Compositions and methods using a combination of oleuropein and quercetin for use in cartilage degeneration.

Abstract

The present invention relates to the use of a composition comprising an effective amount of oleuropein and / or its metabolites in combination with quercetin and / or derivatives for maintaining joint health or preventing or treating joint disorders in an individual. In particular, the present invention relates to a composition comprising an effective amount of oleuropein and / or its metabolites in combination with quercetin and / or derivatives for use in preventing or treating cartilage degeneration in an individual.

Description

Technical Field

[0001] The present invention relates to joint health, and in particular to the use of a composition comprising oleuropein and / or its metabolites and quercetin and / or its metabolites for the prevention or treatment of joint disorders or the maintenance of joint health.

Background Art

[0002] Osteoarthritis (OA) is a disease with a high morbidity rate that has a significant socioeconomic impact. Osteoarthritis is a degenerative disease of the articular cartilage of joints and is the most common form of arthritis, affecting 10% of the adult population. OA is the main cause of physical disability and medical costs for the elderly worldwide. Progressive degeneration and reduction of articular cartilage belong to the main pathological features, and are accompanied by other changes in joint structures such as synovial hyperplasia, subchondral bone sclerosis and thickness, osteophyte formation around the joint, ligament relaxation, and muscle atrophy (all of which contribute to the clinical symptoms of OA). These symptoms include severe pain, stiffness, reduced joint movement, and impairment. Since articular cartilage depends only on its resident cells, chondrocytes, for the maintenance of the extracellular matrix, impairment of chondrocyte function and survival leads to joint cartilage dysfunction.

[0003] Recent ex vivo studies have reported mitochondrial dysfunction in human osteoarthritis (OA) chondrocytes. Analysis of mitochondrial electron transport chain activity in these cells revealed decreased activity of complexes I, II, and III, as well as low ATP production, compared to normal chondrocytes. This mitochondrial dysfunction can affect several pathways involved in cartilage degeneration, including oxidative stress, impaired chondrocyte biosynthesis and proliferation responses, increased cytokine-induced chondrocyte inflammation and matrix catabolism, cartilage matrix calcification, and increased chondrocyte apoptosis (Blanco et al. "The role of mitochondria in osteoarthritis" Nat. Rev. Rheumatol. 7, 161-169 (2011)). Mitochondria are the primary source of aerobic energy production in mammalian cells and maintain a large Ca2+ gradient across the inner membrane, providing signaling potentials for this molecule. Furthermore, mitochondrial Ca2+ may play a role in regulating ATP production within mitochondria and contributing to the orchestration of cellular metabolic homeostasis. (Glancy, B. and RSBalaban (2012). "Role of mitochondrial Ca2+ in the regulation of cellular energetics." Biochemistry 51(14):2959-2973).

[0004] Although the number of individuals suffering from osteoarthritis (OA) is increasing, there is still no cure, and current drug therapies are still only symptomatic, focusing on alleviating symptoms. For example, pain and inflammation are treated with analgesics (such as acetaminophen) and nonsteroidal anti-inflammatory drugs (NSAIDs). Furthermore, the use of these drugs is often accompanied by side effects such as gastrointestinal or cardiovascular risks. Since current treatments for OA do not prevent or cure OA, chondrocyte apoptosis is an effective target for modulating cartilage degeneration.

[0005] [Overview of the prefecture] The inventors have surprisingly demonstrated that the combination of oleuropein (or oleuropein aglycone) and quercetin synergistically activates mitochondrial function at the cellular level through an increase in mitochondrial calcium.

[0006] Therefore, an object of the present invention relates to providing compositions for use in improving joint health. In particular, an object of the present invention is to provide compositions that improve joint health by preventing or treating cartilage degeneration and that solve the aforementioned problems of the prior art relating to side effects such as gastrointestinal and / or cardiovascular risks.

[0007] Accordingly, one aspect of the present invention relates to a composition comprising an effective amount of oleuropein and / or its metabolites and quercetin and / or its derivatives for use in the prevention or treatment of cartilage degeneration in an individual.

[0008] Another aspect of the present invention relates to a method for producing compositions for use according to the present invention.

[0009] In the final aspect, the present invention relates to a kit comprising, in one or more containers, an effective amount of a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives.

[0010] Further features and advantages are described herein and will become apparent from the following drawings and embodiments for carrying out the invention. [Brief explanation of the drawing]

