Breast milk-derived polypeptides and their application in preventing and treating childhood obesity

Abstract

The present application belongs to the field of biomedical technology, and specifically relates to a breast milk-derived polypeptide and its application in preventing and treating childhood obesity. The breast milk-derived polypeptide provided in the present application has an amino acid sequence as shown in SEQ ID NO.1, and the breast milk-derived polypeptide has significant effects in enhancing thermogenesis of brown adipose tissue in lactating mice, reducing the body weight of obese mice mediated by a high-fat diet, improving glucose metabolism, enhancing mitochondrial quality, and reducing mitochondrial autophagy. The breast milk-derived polypeptide and its pharmaceutical composition have good application prospects in the preparation of drugs for preventing and treating childhood obesity.

Description

Technical Field

[0001] The present application belongs to the field of biomedical technology, and specifically relates to breast milk-derived polypeptides and their application in preventing and treating childhood obesity. Background Art

[0002] Childhood obesity is a global public health problem. With the rapid economic development and changes in lifestyle, the prevalence of childhood obesity is increasing. Childhood obesity not only affects children's physical health, but may also increase the risk of chronic diseases in adulthood, such as cardiovascular and cerebrovascular diseases, type 2 diabetes, metabolic syndrome, etc. The occurrence of childhood obesity is the result of the combined effects of multiple factors such as genetics, environment and dietary behavior.

[0003] The diagnosis of childhood obesity usually uses body mass index (BMI) as an indicator. Children of different ages have different BMI cutoffs to determine overweight and obesity. Intervention measures for childhood obesity include lifestyle intervention, psychological and behavioral intervention, drug therapy, and metabolic weight loss surgery. Among them, physical activity and nutritional intervention are considered the best intervention methods for childhood obesity.

[0004] It has become a consensus in the industry that breastfeeding reduces the risk of childhood obesity. Children rely on brown adipose tissue as the main source of heat production, so breast milk's regulation of the functional phenotype of brown adipose tissue in offspring is an important biological basis for resisting childhood obesity.

[0005] However, the current prevention and treatment of childhood obesity mainly includes improving eating habits and increasing physical activity, controlling the total amount of food, and adjusting the dietary structure and dietary behavior. There are no relevant reports on the use of breast milk peptides in the prevention and treatment of childhood obesity. Summary of the invention

[0006] Based on this, one or more embodiments of the present application provide a breast milk-derived polypeptide and its application in preventing and treating childhood obesity.

[0007] On the one hand, the present application provides a breast milk-derived polypeptide, whose amino acid sequence is shown in SEQ ID NO.1.

[0008] On the other hand, the present application provides a pharmaceutical composition comprising the breast milk-derived polypeptide.

[0009] In one embodiment, the drug includes the human milk-derived polypeptide and pharmaceutically acceptable excipients.

[0010] In one embodiment, the concentration of the breast milk-derived polypeptide at which the biological activity is exerted is 8 mg / kg to 12 mg / kg.

[0011] On the other hand, the present application provides the use of the breast milk-derived polypeptide or the pharmaceutical composition in the preparation of a drug for preventing and treating childhood obesity.

[0012] In one embodiment, the childhood obesity includes secondary obesity.

[0013] In one embodiment, the dosage form of the drug includes one or more of tablets, capsules, granules, oral solutions and injections.

[0014] In one embodiment, the subject of the drug is a mammal.

[0015] In one embodiment, the subject of the drug is a primate.

[0016] Optionally, the subject of the drug is a human.

[0017] On the other hand, the present application provides a health product that helps regulate body fat function, and the health product includes the breast milk-derived polypeptide. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the technical solutions in the embodiments of the present application and to more completely understand the present application and its beneficial effects, the following is a brief introduction to the drawings required for the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present application, and those skilled in the art can obtain other drawings based on these drawings without creative work.

