS is measured by summing the level of all measured HS disaccharide forms listed above (preferably the 8 main sulfated and unsulfated forms, i.e. excluding the rare sulfation form with sulfation in position 3 of the GIcNAc). Thus, total HS may be the sum of the levels of all measured HS disaccharide forms listed above (preferably the sum of the 8 main sulfated and unsulfated forms, i.e. excluding the rare sulfation form with sulfation in position 3 of the GIcNAc). In some embodiments, the total HS is measured by summing the level of all of the HS disaccharide forms that are measured at (or above) an analytically detectable level, or at (or above) a level that has been selected as a minimum threshold level (a minimum threshold level that is considered analytically detectable or analytically meaningful, for example, the limit of detection or the lower limit of quantification for such HS disaccharide form). For example, the total HS may be the sum of all of the HS disaccharide forms that are measured at a level at, or above, a given analytically detectable level, or at, or above, a selected minimum threshold level (e.g. at a concentration of >0.1pg / ml). The level of HS disaccharide forms below a given analytically detectable level or below a minimum threshold level would typically provide only a negligible (or de minimis) contribution to the overall HS concentration, and so such HS disaccharide forms may, if desired, be excluded from the summing of HS disaccharide forms that is done to determine or measure Total HS concentration. In some embodiments the total concentration of HS is not measured. “Charge HS” is another GAG form or property which may be measured in the present invention, e.g. as part of the GAG profile. “Charge HS” refers to the total fraction of sulfated disaccharides of HS, i.e. the fraction of sulfated disaccharides of HS present or measured in a sample out of the total HS disaccharides present or measured in a sample (i.e. sulfated HS disaccharides / sulfated + unsulfated HS disaccharides). In some embodiments, “Charge HS” refers to the weighted sum of the concentration of all HS disaccharides divided by the total HS, where the weight is the count of sulfo groups in that disaccharide, i.e. 0 for OS HS, 1 for NS HS, 6S HS, 2S HS, 2 for 2S6S HS, NS6S HS, and NS2S HS, and 3 for Tris HS (and this is the definition of “Charge HS” used in connection with the term “Charge HS” in the Example section herein). As the measurement of “charge HS” is dependent on the measurement of other properties, i.e. the measurement of sulfated and unsulfated HS disaccharides, this property is not referred to herein as an independent GAG property or HS property. In some embodiments it is preferred to measure up to 8 (e.g. 1, 2, 3, 4, 5, 6, 7 or 8) or all 8 of the HS main sulfation forms (i.e. the sulfated and unsulfated forms listed above excluding the sulfation form with sulfation in position 3 of the GIcNAc), together with total HS. Charge HS may additionally be measured in some embodiments. In preferred embodiments, at least one (or at least 2, 3, 4, 5, 6, 7 or 8) HS sulfation form is measured. In some embodiments, the 8 main sulfated and unsulfated HS forms, total HS, the 8 main sulfated and unsulfated CS forms, and total CS are measured, i.e. 18 GAG properties. In some embodiments, the 8 main sulfated and unsulfated HS forms and the 8 main sulfated and unsulfated CS forms are measured, i.e. 16 GAG properties. In some embodiments, the level and / or chemical composition of hyaluronic acid (HA) may be additionally determined in a body fluid sample. Hyaluronic acid (HA) is typically nonsulfated. Accordingly, when HA is measured in accordance with the invention, it is typically and preferably the level (total level or total concentration) of HA that is measured (which may also be referred to as Total HA). This is typically measured as a concentration, e.g. in pg / ml, as described elsewhere herein. In some embodiments, HA is not measured. In some embodiments, the 8 main sulfated and unsulfated HS forms, total HS, the 8 main sulfated and unsulfated CS forms, total CS and total HA are measured, i.e. 19 GAG properties. CS (Total CS) and HS (Total HS) is typically measured in terms of an absolute concentration, e.g. in pg / ml, as described elsewhere herein. The various CS sulfation forms and HS sulfation forms may also be measured in terms of an absolute concentration, e.g. in pg / ml. Thus, in some embodiments the level (or concentration) of a given CS sulfation form or a given HS sulfation form is an absolute level or absolute concentration of a given CS sulfation form or a given HS sulfation form. In some embodiments, absolute levels or absolute concentrations are preferred. However, the various CS sulfation forms and HS sulfation forms may alternatively, or additionally, be measured in terms of a relative concentration (or relative level). Thus, in some embodiments the level (or concentration) of a given CS sulfation form or a given HS sulfation form is the relative level or relative concentration. Thus the “level” or “concentration” of GAG sulfation forms may be its absolute concentration or its relative concentration. In some embodiments, both the absolute concentration and the relative concentration of one or more sulfated GAG forms is measured (or determined). The “relative concentration” may be considered the mass fraction (e.g. in %) of a given CS sulfation form or a given HS sulfation form that is obtained (or calculated or determined) by normalizing its absolute concentration to the total concentration of the relevant GAG class, i.e. by normalizing its absolute concentration to the total CS concentration or total HS concentration (as appropriate). Thus, the relative concentration of a given CS sulfation form may be considered the mass fraction (e.g. in %) of said given CS sulfation form that is obtained (or calculated or determined) by normalizing its absolute concentration by the total CS concentration. The relative concentration of a given HS sulfation form may be considered the mass fraction (e.g. in %) of said given HS sulfation form that is obtained (or calculated or determined) by normalizing its absolute concentration by the total HS concentration. Relative concentration may be expressed in terms of a percentage (%). Thus, in some embodiments the level (or concentration) of a given CS sulfation form or a given HS sulfation form may be the absolute concentration and / or the relative concentration of the given CS sulfation form or the given HS sulfation form. In some embodiments the level (or concentration) of a given CS sulfation form or a given HS sulfation form is the absolute concentration of the given CS sulfation form or the given HS sulfation form. In some embodiments the level (or concentration) of a given CS sulfation form or a given HS sulfation form is the relative concentration of the given CS sulfation form or the given HS sulfation form. A relative concentration may be alternatively viewed as a “fraction” or “mass fraction” or “proportion" or “relative measurement" as discussed below. As discussed above, GAG properties or GAG forms, e.g. disaccharide sulfation forms (with the exception of total CS or total HS) may be measured as a fraction size or fraction or mass fraction (e.g. pg / pg) or proportion or relative measurement, rather than as absolute levels or concentrations, for example are given a value of less than 1 or are normalised to 1 depending on the levels of all the sulfation forms for the relevant GAG class (or all the main sulfation forms for the relevant GAG class) measured (or detectable) in the sample (or are expressed in terms of a %). In other words, the level of each of the desired sulfation forms is measured independently and then normalised to 1. In other words, the level of each of the desired sulfation forms is measured independently and then its mass fraction or volume fraction or mole fraction is computed. These fractions may also be expressed as percentage. In other words, these fractions may also be normalised to 100. For example, in some embodiments, the fraction size of a given sulfated CS form or unsulfated CS form may be determined by measuring the level of the given sulfated CS form or unsulfated CS form and dividing this by the sum of the levels of all of the CS sulfation forms (or all of the main sulfation forms) and the unsulfated CS form measured (or present or detectable) in the sample. In some embodiments, the fraction size of a given sulfated HS form or unsulfated HS form may be determined by measuring the level of the given sulfated HS form or unsulfated HS form and dividing this by the sum of the levels of all of the HS sulfation forms (or main sulfation forms) and the unsulfated HS form measured (or present or detectable) in the sample. When calculating such fractions, it is preferred that at least the main sulfation forms of CS or HS are measured in order to be able to normalise the fraction of the particular individual sulfation form to 1. Relative measurements may be easier to interpret, for example, a measurement of 0s HS of 0.6 indicates that 60% of the measured HS disaccharides are unsulfated. However, absolute levels can also be measured. Indeed, in some embodiments it is preferred to measure absolute concentrations of one or more sulfation forms. In accordance with the present invention, the level of one or both of the GAG properties 4S CS and 4S6S CS is measured or determined. Typically, the disaccharide composition (for example the specific sulfation patterns (e.g. sulfation forms)) of more than one (e.g. all) of the disaccharides making up CS (and optionally also making up HS) is measured or determined. In some embodiments, more than one (e.g. all) sulfation properties or forms (or GAG features or properties or GAGome features or properties) of CS (and optionally also of HS) such as those outlined above (e.g. OS CS, 2S CS, etc), are measured or determined. Appropriate methods of doing this would be well known to a skilled person in the art and any of these could be used. However, a convenient method to achieve such quantification of disaccharide composition or the appropriate properties or forms of CS or HS (and separation of the disaccharide forms) is to use electrophoresis, in particular capillary electrophoresis, e.g. capillary electrophoresis with fluorescence detection, e.g. capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). An alternative method is liquid chromatography, preferably HPLC (high-performance liquid chromatography), for example SAX HPLC. Preferably mass spectrometry is also used (e.g. HPLC-MS), for example electrospray ionization mass spectrometry (ESI-MS). Alternatively, mass spectrometry can be used without chromatography, e.g. liquid chromatography. One example is capillary electrophoresis with laser-induced fluorescence detection. Another example is HPLC ESI-MS. Preferred methods involve high performance liquid chromatography (HPLC), preferably ultra-HPLC (UHPLC), in combination with mass spectrometry, e.g. MS / MS or triple quadrupole mass spectrometry. Preferred methods comprise ultra-high-performance liquid chromatography (UHPLC) coupled with electrospray ionization triple-quadrupole mass spectrometry. Particularly preferred methods are outlined in the Examples. In some methods of the invention where the levels of one or more individual disaccharide forms are measured, the GAG(s) are subjected to a processing step, for example a step of fragmentation or cleavage or digestion, e.g. by chemical digestion or enzyme treatment in order to obtain the disaccharide units which are then analysed. The enzyme may be a GAG lyase, e.g. a chondroitinase or a heparinase, or a combination of chondrotinases, or a combination of heparinases, or a combination of one or more chondroitinases and one or more heparinases. Preferably, the chondroitinase is Chondroitinase ABC or Chondroitinase AC or Chondroitinase A or Chondroitinase B or Chondroitinase C. Preferably the heparinase is Heparinase l-ll-lll. In preferred embodiments one or more chondroitinases and one or more heparinases are used, preferably Chondroitinase ABC and Heparinase l-ll-lll. In some methods of the invention the GAGs in the sample are subjected to a step of extraction (e.g. using a proteolytic agent such as a protease, e.g. a non-specific protease, e.g. proteinase K) and / or purification, e.g. using an anion-exchange resin (or other means to purify GAGs based on the negative charge of the GAGs). However, in preferred methods of the invention one or both of these steps is not carried out (i.e. there is no such extraction and / or no such purification). For example, in preferred embodiments of the invention, e.g. where the chemical composition of the protein-free fraction of CS (and optionally also of HS) is determined, then no such protein digestion (extraction) step is carried out. In other words, said methods do not involve a processing step in which samples are contacted with a proteolytic agent, such as for example a protease, e.g. proteinase K. As discussed elsewhere herein, omitting a processing step in which samples are contacted with a proteolytic agent means that the protein-free fraction of GAGs can be specifically analysed. In other preferred embodiments of the invention, said methods do not involve a processing step in which the GAGs are purified from the sample based on the negative charge of said GAGs, e.g. using an anion-exchange resin. Without wishing to be bound by theory, omitting a processing step in which the GAGs are purified from the sample based on the negative charge of said GAGs (e.g. using an anion-exchange resin) simplifies the method and can lead to efficiencies in terms of the yield of GAGs obtained during processing of the body fluid sample. In preferred methods of the present invention, methods do not involve a processing step in which samples are contacted with a proteolytic agent, such as for example a protease, e.g. proteinase K, and do not involve a processing step in which the GAGs are purified from the sample based on the negative charge of said GAGs, e.g. using an anion-exchange resin (or other means to purify GAGs based on the negative charge of the GAGs). In some methods of the invention the GAGs in the sample (e.g. various different GAG forms in the sample) are subjected to a step of separation and / or quantification, as described elsewhere herein. For example, as discussed elsewhere herein, HPLC in combination with mass spectrometry may be used in preferred embodiments. Particularly preferred methods comprise ultra-high-performance liquid chromatography (LIHPLC) coupled with electrospray ionization triple-quadrupole mass spectrometry. Other methods which might be used are known in the art. However, examples are analytical techniques involving the use of antibodies to various GAG forms, e.g. techniques such as Western blot, ELISA or FACS, or methods involving agarose gel electrophoresis (e.g. fluorophore-assisted carbohydrate electrophoresis (FACE)) or polyacrylamide gel electrophoresis (PAGE). In any embodiments which refer to the determination of the level of one or more from a certain list of GAG properties (or GAG forms or GAG features or GAGome features) in accordance with the invention, in some such embodiments the level of all the listed GAG forms (or GAG properties or GAG features or GAGome features) in accordance with the invention may be determined. In some embodiments, the level of a single GAG property (GAG form or GAG feature or GAGome feature) in accordance with the invention is determined. In other embodiments of the present invention, the level of more than one GAG property (GAG form) is determined (e.g. the level of two or more GAG forms, or three or more GAG forms, or four or more GAG forms, or five or more GAG forms is determined). By “more than one” is meant 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, etc. In any list of markers or GAG properties provided herein, in some embodiments all are measured (or determined). Also, a determination of the level of each and every possible combination of the GAG forms can be performed. Thus, in some embodiments multi-marker methods are performed. Determining the level of multiple of the GAG properties (biomarker multiplexing) in accordance with the invention may improve screening (e.g. diagnostic) accuracy. Thus, although markers (GAG forms or GAG properties or features or GAGome features) in accordance with the invention can be used in the methods of the invention individually, they can also be used in combination, e.g. in the form of a multi-marker assay. In some embodiments, the level of a single GAG form (GAG property) in accordance with the invention is used for screening for NASH or used as the basis of the discriminating between NASH with liver fibrosis and NASH without liver fibrosis, e.g. a discrimination (or distinguishing) between NASH with liver fibrosis and NASH without liver fibrosis may be made on the basis of the level of a single GAG form in accordance with the invention in some embodiments (i.e. on the basis of the level of 4S OS or 4S6S CS). In other embodiments, more than one GAG form in accordance with the invention is used for the basis of the screening for NASH or used as the basis of the discriminating between NASH with liver fibrosis and NASH without liver fibrosis, e.g. a discrimination (or distinguishing) between NASH with liver fibrosis and NASH without liver fibrosis may be made on the basis of the level of more than one GAG form in accordance with the invention (i.e. on the basis of the level of both of 4S CS and 4S6S CS). In some embodiments, where both GAG properties in accordance with the invention is used for the basis of the screening for NASH (e.g. for discriminating or distinguishing between NASH with liver fibrosis and NASH without liver fibrosis), the level of one or more (or all) of the other GAG properties (or GAG forms) described herein may be additionally determined or measured. Based on the observed alterations in the levels of one or more GAG property in accordance with the invention, if desired, scoring methods, scoring systems, markers or formulas can be designed which use such levels of GAG properties in order to arrive at an indication, e.g. in the form of a value or score, which can then be used for screening (e.g. discriminating, diagnosis, etc.). Appropriate scoring systems and parameters (e.g. GAG properties) to be measured can readily be designed based on (or including) one or more of the individual GAG properties in accordance with the present invention. In some embodiments in accordance with the present invention, the chemical composition may be expressed in terms of score (or GAG score), said score being based on (or derived or calculated by using) the measured level of one or both of the GAG properties in accordance with the invention. Thus, a score may be based on (or derived or calculated or computed using) one or both measured (or determined) GAG properties selected from the group consisting of: 4S OS and 4S6S OS. In some embodiments, a score may be based on (or derived or calculated or computed using) one or both measured (or determined) GAG properties in accordance with the present invention (4SCS and / or 4S6S CS) and one more other (e.g. 2, 3, 4, 5, 6, 7, 8, etc., or all) GAG properties described herein. In some embodiments, a score may be based on (or derived or calculated or computed using) measured levels of one or more GAG properties in more than one type of body fluid sample. In some such embodiments, the GAG properties measured in each type of body fluid may be the same or they may be different. In some embodiments, appropriate threshold or cut-off values (e.g. used to declare a sample as being NASH (or indicative of NASH), or to discriminate between NASH with liver fibrosis and NASH without liver fibrosis (or indicate likely NASH with liver fibrosis or likely NASH without liver fibrosis), as appropriate, for use with scores can be designed by a person skilled in the art. By way of example, a cut-off (or threshold) value may be calculated based on ROC curves. In some cases, maximally selected rank statistics may be used to identify a cut-off value (or cut-off score). In some embodiments, screening (e.g. diagnosis, or discriminating, etc.) may be done by comparing a given score for a sample (or subject) to be screened (e.g. diagnosed) with a threshold or cut-off value, and assigning a result (e.g. an indication of NASH, or an indication of NASH with liver fibrosis or an indication of NASH without liver fibrosis) based on whether the score determined is above or below (e.g. significantly above or significantly below) a cutoff value. Any scoring methods, scoring systems, markers or formulas can be used that comprise a GAG property (or combination of GAG