[0011] [Figure 1]This graph shows that oleuropein (Ole) and quercetin (Q) synergistically activate mitochondria during HeLa cell stimulation by increasing mitochondrial Ca2+. The inset shows the effects of oleuropein (3 μM, black), quercetin (3 μM, gray), and the combination of 3 μM oleuropein + 3 μM quercetin (Ole / Q) on the increase in integrated mitochondrial calcium induced by 100 μM histamine. Using the data from the inset, the expected theoretical effect (sum of oleuropein effect and quercetin effect, Ole / Q) and the actually measured effect were determined in this figure, and the synergistic effect was represented. Results are expressed as the mean + / - SEM of n=6-9 experiments. * indicates a statistically significant difference between the measured and theoretical values ​​in mitochondrial calcium with P<0.05 (Student's t-test). [Figure 2] This graph shows that in stimulated HeLa cells, oleuropein aglycone (Oea) synergistically acts with quercetin (Q) to activate mitochondria through an increase in mitochondrial Ca2+. The inset shows the effects of oleuropein aglycone (3 μM, black), quercetin (3 μM, gray), and the combination of 3 μM Oea + 3 μM Q (Oea / Q) on the increase in integrated mitochondrial calcium induced by 100 μM histamine. Using the data from the inset, the expected theoretical effect (sum of Oea effect and quercetin effect) and the actually measured effect for each combination (Oea + quercetin, Oea / Q) were determined in this figure, and the synergistic effect was represented. The results are expressed as the mean + / - SEM of n=6-9 experiments. * indicates a statistically significant difference between the measured and theoretical values ​​in mitochondrial calcium with P<0.05 (Student's t-test). [Figure 3]This graph shows that in stimulated HeLa cells, oleuropein (Ole) does not synergistically interact with oleuropein aglycone (Oea) and does not activate mitochondria by increasing mitochondrial Ca2+. The bar graph shows the effects of oleuropein (3 μM, black), oleuropein aglycone (3 μM, gray), and the combination of 3 μM Ole + 3 μM Oea (Ole / Oea) on the increase in integrated mitochondrial calcium induced by 100 μM histamine. Results are presented as the mean + / - SEM of n=6 to 9 experiments. * indicates a statistically significant difference between measured and theoretical values ​​of mitochondrial calcium with P<0.05 (one-way ANOVA test). [Figure 4] This graph shows that oleuropein (Ole) and oleuropein aglycone (Oea) promote the same level of synergistic effect when combined with quercetin (Q). The synergistic effect was calculated as shown in Figures 1 and 2. The results are expressed as the mean + / - SEM of n=6 experiments. NS (Not Significant) indicates that there is no statistically significant difference between the two groups (combinations) with P<0.05 (Student's t-test). [Figure 5] The "in vitro" model of osteoarthritis chondrocytes (SW1353 cells treated with the pro-inflammatory cytokine interleukin-1β, IL-1β) demonstrates dysfunctional chondrocytes. Therefore, Figure 5A is a graph showing a significant decrease in collagen-IIα1 content in this model compared to control chondrocytes. In addition, Figures 5B and 5C are two graphs showing increased expression of metalloproteinases MMP3 (Figure 5B) and MMP13 (Figure 5C) in IL-1β-treated cells. Results are expressed as the mean + / - SEM of experiments with n=5 (Figure 5A), n=3 (Figure 5B), or n=4 (Figure 5C) for each condition. * indicates a statistically significant difference between the control and SW1353 cells treated with IL-1β for 24 or 48 hours, as shown (P<0.05, one-way ANOVA test). [Figure 6]This study demonstrates that an "in vitro" model of osteoarthritis chondrocytes (SW1353 cells treated with IL-1β) shows increased cell death in SW1353 chondrocytes. Therefore, annexin-V positive cells significantly increased 5 days after IL-1β treatment, as shown. Results are presented as mean + / - SEM values ​​for n=11 experiments per condition. * indicates a statistically significant difference between control cells and IL-1β-treated SW1353 cells at the time of presentation (P<0.05, Student's t-test). [Figure 7] This study demonstrates that an "in vitro" model of chondropathy cells (SW1353 cells treated with IL-1β) exhibits dysfunctional mitochondria with damaged mitochondrial membrane potential. Therefore, the fluorescence ratio of the mitochondrial membrane potential sensor JC10 (590 / 525 nm) significantly decreased 24 hours after IL-1β treatment, indicating reduced mitochondrial activation. Results are presented as the mean value + / - SEM of n=8 experiments for each condition. * indicates a statistically significant difference between control cells and IL-1β-treated SW1353 cells at the time of presentation (P<0.05, one-way ANOVA test). [Figure 8] This study demonstrates that an "in vitro" model of osteoarthritis chondrocytes (SW1353 cells treated with IL-1β) exhibits dysfunctional mitochondria with impaired mitochondrial calcium uptake during stimulation. The graph shows the increase in mitochondrial Ca2+ stimulated by the agonist (histamine) and the decrease in SW1353 cells treated with IL-1β at the time of presentation. Results are presented as the mean + / - SEM of experiments n=46-96 for each condition. * indicates a statistically significant difference between the control and IL-1β-treated SW1353 cells at the time of presentation (P<0.05, one-way ANOVA test). [Figure 9]This shows the SW1353 cell line, an "in vitro" genetic model of chondrocytes. Mitochondrocyte calcium uptake deficiency (MCU-knockdown, MCU-kd) indicates impaired chondrocyte function. Figure 9A is a Western blot showing a decrease in MCU expression in MCU-depleted SW1353 cells (MCU-kd). As shown in Figures 9B and 9C, collagen-IIα1 content and aggrecan content decrease significantly when the expression of MCU, a transporter that diverts calcium uptake in mitochondria, decreases. Furthermore, Figure 9D is a graph showing an increase in metalloproteinase MMP3 expression in MCU-depleted cells. The results are expressed in each graph (Figures 9B, C, and D) as the mean + / - SEM from n=3 experiments for each condition. * indicates a statistically significant difference between control cells and MCU-depleted cells, as indicated by P<0.05 (one-way ANOVA test). [Figure 10] The graph shows that in the cell model of osteoarthritis chondrocytes (SW1353 cells treated with IL-1βB) described in the previous figure, oleuropein aglycone (Oea) synergistically interacts with quercetin (Q) to activate mitochondria via an increase in mitochondrial Ca2+. The inset shows the effects of oleuropein aglycone (0.3 μM, black), quercetin (3 μM, gray), and the combination of 0.3 μM Oea + 3 μM Q (Oea / Q) on the integrated increase in mitochondrial calcium induced by 100 μM histamine. Using the data from the inset, the expected theoretical effect (sum of Oea effect and quercetin effect) and the actually measured effect for each combination (Oea + quercetin, Oea / Q) were determined in this figure. * indicates a statistically significant difference between the measured and theoretical values ​​in mitochondrial calcium with P < 0.05 (Student's t-test). [Figure 11]This graph shows that several combinations of oleuropein aglycone (Oea) and quercetin (Q) synergistically activate mitochondria via increased mitochondrial Ca2+ in a cell model of osteoarthritis chondrocytes (SW1353 cells treated with IL-1βB), as shown in the previous figure. The amounts of Oea and Q are shown at the top of each panel (A, B, C) and expressed in micromoles (μM). The expected theoretical effects (sum of Oea effect and quercetin effect) and the actually measured effects of combinations (Oea + quercetin, Oea / Q) were measured as shown in Figure 10 and compared here to represent the synergistic effect. The results are expressed as the mean + / - SEM of n=7 experiments for each condition. * indicates a statistically significant difference between the measured and theoretical values ​​in mitochondrial calcium with P<0.05 (Student's t-test). [Modes for carrying out the invention]

[0012] definition Before discussing the present invention in further detail, the following terms and general common technical knowledge are first defined.

[0013] In the background of this invention, unless otherwise specified, the percentages referred to are weight / weight percentage.

[0014] The term "and / or" used in the context of "X and / or Y" should be interpreted as "X" or "Y," or "X and Y."

[0015] The numerical ranges used herein are intended to include all digits and subsets of digits that fall within that range, whether expressly disclosed or not. Furthermore, these numerical ranges should be interpreted as providing support for claims that cover any digit or subset of digits within that range. For example, a disclosure of 1 to 10 should be interpreted as supporting ranges such as 1 to 8, 3 to 7, 4 to 9, 3.6 to 4.6, and 3.5 to 9.9.

[0016] As used herein, the terms "prevent" and "prevention" mean administering to a subject that does not exhibit any symptoms of a condition a composition disclosed herein to suppress or prevent the occurrence of at least one symptom associated with that condition. Further, "prevention" includes reducing the risk, incidence, and / or severity of a condition or disorder.

[0017] As used herein, an "effective amount" is an amount that treats or prevents a deficiency, treats or prevents a disease or medical condition, or more generally, alleviates symptoms, manages disease progression, or provides a nutritional, physiological, or medical benefit to an individual.