[0019] Figure 1 MDPAO1 promotes the thermogenic phenotype of brown adipose tissue in lactating mice;

[0020] Figure 2 MDPAO1 reduces body weight and improves glucose metabolism in HFD-induced obese mice;

[0021] Figure 3 Enhanced mitochondrial mass for MDPAO

[0022] Figure 4 MDPAO1 reduces mitophagy;

[0023] MDPAO1 is Milk-Derived Peptide Associated with Obesity 1, i.e., human breast milk-derived obesity-associated polypeptide, i.e., SEQ ID NO.1; HFD group is High-fat diet group; GTT is Glucosetolerance test; ITT is Insulin tolerance test; energy expenditure is energy expenditure; fat cell inner diameter is fat cell inner diameter; interscapular temperature is interscapular temperature; glucose is glucose. DETAILED DESCRIPTION

[0024] The present application will be further described in detail below in conjunction with the accompanying drawings, embodiments and examples. It should be understood that these embodiments and examples are only used to illustrate the present application and are not used to limit the scope of the present application. The purpose of providing these embodiments and examples is to make the understanding of the disclosure of the present application more thorough and comprehensive. It should also be understood that the present application can be implemented in many different forms, is not limited to the embodiments and examples described herein, and those skilled in the art can make various changes or modifications without violating the connotation of the present application, and the equivalent form obtained also falls within the protection scope of the present application. In addition, in the description below, a large number of specific details are given in order to provide a more comprehensive understanding of the present application, and it should be understood that the present application can be implemented without one or more of these details.

[0025] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art to which this application belongs. The terms used herein in the specification of this application are only for the purpose of describing implementation modes and embodiments and are not intended to limit this application.

[0026] the term

[0027] Unless otherwise specified or incompatible herewith, the terms and phrases used herein shall have the following meanings:

[0028] The terms "and / or", "or / and", and "and / or" used in this article include any one of two or more related listed items, and also include any and all combinations of related listed items, and the arbitrary and all combinations include any two related listed items, any more related listed items, or a combination of all related listed items. It should be noted that when at least three items are connected by at least two conjunctions selected from "and / or", "or / and", and "and / or", it should be understood that in this application, the technical solution undoubtedly includes technical solutions that are all connected by "logical and", and undoubtedly includes technical solutions that are all connected by "logical or". For example, "A and / or B" includes three parallel solutions of A, B and A+B. For example, the technical solution of "A, and / or, B, and / or, C, and / or, D" includes any one of A, B, C, and D (that is, the technical solution that is all connected by "logical OR"), and also includes any and all combinations of A, B, C, and D, that is, the combination of any two or any three of A, B, C, and D, and also includes the combination of four of A, B, C, and D (that is, the technical solution that is all connected by "logical AND").

[0029] In the present application, "plurality", "multiple", "multiple times", "multiples", etc., unless otherwise specified, refer to a number greater than 2 or equal to 2. For example, "one or more" means one or greater than or equal to two.

[0030] As used herein, "combination thereof", "any combination thereof", "any combination thereof" etc. include all suitable combinations of any two or more of the listed items.

[0031] Herein, the “suitable” mentioned in “suitable combination”, “suitable method”, “any suitable method”, etc., shall be based on the ability to implement the technical solution of this application, solve the technical problems of this application, and achieve the expected technical effects of this application.

[0032] Herein, “preferred”, “better”, “more preferred” and “suitable” are merely used to describe implementation methods or examples with better effects, and it should be understood that they do not constitute limitations on the scope of protection of this application.

[0033] In the present application, "further", "further", "particularly" and the like are used for descriptive purposes to indicate differences in content, but should not be construed as limiting the scope of protection of the present application.

[0034] In this application, "optionally", "optional", and "optional" mean optional or dispensable, that is, any one of the two parallel schemes of "yes" or "no". If multiple "options" appear in a technical solution, unless otherwise specified and there is no contradiction or mutual restriction, each "optional" is independent.