properties) in accordance with the invention in order to arrive at an indication, e.g. in the form of a value or score, which can then be used for screening (e.g. diagnosis) of NASH or for discriminating between NASH with liver fibrosis and NASH without liver fibrosis). For example, said methods etc., can be an algorithm that comprises a GAG property (or combination of GAG properties) in accordance with the invention as input, to e.g. perform pattern recognition of the samples, in order to arrive at an indication, e.g. in the form of a value or score, which can then be used for screening (e.g. diagnosis) of NASH or for discriminating between NASH with liver fibrosis and NASH without liver fibrosis. Non-limiting examples of such algorithms include machine learning or deep learning algorithms that implement classification (algorithmic classifiers), such as linear classifiers (e.g. Fisher’s linear discriminant, logistic regression, naive Bayes classifier, perceptron); support vector machines (e.g. least squares support vector machines); quadratic classifiers; kernel estimation (e.g. k-nearest neighbor); boosting; decision trees (e.g. random forests); neural networks; learning vector quantization. The use of such classifiers, e.g. machine learning classifiers, e.g. random forest classifiers, would be within the skill of a person skilled in the art. In some embodiments, a multivariable logistic regression model may be used to compute the GAG score. As described elsewhere herein, discriminating between for NASH with liver fibrosis and NASH without liver fibrosis (or analogous methods) in accordance with the present invention may involve using a score, or expressing the chemical composition in accordance with the invention using a score. As described elsewhere herein, in some such embodiments, an altered score (e.g. increased or decreased as the case may be) in comparison to a control score (or cut-off level or threshold level) is indicative of NASH with liver fibrosis or NASH without liver fibrosis (as appropriate) in said subject. As described elsewhere herein screening for NASH in accordance with the present invention may involve using a score, or expressing the chemical composition in accordance with the invention using a score. As described elsewhere herein, in some such embodiments, an altered score (e.g. increased or decreased as the case may be) in comparison to a control score (or cut-off level or threshold level) is indicative of NASH in said subject. As discussed above, the present invention provides a method of discriminating between NASH with liver fibrosis and NASH without liver fibrosis in a subject. The present invention also provides a method of screening for NASH with liver fibrosis or NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the level and / or chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS; and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). The method of screening for NASH with liver fibrosis or NASH without liver fibrosis in accordance with the present invention can be used, for example, for diagnosing NASH with liver fibrosis or NASH without liver fibrosis. The present invention therefore also provides a method of diagnosing NASH with liver fibrosis or NASH without liver fibrosis in a subject. Thus, in one aspect the present invention provides a method for diagnosing NASH with liver fibrosis or NASH without liver fibrosis in a subject. In some embodiments, a positive diagnosis (i.e. the presence of NASH with liver fibrosis or the presence of NASH without liver fibrosis as the case may be) is made if the level of one or more of the GAG properties in accordance with the invention in the sample is altered (increased or decreased as the case may be) in comparison to an appropriate control level (e.g. as discussed elsewhere herein). Thus, a further aspect, the present invention provides a method of diagnosing NASH with liver fibrosis or NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS. wherein said sample has been obtained from said subject. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). As discussed above, the present invention provides a method of discriminating between NASH with liver fibrosis and NASH without liver fibrosis in a subject. Alternatively viewed, the present invention provides a method of distinguishing between (or differentiating between) NASH with liver fibrosis and NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). The level of one or more GAG property in accordance with the present invention may be used in (or for) the differential diagnosis of NASH with liver fibrosis and NASH without liver fibrosis (NASH with liver fibrosis versus NASH without liver fibrosis), or as a part of a differential diagnosis process for NASH with liver fibrosis and NASH without liver fibrosis (NASH with liver fibrosis versus NASH without liver fibrosis). The concept of “differential diagnosis” is well understood in the art. Thus, in another aspect, the present invention provides a method of differential diagnosis of NASH with liver fibrosis and NASH without liver fibrosis (NASH with liver fibrosis versus NASH without liver fibrosis) in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluid, preferred control levels, etc.). As described elsewhere herein, methods in accordance with the present invention may be used in conjunction with other diagnostic tests or methods. Thus, in another aspect, the present invention provides a method of confirming (or validating) a prior diagnosis of NASH (e.g. NASH with liver fibrosis or NASH without liver fibrosis) in a subject, said method comprising: determining the chemical composition the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Embodiments of methods described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). In another aspect, the present invention provides a method of discriminating between NASH with liver fibrosis or NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample, wherein said sample has been obtained from said subject. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). In another aspect, the present invention provides a method of screening for NASH with liver fibrosis or NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample, wherein said sample has been obtained from said subject. Embodiments of methods of screening for NASH with liver fibrosis or NASH without liver fibrosis of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). In another aspect, the present invention provides a method of diagnosing NASH with liver fibrosis or NASH without liver fibrosis in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample, wherein said sample has been obtained from said subject. Embodiments of methods of screening (e.g. diagnosing) NASH with liver fibrosis or NASH without liver fibrosis of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). Unless otherwise clear from the context, the features and discussion herein in relation to methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (e.g. in relation to preferred GAG properties or combinations thereof, or scores for measurement, preferred processing steps, preferred body fluids, preferred control levels, etc.) apply, mutatis mutandis, to other related methods of present invention (e.g. to methods of screening for (e.g. diagnosing) NASH with liver fibrosis or NASH without liver fibrosis, differentially diagnosing NASH with liver fibrosis or NASH without liver fibrosis, distinguishing (or differentiating) between NASH with liver fibrosis and NASH without liver fibrosis, or confirming a (prior) diagnosis of NASH with liver fibrosis or NASH without liver fibrosis, etc.). In some embodiments, the invention provides the use of the methods of the invention (e.g. discriminating methods, screening methods, diagnostic methods, or methods of determining progression, etc., as described herein) in conjunction with other known screening, diagnostic or progression monitoring methods for NASH (e.g. NASH with fibrosis or NASH without fibrosis), such as imaging or scanning methods (e.g. ultrasound scan, Magnetic Resonance Imaging (MRI) scan or computed tomography (CT) scan)) and / or analysis of liver biopsies. Thus, for example, methods of the invention can be used to confirm a diagnosis (confirm a prior diagnosis) of NASH (e.g. NASH with fibrosis or NASH without fibrosis) in a subject. In some embodiments the methods of the present invention are used alone. The level of the GAG form (or GAG property) in question can be determined or measured by analyzing the sample which has been obtained from or removed from the subject by an appropriate means. The determination is typically carried out in vitro. Levels of one or more of the GAG forms in the sample can be measured (determined) by any appropriate assay or technique or method, a number of which are well known and documented in the art. Electrophoresis, e.g. agarose gel electrophoresis or capillary electrophoresis (in particular capillary electrophoresis with fluorescence detection such as CE-LIF) is a technique that can be used for measuring (determining) the levels of one or more of the GAG forms. Liquid chromatography, in particular HPLC (high-performance liquid chromatography) in combination with mass spectrometry (MS) are preferred techniques for measuring (determining) the levels of one or more GAG forms. Suitable electrophoresis, e.g. capillary electrophoresis, and liquid chromatography, e.g. HPLC techniques for GAG form analysis, together with appropriate mass spectrometry methods (and associated data processing techniques) are well known and documented in the art. One method that may be used in the invention is capillary electrophoresis with laser-induced fluorescence detection, CE-LIF (e.g. as described in Galeotti et al., 2014, Electrophoresis 35: 811-818; and Kottler et al., 2013, Electrophoresis 34: 2323-2336). HPLC combined with post column derivatization and fluorimetric detection can also be used, e.g. as described in Volpi 2006, Curr Pharm Des 12:639-658, as can HPLC combined with ESI-MS (electrospray ionization-mass spectrometry), e.g. as described in Volpi and Linhardt, 2010, Nature protocols 5:993-1004, also with fluorimetric detection, e.g. as described in Galeotti and Volpi, 2011, Anal Chern 83:6770-6777, or Volpi et al., 2014, Nature Protocols 9:541-558. Agarose gel electrophoresis can also be used, e.g. FACE (fluorophore assisted carbohydrate electrophoresis) as described in Volpi and Maccari, 2006, AnalytTechnol Biomed Life Sci, 834:1-13; and Volpi and Maccari, 2002, Electrophoresis 23:4060-4066. A particularly preferred method for determining the level of one or more of the GAG forms in the sample is described herein in the Examples. Thus, preferred methods may involve high performance liquid chromatography (HPLC), preferably ultra-HPLC (UHPLC), in combination with mass spectrometry, e.g. MS / MS or triple quadropole mass spectrometry. Particularly preferred methods comprise ultra-high-performance liquid chromatography (UHPLC) coupled with electrospray ionization triple-quadrupole mass spectrometry system. An example of such methods is described in Tamburro et al. (Journal of Chromatography B, 1177 (2021) 122761). Certain methods of sample preparation (or processing), e.g. GAG extraction and purification are also known and described in the art, for example Volpi and Maccari, 2005, Biomacromolecules 6:3174-3180 and Clin Chim Acta 356:125-133, Coppa etal., 2011 Glycobiology 21:295-303. Such reported art based methods of sample preparation (or processing) involve a protease treatment (protease extraction) and purification step based on using an anion-exchange resin. In some methods of the present invention such a protease treatment step and / or purification step using an anion-exchange resin may be performed. However, as discussed elsewhere herein, in preferred methods a step of protease treatment and / or a purification step using an anion-exchange resin is not performed. In particular, in preferred methods where the protein-free fraction of the GAG(s) is analysed, then a step of protease treatment is not performed. In some embodiments HPLC and mass spectrometry (and associated data processing techniques) is used to obtain a fraction of the level of one or more particular GAG forms (e.g. the sulfated or unsulfated disaccharide forms) in the sample in comparison to the total amount. For example, after sample preparation, GAGs can be digested using enzymes, separated in an HPLC column and characterized using mass spectrometry (MS). As described elsewhere herein, the quantities of one or more individual GAG forms (e.g. a particular sulfated or unsulfated disaccharide form) may be conveniently normalised (i.e. divided) by the sum of all the quantities of individual GAG forms measured, to yield fractions (or proportions or relative concentrations). However, absolute concentrations (or absolute levels) of individual GAG forms (e.g. GAG sulfation forms) may alternatively, or additionally, be measured. In accordance with the present invention, a quantitative, semi-quantitative or qualitative assessment (determination) of the level of one or more of the GAG forms can be made. Appropriate methods of doing this would be well known to a skilled person in the art and any of these could be used. However, a convenient method to achieve such quantification of disaccharide composition or the appropriate properties or forms of CS or HS (and separation of the disaccharide forms) is to use electrophoresis, in particular capillary electrophoresis, e.g. capillary electrophoresis with fluorescence detection, e.g. capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) (e.g. as described in Galeotti 2014, supra, or Kottler 2013, supra). An alternative method, which is preferred in some embodiments, is to use liquid chromatography, preferably HPLC (high-performance liquid chromatography), for example SAX HPLC or for example as described in Volpi 2006, supra, Galeotti and Volpi 2011, supra, Volpi et al., 2014, supra or Volpi and Linhardt, 2010, supra. Preferably mass spectrometry is also used (HPLC-MS), for example electrospray ionization mass spectrometry (ESI-MS), e.g. HPLC ESI-MS. Particularly preferred methods are outlined in the Examples. One example of a particular method is capillary electrophoresis (e.g. for example, capillary electrophoresis with laser-induced fluorescence detection). Another particularly preferred example would be HPLC followed by MS (HPLC-MS), e.g. HPLC ESI-MS. Preferred HPLC-MS methods are discussed elsewhere herein. Thus, in preferred methods of the invention said level or chemical composition of said GAG or GAG property is determined by HPLC and mass spectrometry. Preferably said HPLC is ultra-HPLC and / or said mass spectrometry is triple-quadrupole mass spectrometry. In certain preferred methods, said level or chemical composition of said GAG or GAG property is determined by high performance liquid chromatography (HPLC), preferably ultra-HPLC (UHPLC), in combination with mass spectrometry, e.g. MS / MS or triple quadrupole mass spectrometry. Preferred methods comprise ultra-high-performance liquid chromatography (UHPLC) coupled with (or in combination with) electrospray ionization triplequadrupole mass spectrometry. Generally, the determination of the GAG properties (or forms or features) in accordance with the present invention does not involve the measurement of GAG molecules in the exact same form as found in the body fluid of a subject (e.g. does not involve the measurement of a naturally occurring form of GAG). For example, such native or naturally occurring GAG molecules are often found in biological samples, e.g. body fluid samples, in the form of long sugar chains which can either be attached to proteins (also referred to herein as protein-bound GAGs or proteoglycan GAGs), or not attached to proteins (also referred to herein as free GAGs or protein-free GAGs). In some embodiments, methods of the invention may include a step of processing a sample. In some embodiments, the methods of the invention may thus be performed on such processed samples or materials derived from such processed samples. Thus, generally the methods of the invention are carried out on samples which have been processed in some way (e.g. are man-made rather than native samples). Processing steps may include, but are not limited to, extraction or purification of GAGs from the sample, steps of fragmentation or cleavage or digestion of proteins present in the sample, e.g. as a means of separating or extracting or removing GAGs from the protein to which they are attached, e.g. through the use of a protease such as proteinase K, purification of GAGs, e.g. using an anion-exchange resin, isolating cells from the sample, isolating cell components from the sample, extracting (e.g. isolating or purifying) proteins / peptides from the sample. Said processing steps thus also include steps carried out on a body fluid sample to prepare it for analysis, e.g. in the case of a urine sample, the removal of cells or other impurities may be done. A processing step may involve one or more of digestion, extraction, purification, boiling, filtration, lyophilization, fractionation, centrifugation, concentration, dilution, inactivation of interfering components, addition of reagents, derivatization, complexation and the like. Exemplary processing steps are described in the Examples. Although in certain methods of the invention steps of fragmentation or cleavage or digestion of proteins present in the sample (e.g. as a means of separating or extracting or removing GAGs from the protein to which they are attached, e.g. through the use of a protease such as proteinase K) and / or purification of GAGs (e.g. using an anion-exchange resin) may be done, as is evident from the discussion elsewhere herein in certain preferred methods a step of fragmentation / cleavage / digestion of proteins and / or a step of purification (e.g. using an anion-exchange resin) is not performed. In particular, in preferred methods where the chemical composition of the protein-free fraction of the GAG(s) is determined, a step of fragmentation / cleavage / digestion of proteins is not performed. In general, the GAG containing body fluid sample that has been obtained from a subject is subjected to at least one processing step prior to determining the chemical composition in accordance with the methods of the present invention. In particular, in methods wherein the levels of one or more of the specific sulfated or unsulfated forms of CS or HS disaccharides are determined, the GAGs are preferably subjected to a processing step to obtain the disaccharide units for analysis. In some such methods of the invention where the levels of certain individual disaccharide forms are measured, the GAGs, e.g. the full length GAG molecules, or polymerised polysaccharide chains of GAGs, or chains of repeating disaccharide units of GAGs, are subjected to a processing step, for example a step of fragmentation or cleavage or digestion, e.g. by chemical digestion or enzyme treatment. Appropriate methods of digestion or enzyme treatment would be known to a person skilled in the art, e.g. the use of one or more GAG lyase enzymes, e.g. one or more chondroitinase enzymes such as Chondroitinase ABC or Chondroitinase B, and / or the use of one or more heparinase enzymes such as Heparinase l-ll-lll, in order to obtain the disaccharide units which are then analysed. Other methods to determine compositions of GAGs which might be used are known in the art. However, examples are analytical techniques involving the use of antibodies to various GAG forms, e.g. techniques such as Western blot, ELISA or FACS, or methods involving agarose gel electrophoresis (e.g. fluorophore-assisted carbohydrate electrophoresis (FACE)) or polyacrylamide gel electrophoresis (PAGE). In some embodiments, the level of one or more GAG forms (e.g. specific sulfated or unsulfated forms of CS or HS disaccharides, which have for example been derived from the full length GAG molecule or a chain of repeating disaccharide units of a GAG molecule by fragmentation, cleavage or digestion) in association with (e.g. physical association with or in complex with or derivatized with or labelled with) a reagent (e.g. 2-aminoacridone) that is being used to detect the GAG form is determined. Thus, in some embodiments the level of a complex of a GAG form and the reagent used to detect the GAG form is determined. Reagents suitable for detecting particular GAG forms are discussed elsewhere herein, but include antibodies, or some kind of fluorophore (or other detectable label or dye) attached to (or used to derivatize) the GAG form in question, for example to make it detectable by a fluorimeter (or other detection device). Thus, purely by way of example, in some embodiments the level of a