[0018] "Animal" includes, but is not limited to, mammals such as rodents, aquatic mammals, dogs, cats, and other household pets such as sheep, pigs, cows, and horses, and humans. When the terms "animal," "mammal," or their plurals are used, these terms also apply to any animal in which the indicated or intended effect can be demonstrated, for example, by the benefit to the animal from the improvement of mitochondrial calcium transport. The terms "individual" or "subject" are often used herein to refer to a human, but the disclosure is not so limited. Thus, the terms "individual" or "subject" refer to any animal, mammal, or human that can benefit from the methods and compositions disclosed herein.

[0019] The term "pet" means any animal that can benefit from or tolerate a composition provided by the present disclosure. For example, a pet may be an animal such as a bird, bovine, canine, equine, feline, goat, lupine, murine, ovine, or swine, but a pet can be any suitable animal. The term "companion animal" means a dog or a cat.

[0020] The "subject" or "individual" is a mammal, preferably a human. The term "elderly" in the context of humans means an age of at least 60 years, preferably over 63 years, more preferably over 65 years, and most preferably over 70 years. The term "older adult" in the context of humans means a postnatal age of 45 years or older, preferably over 50 years, more preferably over 55 years, and includes elderly individuals. The term "older adult" in the context of humans means a postnatal age of 45 years or older, preferably over 50 years, more preferably over 55 years, and includes elderly individuals.

[0021] An "oral nutritional supplement" or "ONS" is a composition containing at least one macronutrient and / or at least one micronutrient, and is, for example, in the form of a sterile liquid, semi-solid or powder, and is intended to supplement other nutritional intakes such as nutritional intake from food. Non-limiting examples of commercially available ONS products include MERITENE®, BOOST®, NUTREN®, and SUSTAGEN®. In some embodiments, the ONS can be ingested without further adding a liquid, for example, it can be a beverage in liquid form where the liquid volume is for one serving of the composition.

[0022] A "kit" means that the components of the kit are physically associated in or with one or more containers and are considered as one unit for manufacture, distribution, sale, or use. Containers include, but are not limited to, bags, boxes, cartons, bottles, packages of any type or design or material, overwraps, shrink wraps, attached components (such as stapled or glued ones), or combinations thereof.

[0023] Unless otherwise specified or expressly implied to be incompatible by the context in which they are referred, any reference to a single feature or single limitation of the Invention shall include a corresponding set of features or limitations, and any reference to a set of features or limitations shall include a corresponding single feature or single limitation.

[0024] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art.

[0025] Composition for use Joint diseases can involve inflammation to varying degrees. In some diseases, such as rheumatoid arthritis (RA), inflammation is the most important factor. In other diseases, such as osteoarthritis (OA), inflammation is not as pronounced. However, in all of these diseases, the articular cartilage is destroyed by catabolic elements.

[0026] The inventors have shown that providing a combination of oleuropein and / or its metabolites with quercetin and / or derivatives synergistically improves altered mitochondrial function, for example, in osteoarthritis.

[0027] In other words, the present invention relates in a first aspect to a composition comprising an effective amount of a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives for use in improving joint health, for example, in preventing or treating cartilage degeneration in an individual.

[0028] In another form, this aspect of the present invention may be described as the use of an effective amount of a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives in the manufacture of a pharmaceutical for the prevention or treatment of cartilage degeneration in an individual.

[0029] Use for the prevention or treatment of cartilage degeneration is synonymous with use for inhibiting or reducing cartilage degeneration.

[0030] Accordingly, embodiments of the present invention include compositions comprising an effective amount of a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives for use in preventing or treating cartilage degeneration.

[0031] Further embodiments of the present invention include compositions for use according to the present invention, the compositions further comprising calcium.

[0032] Raw materials: Major bioactive compounds Oleuropein and quercetin are the main biologically active molecules according to the present invention.

[0033] Oleuropein is a polyphenol found in the fruits, roots, trunks, and more specifically, leaves of plants belonging to the Oleaceae family, particularly olives (Olea europaea).

[0034] In one embodiment, at least a portion of oleuropein is obtained by extraction from plants belonging to the Oleaceae family, for example, by extraction from one or more of the stems, leaves, fruits, or kernels of plants belonging to the Oleaceae family, preferably such as olive (olive tree), plants of the genus Ligustrum, plants of the genus Syringa, plants of the genus Fraximus, plants of the genus Jasminum, and plants of the genus Osmanthus. Additionally or alternatively, at least a portion of oleuropein and / or its metabolites can be obtained by chemical synthesis.

[0035] Non-limiting examples of suitable metabolites of oleuropein include oleuropein aglycone, hydroxytyrosol, elenolic acid, homovanillyl alcohol, isohomovanillyl alcohol, their glucuronidated forms, their sulfate forms, their derivatives, and mixtures thereof.

[0036] Quercetin is the aglycone form of many other flavonoid glycosides, such as rutin and quercitrin, which are found in citrus fruits, buckwheat, and onions. Quercetin is derived from the glycosides quercitrin and rutin, which are associated with rhamnose and rutinose, respectively. Similarly, guaijavelin is a 3-O-arabinoside, hyperoside is a 3-O-galactoside, isoquercitin is a 3-O-glucoside, and spireoside is a 4'-O-glucoside. Miquelianin is quercetin 3-O-β-D-glucuronopyranoside.

[0037] In preferred embodiments, quercetin derivatives may be selected from the group consisting of quercetin 3-O-galactoside, quercetin 3-O-glucoside (isoquercetin), quercetin 3-O-xyloside, quercetin 3-O-rhamnoside (quercitrin), quercetin 3-O-glucuronide, quercetin 7-O-glucoside, quercetin 3-O-diglucoside, quercetin 3,4'-diglucoside, quercetin 3-O-rhamnoside-7 O-glucoside, quercetin 3-O-rutinoside (rutin), quercetin 3-O-6''-acetylglucoside, quercetin 3-methyl ether, quercetin 3,3'-dimethyl ether, and mixtures thereof.

[0038] Quercetin may be derived from any suitable raw material and may be isolated or chemically synthesized.

[0039] In preferred embodiments, oleuropein, quercetin, and their derivatives are obtained from plant materials. For example, oleuropein can be obtained from olive plants, rutin from onions, and quercetin from onions, green tea, apples, berries, ginkgo biloba, St. John's wort, elderberry, buckwheat tea, etc.

[0040] The effective doses of oleuropein and / or its metabolites and quercetin and / or its derivatives vary depending on the specific composition, the age and condition of the patient, and the specific disorder or disease being treated. However, in general embodiments, an individual may be administered 0.001 mg to 1.0 g / day, preferably 0.01 mg to 0.9 g / day, more preferably 0.1 mg to 750 mg / day, more preferably 0.5 mg to 500 mg / day, and most preferably 1.0 mg to 200 mg / day. Furthermore, the inventors have found that the active dose of oleuropein or its derivative in a combination can be reduced to obtain equivalent efficacy.