[0035] In the present application, the terms "first", "second", "third", "fourth", etc. in "the first aspect", "the second aspect", "the third aspect", "the fourth aspect", etc. are used only for descriptive purposes and cannot be understood as indicating or implying relative importance or quantity, nor can they be understood as implicitly indicating the importance or quantity of the indicated technical features. Moreover, "first", "second", "third", "fourth", etc. only serve the purpose of non-exhaustive enumeration and description, and it should be understood that they do not constitute a closed limitation on quantity.

[0036] In the present application, the technical features described in an open manner include closed technical solutions composed of the listed features, and also include open technical solutions containing the listed features.

[0037] In the present application, when it comes to numerical intervals (i.e., numerical ranges), unless otherwise specified, the optional numerical distribution is considered continuous within the above numerical interval, and includes the two numerical endpoints (i.e., the minimum and maximum values) of the numerical range, and each numerical value between the two numerical endpoints. Unless otherwise specified, when the numerical interval only refers to the integers within the numerical interval, it includes the two endpoint integers of the numerical range, and each integer between the two endpoints. In this article, it is equivalent to directly listing each integer, such as t is an integer selected from 1 to 10, indicating that t is any integer selected from the integer group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. In addition, when multiple ranges are provided to describe features or characteristics, these ranges can be combined. In other words, unless otherwise specified, the ranges disclosed herein should be understood to include any and all sub-ranges included therein.

[0038] The temperature parameters in this application, unless otherwise specified, are allowed to be either constant temperature treatment or to vary within a certain temperature range. It should be understood that the constant temperature treatment allows the temperature to fluctuate within the accuracy range controlled by the instrument. Fluctuations within the range of ±5°C, ±4°C, ±3°C, ±2°C, and ±1°C are allowed.

[0039] In the present application, % (w / w) and wt% both represent weight percentage, % (v / v) refers to volume percentage, and % (w / v) refers to mass volume percentage.

[0040] The term "MDPAO1 (Milk-Derived Peptide Associated with Obesity 1)" refers to an obesity-associated peptide derived from human breast milk.

[0041] Breast milk is rich in various nutrients. In addition to providing essential nutrients for the growth and development of offspring, the peptides rich in breast milk have great potential in regulating offspring's adipose tissue and reducing obesity.

[0042] On the one hand, the present application provides a breast milk-derived polypeptide, whose amino acid sequence is shown in SEQ ID NO.1.

[0043] IPFFDPQIPKLTDLEN (SEQ ID NO.1)

[0044] The breast milk-derived polypeptide provided in the present application has an amino acid sequence as shown in IPFFDPQIPKLTDLEN. The breast milk-derived polypeptide has significant effects in enhancing thermogenesis in brown adipose tissue of lactating mice, reducing the body weight of obese mice mediated by a high-fat diet and improving glucose metabolism, enhancing mitochondrial quality and reducing mitochondrial autophagy. The breast milk-derived polypeptide and its pharmaceutical composition have good application prospects in the preparation of drugs for preventing and treating childhood obesity.

[0045] Optionally, the breast milk-derived polypeptide of the present application is synthesized by biosynthesis.

[0046] On the other hand, the present application provides a pharmaceutical composition comprising the breast milk-derived polypeptide.

[0047] In a specific example, the drug includes the human milk-derived polypeptide and pharmaceutically acceptable excipients.

[0048] Optionally, the concentration of the breast milk-derived polypeptide that exerts biological activity is 8 mg / kg to 12 mg / kg.

[0049] The present application also provides the use of the breast milk-derived polypeptide or the pharmaceutical composition in the preparation of a drug for preventing and treating childhood obesity.

[0050] In this application, "prevention" and "prevention and / or treatment" have the same meaning and can be used interchangeably. In the present invention, "prevention" includes prevention, treatment, adjuvant therapy and other aspects. As used herein, "prevention" means to alleviate, delay the progression, attenuate, prevent, or maintain an existing disease or condition. "Prevention" also includes curing one or more symptoms of a disease or condition, preventing its development, or alleviating it to a certain extent.