GAG form in association with (e.g. in complex with or derivatized with) an antibody or fluorophore or the like may be determined. In some embodiments the level of a GAG form in association with (e.g. in complex with or derivatized with) 2-aminoacridone may be determined. As certain preferred methods of the invention comprise the step of determining the chemical composition of the protein-free fraction of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally also of the GAG heparan sulfate (HS)) in a body fluid sample, advantageously there is no need for the GAG molecules to be separated or extracted from the proteins to which they are attached. Instead, in such embodiments, the protein-free fraction (only the protein-free fraction) of the GAGs in a body fluid sample can be analysed from the sample without any need for such processing to separate the GAGs from the protein, e.g. by digesting the protein. Thus, preferred methods do not comprise a processing step in which said samples are contacted with a proteolytic agent such as a protease. Other preferred methods do not comprise a step in which the GAGs are purified from the sample based on the negative charge of said GAGs (e.g. using an anion-exchange resin). Thus, in preferred methods of the present invention, said sample has been obtained from said subject and has been subjected to processing prior to determining said level and / or chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units; and (b) does not comprise prior to (a) at least one of: (i) contacting said sample with a proteolytic agent; and (ii) purifying said GAG(s) in said sample based on the negative charge of said GAG(s). A yet further aspect of the invention provides a method of discriminating between NASH with fibrosis and NASH without fibrosis in a subject having NASH, or in a subject suspected of having NASH, said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally also of the GAG HS) in a body fluid sample, wherein said determining the chemical composition of said GAG(s) comprises determining the level of one or more GAG property selected from the group consisting of (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject and has been subjected to processing prior to determining said chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units; and (b) does not comprise prior to (a) at least one of: (i) contacting said sample with a proteolytic agent; and (ii) purifying said GAG(s) in said sample based on the negative charge of said GAGs. Embodiments of other aspects of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG properties (or groups of GAG properties), preferred processing steps, preferred body fluids, etc.). In preferred methods, said fragmenting of (a) is conveniently performed by contacting said GAG(s) with one or more GAG lyase enzymes (e.g. as discussed elsewhere herein). For example, said fragmenting of (a) may be performed by contacting said GAG(s) with one or more chondroitinase enzymes (and optionally one or more heparinase enzymes). In art based methods, said contacting step of (b)(i) is conveniently performed by contacting said sample with one or more protease enzymes, e.g. proteinase K. Thus, in certain preferred methods of the invention such a step is not carried out. The proteolytic agent of (b)(i) may be a protease (e.g. a non-specific protease such as proteinase K). Thus, certain preferred methods of the invention do not comprise prior to (a) a step of contacting the sample with a protease (e.g. a non-specific protease e.g. proteinase K). In art based methods, said purifying step of (b)(ii) is conveniently performed by using an anion-exchange resin. Thus, in certain preferred methods of the invention such a step is not carried out. Thus, certain preferred methods of the invention do not comprise prior to (a) a step of purifying said GAG(s) in said sample using an anion-exchange resin. In preferred methods of the invention, the method does not comprise the contacting of (b)(i). In preferred methods of the invention, the method does not comprise the purifying of (b)(ii). In other preferred methods of the invention neither step (b)(i) nor (b)(ii) are carried out. Thus, in particularly preferred embodiments the method does not comprise the contacting of (b)(i) or the purifying of (b)(ii). As mentioned above, in some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention an altered level (increased or decreased as the case may be) of one or more of the GAG properties in accordance with the invention (4S CS (e.g. absolute concentration of 4S CS) and / or 4S6S CS (e.g. relative concentration of 4S6S CS)) in comparison to a control level is indicative of either NASH with liver fibrosis or NASH without liver fibrosis in said subject (indicative of whether said subject has either NASH with liver fibrosis or NASH without liver fibrosis). The skilled person would be readily able to establish appropriate control levels for use in accordance with the invention. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being different than (or altered (e.g. increased or decreased, preferably increased) in comparison to) a control level of the same one or more GAG property is indicative of NASH with liver fibrosis, wherein said control level is within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of subjects (or from a reference population) without (or not having) liver fibrosis; or (b) the level of one or more GAG property in accordance with the invention being different than (or altered (e.g. increased or decreased, preferably decreased) in comparison to) a control level of the same one or more GAG property is indicative of NASH without liver fibrosis, wherein said control level is comprised within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of subjects (or from a reference population) with (or having) liver fibrosis. In some embodiments, the group of subjects (or reference population) without (or not having) liver fibrosis may be a group of subjects (or a reference population) having NASH without fibrosis. In some embodiments, the group of subjects (or a reference population) with (or having) liver fibrosis may be a group of subjects (or a reference population) having NASH with fibrosis. The concept of “reference intervals” (which may also be referred to as “reference ranges”) is well known in the art, particularly in medicine and health related areas. A reference interval is the range (or interval) of values (e.g. levels) for a physiological measurement that has been established or determined for (or established or determined as being characteristic, or indicative of) a particular health state in a reference population (or reference group of subjects). In some cases, a reference interval is the range (or interval) of values (e.g. levels) for a physiological measurement that has been established or determined for (or established or determined as being characteristic of, or indicative of) a particular disease (or disease state or condition) state in a reference population having a given disease or disease state or condition. Reference intervals for a particular measurement or value (e.g. the level of one or more of the GAG properties in accordance with the invention) define an interval between which (or a range within which) a certain proportion (e.g. %) of measurements (or values) in a reference population fall into. The proportion may be for example 80% to 99%. For example the proportion may be 80%, 85%, 90% or 95%. Preferably, the proportion is 95%. Typically and preferably, the proportion is the central proportion of measurements (or values) in a reference population. By way of an example, when the proportion is 95% (preferably the central proportion is 95%), a reference interval for a particular measurement or value (e.g. the level of one or more of the GAG properties in accordance with the invention) is the interval between which (or range within which) 95% of the measurements (or values) for a reference population fall into. This means that 5% of measurements in such a reference population would be outside this reference interval. When the 95% proportion is the central proportion of the reference population (which is preferred), this means that 2.5% of measurements (or values) would be less than the lower limit of the reference interval and 2.5% of measurements (or values) would be higher than the upper limit of the reference interval. When an 80% proportion is the central proportion of the reference population, this means that 10% of measurements (or values) would be less than the lower limit of the reference interval and 10% of measurements (or values) would be higher than the upper limit of the reference interval. When an 85% proportion is the central proportion of the reference population, this means that 7.5% of measurements (or values) would be less than the lower limit of the reference interval and 7.5% of measurements (or values) would be higher than the upper limit of the reference interval. When a 90% proportion is the central proportion of the reference population, this means that 5% of measurements (or values) would be less than the lower limit of the reference interval and 5% of measurements (or values) would be higher than the upper limit of the reference interval. The upper limit of a reference interval is typically referred to as an upper reference limit (URL). The lower limit of a reference interval is typically referred to as a lower reference limit or (LRL). In some embodiments, a URL or a LRL may be used a cut-off or threshold level to provide an indication of NASH with liver fibrosis or NASH without liver fibrosis (as appropriate). In some embodiments, an altered level (increased level or decreased level as the case may be) of one or more GAG property in accordance with the invention in comparison to (or relative to) a URL or a LRL of a reference interval is indicative of NASH with liver fibrosis or NASH without liver fibrosis (as appropriate). The skilled person is familiar with reference intervals, and now that the present inventors have found that the levels of certain GAG properties are useful in discriminating between NASH with liver fibrosis and NASH without liver fibrosis in accordance with the present invention, the skilled person would readily be able to establish appropriate reference intervals (and associated URLs and LRLs) from (or of) appropriate reference populations for use in accordance with the present invention. As mentioned above, the concept of reference intervals is well-established in the art. For example, the Clinical and Laboratory Standards Institute publication “EP28-A3c Defining, Establishing, and Verifying Reference Intervals in the Clinical Laboratory; Approved Guideline - Third Edition" (2010, Vol. 28. No. 30; ISBN 1-56238-682-4) provides detailed discussion and guidance in this regard. A reference interval may be based on measurements (or values) of (or taken from) any appropriate number of subjects in a reference population (i.e. the reference population may be of any appropriate size). The skilled person in readily able to select an appropriately sized reference population. Guidance on appropriate reference population sizes is provided in the above mentioned Clinical and Laboratory Standards Institute publication. The reference population may have at least 5 subjects (e.g. at least 5, 6, 7, 8, 9 or 10 subjects). Preferably, the reference population has at least 10 subjects. In some embodiments, there may be at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 120, at least 200, at least 500 subjects or at least 1000 subjects in a reference population. In some embodiments, there may be up to 50, up to 100, up to 200 or up to 500 subjects in a reference population. In some embodiments, there may be up to 10 to 20, 10 to 50, 10 to 100, 10 to 200, 10 to 500 or 10 to 1000 subjects in a reference population. As is evident from the above, in some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention that provide an indication of NASH with liver fibrosis, a reference interval is a reference interval for a given GAG property in accordance with the invention that has been determined for the same sample type obtained from a reference population group of subjects not having liver fibrosis (e.g. having NASH without liver fibrosis). As is also evident from the above, in some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention that provide an indication of NASH without liver fibrosis, a reference interval is a reference interval for a given GAG property in accordance with the invention that has been determined for the same sample type obtained from a reference population group of subjects having liver fibrosis (e.g. having NASH with liver fibrosis). In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being higher than (or increased in comparison to) a control level of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level is within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of subjects (or from a reference population) without (or not having) liver fibrosis; or (b) the level of one or more GAG property in accordance with the invention being lower than (or decreased in comparison to) a control level of the same one or more GAG property is indicative of NASH without fibrosis, wherein said control level is comprised within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of subjects (or from a reference population) with (or having) liver fibrosis. In some embodiments, the group of subjects (or reference population) without (or not having) liver fibrosis may be a group of subjects (or a reference population) having NASH without fibrosis. In some embodiments, the group of subjects (or a reference population) with (or having) liver fibrosis may be a group of subjects (or a reference population) having NASH with fibrosis. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being higher than a control level (or increased in comparison to a control level) of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level is an upper reference limit of a reference interval that has been determined for the same sample type obtained from a group of subjects (or reference population) without (or not having) liver fibrosis; or (b) the level of one or more GAG property in accordance with the invention being lower than a control level (or decreased in comparison to a control level) of the same one or more GAG property is indicative of NASH without fibrosis, wherein said control level is an upper reference limit of a reference interval that has been determined for the same sample type obtained from a group of subjects (or reference population) without (or not having) liver fibrosis. In some embodiments, the group of subjects (or reference population) without (or not having) liver fibrosis may be a group of subjects (or a reference population) having NASH without fibrosis. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level one or more GAG property in accordance with the invention being lower than a control level (or decreased in comparison to a control level) of the same one or more GAG property is indicative of NASH with liver fibrosis, wherein said control level is an upper reference limit of a reference interval that has been determined for the same sample type obtained from a group of subjects (or reference population) with (or having) NASH with liver fibrosis; or (b) the level of one or more GAG property in accordance with the invention being lower than a control level (or decreased in comparison to a control level) of the same one or more GAG property is indicative of NASH without liver fibrosis, wherein said control level is a lower reference limit of a reference interval that has been determined for the same sample type obtained from a group of subjects (or reference population) with (or having) NASH with liver fibrosis. As described above, upper reference limits and lower reference limits may in some cases be used as, or in some cases may be alternatively viewed as, cut-off levels or threshold levels, with an indication of NASH with liver fibrosis or NASH without liver fibrosis being reached (or provided) based on the measured level of one or more GAG property in accordance with the invention relative to (or as compared with) such a cut-off limit. In some preferred embodiments, reference intervals for a particular measurement or value (e.g. the level of one or more of the GAG properties in accordance with the invention) define an interval between which (or a range within which) a 95% of measurements (or values) in a reference population fall into. In certain alternative embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, a reference (or control) range could be generated (or calculated or established) based on 95% confidence intervals (e.g. 95% confidence around a mean measurement in a group (or population) of NASH with fibrosis subjects or NASH without fibrosis subjects, as appropriate). Thus, in some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) a level of a GAG property in accordance with the present invention that is in (or that falls within) a reference range that has been generated based on 95% confidence intervals around a mean measurement (of the level of said GAG property) in the same sample type in a group (or population) of NASH with fibrosis subjects, is indicative of NASH with fibrosis; or (b) a level of a GAG property in accordance with the present invention that is in (or that falls within) a reference range that has been generated based on 95% confidence intervals around a mean measurement (of the level of said GAG property) in the same sample type in a group (or population) of NASH without fibrosis subjects, is indicative of NASH without fibrosis; or (c) a level of a GAG property in accordance with the present invention that is not in (or that falls outside) a reference range that has been generated based on 95% confidence intervals around a mean measurement (of the level of said GAG property) in the same sample type in a group (or population) of NASH without fibrosis subjects, is indicative of NASH with fibrosis. In certain alternative embodiments, other confidence intervals could be more appropriate, such as 80% or 90% or 99%. In certain embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, a reference range or reference interval for (or from) a given group or population of subjects (e.g. for a group or population of subjects having NASH with fibrosis, or for a group or population of subjects having NASH without fibrosis) could be generated (or determined or calculated or established) within which there is a 95% probability that a given GAG property in accordance with the invention has its mean (or standardized mean). Such an interval (or range) may be referred to as a high-density interval (HDI). Thus, in some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) a level of a GAG property in accordance with the present invention is indicative of NASH with fibrosis when said level is in (or when it falls within) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects having NASH with fibrosis, and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval; or (b) a level of a GAG property in accordance with the present invention is indicative of NASH without fibrosis when said level is in (or when it falls within) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects having NASH without fibrosis, and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval; or (c) a level of a GAG property in accordance with the present invention is indicative of NASH with fibrosis when said level is not in (or when it falls outside) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects having NASH without fibrosis, and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval. In certain alternative embodiments, other high-density intervals could be more appropriate, such as the probability is 80% or 90% or 99%. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being higher than a control level of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level has been determined in the same sample type obtained from one or more subjects (e.g. group or population of subjects) without (or not having) liver fibrosis; or (b) the level of one or more GAG property in accordance with the invention being lower than a control level of the same one or more GAG property is indicative of NASH without fibrosis, wherein said control level has been determined in the same sample type obtained from one or more subjects (e.g. group or population of subjects) with (or having) liver fibrosis. In some embodiments, the one or more subjects (e.g. group or population of subjects) without (or not having) liver fibrosis may be one or more subjects (e.g. group or population of subjects) having NASH without liver fibrosis. In some embodiments, the one or more subjects (e.g. group or population of subjects) with (or having) liver fibrosis may be one or more subjects (e.g. group or population of subjects) having NASH with liver fibrosis. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being higher than a control level of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more subjects (e.g. a population of subjects) having NASH without fibrosis and from one or more subjects (e.g. a population of subjects) having NASH with fibrosis, wherein the control level is a cutoff level that has been derived (or established or determined) to distinguish (or discriminate) between samples from NASH with fibrosis subjects as opposed to samples from NASH without fibrosis subjects; or (b) the level of one or more GAG property in accordance with the invention being lower than a control level of the same one or more GAG property is indicative of NASH without fibrosis, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more subjects (e.g. a population of subjects) having NASH without fibrosis and from one or more subjects (e.g. a population of subjects) having NASH with fibrosis, wherein the control level is a cut-off level that has been derived (or established or determined) to distinguish (or discriminate) between samples from NASH with fibrosis subjects as opposed to samples from NASH without fibrosis subjects. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being altered in comparison to (preferably being higher than) a control level of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more (e.g. a population of) reference subjects (e.g. reference subjects not having liver fibrosis, such as reference subjects having NASH without fibrosis), wherein the control level is a cut-off level (or threshold level) that has been derived (or established or determined) to provide an indication of NASH with fibrosis when the level of the one or more GAG property in accordance with the invention is altered (preferably higher) in comparison thereto; or (b) the level of one or more GAG property in accordance with the invention being altered in comparison to (preferably being lower than) a control level of the same one or more GAG property is indicative of NASH without fibrosis, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more (e.g. a population of) reference subjects (e.g. reference subjects having liver fibrosis, such as reference subjects having NASH with fibrosis), wherein the control level is a cut-off level that has been derived (or established or determined) to provide an indication of NASH without fibrosis when the level of the one or more GAG property in accordance with the invention is altered (preferably lower) in comparison thereto. In some embodiments of the invention, a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence in (or for) the indication of NASH with liver fibrosis or NASH without liver fibrosis (as the case may be) in accordance with the invention, for example an (or at least an) 80%, 85%, 90% or 95% level of confidence in (or for) the indication of NASH with liver fibrosis or NASH without liver fibrosis. Thus, in some embodiments of the invention, a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence in (or for) the indication of NASH with liver fibrosis or NASH without liver fibrosis (as the case may be) in accordance with the invention when an altered level (increased (or higher) or decreased (or lower) as the case may be) of one or more GAG properties in accordance with the invention is determined (or observed) in comparison to a control level (e.g. a cut-off or threshold level). For example, a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence (e.g. an (or at least an) 80%, 85%, 90% or 95% level of confidence) that an indication (e.g. diagnosis) of NASH with liver fibrosis or of NASH without liver fibrosis is accurate (or correct). In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being within a range (within a range of levels) that has been established as being indicative of the sample being from a subject having NASH with fibrosis as opposed to being from a subject having NASH without fibrosis, is indicative of NASH with fibrosis; or (b) the level one or more GAG property in accordance with the invention being within a range (within a range of levels) that has been established as being indicative of the sample being from a subject having NASH without fibrosis as opposed to being from a subject having NASH with fibrosis, is indicative of NASH without fibrosis. In some such embodiments, (i) said range of (a) is a range of levels that has been determined by determining the level of said one or more GAG property in the same sample type obtained from a population of subjects having NASH without fibrosis and from a population of subjects having NASH with fibrosis and establishing a range of levels that is indicative of samples being from subjects having NASH with fibrosis as opposed to being from subjects having NASH without fibrosis; and / or (ii) said range of (b) is a range of levels that has been determined by determining the level of said one or more GAG property in the same sample type obtained from a population of subjects having NASH without fibrosis and from a population of subjects having NASH with fibrosis and establishing a range of levels that is indicative of samples being from subjects having NASH without fibrosis as opposed to being from subjects having NASH with fibrosis. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being within a reference interval (or range) that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH with fibrosis, is indicative of NASH with fibrosis; or (b) the level of one or more GAG property in accordance with the invention being within a reference interval (or range) that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH without fibrosis, is indicative of NASH without fibrosis. In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH with fibrosis, is indicative of NASH with fibrosis; or (b) the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH without fibrosis, is indicative of NASH without fibrosis; In some embodiments of methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or other related methods) of the present invention, (a) the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject z|. 1 having NASH with fibrosis as opposed to from a subject having NASH without fibrosis, is indicative of NASH with fibrosis; or (b) the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH without fibrosis as opposed to from a subject having NASH with fibrosis, is indicative of NASH without fibrosis. In embodiments of the invention that refer to a population, or a population of subjects, or a group, or a group of subjects (e.g. a reference population or a reference group of subjects), or similar terms, the population or group (e.g. reference or control population) may have at least 5 subjects (e.g. at least 5, 6, 7, 8, 9 or 10 subjects). Preferably, the population or group has at least 10 subjects. In some embodiments, there may be at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 120, at least 200, at least 500 subjects or at least 1000 subjects in a population or group (e.g. reference or control population). In some embodiments, there may be up to 50, up to 100, up to 200 or up to 500 subjects in a population or group (e.g. reference or control population). In some embodiments, there may be up to 10 to 20, 10 to 50, 10 to 100, 10 to 200, 10 to 500 or 10 to 1000 subjects in a population or group (e.g. reference or control population). As mentioned above, the present invention also provides method of screening for non-alcoholic steatohepatitis (NASH) in a subject, said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (OS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS, and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Thus, in some embodiments, the present invention provides a method of screening for NASH irrespective of whether the NASH is NASH with fibrosis or NASH without fibrosis. In some embodiments, the NASH may be NASH with fibrosis. In some embodiments, the NASH may be NASH without fibrosis. Unless otherwise clear from the context, features described elsewhere herein in relation to other aspects of the invention (e.g. preferred body fluids, GAG properties, processing steps, subjects, etc.) may be applied, mutatis mutandis, to methods of screening for NASH in accordance with the present invention. For example, in some embodiments of methods of screening for NASH said method comprises determining the level of 4S CS. In some embodiments of methods of screening for NASH said method comprises determining the level of 4S6S CS. In some embodiments of methods of screening for NASH said method comprises determining the absolute concentration of 4S CS. In some embodiments of methods of screening for NASH said method comprises determining the relative concentration of 4S6S CS. In some embodiments, methods of screening for NASH of the present invention comprise determining the absolute concentration of 4S CS or the relative concentration of 4S6S CS. In some embodiments, methods of screening for NASH of the present invention comprise determining the absolute concentration of 4S CS and the relative concentration of 4S6S CS. In some embodiments of methods of screening for NASH, the body fluid sample is blood (e.g. plasma). In some embodiments of methods of screening form NASH, the body fluid is urine. In some embodiments of methods of screening for NASH, the body fluid is blood (e.g. plasma) and the method comprises determining the level of 4S CS (preferably the absolute concentration of 4S CS). In some embodiments of methods of screening for NASH, the body fluid is urine and the method comprises determining the level of 4S6S CS (preferably the relative concentration of 4S6S CS). In some embodiments, methods of screening for NASH comprise determining the chemical composition of CS in accordance with the invention (and optionally also the chemical composition of HS) in more than one (e.g. two) type of body fluid sample (e.g. in a blood (e.g. plasma) sample and in a urine sample). Put another way, in some embodiments of methods of screening for NASH, methods may comprise determining the chemical composition of CS in accordance with the invention (and optionally also the chemical composition of HS) in different (e.g. two different) body fluid sample types (e.g. in a blood (e.g. plasma) sample and in urine sample). Thus, in some embodiments of methods of screening for NASH, methods of the present invention may comprise determining the level of 4S CS (preferably the absolute concentration of 4S CS) in a blood (e.g. plasma) sample) and determining the level of 4S6S CS (preferably the relative concentration of 4S6S CS) in a urine sample. In some embodiments of methods of screening for NASH of the present invention, (i) said body fluid sample is blood (e.g. plasma) and the level of 4S CS is determined and said level of 4S CS is the absolute concentration of 4S CS; and / or (ii) said body fluid sample is urine and the level of 4S6S CS is determined and said level of 4S6S CS is the relative concentration of 4S6S CS. The present invention also provides a method of screening for non-alcoholic steatohepatitis (NASH) in a subject, said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally also of the GAG HS) in a body fluid sample, wherein said determining the chemical composition of said GAG(s) comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS, and (ii) 4S6S CS, wherein said sample has been obtained from said subject and has been subjected to processing prior to determining chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units; and (b) does not comprise prior to (a) at least one of: (i) contacting said sample with a proteolytic agent; and (ii) purifying said GAG(s) in said sample based on the negative charge of said GAG(s). Unless otherwise clear from the context, embodiments of other aspects of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG properties (or groups of GAG properties), preferred processing steps, preferred body fluids, etc.). In certain preferred embodiments of methods of screening for NASH, an altered level (e.g. an increased level or a decreased level) of one or more GAG property in accordance with the invention in said sample in comparison to a control level is indicative of NASH. In methods of screening for NASH in accordance with the present invention a control level may be the level of a GAG form (GAG property) in a control subject or control population (e.g. in a sample that has been obtained from a control subject or population). Appropriate control subjects or samples or populations for use in the methods of screening for NASH in accordance with the invention would be readily identified by a person skilled in the art, for example an appropriate control group is as described in the Examples. Such subjects might also be referred to as "control" subjects or as a reference population. Examples of appropriate populations of control subjects would include subjects having simple steatosis. Simple steatosis (or simple fatty liver) is a largely harmless build-up of fat in the liver. Other examples of appropriate populations of control subjects would include healthy subjects, for example, individuals who have no history of any form of liver disease and no other concurrent disease. Other control subjects would include individuals who are not suffering from, and preferably have no history of, liver disease (e.g. not suffering from, and preferably have no history of, liver disease). In some embodiments, control subjects for methods of screening for NASH in accordance with the present invention are not regular users of any medication. In some embodiments, control subjects for methods of screening for NASH in accordance with the present invention are healthy subjects. In some embodiments, control subjects for methods of screening for NASH in accordance with the present invention are subjects having simple steatosis. In some embodiments of methods of screening for NASH in accordance with the present invention, the control level is the level of said GAG in a sample(s) obtained from one or more subjects (e.g. a population or group of subjects) having simple steatosis or from one or more (e.g. a population or group of subjects) healthy subjects. Thus, in some embodiments of methods of screening for NASH in accordance with the present invention, the control level has been determined in the same sample type (e.g. blood (e.g. plasma) or urine) obtained from one or more subjects (e.g. a population or group of subjects) having simple steatosis or from one or more healthy subjects. In some embodiments of methods of screening for NASH in accordance with the present invention, the control level has been determined in the same sample type (e.g. blood (e.g. plasma) or urine) obtained from one or more subjects (e.g. a population or group of subjects) having simple steatosis. In some embodiments of methods of screening for NASH of the present invention, an altered level (e.g. increased or decreased level) of 4S OS (preferably the absolute concentration of 4S CS) in said body fluid sample (preferably a blood (e.g. plasma) sample) in comparison to a control level is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, an increased level of 4S CS (preferably the absolute concentration of 4S CS) in said body fluid sample (preferably a blood (e.g. plasma) sample) in comparison to a control level is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, a decreased level of 4S CS (preferably the absolute concentration of 4S CS) in said body fluid sample (preferably a blood (e.g. plasma) sample) in comparison to a control level is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, an altered level (e.g. increased or decreased level) of 4S6S CS (preferably the relative concentration of 4S6S CS) in said body fluid sample (preferably a urine sample) in comparison to a control level is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, an increased level of 4S6S CS (preferably the relative concentration of 4S6S CS) in said body fluid sample (preferably a urine sample) in comparison to a control level is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being different than (or altered (e.g. increased or decreased)) in comparison to) a control level of the same one or more GAG property is indicative of NASH, wherein said control level is within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of control subjects (or from a reference or control population). Suitable control levels and subjects and populations of use in methods of screening for NASH of the present invention are discussed elsewhere herein. Certain principles in relation to reference intervals are discussed elsewhere herein. The upper limit of a reference interval is typically referred to as an upper reference limit (URL). The lower limit of a reference interval is typically referred to as a lower reference limit or (LRL). In some embodiments, a URL or a LRL may be used a cut-off or threshold level to provide an indication of NASH. In some embodiments of methods of screening for NASH of the present invention, an altered level (increased level or decreased level as the case may be) of one or more GAG property in accordance with the invention in comparison to (or relative to) a URL or a LRL of a reference interval is indicative of NASH. Now that the present inventors have found that the levels of certain GAG properties are useful in screening for NASH in accordance with the present invention, the skilled person would readily be able to establish appropriate reference intervals (and associated URLs and LRLs) from (or of) appropriate reference populations for use in accordance with the present invention. As mentioned above, the concept of reference intervals is well-established in the art. For example, the Clinical and Laboratory Standards Institute publication “EP28-A3c Defining, Establishing, and Verifying Reference Intervals in the Clinical Laboratory; Approved Guideline - Third Edition” (2010, Vol. 28. No. 30; ISBN 1-56238-682-4) provides detailed discussion and guidance in this regard. In some embodiments of methods of screening for NASH of the present invention, a reference interval, may be a reference interval for a given GAG property in accordance with the invention that has been determined for the same sample type obtained from a control population (or group) of subjects. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being higher than (or increased in comparison to) a control level of the same one or more GAG property is indicative of NASH, wherein said control level is within (or comprised within) a reference interval that has been determined for the same sample type obtained from a group of control subjects (or from a reference or control population). Suitable control levels and subjects and populations of use in methods of screening for NASH of the present invention are discussed elsewhere herein. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being higher than a control level (or increased in comparison to a control level) of the same one or more GAG property is indicative of NASH with fibrosis, wherein said control level is an upper reference limit of a reference interval that has been determined for the same sample type obtained from a group of control subjects (or from a reference or control population). Suitable control levels and subjects and populations of use in methods of screening for NASH of the present invention are discussed elsewhere herein. In some embodiments of methods of screening for NASH of the present invention, the level one or more GAG property in accordance with the invention being lower than a control level (or decreased in comparison to a control level) of the same one or more GAG property is indicative of NASH, wherein said control level is an upper reference limit of a reference interval that has been determined for the same sample type obtained from a group of subjects (or a reference population) with (or having) NASH. As described above, upper reference limits and lower reference limits may in some cases be used as, or in some cases may be alternatively viewed as, cut-off levels or threshold levels, with an indication of NASH in said subject being reached (or provided) based on the measured level of one or more GAG property in accordance with the invention relative to (or as compared with) such a cut-off limit. In some preferred embodiments, reference intervals for a particular measurement or value (e.g. the level of one or more of the GAG properties in accordance with the invention) define an interval between which (or a range within which) 95% of measurements (or values) in a reference population fall into. In certain alternative embodiments, other confidence intervals could be more appropriate, such as 80% or 90% or 99%. In certain alternative embodiments of methods of screening for NASH of the present invention, a reference (or control) range could be generated (or calculated or established) based on 95% confidence intervals (e.g. 95% confidence around a mean measurement in a group (or population) of NASH subjects). Thus, in some embodiments of methods of screening for NASH of the present invention, a level of a GAG property in accordance with the present invention that is in (or that falls within) a reference range that has been generated based on 95% confidence intervals around a mean measurement (of the level of said GAG property) in the same sample type in a group (or population) of subjects having NASH, is indicative of NASH. In certain embodiments of methods of screening for NASH of the present invention, a reference range or reference interval for (or from) a group or population of subjects having NASH could be generated (or determined or calculated or established) within which there is a 95% probability that a given GAG property in accordance with the invention has its mean (or standardized mean). Such an interval (or range) may be referred to as a high-density interval (HDI). Thus, in some embodiments of methods of screening for NASH of the present invention, a level of a GAG property in accordance with the present invention is indicative of NASH when said level is in (or when it falls within) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects having NASH, and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval. In alternative embodiments of methods of screening for NASH of the present invention, a level of a GAG property in accordance with the present invention that is not in (or that falls outside) a reference range that has been generated based on 95% confidence intervals around a mean measurement (of the level of said GAG property) in the same sample type in a group (or population) of subjects without NASH (for example, healthy subjects or subjects with simple steatosis), is indicative of NASH. In certain embodiments of methods of screening for NASH of the present invention, a reference range or reference interval for (or from) a group or population of subjects without NASH could be generated (or determined or calculated or established) within which there is a 95% probability that a given GAG property in accordance with the invention has its mean (or standardized mean). Such an interval (or range) may be referred to as a high-density interval (HDI). In certain alternative embodiments, other high-density intervals could be more appropriate, such as the probability is 80% or 90% or 99%. Thus, in some embodiments of methods of screening for NASH of the present invention, a level of a GAG property in accordance with the present invention is indicative of NASH when said level is in (or when it falls within) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects having NASH, and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval. In alternative embodiments of methods of screening for NASH of the present invention, a level of a GAG property in accordance with the present invention is indicative of NASH when said level is not in (or when it falls outside) a reference range or reference interval, wherein said reference range or reference interval has been generated based on the level of said GAG property in the same sample type from a group (or population) of subjects without NASH (for example, healthy subjects or subjects with simple steatosis), and wherein said reference range or reference interval has been generated on the basis that (or such that) there is a 95% probability that said GAG property in accordance with the invention has its mean (or standardized mean) level within said reference range or reference interval. Reference ranges could, for example, be derived from data in Table 1 herein. For the avoidance of doubt, such reference ranges based on Table 1 are purely exemplary, and the methods of the present invention are not limited thereto. Certain embodiments of methods of the present invention (e.g. methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis, and methods of screening for NASH) refer to “95% confidence intervals”, “95% probability” or “95% of measurements”. The present invention also provides certain alternative embodiments that are identical to embodiments that refer to “95% confidence intervals”, “95% probability” or “95% of measurements”, with the exception that these alternative embodiments refer to “80% confidence intervals”, “80% probability” or “80% of measurements”. The present invention also provides certain alternative embodiments that are identical to embodiments that refer to “95% confidence intervals”, “95% probability” or “95% of measurements”, with the exception that these alternative embodiments refer to “90% confidence intervals”, “90% probability” or “90% of measurements”. The present invention also provides certain alternative embodiments that are identical to embodiments that refer to “95% confidence intervals”, “95% probability” or “95% of measurements”, with the exception that these alternative embodiments refer to “99% confidence intervals”, “99% probability” or “99% of measurements”. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being altered in comparison to (preferably higher than) a control level of the same one or more GAG property is indicative of NASH, wherein said control level has been determined in the same sample type obtained from one or more control subjects (e.g. group or population of control subjects). Suitable control levels and subjects and populations of use in methods of screening for NASH of the present invention are discussed elsewhere herein. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being altered in comparison to (preferably higher than) a control level of the same one or more GAG property is indicative of NASH, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more control subjects (e.g. group or population of control subjects) and from one or more subjects having NASH (e.g. a group or population of subjects having NASH), wherein the control level is a cut-off level that has been derived (or established or determined) to distinguish (or discriminate) between samples from NASH subjects as opposed to samples from control subjects. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being altered in comparison to (preferably being higher than) a control level of the same one or more GAG property is indicative of NASH, wherein said control level is a level that has been determined by determining the level of said one or more GAG property in the same sample type obtained from one or more control subjects (e.g. group or population of control subjects), wherein the control level is a cut-off level (or threshold level) that has been derived (or established or determined) to provide an indication of NASH when the level of the one or more GAG property in accordance with the invention is altered (preferably higher) in comparison thereto. In some embodiments of methods of screening for NASH of the present invention a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence in (or for) the indication of NASH in accordance with the invention, for example an (or at least an) 80%, 85%, 90% or 95% level of confidence in (or for) the indication of NASH. Thus, in some embodiments of the invention, a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence in (or for) the indication of NASH in accordance with the invention when an altered level (increased (or higher) or decreased (or lower) as the case may be) of one or more GAG properties in accordance with the invention is determined (or observed) in comparison to a control level (e.g. a cut-off or threshold level). For example, a control level, e.g. cut-off level (or threshold level), can be derived (or determined or established) to provide a level of confidence (e.g. an (or at least an) 80%, 85%, 90% or 95% level of confidence) that an indication (e.g. diagnosis) of NASH is accurate (or correct). In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being within a range (within a range of levels) that has been established as being indicative of the sample being from a subject having NASH (e.g. as opposed to being from control subject), is indicative of NASH. In some such embodiments, said range is a range of levels that has been determined by determining the level of said one or more GAG property in the same sample type obtained from a population of subjects having NASH and from a population of control subjects and establishing a range of levels that is indicative of samples being from subjects having NASH (e.g. as opposed to being from a control subject). In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being within a reference interval (or range) that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH, is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH, is indicative of NASH. In some embodiments of methods of screening for NASH of the present invention, the level of one or more GAG property in accordance with the invention being a level that has been established as being (or has been determined to be) indicative of the sample being from a subject having NASH as opposed to from a control subject, is indicative of NASH. As is evident from discussion above, in some embodiments of methods of screening for NASH of the present invention based on the observed alterations in the levels of one or more GAG property in accordance with the invention, if desired, scoring methods, scoring systems, markers or formulas can be designed which use such levels of GAG properties in order to arrive at an indication, e.g. in the form of a value or score, which can then be used for screening (e.g. diagnosis, etc.). Thus, discussion of scores, etc., elsewhere herein may also be applied, mutatis mutandis, to methods of screening for NASH of the present invention. In another aspect, the present invention provides a method of screening for (e.g. diagnosing) NASH in a subject, said method comprising: determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S 6S in a body fluid sample, wherein said sample has been obtained from said subject. Embodiments of methods of screening for NASH may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). As discussed above, in some aspects, the present invention provides a method of screening for NASH in a subject. Methods of screening may be used for diagnosing (or for confirming a diagnosis of) NASH. Thus, the present invention also provides a method of diagnosing NASH in a subject. The present invention also provides a method of monitoring for the occurrence of NASH in a subject at risk. The present invention also provides a method for monitoring the progression of NASH in a subject. The present invention also provides a method of screening for NASH in a population at risk of having or developing NASH (e.g. individuals presenting risk factors (e.g. being overweight or obese or having Type 2 diabetes) or individuals presenting symptoms). Thus, the method of screening for NASH in accordance with the present invention can be used, for example, for diagnosing NASH, for monitoring for the occurrence of NASH in a subject at risk, for monitoring the progression of NASH, for screening for NASH in a population at risk of having or developing NASH (such as individuals presenting risk factors (e.g. being overweight or obesity or Type 2 diabetes) or individuals presenting symptoms). Thus, in one aspect, the present invention provides a method for diagnosing NASH in a subject. The present invention provides a method of diagnosing non-alcoholic steatohepatitis in a subject, said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS, and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Unless otherwise clear from the context, embodiments of methods of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG properties (or groups of GAG properties), preferred processing steps, preferred body fluids, preferred control levels, etc.). In some embodiments, a positive diagnosis (e.g. the presence of NASH) is made if the level of one or more of the GAG properties in accordance with the invention in the sample is altered (increased or decreased, preferably increased) in comparison to a control level. In some embodiments, a scoring system or method (e.g. a GAG score) may be used to make a diagnosis. In some embodiments when a scoring system (e.g. a GAG score) is used, a positive diagnosis (i.e. the presence of NASH) may be made if the score is altered (or significantly altered) in comparison to a control score or cut-off (or threshold) level. An altered (increased or decreased as the case may be) level (or composition or score) of one or more of the GAG forms (GAG properties) in accordance with the invention (e.g. in methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or related methods) or methods of screening for NASH) includes any measurable alteration or change of the GAG form (biomarker) (or score) in question when the GAG property (or GAG form) in question is compared with a control level (e.g. reference limit or cut-off or threshold level). An altered level (or score) includes an increased or decreased level (or score). Preferably, the level (or score) is significantly altered, compared to the level (or score or cut-off level) found in an appropriate control (e.g. control sample or subject or population). More preferably, the significantly altered levels or compositions or scores are statistically significant, preferably with a p-value of <0.05. In some embodiments, an alteration in level (or score) of 2%, Ss 3%, >5%, >10%, >25%, >50%, >75%, >100%, >200%, >300%, >400%, >500%, >600%, >700%, >800%, >900%, >1000%, >2000%, >5000%, or >10,000% compared to the level (or score) found in an appropriate control sample or subject or population (i.e. when compared to a control level) may be indicative of the presence of NASH in accordance with the invention, or may be indicative of NASH with fibrosis or NASH without fibrosis (as appropriate) in accordance with the invention. The "increase" in the level or "increased" level of one or more of the GAG forms (GAG properties) or scores as described herein includes any measurable increase or elevation of the GAG form (biomarker) (or score) in question when the GAG form (or score) in question is compared with a control level (or control score or cut-off level or reference limit). Preferably, the level (or score) is significantly increased, compared to the level (or score or cut-off level) found in an appropriate control (e.g. control sample or subject or population). More preferably, the significantly increased levels (or scores) are statistically significant, preferably with a p-value of <0.05. In some embodiments, an increase in level (or score) of >2%, >3%, >5%, >10%, >25%, >50%, >75%, >100%, >200%, >300%, >400%, >500%, >600%, >700%, >800%, >900%, >1000%, >2000%, >5000%, or >10,000% compared to the level (or score) found in an appropriate control sample or subject or population (i.e. when compared to a control level or control score or cut-off level) may be indicative of the presence of NASH in accordance with the invention, or may be indicative of NASH with fibrosis or NASH without fibrosis (as appropriate) in accordance with the invention. The "decrease" in the level or "decreased" level of one or more of the GAG forms (GAG properties) or scores as described herein includes any measurable decrease or reduction of the GAG form (biomarker) (or score) in question when the GAG form in question is compared with a control level (or control score or cut-off level). Preferably, the level (or score) is significantly decreased, compared to the level (or score or cut-off level) found in an appropriate control (e.g. control sample or subject or population). More preferably, the significantly decreased levels (or scores) are statistically significant, preferably with a p-value of <0.05. In some embodiments, a decrease in level (or score) of >2%, >3%, >5%, >10%, >25%, >50%, >75%, >80%, >90%, >95% or >99% compared to the level (or score) found in an appropriate control sample or subject or population (i.e. when compared to a control level or control score or cut-off level) is indicative of the presence of NASH in accordance with the invention, or may be indicative of the NASH with fibrosis or NASH without fibrosis (as appropriate) in accordance with the invention. “Control levels” are discussed elsewhere herein. A “control level” may be the level of a relevant GAG property in a control subject or population (e.g. in a sample(s) that has been obtained from a control subject or population). A “control level” may be a level of a relevant GAG property where the level is derived from (or established based on or calculated from) the level in a control subject or population (e.g. in a sample(s) that has been obtained from a control subject or population). Such a population may be referred to as a reference population. Appropriate control subjects (or populations) or samples for use in methods of the invention would be readily identified by a person skilled in the art. Suitable control levels are described elsewhere herein. The control level may correspond to the level of the equivalent (corresponding) GAG form in appropriate control subjects or samples or populations, e.g. may correspond to a cut-off or threshold level or range found in a control or reference population. Control levels may also be referred to as "reference" levels. The control level may be a discrete figure or a range. Although the control level for comparison could be derived by testing an appropriate control subject or set of control subjects (or a control population), the methods of the invention would not necessarily involve carrying out active tests on control subjects as part of the methods of the present invention but would generally involve a comparison with a control level or control range which had been determined previously from control subjects (or a control population) and was known to the person carrying out the methods of the invention. A “control chemical composition” is the chemical composition in a control subject or population (e.g. in a sample that has been obtained from a control subject or population). The discussion above in relation to a “control level” (e.g. appropriate control subjects, control samples, control populations, etc.) may be applied, mutatis mutandis, to “a control chemical composition”. As described elsewhere herein, screening for NASH, and discriminating between NASH with fibrosis and NASH without fibrosis, in accordance with the present invention may involve using a score (or a GAG score), or expressing the chemical composition determined in accordance with the invention using a score (or GAG score). In some such embodiments, an altered score (e.g. increased or decreased as the case may be) in comparison to a “control score” (or cut-off level or threshold level) is indicative of NASH, or is indicative of NASH with fibrosis or of NASH without fibrosis (as appropriate) in said subject. The discussion elsewhere herein in relation to a “control level” (e.g. appropriate control subjects, control samples, control populations, etc.) may be applied, mutatis mutandis, to a “control score”. As described elsewhere herein, in some preferred methods of the present invention, the method comprises determining the chemical composition in accordance with the invention of the protein-free fraction of the GAG chondroitin sulfate (CS) (and optionally also of the GAG heparan sulfate (HS)) (or a score based thereon). In some such embodiments, an altered chemical composition of the protein-free fraction (or a score based thereon) in comparison to a chemical composition of the protein-free fraction of said GAG(s) in a control (e.g. control sample or control score or cut-off level) is indicative of is indicative of NASH, or is indicative of NASH with fibrosis or of NASH without fibrosis (as appropriate) in said subject. The discussion elsewhere herein in relation to a “control level” or “control chemical composition” or “control score” (e.g. appropriate control subjects, control samples, control populations, etc.) may be applied, mutatis mutandis, to embodiments of the invention that comprise determining chemical composition in accordance with the invention of the protein-free fraction of said GAG(s) (or a score based thereon). Methods of the present invention can also be used to monitor NASH progression (e.g. to monitor progression from NASH without liver fibrosis to NASH with fibrosis). Such monitoring can take place before, during or after treatment of NASH by therapy, e.g. pharmaceutical therapy. Thus, in another aspect the present invention