[0041] In some embodiments, a combination of oleuropein or a metabolite and quercetin or a derivative is administered in a composition further containing calcium. At least a portion of the calcium may be one or more calcium salts, such as calcium acetate, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium gluconate, calcium lactate, or a mixture thereof. In general embodiments, an individual is administered 0.1 g to 1.0 g of calcium per day, preferably 125 mg to 950 g of calcium per day, more preferably 150 mg to 900 mg of calcium per day, more preferably 175 mg to 850 mg of calcium per day, and most preferably 200 mg to 800 mg of calcium per day.

[0042] In an alternative embodiment, the combination of oleuropein and quercetin may be administered sequentially with calcium in separate compositions. The term “sequentially” means administering calcium and at least one of oleuropein or its metabolites sequentially, such that in a first time period, oleuropein or at least one of its metabolites is administered without calcium, and in a second time period (before or after the first time period), calcium is administered without the combination of oleuropein and quercetin. The time between sequential administrations may be, for example, one second or a few seconds, a few minutes or a few hours on the same day; one day or a few days or a few weeks on the same month; or one month or a few months on the same year.

[0043] In some embodiments, oleuropein or its metabolites and quercetin or its metabolites are the only polyphenols in the composition and / or the only polyphenols administered to the individual.

[0044] The composition may contain at least one effective amount of oleuropein or its metabolites. For example, a single supply or dose of the composition may contain an effective amount, and a package may contain one or more supplies or doses. Optionally, the composition may further contain calcium.

[0045] In another embodiment, oleuropein and / or derivatives may be provided by any of the compositions and methods disclosed in International Publication Nos. 2019 / 092068 and 2019 / 092066, titled “Biological Conversion of Oleuropein” and “Method for Selecting Probiotics,” respectively, and in International Publication No. 2019 / 092069, titled “Homovanillyl Alcohol (HVA), HVA Isomers, Method for Producing Compositions Containing Such Compounds, and Method for Using Such Compounds,” the wholes of which are incorporated herein by reference.

[0046] Raw materials Further bioactive compounds The compositions for use according to the present invention may also contain at least one further bioactive compound selected from the group consisting of antioxidants, anti-inflammatory compounds, glycosaminoglycans, prebiotics, fibers, probiotics, fatty acids, enzymes, minerals, trace elements and / or vitamins.

[0047] In the context of this application, the term "bioactive" means that a compound contributes to the health of an individual or has an effect on the human body beyond its effect of meeting basic nutritional requirements.

[0048] At least one further bioactive compound may be derived from a natural source; that is, the compound may be obtained from an extract of plants, animals, fish, fungi, algae, or microbial fermentation products. Minerals are also considered to be derived from natural sources within this definition.

[0049] In preferred embodiments, the enzyme may be a protease such as trypsin, or an enzyme extract such as bromelain.

[0050] Nutritional composition The compositions for use according to the present invention may be nutritional compositions or pharmaceutical compositions, and may be for human or veterinary use.

[0051] Therefore, in a preferred embodiment, the composition for use according to the present invention is a nutritional composition.

[0052] In the context of this application, "nutritional composition" means a composition that serves as a source of nutrition for an individual.

[0053] The nutritional product or composition of the present invention can serve as a complete or near-complete source of nutrition.

[0054] As used herein, “complete nutrition” encompasses nutritional products and compositions containing sufficient types and levels of major nutrients (proteins, fats, and carbohydrates) and sufficient micronutrients to make the composition the sole nutritional source for the animal to which it is administered. The patient can obtain 100% of their nutritional requirements from such complete nutrition composition.

[0055] As used herein, “near-complete nutrition” includes nutritional products or compositions that do not contain sufficient levels of major nutrients (proteins, fats, and carbohydrates) or sufficient micronutrients to make the composition the sole nutritional source for an animal administered the composition. Partial or near-complete nutritional compositions can be used as nutritional supplements.

[0056] The combination of oleuropein and quercetin can be administered in any composition suitable for human and / or animal intake. In preferred embodiments, the combination is administered orally or enterally (e.g., via tube feeding) to the individual. For example, the combination can be administered to the individual as a beverage, food product, capsule, tablet, powder, or suspension.

[0057] Non-limiting examples of suitable compositions include food compositions, dietary supplements, dietary supplements (e.g., liquid ONS), complete nutrition compositions, beverages, pharmaceuticals, oral nutritional supplements, medical foods, nutraceuticals, foods for specific medical purposes (FSMPs), powdered nutritional products that are reconstituted with water or milk before ingestion, food additives, pharmaceuticals, drinks, pet food, and combinations thereof.

[0058] Raw materials for nutritional compositions Protein source In one embodiment, the composition for use according to the present invention comprises a protein source. This protein source may be a food protein, including, but is not limited to, animal proteins (such as milk protein, meat protein, and egg protein), plant proteins (such as soy protein, wheat protein, rice protein, and pea protein), or a combination thereof. In one embodiment, the protein is selected from the group consisting of whey, chicken, corn, casein salt, wheat, flax, soybeans, carob beans, peas, or a combination thereof.

[0059] Carbohydrate sources In one embodiment, the composition contains a carbohydrate source. The composition of the present invention may contain any suitable carbohydrate, but is not limited to starch, sucrose, lactose, glucose, fructose, corn syrup solid, maltodextrin, modified starch, amylose starch, tapioca starch, corn starch, xylitol, sorbitol, or a combination thereof.

[0060] fat source In one embodiment, the composition includes a fat source. The fat source may include any suitable fat or fat mixture. For example, the fat source may include, but is not limited to, vegetable fats (e.g., olive oil, corn oil, sunflower oil, high oleic acid oil, rapeseed oil, canola oil, hazelnut oil, soybean oil, palm oil, coconut oil, blackcurrant seed oil, borage oil, lecithin, etc.), animal fats (e.g., milk fat), or combinations thereof. The fat source may also be a crude product of the above fats (e.g., polyphenol-containing olive oil).

[0061] Fragrance etc. In addition, compositions for use according to the present invention may also contain natural or synthetic flavors, such as fruit flavors like banana, orange, peach, pineapple, or raspberry, or other plant-based flavors like vanilla, cocoa, or coffee.

[0062] Nutrition composition format This nutritional composition may optionally contain, in addition to the main bioactive ingredient and any further bioactive ingredients, and optionally one or more of the protein, carbohydrate, and fat sources, any number of additional food ingredients, including conventional food additives (synthetic or natural), such as one or more acidulants, additional thickeners, buffers or pH adjusters, chelating agents, colorants, emulsifiers, additives, flavorings, minerals, penetrating agents, pharmaceutically acceptable carriers, preservatives, stabilizers, sugars, sweeteners, quality improvers, and / or vitamins. Any selected ingredients may be added in any appropriate amount.