[0051] One or more embodiments of the present application also provide a drug, which includes the polypeptide and pharmaceutically acceptable excipients.

[0052] In this application, "drug" includes any agent, compound, composition or mixture that provides a pharmacological effect in vivo or in vitro, and often provides a beneficial effect. The scope of the pharmacological effect of the "drug" in vivo is not particularly limited, and it can be a systemic effect or only a local effect. The activity of the "drug" is not particularly limited, and it can be an active substance that can interact with other substances, or it can be an inert substance that does not interact.

[0053] As used herein, "pharmaceutically acceptable" refers to those ligands, materials, compositions, and / or dosage forms that are suitable for administration to a patient within the scope of sound medical judgment and commensurate with a reasonable benefit / risk ratio.

[0054] In a specific example, the childhood obesity is secondary obesity.

[0055] Secondary obesity refers to obesity caused by other known diseases or specific reasons, which is different from primary obesity (also called simple obesity), which is usually related to genetics, diet and exercise habits. Secondary obesity may be caused by a variety of factors, including but not limited to endocrine disorders, drug side effects, neuropsychiatric factors, etc.

[0056] In a specific example, the pharmaceutical composition and pharmaceutically acceptable excipients are prepared together into a preparation.

[0057] In this application, "pharmaceutically acceptable excipient" refers to a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material. As used herein, the phrase "pharmaceutically acceptable excipient" includes buffers, sterile water for injection, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are compatible with drug administration. Each carrier must be "pharmaceutically acceptable" in the sense of being compatible with the other ingredients in the formulation and not harmful to the patient. Suitable examples include, but are not limited to: (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch, potato starch and substituted or unsubstituted β-cyclodextrins; (3) cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and suppository waxes; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn starch, etc. Rice oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerol, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffers, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) phosphate buffered saline; and (21) other nontoxic compatible substances used in pharmaceutical formulations.

[0058] In the present application, "excipients" include but are not limited to mannitol, sorbitol, sodium metabisulfite, sodium bisulfite, sodium thiosulfate, cysteine ​​hydrochloride, thioglycolic acid, methionine, vitamin C, disodium ethylenediaminetetraacetic acid (disodium EDTA), calcium sodium EDTA, carbonates, acetates, phosphates or aqueous solutions of monovalent alkali metals, hydrochloric acid, acetic acid, sulfuric acid, phosphoric acid, amino acids, sodium chloride, potassium chloride, sodium lactate, xylitol, maltose, glucose, fructose, oligofructose, dextran, glycine, starch, sucrose, dextrin (such as maltodextrin), lactose, mannitol, silicon derivatives, cellulose and its derivatives, alginate, gelatin, polyvinyl pyrrolidone, glycerol, Tween 80, agar, calcium carbonate, calcium bicarbonate, surfactants, polyethylene glycol, cyclodextrin, phospholipid materials, kaolin, talc, calcium stearate, and magnesium stearate.

[0059] One or more embodiments of the present application also provide a method for preventing and treating childhood obesity, comprising administering a therapeutically effective amount of the human milk-derived polypeptide of the present application to a subject.

[0060] In some embodiments, the administration of the drug includes, but is not limited to, oral, rectal, parenteral (intravenous, intramuscular or subcutaneous) injection, and topical administration, inhalation.

[0061] In some of these embodiments, the drug may be administered orally, by enema, or parenterally.

[0062] In some of these embodiments, the drug administration cycle can be intermittent administration, periodic administration, continuous administration or long-term administration.

[0063] Solid dosage forms for oral administration may include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active ingredient is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose, and acacia; (c) humectants, for example, glycerol; (d) disintegrants, for example, agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) solubilizers, for example, paraffin; (f) absorption accelerators, for example, quaternary ammonium compounds; (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents.