provides a method for monitoring the progression of NASH in a subject having NASH. In such methods for monitoring the progression of NASH in a subject, the level of one or more of the GAG properties in accordance with the present invention (or a score derived or based thereon) is indicative of the progression of NASH. Thus, in another aspect, the present invention provides a method of monitoring the progression of NASH in a subject having NASH, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Preferred aspects of other methods of the invention described herein (e.g. preferred GAG properties, body fluids, processing steps, etc.) may be applied, mutatis mutandis, to this aspect of the invention. In preferred embodiments of methods of monitoring NASH progression of the invention, an altered level (increased level or decreased level as the case may be) of one or more of the GAG properties in accordance with the invention (4S CS (e.g. absolute concentration of 4S CS) or4S6S CS (e.g. relative concentration of 4S6S CS)), in comparison to a control level is indicative of NASH progression in said subject. In some embodiments of methods of monitoring the progression of NASH in accordance with the present invention, progression of NASH is monitored and an altered (or different) level, preferably an increased (or higher) level, of one or more GAG property in accordance with the invention over time as compared to a control level of the same one or more GAG property is indicative of progression of NASH, wherein said control level has been determined in the same sample type obtained from one or more subjects not having NASH (or not having any liver disease) or wherein said control level has been determined in the same sample type obtained from an earlier sample taken from the subject whose NASH progression is being monitored. In some embodiments of methods of monitoring progression of NASH in accordance with the invention, the control level may be a level of one or more GAG property in accordance with the invention determined in the same sample type obtained from one or more subjects (e.g. a population (e.g. a reference population) of subjects) not having NASH (or not having any liver disease). In some embodiments, of methods of monitoring progression of NASH in accordance with the invention, the control level may be a level of one or more GAG property in accordance with the invention determined in the same sample type obtained from one or more healthy (or normal) subjects (e.g. a population (e.g. a reference population) of healthy (or normal) subjects), or the control level may be a level of one or more GAG property in accordance with the invention determined in the same sample type obtained from one or more subjects having simple steatosis. In some preferred embodiments of methods of monitoring progression in accordance with the present invention, progression of NASH is monitored and an increasing level of one or more GAG property in accordance with the invention over time as compared to a control level of the same one or more GAG property is indicative of progression of NASH, wherein said control level has been determined in the same sample type obtained from an earlier sample taken from the subject whose NASH progression is being monitored. Thus, in some embodiments of methods of monitoring progression of NASH in accordance with the invention, the control level is a level that has been determined in the same sample type obtained from an earlier (e.g. a first) sample taken from the subject whose NASH progression is being monitored. A level from an earlier (e.g. first) sample from the subject whose NASH progression is being monitored may be considered a “baseline” level in the subject. This type of control level (i.e. a control level from an individual subject) is particularly useful for embodiments of the invention where serial or periodic measurements of GAG form(s) in individuals are taken looking for changes in the levels of the GAG property(ies). In this regard, an appropriate control level can be the individual's own baseline, stable, nil, previous or dry value (as appropriate) as opposed to a control or cut-off level found in the general control population. Thus, a control level for methods of monitoring progression of NASH in accordance with the invention may correspond to the level of the marker (GAG property) in question in the same individual subject, or a sample from said subject, measured at an earlier time point (e.g. a "baseline" level in that subject). In some embodiments, the level of one or more of the GAG properties in accordance with the invention is indicative of NASH progression, preferably with high (or higher) or increased (or increasing) levels (e.g. as measured over time e.g. by taking serial or periodical measurements) being indicative of NASH progression (NASH worsening). In some embodiments of methods of monitoring progression in accordance with the invention, the body fluid sample is a blood (e.g. plasma) sample (ora processed blood (e.g. plasma) sample). In some embodiments of methods of monitoring progression in accordance with the invention, the body fluid sample is a urine sample (or a processed urine sample). In some embodiments of methods of monitoring the progression of NASH in accordance with the present invention, the level of 4S OS (e.g. absolute concentration of 4S CS) is determined (or measured) in a blood (e.g. plasma) sample and / or the level of 4S6S OS (e.g. relative concentration of 4S6S CS) is determined (or measured) in a urine sample. In another aspect, the present invention provides a method of monitoring the progression of NASH in a subject having NASH, said method comprising: determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Features of other methods of the invention described herein (e.g. preferred GAG properties, body fluids, processing steps, etc.) may be applied, mutatis mutandis, to this aspect of the invention. Preferred GAG properties (or groups of GAG properties) for use in connection with methods of monitoring progression in accordance with the invention are discussed elsewhere herein in connection with other aspects of the invention. Sample processing step and / or detection (or measurement methods, etc.) for use in connection with methods of monitoring progression in accordance with the invention are discussed elsewhere herein in connection with other aspects of the invention. Indeed, unless otherwise clear from the context, features described elsewhere herein in connection with other methods of the invention may also be applied to methods of monitoring progression of the invention. In a further aspect, the present invention provides a method of providing information that is useful for monitoring progression of NASH in a subject having NASH, said method comprising: determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAGs comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS, (ii) 4S6S CS, wherein said sample has been obtained from said subject. Said information is of course typically the determined level of one or more of said GAG properties. Features of other methods of the invention described herein (e.g. preferred GAG properties, body fluids, processing steps, etc.) may be applied, mutatis mutandis, to this aspect of the invention. Methods of the present invention can be used in the active monitoring of patients which have not been subjected to therapy, e.g. to monitor the progression of NASH in untreated patients. Again, serial measurements can allow an assessment of whether or not, or the extent to which, the NASH is worsening, thus, for example, allowing a more reasoned decision to be made as to whether intervention (e.g. therapeutic intervention) is necessary or advisable. In another aspect, the present invention provides a method for determining the clinical severity of NASH in a subject. In such methods the level of one or more of the GAG forms in accordance with the invention in the sample (or a score derived therefrom or based thereon) shows an association with the severity of the NASH. Thus, the level of one or more of the GAG forms in accordance with the present invention can be indicative of the severity of the NASH. In some embodiments, the more altered (e.g. the more increased) the level of one or more of the GAG forms in accordance with the invention in comparison to a control level (e.g. a control level as described elsewhere herein), the greater the likelihood of a more severe form of NASH. In some embodiments, the methods of the invention can thus be used in the selection of patients for therapy. Serial (periodical) measuring of the level of one or more of the GAG forms in accordance with the invention (biomarkers) (or a score derived therefrom or computed based thereon) may also be used to monitor the severity of NASH looking for either increasing or decreasing levels over time. Observation of altered levels (increase or decrease as the case may be) may also be used to guide and monitor therapy (or other clinical decisions), for example in the situation of "watchful waiting" before treatment, e.g. before initiation of pharmaceutical therapy, or during or after treatment to evaluate the effect of treatment and look for signs of therapy failure. The present invention also provides a method of patient selection or treatment selection as it provides a means of distinguishing (or discriminating between) patients with NASH from patients with no NASH. The present invention also provides a method of patient selection or treatment selection as it provides a means of distinguishing (or discriminating between) patients having NASH with liver fibrosis from patients having NASH without fibrosis. Such methods may guide appropriate treatment. In some embodiments, the invention provides a method of monitoring (e.g. continuously monitoring or performing active surveillance of) a subject having NASH. Such monitoring may guide which treatment to use or whether no treatment should be given. In some embodiments, patients with less severe NASH may be put under watchful waiting or active surveillance and may not be given treatment (e.g. pharmaceutical therapy). In some embodiments, patients with severe or more severe NASH may be given treatment. The present invention also provides a method of determining (or monitoring) the efficacy of a therapeutic regime being used to treat NASH, in other words following or monitoring a response to treatment. In such methods, an alteration (increase or decrease as the case may be) in the level (or scores) of one or more of the GAG properties in accordance with the present invention indicates the efficacy of the therapeutic regime being used. For example, in some embodiments, if the level of one or more of the GAG forms in accordance with the present invention (or a score derived therefrom (or based thereon)) for which an increased level (or score) is indicative of NASH is reduced during (or after) therapy, this is indicative of an effective therapeutic regime. The methods of the present invention can be carried out on any appropriate body fluid sample. Typically the sample has been obtained from (removed from) a subject (e.g. as described elsewhere herein), preferably a human subject. In other aspects, the method further comprises a step of obtaining a sample from the subject. Reference herein to "body fluid" includes reference to all fluids derived from the body of a subject. Exemplary fluids include blood (including all blood derived components, for example plasma, serum, etc.), urine, saliva, tears, bronchial secretions or mucus. Preferably, the body fluid is a circulatory fluid (especially blood or a blood component) or urine. Especially preferred body fluids are blood (e.g. plasma) or urine. Particularly preferred body fluids are plasma or urine. In some preferred embodiments the sample is a blood sample (e.g. a plasma sample or serum sample). In some preferred embodiments the sample is a plasma sample. In some embodiments a plasma sample is a platelet-poor plasma sample. In some embodiments the sample is a serum sample. In some preferred embodiments the sample is a urine sample. In some embodiments, the sample is not a urine sample. In some embodiments, the sample is not a blood sample (e.g. not a plasma sample). The body fluid or sample may be in the form of a liquid biopsy. The term "sample" also encompasses any material derived by processing a body fluid sample (e.g. derived by processing a blood (e.g. plasma) or urine sample). Thus the term “sample” includes a processed sample (e.g. a processed blood (e.g. plasma) or urine sample). Processing of biological samples to obtain a test sample may involve one or more of: digestion, boiling, filtration, distillation, centrifugation, lyophilization, fractionation, extraction, concentration, dilution, purification, inactivation of interfering components, addition of reagents, derivatization, complexation and the like, e.g. as described elsewhere herein. Suitable processing steps can be selected depending on the features of the method being performed. Any suitable method for isolating body fluid samples (e.g. blood or urine samples) may be employed. Any sample that could be directly or indirectly affected by NASH may be used. Samples (e.g. original or unprocessed samples) typically comprise a protein-free fraction of GAGs and a protein-bound fraction of GAGs (as discussed elsewhere herein). In preferred methods of the present invention, the chemical composition of the protein-free fraction of the GAG chondroitin sulfate (CS) (and optionally also of the GAG heparan sulfate (HS)) is determined. Thus, in certain preferred embodiments a body fluid sample has been processed (or is processed) such that only (or essentially only) the protein-free fraction of said GAG(s) (typically disaccharide units derived therefrom) is subsequently analysed (i.e. processed such that the chemical composition of the protein-free fraction of said GAG(s) (typically disaccharide units derived therefrom) is subsequently determined). In other methods of the invention, the sample has been processed (or is processed) such that the entire (protein-free plus protein-bound) fraction or pool of said GAG(s) is subsequently analysed (i.e. processed such that the chemical composition of the entire (protein-free plus protein-bound) fraction or pool of said GAG(s) (typically disaccharide units derived therefrom) is subsequently determined). The term "sample" also encompasses any material derived by processing (e.g. as described above) a biological sample. Derived materials include disaccharide units (or a population of disaccharide units) derived by processing GAGs (e.g. as described elsewhere herein). In some embodiments, methods of the invention may include a step of processing a sample. In some embodiments, methods of the invention may thus be performed on such processed samples or materials derived from such processed samples. In some embodiments, methods of the invention may thus be performed on samples that have been processed. Processing steps include, but are not limited to, isolating cells from the sample, isolating cell components from the sample, and extracting (e.g. isolating or purifying) proteins / peptides (although as certain preferred methods of the invention involve the determination of the protein-free GAG fraction, the extraction of proteins or the removal of the protein component from the proteoglycans (protein-bound GAGs) present in the sample, e.g. by digesting or otherwise removing the protein component, is preferably not carried out in some embodiments). A processing step may involve one or more of filtration, distillation, centrifugation, extraction, concentration, dilution, purification, inactivation of interfering components, addition of reagents, derivatization, amplification, adapter ligation, and the like. Samples can be used immediately or can be stored for later use (e.g. at -80°C). The methods of the invention as described herein can be carried out on any type of subject which is capable of suffering from NASH. The methods are generally carried out on mammals, for example humans, primates (e.g. monkeys), laboratory mammals (e.g. mice, rats, rabbits, guinea pigs), livestock mammals (e.g. horses, cattle, sheep, pigs) or domestic pets (e.g. cats, dogs). Preferably the subject is a human. In some embodiments of the present invention, the subject (e.g. a human) is a subject having NASH or a subject suspected of having NASH. In some embodiments of the present invention, the subject (e.g. a human) is a subject having NASH. In some embodiments, the subject is a subject that has been diagnosed with NASH. In other embodiments, the subject (e.g. a human) is a subject suspected of having NASH. In methods of monitoring progression in accordance with the invention the subject is a subject having NASH. In some embodiments of the present invention, the subject (e.g. a human) is a subject having NASH without fibrosis. In some embodiments, the subject is a subject that has been diagnosed with NASH without fibrosis. In other embodiments, the subject (e.g. a human) is a subject suspected of having NASH without fibrosis. In some embodiments of the present invention, the subject (e.g. a human) is a subject having NASH with fibrosis. In some embodiments, the subject is a subject that has been diagnosed with NASH with fibrosis. In other embodiments, the subject (e.g. a human) is a subject suspected of having NASH with fibrosis. In some embodiments of methods of screening for NASH in accordance with the present invention, the subject may be a subject at risk of (e.g. at particular risk of) developing NASH. Such “at risk” subjects would be readily identified by a person skilled in the art, but would include for example subjects that are overweight, obese, have Type 2 diabetes, that have hypothyroidism, that have high blood pressure, that have high cholesterol levels and / or that are >50 years of age. In some embodiments, methods can be carried out on “healthy” patients (subjects) or at least patients (subjects) which are not manifesting any clinical signs or symptoms of NASH. Subjects may be male subjects orfemale subjects. In some embodiments, subjects are female. In some embodiments, subjects are male. NASH is an abbreviation for non-alcoholic steatohepatitis. NASH may be “NASH without liver fibrosis”. “NASH without liver fibrosis” is NASH that is not accompanied by liver fibrosis. “NASH without liver fibrosis” is characterised by the absence of liver fibrosis. “NASH without liver fibrosis” may alternatively be referred to as “non-fibrotic NASH”. Liver fibrosis is scarring of, or around, the liver. Liver fibrosis may also include scarring of blood vessels associated with (e.g. nearby) the liver. NASH without liver fibrosis is a less severe form of NASH as compared to “NASH with liver fibrosis”. As NASH progresses, persistent inflammation can cause liver fibrosis. “NASH with liver fibrosis” is NASH that is accompanied by liver fibrosis. “NASH with liver fibrosis” is thus characterized by (the presence of) liver fibrosis. “NASH with liver fibrosis” may alternatively be referred to as “fibrotic NASH”. “NASH without liver fibrosis” may also be simply referred to as “NASH without fibrosis”. “NASH with liver fibrosis” may also be simply referred to as “NASH with fibrosis”. In some aspects, methods of the invention are provided which further comprise a step of treating NASH (e.g. NASH with fibrosis or NASH without fibrosis) by therapy (e.g. pharmaceutical therapy), or surgery, or other clinical management program. In some embodiments, if the result of a method of the invention is indicative of NASH (e.g. NASH with fibrosis or NASH without fibrosis), or is indicative of NASH progression (worsening), in the subject (e.g. if a positive indication or diagnosis of NASH (e.g. NASH with fibrosis or NASH without fibrosis) is made), then an additional step of treating the NASH by therapy or surgery or providing a clinical management program appropriate for the NASH can be performed. For example, if the result of a method of the invention is indicative of NASH (e.g. NASH with fibrosis or NASH without fibrosis), or is indicative of NASH progression (worsening) in the subject, the subject may be instructed (or advised) to make certain lifestyle changes (e.g. to exercise or more regularly exercise) and / or the subject may be instructed (or advised) to lose weight (e.g. if a subject is overweight). In some embodiments, if the level of one or more GAG properties in accordance with the invention in a sample (or a score based on these levels) is altered by a particular degree in comparison to a control level or score (or cut-off level), then a further step of administering a therapeutically effective amount of a pharmaceutical agent to the patient is performed and / or surgery is