[0063] This nutritional composition may be provided in any preferred form.

[0064] Examples of nutritional composition forms in which compositions for use according to the present invention may be supplied include solutions, ready-for-consumption compositions (e.g., ready-for-consumption beverages or instant beverages), liquid foods, soft drinks, juices, sports drinks, dairy beverages, milkshakes, yogurt drinks, soups, and the like.

[0065] In other embodiments, the nutritional composition may be provided in the form of a concentrate, powder, or granules (e.g., effervescent granules) and may be diluted with water or other liquids, such as milk or fruit juice, to prepare a ready-made composition.

[0066] The formats of the nutritional composition can also include baked goods, dairy products, desserts, confectionery products, cereal bars, and breakfast cereals. Examples of dairy products include milk and milk beverages, yogurt, and other fermented dairy products, ice cream, and cheese. Examples of baked goods include bread, biscuits, and cakes.

[0067] In one embodiment, the compositions for use according to the present invention may also be available in a wide variety of forms designed as animal food, particularly dog ​​or cat food, such as a wet form, a semi-wet form or a dry form, and especially in the form of biscuits.

[0068] Route of administration The nutritional compositions of this disclosure may be administered by any means suitable for administration to humans, and in particular to any site of the gastrointestinal tract. Intestinal administration, oral administration, and administration through tubes or catheters are all included in this disclosure. The nutritional compositions may also be administered by methods selected from oral, enteral, sublingual, sublabial, buccal, topical, etc.

[0069] This nutritional composition can be administered in any known form that is convenient for administration, such as tablets, capsules, liquids, chewables, softgels, sachets, powders, syrups, suspensions, emulsions, and solutions. In soft capsules, the active ingredient is preferably dissolved or suspended in a suitable liquid such as fatty oil, paraffin oil, or liquid polyethylene glycol. Stabilizers may be added as desired.

[0070] When this nutritional composition is administered via enteral nutrition, it can be used for short-term or long-term enteral nutrition.

[0071] Inhibition or reduction of cartilage degeneration Cartilage degeneration can be the result of pathology (either chronic or acute), trauma, or a combination thereof.

[0072] Cartilage degeneration occurs in both conditions dominated by inflammation (e.g., rheumatoid arthritis) and conditions in which inflammation is not so pronounced (e.g., osteoarthritis).

[0073] Trauma can also trigger the onset of cartilage degeneration processes. For example, a ligament tear in the knee can lead to knee joint instability, initiating the degenerative process.

[0074] In the context of this application, trauma refers to physiological damage caused by external factors such as falls or impacts from vehicles. Trauma may also refer to the accumulation of small injuries over time, so-called "wear and tear."

[0075] While surgical treatment is often preferred for treating injuries, in one embodiment, the present invention relates to a treatment method for treating injuries by surgical treatment and by administering the composition of the present invention.

[0076] Therefore, embodiments of use according to the present invention include use to inhibit or reduce cartilage degeneration, which is a result of pathology or trauma.

[0077] Examples of medical conditions involving cartilage degeneration, for which the compositions of the present invention may be useful, include osteoarthritis, rheumatoid arthritis, gout and pseudogout, septic arthritis, ankylosing spondylitis, juvenile idiopathic arthritis, Still's disease, psoriasis (psoriatic arthritis), reactive arthritis, Ehlers-Danlos syndrome, hemochromatosis, hepatitis, Lyme disease, inflammatory bowel disease (including Crohn's disease and ulcerative colitis), Henoch-Schönlein purpura, hyperimmunoglobulinemia D with recurrent fever, sarcoidosis, TNF receptor-associated periodic syndromes, Wegener's granulomatosis (and many other vasculitis syndromes), familial Mediterranean fever, and systemic lupus erythematosus.

[0078] In a preferred embodiment, the composition of the present invention is intended for use in inhibiting or reducing cartilage degeneration in RA and / or OA.

[0079] In a more preferred embodiment, the composition of the present invention is intended for use in inhibiting or reducing cartilage degeneration in OA.

[0080] Furthermore, while we do not wish to be bound by theory, it has been observed that while inflammation often leads to cartilage degeneration in joints, cartilage degeneration can also occur in situations where the inflammatory component is far less, and perhaps even negligible.

[0081] For example, joint trauma can sufficiently initiate cartilage degeneration even without significant inflammatory components present in, for instance, rheumatoid arthritis (RA). Trauma may include, for example, ligament rupture or impact trauma to a joint (e.g., knee, finger).

[0082] In another example, osteoarthritis (OA) is primarily a degenerative joint disease with fewer inflammatory components.

[0083] Accordingly, in one embodiment, the present invention relates to a composition for use according to the present invention for inhibiting or reducing cartilage degeneration, wherein the cartilage degeneration occurs in association with a condition that involves little or no inflammatory elements, such as trauma or osteoarthritis.

[0084] Use to counter initial denaturation events The hypertrophy suggests catabolic activity in chondrocytes that does not exhibit a normal phenotype.

[0085] Accordingly, the present invention relates in one embodiment to a composition comprising a combination of oleuropein and / or its metabolites and quercetin and / or derivatives for use in inhibiting or reducing chondrocyte hypertrophy, which is an initial event indicating cartilage degeneration.

[0086] Treatment or prevention of age-related decline in mobility The compositions for use according to the present invention have been shown to inhibit or reduce proteolytic activity.

[0087] Aging leads to cartilage degeneration.

[0088] Therefore, the present invention relates to a composition for use in inhibiting or reducing cartilage degeneration associated with aging.

[0089] In another embodiment, the present invention relates to compositions for use in inhibiting or reducing collagen degeneration in aging-related cartilage degeneration, for example, for use in inhibiting or preventing collagen II degeneration in aging-related cartilage.

[0090] Cartilage degeneration can contribute to joint stiffness and pain, leading to reduced mobility in patients.

[0091] In other embodiments, compositions for use according to the present invention may be used to: i) maintain or improve joint function, including cartilage function, in aging; and ii) reduce joint pain, including inflammatory and / or nociceptive pain.

[0092] In further embodiments, the present invention relates to compositions for use according to the present invention for improving mobility in subjects, for example, adult or aged mammals.

[0093] Therefore, in a preferred embodiment, the composition according to the present invention may be used, for example, to improve the activity and / or mobility of an individual by preventing or treating osteoarthritis and / or inhibiting or reducing cartilage degeneration.