[0064] Solid dosage forms such as tablets, pills, capsules, pills and granules can be prepared using coatings and shell materials, such as enteric coatings and other materials known in the art. They may contain opacifiers, and the release of the active ingredient in such compositions can be delayed in a certain part of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes. If necessary, the active ingredient can also be formed into microcapsules with one or more of the above-mentioned excipients.

[0065] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active ingredient, the liquid dosage form may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, specifically, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide and oils, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or mixtures of these substances. In addition to these inert diluents, the composition may also contain adjuvants, such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and spices. For example, the suspension may contain a suspending agent, specifically, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and dehydrated sorbitan esters, microcrystalline cellulose, aluminum methylate and agar or mixtures of these substances.

[0066] Compositions for parenteral injection may include physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous or non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.

[0067] Dosage forms for topical administration include ointments, powders, patches, sprays and inhalants, which are prepared by mixing the active ingredient with pharmaceutically acceptable excipients and any preservatives, buffers, or propellants that may be required under sterile conditions.

[0068] It is to be understood that the drugs of the embodiments of the present invention can be added with different pharmaceutically acceptable carriers to prepare suitable clinical dosage forms, which include but are not limited to the dosage forms described above.

[0069] In the present application, "subject" refers to a patient who takes a polypeptide or a pharmaceutical composition.

[0070] In the present application, "subject" is an animal, preferably a mammal, more preferably a human, and the subject includes but is not limited to consumers of health products and patients with diseases, disorders and / or symptoms. The subject in the present invention is preferably a mammal. The term "mammal" mainly refers to warm-blooded vertebrate mammals, including but not limited to: such as cats, dogs, rabbits, bears, foxes, wolves, monkeys, deer, mice (such as rats, mice), pigs, cattle, sheep, horses, humans, etc., preferably primates, more preferably humans.

[0071] In some embodiments, the drug is suitable for humans or other mammals. In the present invention, "other mammals" are not humans, and non-limiting examples of "other mammals" include cats, dogs, rabbits, bears, foxes, wolves, monkeys, deer, mice (such as rats, mice), pigs, cattle, sheep, horses, etc., and further include mice or rats; non-limiting examples of "other mammals" include primates.

[0072] In some embodiments, the subject is a mammal.

[0073] In some embodiments, the subject is a human, rat, or mouse.

[0074] The embodiments of the present application will be described in detail below in conjunction with examples. It should be understood that these examples are only used to illustrate the present application and are not intended to limit the scope of the present application. The experimental methods for which specific conditions are not specified in the following examples are preferably referred to the guidance provided in the present application, and can also be based on the experimental manual or normal conditions in this area, can also be based on the conditions recommended by the manufacturer, or refer to experimental methods known in the art.

[0075] In the following specific embodiments, the measured parameters of raw material components may have slight deviations within the range of weighing accuracy unless otherwise specified. For temperature and time parameters, acceptable deviations caused by instrument test accuracy or operation accuracy are allowed.

[0076] It should be understood that in the various embodiments of the present application, the size of the serial numbers of the above-mentioned processes does not mean the order of execution. The execution order of each process should be determined by its function and internal logic, and should not constitute any limitation on the implementation process of the embodiments of the present application.

[0077] Example 1

[0078] It is to be understood that the MDPAO1 mentioned below is the maternal polypeptide shown in SEQ ID NO.1 of the present application.

[0079] 1. MDPAO1 (Milk-Derived Peptide Associated with Obesity 1) enhances thermogenesis in brown adipose tissue of lactating mice

[0080] Two-week-old mice were randomly divided into two groups: normal breast milk feeding group (Ctrl group) and breast milk peptide intervention group (MDPAO1 group), with 6 mice in each group, and the experiment was repeated 3 times. Ctrl group: 2-week-old mice were fed normally by their mothers. MDPAO1 group: 2-week-old mice were fed normally by their mothers and MDPAO1 intervention was performed. 10 mg / kg MDPAO1 was injected subcutaneously in the scapular area once a day for 7 days. The overall metabolic level was monitored on the 6th day, and the temperature of the interscapular area was measured 1 hour after cold stimulation on the 7th day. The euthanized mice were removed and brown adipose tissue was removed for hematoxylin-eosin (H&E) staining and immunohistochemistry.