performed and / or the provision of a clinical management program appropriate for the NASH is made. Preferred degrees of alteration are discussed elsewhere herein. In some aspects, methods of the invention are provided which further comprise a step of carrying out (or performing) an additional diagnostic or screening procedure for NASH (e.g. NASH with liver fibrosis or NASH without fibrosis, as appropriate), e.g. by liver biopsy. A yet further aspect provides a kit for screening for NASH (e.g. for diagnosing or for determining severity or progression of NASH), which comprises one or more agents suitable for determining the level of one or more of the GAG properties (GAG forms) described herein in accordance with the invention, in a sample. A yet further aspect provides a kit for screening for NASH (e.g. for diagnosing or for determining severity or progression of NASH), which comprises one or more reagents (or components) for processing a body fluid sample (such as urine or blood (e.g. plasma)) that comprises chondroitin sulfate (OS), and optionally further comprises heparan sulfate (HS). In preferred aspects said kits are for use in the methods of the invention as described herein. Preferably said kits comprise instructions for use of the kit components, for example in screening (e.g. diagnosis) in accordance with the invention. A yet further aspect provides a kit for discriminating between NASH with liver fibrosis and NASH without liver fibrosis, which comprises one or more agents suitable for determining the level of one or more of the GAG properties (GAG forms) described herein in accordance with the invention, in a sample. A yet further aspect provides a kit for discriminating between NASH with liver fibrosis and NASH without liver fibrosis, which comprises one or more reagents (or components) for processing a body fluid sample (such as urine or blood (e.g. plasma)) that comprises chondroitin sulfate (OS) and optionally further comprises heparan sulfate (HS). In preferred aspects said kits are for use in the methods of the invention as described herein. Preferably said kits comprise instructions for use of the kit components, for example in discriminating between NASH with liver fibrosis and NASH without liver fibrosis in accordance with the invention. A yet further aspect provides a kit for screening for NASH, which comprises one or more agents suitable for determining the level of one or more of the GAG properties (GAG forms) described herein in accordance with the invention, in a sample. A yet further aspect provides a kit for screening for NASH, which comprises one or more reagents (or components) for processing a body fluid sample (such as urine or blood (e.g. plasma)) that comprises chondroitin sulfate (CS) and optionally further comprises heparan sulfate (HS). In preferred aspects said kits are for use in the methods of the invention as described herein. Preferably said kits comprise instructions for use of the kit components, for example in screening for NASH in accordance with the invention. In another aspect, the present invention provides a method of determining (or detecting) the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from a subject having, or suspected of having, NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis). In another aspect, the present invention provides a method of determining (or detecting) the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample, wherein said method comprises determining (or detecting) the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample wherein said sample has been obtained from a subject having, or suspected of having, NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis). In one aspect, the present invention provides a method of detecting (or determining) the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS), said method comprising: (a) obtaining a body fluid sample from a human patient having, or suspected of having, NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis); and (b) detecting (or determining) the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in said sample, wherein said detecting (or determining) the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS. In one aspect, the present invention provides a method of determining (or detecting) the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in a body fluid sample, said method comprising: (a) obtaining a body fluid sample from a human patient having, or suspected of having, NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis); and (b) detecting (or determining) the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS in said sample. A yet further aspect of the invention provides a method of detecting (or determining) the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally also of the GAG HS) in a body fluid sample, wherein said detecting (or determining) the chemical composition of said GAG(s) comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject having NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis), or suspected of having NASH (e.g. NASH with liver fibrosis and NASH without liver fibrosis), and has been subjected to processing prior to determining said chemical composition, wherein said processing (a) comprises fragmenting said GAG(s) into disaccharide units; and (b) does not comprise prior to (a) at least one of: (i) contacting said sample with a proteolytic agent; and (ii) purifying said GAG(s) in said sample based on the negative charge of said GAG(s). The features and discussion herein in relation to the methods of discriminating between NASH with liver fibrosis and NASH without liver fibrosis and methods of screening for NASH (e.g. methods of diagnosing, etc.) and other methods described herein, for example in relation to preferred GAG forms or combinations thereof for measurement, can be applied, mutatis mutandis, to methods of detecting (or determining) of the present invention. In a further aspect, the present invention provides a method of providing information that is useful in (or for) screening for NASH (e.g. diagnosing NASH or confirming a diagnosis of NASH) or useful in (or for) discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or diagnosing NASH with liver fibrosis or NASH without liver fibrosis, differentially diagnosing NASH with liver fibrosis or NASH without liver fibrosis, or confirming a diagnosis of NASH with liver fibrosis or NASH without liver fibrosis) in a subject having NASH or in a subject suspected of having NASH, said method comprising: determining chemical composition the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample, wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Said information is of course typically the determined (or measured) level of one or more of said GAG properties. Embodiments of methods of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). In a further aspect, the present invention provides a method of providing information that is useful in (or for) screening for NASH (e.g. diagnosing NASH or confirming a diagnosis of NASH) or useful in (or for) discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or diagnosing NASH with liver fibrosis or NASH without liver fibrosis, differentially diagnosing NASH with liver fibrosis or NASH without liver fibrosis, or confirming a diagnosis of NASH with liver fibrosis or NASH without liver fibrosis) in a subject having NASH or in a subject suspected of having NASH, said method comprising determining the level of one or more GAG property selected from the group consisting of: (i) 4S CS and (ii) 4S6S CS, wherein said sample has been obtained from said subject. Said information is of course typically the determined (or measured) level of one or more of said GAG properties. Embodiments of methods of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, etc.). A yet further aspect of the invention provides a method of screening for NASH (e.g. diagnosing NASH or confirming a diagnosis of NASH) in a subject, said method comprising analysing disaccharide units that have been derived the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally further comprising analysing disaccharide units that have been derived from the glycosaminoglycan heparan sulfate (HS)) in a body fluid sample, wherein said method comprises determining (or measuring) the level of one or more GAG property in accordance with the invention. Embodiments of methods of screening of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluid, preferred control levels, preferred subjects, etc.). A yet further aspect of the invention provides a method of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or diagnosing NASH with liver fibrosis or NASH without liver fibrosis, differentially diagnosing NASH with liver fibrosis or NASH without liver fibrosis, or confirming a diagnosis of NASH with liver fibrosis or NASH without liver fibrosis) in a subject, said method comprising analysing disaccharide units that have been derived from the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally further comprising analysing disaccharide units that have been derived from the glycosaminoglycan heparan sulfate (HS)) in a body fluid sample, wherein said method comprises determining (or measuring) the level of one or more GAG property in accordance with the invention. Embodiments of methods of discriminating of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluid, preferred control levels, preferred subjects, etc.). A yet further aspect of the invention provides a method of screening for NASH (e.g. diagnosing NASH or confirming a diagnosis of NASH) in a subject, said method comprising analysing a population of disaccharide units consisting essentially of disaccharide units that have been derived from the non-proteoglycan fraction (or protein-free fraction) of the glycosaminoglycan (GAG) chondroitin sulfate (OS) (and optionally further comprising analysing a population of disaccharide units consisting essentially of disaccharide units that have been derived from the non-proteoglycan fraction (or protein-free fraction) of the glycosaminoglycan (GAG) heparan sulfate (HS)) in a body fluid sample, wherein said method comprises determining the level of one or more GAG property in accordance with the invention. Embodiments of methods of screening of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, preferred subjects, etc.). A yet further aspect of the invention provides a method of discriminating between NASH with liver fibrosis and NASH without liver fibrosis (or diagnosing NASH with liver fibrosis or NASH without liver fibrosis, differentially diagnosing NASH with liver fibrosis or NASH without liver fibrosis, or confirming a diagnosis of NASH with liver fibrosis or NASH without liver fibrosis) in a subject, said method comprising analysing a population of disaccharide units consisting essentially of disaccharide units that have been derived from the non-proteoglycan fraction (or protein-free fraction) of the glycosaminoglycan (GAG) chondroitin sulfate (CS) (and optionally further comprising analysing a population of disaccharide units consisting essentially of disaccharide units that have been derived from the non-proteoglycan fraction (or protein-free fraction) of the glycosaminoglycan (GAG) heparan sulfate (HS)) in a body fluid sample, wherein said method comprises determining the level of one or more GAG property in accordance with the invention. Embodiments of methods of screening of the invention described elsewhere herein may be applied, mutatis mutandis, to this aspect of the invention (e.g. preferred GAG forms (or groups of GAG forms), preferred processing steps, preferred body fluids, preferred control levels, preferred subjects, etc.). Where the terms “comprise”, “comprises”, “comprising”, “has” or “having”, or other equivalent terms are used herein, then in some more specific embodiments these terms include the term “consists of” or “consists essentially of”, or other equivalent terms. The invention will be further described with reference to the following non-limiting Example with reference to the following drawing: Figure 1: Boxplots of plasma 4S CS absolute concentration (left) and urine 4S6S CS relative concentration (right) in patients with NASH without fibrosis (NASH (without fibrosis)), NASH with fibrosis (NASH (fibrosis)), and simple steatosis. Point shape indicate patient sex. Key: F - female; M - male. EXAMPLE Introduction We investigated glycosaminoglycans (GAGs) profiles - or GAGomes - comprising chondroitin sulfate (CS) disaccharides and heparan sulfate (HS) disaccharides as biomarkers for screening for non-alcoholic steatohepatitis (NASH) in a subject having or suspected of having non-alcoholic steatohepatitis as well as discriminating between nonalcoholic steatohepatitis with liver fibrosis and non-alcoholic steatohepatitis without liver fibrosis in a subject having non-alcoholic steatohepatitis or in a subject suspected of having non-alcoholic steatohepatitis. In this study, the GAGome of the protein-free fraction of GAGs (or free GAGome) of plasma or urine samples was studied. Patients and Methods and Results Plasma and urine was collected from 28 patients. The median age was 62.5 years old, 67.9% were females, and the median body-mass-index (BMI) was 29. There were 12 patients with simple steatosis and 16 with NASH (10 without liver fibrosis and 6 with liver fibrosis). Patient characteristics are summarized in Table 2. We measured the plasma and urine GAGome using a standardized UHPLC-MS / MS method (D. Tamburro, S. Bratulic, S. A. Shameh, N. K. Soni, A. Bacconi, F. Maccari, F. Galeotti, K. Mattsson, N. Volpi, J. Nielsen, F. Gatto, Journal of Chromatography B, 1177 (2021) 122761) in a single blinded laboratory. Briefly, GAGome extraction from each sample was performed following Elypta MIRAM® Free Glycosaminoglycan Kit instructions for use. All reagents and consumables used were contained in the kit. The method for GAGome extraction included an enzymatic digestion assay using Chondroitinase ABC and Heparinase l-ll-lll to depolymerize GAGs in the sample into disaccharides. Note that compared to other methods described in the art (see Volpi et al., Nature Protocols, 9, 541-558 (2014)) where use of a non-specific protease is recommended for biological fluid analysis, the present method omitted the addition of a protease and thus the analysis was limited to the protein-free fraction of GAGs. Note that as compared to Volpi et al., the method used in the present study omitted the step of purifying the GAGs using an anion-exchange resin. GAG disaccharides were subsequently labeled using 2-aminoacridone. The processed samples were then injected into an ultra-high-performance liquid chromatography (UHPLC) coupled with electrospray ionization triple-quadrupole mass spectrometry system (ESI-MS / MS, Waters® 6 Acquity l-class Plus Xevo TQ-S micro) for disaccharide separation and detection. The peaks of GAG disaccharides (listed below) were acquired at pre-specified retention times across six transitions using multiple reaction monitoring (MRM) analysis implemented in the mass spectrometry software (Waters® 9 TargetLynx). We used the mass spectrometry software (Waters® TargetLynx) for peak integration, construction of calibration curves, and quantification. The measured GAG profiles (GAGomes) consisted of absolute concentrations for 17 GAG disaccharides, corresponding to 8 different sulfation patterns of chondroitin sulfate (OS) and heparan sulfate (HS), and hyaluronic acid (HA) disaccharide. Specifically, we quantified 8 OS disaccharides (OS CS, 2S OS, 6S CS, 4S CS, 2S6S OS, 2S4S OS, 4S6S OS, Tris OS) and 8 HS disaccharides (OS HS, 2S HS, 6S HS, NS HS, NS6S HS, NS2S HS, 2S6S HS, Tris HS). The GAGome was expanded to include an additional 22 dependent features: the total CS and total HS concentration as the sum of the corresponding disaccharide concentrations, the CS and HS charge, two ratios (4S CS / OS CS and 6S CS / OS CS), and the relative concentration (or mass fraction, in %) of each of the 16 CS and HS disaccharide by normalizing its absolute concentration by the total CS and HS concentration, respectively. For each sample, the plasma or urine GAGome, therefore, consisted of 39 GAG features. We fitted a Bayesian linear model to compare each GAG feature (response variable, standardized by mean-centering and standard-deviation-scaling) in patients with NASH with fibrosis (“fibrosis” group) versus patients with NASH with or without fibrosis (“NASH” group) versus patients with simple steatosis (control group) adjusted by the patient. We also analysed each GAG feature in the NASH without fibrosis group. We used the Region of Practical Equivalence (ROPE) framework to determine if a GAG feature was significantly altered in the NASH group - and specifically in the NASH with fibrosis group - versus the simple steatosis group (“control”) (Kruschke, J. Exp Psychol Gen, 142, 573-603 (2013)). In short, a GAG feature was deemed significantly altered in the “fibrosis” group versus the control group if less than 5% of the posterior samples fell inside the pre-specified ROPE interval (% in ROPE <5%), where the interval boundaries were centered on 0 + / - 20% of the GAG feature standardized mean. Intuitively, this means that if, for example, a GAG feature in the “fibrosis” group is found to be within 20% of the mean measurement in the control group in >5% of the posterior probability distribution (i.e., the % in ROPE), then that GAG feature has practical equivalent measurements in both the “fibrosis” and control group so it cannot be considered significantly altered in NASH with fibrosis. Conversely, if the % in ROPE is <5%, then the GAG feature has no practical equivalent measurements in the “fibrosis” vs control group, and it can be considered significantly altered in NASH with fibrosis. In addition to the ROPE, a High-Density Interval (HDI) was determined for each GAG feature as the interval where there is 95% probability that a GAG feature has its standardized mean for a given group. For example, an HDI ranging -0.12 to 1.91 for the GAG feature “4S CS plasma concentration [ug / ml]” indicate that that group has a 95% probability to correlate with a change in 4S CS plasma concentration between -0.12 standard deviations below the mean in the control group and +1.91 standard deviations above the mean in the control group. Bayesian estimation and ROPE analysis was carried out using the brms (2.14.4) package (Burkner, Journal of Statistical Software, 80, 1-28 (2017); Burkner, The R journal, 10, 395-411 (2018)), in R (4.0.3). In Table 1, the results are presented for each GAG feature detected in either the plasma or urine GAGome in this study. Two features were significantly altered in the “fibrosis” group (% in ROPE <5%): the plasma 4S CS absolute concentration and the urine 4S6S CS relative concentration (Figure 1). Both GAG features were significantly increased compared to the control group. Figure 1 also shows that both GAG features were increased in the NASH with fibrosis group compared to the NASH without fibrosis group. These findings not only indicate that these GAG features can be useful to screen for NASH in subjects suspected of NASH but also to specifically differentiate patients with NASH with fibrosis. Table 1. ROPE analysis for all detectable GAG features in the plasma and urine GAGome of patients with NASH with fibrosis versus the control group. ; GAG feature ; % in ROPE i HDI 95% lower limit : HDI 95% upper limit ; 4S CS plasma concentration [ug / ml] 3,5% -0,12 1,91 j 4S6S CS urine [%] | 4,6% -0,26 | 1,88 ; Total CS plasma ! 6,5% i -0,40 ; 1,79 i NS HS urine [%] 7,5% -0,45 1,73 ; NS HS urine concentration [ug / ml] ! 9,6% | -0,53 H,63 ; 6S CS urine concentration [ug / ml] 9,9% -1,54 0,56 i 4S CS urine [%] 10,2% -0,52 U,54 i 4S CS plasma [%] 10,7% -0,68 h,60 i 6S CS plasma [%] ! 11,2% -1,53 0,61 ; Total HS urine i 11,2% | -0,62 h,46 i OS CS plasma [%] 11,3% -1,60 0,68 i OS CS plasma concentration [ug / ml] 11,3% | -0,69 h,52 i OS HS urine concentration [ug / ml] | 11,6% -0,68 1,47 i OS HS plasma concentration [ug / ml] i 13,2% | -0,94 1,39 i Total HS plasma 13,7% -0,88 M.40 i 2s6S CS urine concentration [ug / ml] i 14,1% | -1,31 0,75 i OS CS urine [%] i 14,2% -1,29 | 0,84 i 6S CS urine [%] 14,6% | -1,32 0,83 i 4S CS urine concentration [ug / ml] 14,6% -1,28 0,89 i OS HS urine [%] ! 14,7% -1,06 1,16 i 6S CS plasma concentration [ug / ml] i 15,0% -1,29 0,95 ; Total CS urine 15,2% -1,13 p,06 i OS CS urine concentration [ug / ml] 15,2% -1,08 1,17 i Total HA urine 15,5% -1,11 1,00 i 2S6S CS urine [%] i 15,7% | -1,08 1,07 4S6S CS urine concentration [ug / ml] 16,6% -0,99 1,08 Table 2: Study population characteristics. wm ?WW SWsfe s&x^ssss- SSWIW ¢ 5 x X. .s? 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Claims