[0094] Other preferred embodiments relate to compositions for use according to the present invention, which are used to prevent or reduce joint pain (inflammatory and / or nociceptive pain) in order to prevent cartilage degeneration and thus maintain healthy joints, or to maintain or improve mobility. In further embodiments, compositions of the present invention may be used to maintain the condition of cartilage.

[0095] target group The target group of compositions for use according to the present invention may be any mammal exhibiting cartilage degeneration due to suffering from one or more of the cartilage degeneration-related conditions mentioned herein. Cartilage degeneration can be detected by visual means such as X-ray imaging. Alternatively, products of cartilage degeneration can be detected in body fluids. For example, one or more collagen II epitopes such as (Coll2-1, Coll2-1 NO2, CTX-II) can be detected in samples such as plasma or urine samples.

[0096] Another group of subjects may be any mammals that have not yet shown cartilage degeneration but are at risk of cartilage degeneration, for example, OA, RA, or any of the cartilage degeneration-related medical conditions referred to herein. In a preferred embodiment, the present invention relates to a composition according to the present invention, comprising a combination of oleuropein or its metabolites and quercetin or a derivative, for use in inhibiting or reducing early cartilage degeneration, and which is administered to this group of subjects.

[0097] Specific embodiments of the present invention relate to compositions for use in improving the activity and / or mobility of an individual, for example, by preventing or treating osteoarthritis, and / or by inhibiting or reducing cartilage degeneration in elderly or aged individuals.

[0098] In further embodiments, the compositions for use according to the present invention may be for use in mammals such as humans or in pets. Examples of pets include cats, dogs, and horses.

[0099] While the present invention may be useful in many different age groups, in preferred embodiments, the compositions for use in improving mobility according to the present invention are intended for elderly populations, particularly healthy aged and / or elderly mammals.

[0100] Method for producing the nutritional composition of the present invention In a further aspect, the present invention relates to a method for producing a nutritional composition for use according to the present invention, wherein the method is The method includes the steps of preparing raw materials for a nutritional composition comprising a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives, and mixing them so that the nutritional composition comprises a combination of oleuropein and / or its metabolites and quercetin and / or its derivatives.

[0101] Pharmaceutical composition for use In further embodiments, the present invention relates to a composition for use in inhibiting or preventing cartilage degeneration according to the present invention, which is a pharmaceutical composition.

[0102] By being classified as a pharmaceutical, it means a composition in which a substance is used on or within the body to prevent, diagnose, alleviate, treat, or cure a disease in humans or animals, unlike a nutritional composition. According to the present invention, this pharmaceutical can be used to suppress or reduce cartilage degeneration.

[0103] The pharmaceutical product may be intended for human use. Alternatively, it may be a veterinary composition suitable for, for example, dogs, cats, or horses, particularly thoroughbred horses.

[0104] In one preferred embodiment, the pharmaceutical composition of the present invention comprises a combination of oleuropein or its metabolites and quercetin or a derivative thereof.

[0105] In another preferred embodiment, the pharmaceutical composition of the present invention comprises oleuropein or its metabolite, quercetin or a derivative, and curcumin.

[0106] The present invention further relates to the use of pharmaceuticals according to the present invention, as described herein as an use of the compositions of the present invention.

[0107] The present invention provides a pharmaceutical composition for use comprising oleuropein or its metabolites in combination with quercetin or its derivatives and / or curcumin, in combination with at least one additive selected from the group consisting of pharmaceutically acceptable additives. Procedures for preparing the pharmaceutical composition according to the present invention can be readily obtained by those skilled in the art, for example, by following the description in the handbook Remington's Pharmaceutical Sciences, Mid. Publishing Co, Easton, Pa., USA. Physiologically acceptable additives, vehicles, and adjuvants are also described in the handbook entitled "Handbook of Pharmaceutical Excipients, Second edition, American Pharmaceutical Association, 1994". To formulate the pharmaceutical composition according to the present invention, it would be advantageous for those skilled in the art to refer to the latest editions of the European Pharmacopoeia or the United States Pharmacopeia (USP). In particular, it would be advantageous for those skilled in the art to refer to the 4th edition "2002" of the European Pharmacopoeia or the USP 25-NF 20 edition of the United States Pharmacopeia (USPharmacopoeia).

[0108] Advantageously, the pharmaceutical compositions as defined above are suitable for oral, parenteral, or intravenous administration. If the pharmaceutical composition for use according to the present invention contains at least one pharmaceutically acceptable or physiologically acceptable additive, in particular, the additive is suitable for administration of the composition via an oral route, or the additive is suitable for administration of the composition via a parenteral route.

[0109] The pharmaceutical compositions for use according to the present invention are available in solid or liquid form without distinction. For oral administration, solid pharmaceutical compositions in the form of tablets, capsules, or gelatin capsules would be preferred.

[0110] If in liquid form, the pharmaceutical composition would preferably be in the form of an aqueous or non-aqueous suspension, or an emulsion of water in oil or oil in water.

[0111] The solid pharmaceutical form may contain, as a vehicle, adjuvant, or additive, at least one diluent, one fragrance, one solubilizer, one lubricant, one suspending agent, one binder, one disintegrant, and one encapsulating agent. Such compounds include, for example, magnesium carbonate, magnesium stearate, talc, lactose, pectin, dextrin, starch, gelatin, cellulosic materials, and cocoa butter. The liquid composition may contain water, optionally as a mixture with propylene glycol or polyethylene glycol, and may also optionally contain colorants, fragrances, stabilizers, and thickeners.

[0112] Combination with known treatments In situations where there are no disease-modifying osteoarthritis medications (DMOADs), alternative treatments and prevention of osteoarthritis can be carried out through nutrition.

[0113] Histological data reveal that oleuropein has a greater impact on the OA score than compounds that primarily affect degeneration. Therefore, the efficacy of oleuropein may be due to a combined effect on inflammation and degeneration. Accordingly, compositions of the present invention, which have been shown to inhibit or reduce degeneration in preferred embodiments, can be combined with treatments to inhibit or reduce inflammation.

[0114] Treatment method The present invention also relates to a method for preventing or treating cartilage degeneration, such as a medical condition resulting in cartilage degeneration or an injury associated with cartilage degeneration, comprising administering an effective amount of a composition according to the present invention to an individual in need. For example, the method comprises administering an effective amount of a composition comprising oleuropein or its metabolites in combination with quercetin or a derivative.

[0115] As used herein, “effective dose” means a quantity that prevents a deficiency, treats a disease or medical condition in an individual, or more generally, alleviates symptoms, controls the progression of a disease, or provides a nutritional, physiological, or medical benefit to the individual.

[0116] The effective amount of the composition of the present invention required to achieve a therapeutic effect may, of course, vary depending on the specific composition, route of administration, the age and condition of the subject, and the specific disorder or disease being treated.