1. A method of discriminating between non-alcoholic steatohepatitis withliver fibrosis and non-alcoholic steatohepatitis without liver fibrosis in a subject having non-alcoholic steatohepatitis or in a subject suspected of having nonalcoholic steatohepatitis, said method comprising:determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample,wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of:(i) 4S CS; and(ii) 4S6S CS,wherein said sample has been obtained from said subject.

2. The method of claim 1, wherein(a) the level of said one or more GAG property being higher than a control level of the same one or more GAG property is indicative of nonalcoholic steatohepatitis with liver fibrosis, wherein said control level has been determined in the same sample type obtained from one or more subjects without liver fibrosis; or(b) the level of said one or more GAG property being lower than a control level of the same one or more GAG property is indicative of nonalcoholic steatohepatitis without liver fibrosis, wherein said control level has been determined in the same sample type obtained from one or more subjects with liver fibrosis.

3. The method of claim 2, wherein in (a) said one or more subjects withoutliver fibrosis are one or more subjects having non-alcoholic steatohepatitis without liver fibrosis, and / or wherein in (b) said one or more subjects with liver fibrosis are one or more subjects having non-alcoholic steatohepatitis with liver fibrosis.

4. The method of any one of claims 1 to 3, wherein said body fluid sampleis blood (e.g. plasma) or urine.

5. The method of any one of claims 1 to 4, wherein said body fluid sampleis blood (e.g. plasma) and wherein the level of 4S CS is determined.

6. The method of any one of claims 1 to 5, wherein said level of 4S CS isthe absolute concentration of 4S CS.

7. The method of any one of claims 1 to 6, wherein said body fluid sampleis urine and wherein the level of 4S6S CS is determined.

8. The method of any one of claims 1 to 7, wherein said level of 4S6S CSis the relative concentration of 4S6S CS.

9. The method of any one of claims 1 to 8, wherein(i) said body fluid sample is blood (e.g. plasma) and the level of 4S CS is determined and the level of 4S CS is the absolute concentration of 4S CS; and / or(ii) said body fluid sample is urine and the level of 4S6S CS is determined and the level of 4S6S CS is the relative concentration of 4S6S CS.

10. The method of any one of claims 1 to 9, wherein said method furthercomprises determining the chemical composition of the glycosaminoglycan (GAG) heparan sulfate (HS) in said body fluid sample.

11. The method of any one of claims 1 to 10, wherein said determining thechemical composition is determining the chemical composition of the protein-free fraction of the glycosaminoglycan(s).

12. The method of any one of claims 1 to 11, wherein said sample has beenobtained from said subject and has been subjected to at least one processing step prior to determining said chemical composition.

13. The method of any one of claims 1 to 12, wherein the GAG(s) is, or hasbeen, subjected to a processing step to obtain disaccharide units for analysis.

14. The method of claim 12 or claim 13, wherein said at least oneprocessing step does not comprise contacting said sample with a proteolytic agent.

15. The method of any one of claims 1 to 14, wherein said sample has beenobtained from said subject and has been subjected to processing prior to determining said chemical composition,wherein said processing(a) comprises fragmenting said GAG(s) into disaccharide units, preferably said fragmenting of (a) is performed by contacting said GAG(s) with one or more GAG lyase enzymes; and(b) does not comprise prior to (a) at least one of:(i) contacting said sample with a proteolytic agent; and(ii) purifying said GAG(s) in said sample based on the negative charge of said GAG(s) (e.g. using an anion exchange resin), preferably, said method does not comprise the contacting of (b)(i) or the purifying of (b)(ii).

16. The method of any one of claims 1 to 15, wherein said level of saidGAG property is determined by HPLC and mass spectrometry, preferably said HPLC is ultra-HPLC and / or said mass spectrometry is triple-quadrupole mass spectrometry.

17. The method of any one of claims 1 to 16, wherein said method is amethod of discriminating between non-alcoholic steatohepatitis with liver fibrosis and non-alcoholic steatohepatitis without liver fibrosis in a subject having non-alcoholic steatohepatitis.

18. A method of screening for non-alcoholic steatohepatitis in a subject,said method comprising determining the chemical composition of the glycosaminoglycan (GAG) chondroitin sulfate (CS) in a body fluid sample,wherein said determining the chemical composition of said GAG comprises determining the level of one or more GAG property selected from the group consisting of:(i) 4S OS, and(ii) 4S6S OS,wherein said sample has been obtained from said subject.

19. The method of claim 18, wherein an altered level of one or both of saidGAG properties in said sample in comparison to a control level is indicative of non-alcoholic steatohepatitis.

20. The method of claim 19, wherein said control level has been determinedin the same sample type obtained from one or more subjects having simple steatosis or from one or more healthy subjects.

21. The method of any one of claims 18 to 20, wherein said method hasone more features as defined in any one of claims 4 to 16.

22. The method of any one of claims 18 to 21, wherein(i) said body fluid sample is blood (e.g. plasma) and the level of 4SCS is determined and said level of 4S CS is the absolute concentrationof 4S CS; and / or(ii) said body fluid sample is urine and the level of 4S6S CS is determined and said level of 4S6S CS is the relative concentration of 4S6S CS.

23. The method of any one of claims 18 to 22, wherein said method is usedfor diagnosing non-alcoholic steatohepatitis.

24. The method of any one of claims 1 to 23, wherein said subject is ahuman subject.

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