[0117] The present invention further provides a method for preventing or treating a disease involving cartilage degeneration, such as OA or RA, inhibiting or reducing cartilage degeneration, inhibiting or reducing collagen degeneration in cartilage, or inhibiting or reducing collagen II degeneration in cartilage, the method comprising administering an effective amount of the composition for use according to the present invention to an individual.

[0118] In one embodiment, the treatment method according to the present invention relates to the prevention or treatment of osteoarthritis.

[0119] The therapeutic method according to the present invention may be used in mammals, such as humans, or in pets, such as dogs, cats, and / or horses.

[0120] In certain embodiments, the composition of the present invention administered in the therapeutic method may be one or more nutritional compositions and / or pharmaceutical compositions of the present invention.

[0121] kit The Disclosure also provides a kit comprising combinations of oleuropein and / or its metabolites and quercetin and / or derivatives in one or more containers. Embodiments of the kit include one or more first containers, each containing oleuropein and / or metabolites separately from quercetin and / or derivatives contained in at least one second container, and the kit further includes instructions for mixing oleuropein with quercetin into unit dosage forms.

[0122] In one embodiment of the kit, the combination may be provided together as one or more pre-packaged unit dosage forms, for example, as separate containers each containing a dry powder, such that each container contains one pre-packaged unit dosage form.

[0123] In another embodiment, the kit may include a plurality of compositions for mixing together to form one or more compositions disclosed herein. For example, the kit may contain two or more dry powders in separate containers relating to each other, each of which contains a portion of the final unit dosage form. In a non-limiting example of such an embodiment, the kit may include one or more first containers containing oleuropein and one or more second containers containing quercetin. The contents of one of the first containers may be mixed with one of the second containers to form at least a portion of the unit dosage form of the composition.

[0124] The above administration examples do not require uninterrupted daily administration. Rather, several short interruptions in administration, such as 2-4 day breaks during the administration period, may be permitted. The ideal duration of administration of this composition can be determined by those skilled in the art.

[0125] Disclosure combinations It should be noted that embodiments and features described in the context of one aspect of the present invention also apply to other aspects of the invention.

[0126] The compositions for use according to the present invention are described herein in terms of different parameters such as raw materials, form of nutritional composition, use, and target group. It should be noted that embodiments and features described in the context of one of the parameters of the compositions for use according to the present invention may be combined with other embodiments and features described in the context of another parameter, unless otherwise expressly specified.

[0127] All patent and non-patent documents cited in this application are incorporated into this application exactly as they appear in the documents.

[0128] The present invention will be described in more detail by the following non-limiting embodiments. [Examples]

[0129] The following non-limiting examples illustrate experimental data supporting the compositions and methods disclosed herein.

[0130] Example 1 To test the effects of oleuropein (or oleuropein aglycone), quercetin, and combinations thereof in living cells, the inventors measured the elevation of mitochondrial calcium in HeLa cells. HeLa cells were purchased from ATCC. HeLa cells were seeded in 96-well plates at a density of 50,000 cells / well in minimal essential medium (DMEM, Gibco), high glucose, +10% fetal bovine serum. Mitochondrial calcium measurements were performed using HeLa cells (Sirion Biotech) infected with an adenovirus expressing mitochondrial-modified aequorin, a calcium sensor targeted in mitochondria (Montero et al., 2004). For aecion reconstitution, 24 hours after infection, cells were incubated for 2 hours at room temperature (22±°C) in standard medium (145 mM NaCl, 5 mM KCl, 1 mM MgCl2, 1 mM CaCl2, 10 mM glucose, and 10 mM Hepes, pH 7.4) supplemented with 1 μM wild-type coelenterazine. During processing, the compound was added directly to the cell culture or myotube culture tissue 2 hours prior to measurement. Luminescence was measured using FLIPR tetraequorin (Molecular Devices). Increased mitochondrial calcium was obtained by stimulating cells with 100 μM histamine. Calibration of luminescence data to calcium concentration was performed using the algorithm described above (Alvarez & Montero, 2002). Quantification was performed using custom module analysis based on Excel (Microsoft) and GhaphPad Prism 7.02 (GhaphPad) software.

[0131] To investigate the effects of the pro-inflammatory cytokine interleukin-1β as an osteoarthritis mimic in a chondrocyte model, and to test the effect of mitochondrial calcium uniporter (MCU) removal on chondrocyte function, the inventors measured chondrocyte and mitochondrial functions in SW1353 cells. SW1353 cells were purchased from ATCC. SW1353 cells were seeded in 96-well plates at a density of 10,000 cells per well in a 100 mm dish (for mitochondrial calcium measurement) or at a density of 1,000,000 cells (for Western blotting). Cells were cultured in high-glucose minimal essential medium (DMEM, Gibco) containing 10% fetal bovine serum and 1% penicillin-streptomycin.

[0132] To evaluate chondrocyte function, collagen-IIα1 expression, agrecan, and metalloproteinases-3 and -13 (MMP3 and MMP13) were analyzed by Western blotting. SW1353 cells were evaluated with or without treatment with 10 ng / mL interleukin-1b for 24 or 48 hours. Protein extracts were prepared in a suitable buffer containing 150 mM NaCl, 1.0% IGEPAL CA-630, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris, pH 8.0, Complete EDTA-free protease inhibitor mixture (Roche), 1 mM PMSF, 1 mM NaVO3, 5 mM NaF, and 3 mM β-glycerophosphate and phosphatase inhibitor (Roche). 40 μg of total protein was packed according to BCA quantification. Proteins were separated by SDS-PAGE electrophoresis on commercially available 4-12% acrylamide gel (Thermo Fisher Scientific) and transferred to PVDF membrane (Thermo Fisher Scientific) by wet electrophoresis. The blots were blocked at room temperature for 1 hour with a TBS-tween solution (0.5M Tris, 1.5M NaCl, 0.01% Tweenn) of 5% fetal bovine serum (Sigma-Aldrich) and incubated overnight with the primary antibody at 4°C. The secondary antibody was incubated at room temperature for 1 hour. The following antibodies were used: anti-collagen-IIα1 (1:1000, Abcam), anti-aggrecan (1:1000, Abcam), anti-MMP-3 (1:1000, Abcam), and anti-MMP-13 (1:1000, Abcam). The secondary HRP conjugate antibody, purchased from Cell Signalling, was used at a 1:5000 dilution. Next, the content of collagen-IIα1, aggrecan, MMP3, and MMP13 was quantified by densitometry and normalized to the GAPDH content (anti-GAPDH, 1:5000, detected by Cell Signaling).

[0133] To eliminate mitochondrial calcium uniporter (MCU) protein expression, SW1353 cells were transfected with an engineered vector for MCU knockdown manufactured by Sirion Biotech. Three days after infection, MCU expression was quantified by Western blotting and densitometry analysis as described above. The MCU content was normalized relative to the mitochondrial protein TOM20 content. The following antibodies were used: anti-MCU (1:1000, Sigma-Aldrich), anti-TOM20 (1:5000, Cell Signaling).

[0134] To quantify the effect of interleukin-1β on cell death in SW1353 cells, apoptosis dynamics experiments were performed using an IncuCyte ZOOM instrument (Essen Bioscience, Ann Arbor, MI, USA). Cells were seeded in DMEM medium in a 96-well plate format at 50% confluence. After 24 hours, cells were incubated with IncuCyte Annexin-V Green (4642) and treated with IL-1β according to the supplier's instructions. Four images per well were collected at presentation time using a 10× objective lens and bandwidth filter (Ex: 440 / 80nm, Em: 504 / 44nm). The data were output as the area covered by Annexin-V-positive material per well (μm2) and normalized for the area covered by cells.

[0135] To measure mitochondrial membrane potential, SW1353 cells were seeded at a density of 8000 cells / well in growth medium (DMEM high glucose, Gibco + 10% fetal bovine serum) in 96-well plates. After 24 hours, the cells were treated with 10 ng / ml IL-1β for the number of days shown in Figure 7. The cells were then loaded with a fluorescent mitochondrial membrane potential sensor JC-10. Fluorescence was acquired using MetaXpress Confocal (Molecular Devices) at the following emission wavelengths, as instructed by the supplier: 590 nm (excited at 540 nm) and 525 nm (excited at 49 nm). The fluorescence ratio at 590 nm / 525 nm is proportional to the change in mitochondrial membrane potential.

[0136] The same procedure used for HeLa cells was followed to measure mitochondrial calcium in SW1353 cells. The synergistic effects of quercetin and oleuropein in the osteoarthritis chondrocyte model were quantified as described for HeLa cells.

[0137] result: As shown in Figure 1, oleuropein synergistically with quercetin to activate mitochondria by increasing mitochondrial calcium elevation in HeLa cells during stimulation. As shown in Figure 2, oleuropein aglycone also synergistically with quercetin to activate mitochondria by increasing mitochondrial calcium elevation in HeLa cells during stimulation. Conversely, as shown in Figure 3, oleuropein does not synergistically with oleuropein aglycone to activate mitochondria via mitochondrial calcium elevation in HeLa cells. As shown in Figure 4, oleuropein and oleuropein aglycone promote the same level of synergy in combination with quercetin.

[0138] As shown in Figure 5, the cell model of osteoarthritis chondrocytes, considering collagen-IIα1 content, shows a significant decrease in dysfunctional chondrocytes compared to controls (non-osteoarthritis chondrocytes, Figure 5A), and increased expression of metalloproteinases MMP3 (Figure 5B) and MMP13 (Figure 5C). Consistently, these dysfunctional osteoarthritis chondrocytes are characterized by increased cell death (Figure 6). As shown in Figure 7, considering the decrease in mitochondrial membrane potential, mitochondrial function is impaired in this osteoarthritis chondrocyte cell model. Indeed, the elevation of mitochondrial calcium is significantly reduced during stimulation in this cell model of osteoarthritis chondrocytes (Figure 8). As shown in Figure 9, a gene model of chondrocytes with dysfunctional mitochondrial calcium uptake (MCU-knockdown, kd, Figure 9A) reveals chondrocyte dysfunction that mimics the effects of osteoarthritis. Therefore, collagen-IIα1 expression (Figure 9B) and aggrecan expression are decreased in MCU-kd cells (Figure 9C), while metalloproteinase MMP3 expression is increased. Finally, as shown in Figure 10, oleuropein aglycone (Oea) synergistically with quercetin (Q) activates mitochondria via mitochondrial Ca2+ elevation in the described cell model of osteoarthritis chondrocytes (IL-1b-treated SW1353 cells). As shown in Figure 11, several combinations of oleuropein aglycone (Oea) and quercetin (Q) synergistically activate mitochondria via mitochondrial Ca2+ elevation in the described cell model of osteoarthritis chondrocytes.

Claims

1. A combination of an effective amount of at least one selected from oleuropein aglycone, elenolic acid, homovanillyl alcohol, and isohomovannillyl alcohol, and quercetin and / or a derivative of quercetin, for use in the prevention or treatment of cartilage degeneration in an individual, A composition in which the quercetin derivative is selected from the group consisting of quercetin 3-O-galactoside, quercetin 3-O-glucoside (isoquercetin), quercetin 3-O-xyloside, quercetin 3-O-rhamnoside (quercitrin), quercetin 3-O-glucuronide, quercetin 7-O-glucoside, quercetin 3-O-diglucoside, quercetin 3,4'-diglucoside, quercetin 3-O-rhamnoside-7O-glucoside, quercetin 3-O-rutinoside (rutin), quercetin 3-O-6''-acetylglucoside, quercetin 3-methyl ether, quercetin 3,3'-dimethyl ether, and mixtures thereof.

2. The composition for use according to claim 1, wherein the combination comprises oleuropein aglycone and quercetin.

3. The composition for use according to claim 1 or 2, which increases mitochondrial calcium in chondrocytes.

4. A composition for use according to any one of claims 1 to 3, further comprising calcium.

5. A composition for use according to any one of claims 1 to 4, further comprising at least one compound selected from the group consisting of antioxidants, anti-inflammatory compounds, glycosaminoglycans, prebiotics, fibers, probiotics, fatty acids, enzymes, minerals, trace elements and / or vitamins.

6. The composition for use according to any one of claims 1 to 5, wherein the composition is selected from the group consisting of food compositions, dietary supplements, nutritional compositions, oral nutritional supplements, medical foods, nutraceuticals, beverages, powdered nutritional products that are reconstituted with water or milk before ingestion, food additives, foods for specific medical purposes (FSMP) pharmaceuticals, drinks, pet food, and combinations thereof.

7. The composition for use according to any one of claims 1 to 6, wherein the composition is in the form of a solid powder, a powder stick, a capsule, or a solution.

8. The composition for use according to any one of claims 1 to 7, wherein the use is to i) maintain or improve joint function, including cartilage function, in aging, and ii) reduce joint pain, including inflammatory and / or nociceptive pain.

9. The composition for use according to any one of claims 1 to 8, wherein the use is for improving the activity and / or mobility of the individual.

10. The composition for use according to any one of claims 1 to 9, wherein the use is for inhibiting or reducing cartilage degeneration in osteoarthritis.

11. The composition for use according to any one of claims 1 to 10, wherein the individual is a middle-aged or elderly person.

Images