[0081] Results from Figure 1 As shown in the figure, 2-week mice were used as the research objects. After MDPAO1 intervention, the temperature of the interscapular area of ​​the MDPAO1 group mice increased after cold stimulation, and the energy consumption and oxygen consumption increased compared with the Ctrl group. In addition, compared with the Ctrl group, H&E staining showed that the size of brown adipocytes in the MDPAO1 group was smaller, and immunohistochemistry results showed that the expression of UCP1 in brown adipose tissue in the MDPAO1 group increased.

[0082] 2. MDPAO1 reduces body weight and improves glucose metabolism in obese mice induced by a high-fat diet

[0083] Six-week-old mice were randomly divided into three groups: control group (Chow diet), high-fat diet group (HFD group), high-fat diet and MDPAO1 intervention group (HFD+MDPAO1 group), with 6 mice in each group, and the experiment was repeated 3 times. Chow diet group: mice were fed normal diet for 6 weeks. HFD group: mice were fed diet containing 60% fat for 6 weeks. HFD+MDPAO1 group: mice were fed diet containing 60% fat and MDPAO1 intervention was performed at the same time for 6 weeks. 10 mg / kg MDPAO1 was injected subcutaneously in the scapular area twice a week for 10 weeks. The whole experiment lasted for 10 weeks. During this period, the body weight, food intake, and water intake of mice in each group were weighed every 3 days. Glucose tolerance test (GTT) and insulin tolerance test (ITT) were performed on the 49th day and 56th day, respectively. Mice were euthanized at week 10 to remove brown adipose tissue, which was measured and imaged with a ruler, and then examined by transmission electron microscopy.

[0084] Results from Figure 2As shown, 6-week-old mice were used as research subjects, and an obese mouse model was constructed using a 60% high-fat diet. The HFD group had a significant increase in body weight compared with the Chow diet group, decreased glucose tolerance and insulin resistance, and decreased brown adipose tissue weight. After MDPAO1 intervention, the body weight of high-fat diet-mediated obese mice was significantly reduced, glucose tolerance and insulin resistance were significantly improved, and the weight of brown adipose tissue increased. In addition, there was no difference in food intake and water intake between the groups. Transmission electron microscopy results showed that after MDPAO1 intervention, the mitochondrial morphology of brown adipose tissue returned to normal.

[0085] 3. MDPAO enhances mitochondrial quality

[0086] Studies have reported that reduced mitochondrial autophagy and increased mitochondrial mass improve the thermogenic function of brown fat. This application further verifies whether MDPAO can enhance mitochondrial mass and reduce mitochondrial autophagy.

[0087] Primary brown adipocytes were used as the research object, and the growth medium was prepared: DMEM / 12 medium, 20% fetal bovine serum, 1% double antibody (penicillin, streptomycin);

[0088] Induction medium: DMEM / 12 medium, 10% fetal bovine serum, 1% double antibody (penicillin, streptomycin), 16.5 μg / mL insulin, 0.25 μM dexamethasone, 0.5 mM IBMX, 2 μM rosiglitazone;

[0089] Differentiation medium: DMEM / 12 medium, 10% fetal bovine serum, 1% double antibody (penicillin, streptomycin), 1 μg / mL insulin, 10 nM T3.

[0090] Primary brown adipocytes were seeded in 6-well plates at 2 × 10 6 cells at 37°C and 5% CO 2 When the incubator culture reached 100% cell density, the induction medium was replaced and cultured for 72 hours. After 72 hours, the differentiation medium was replaced and cultured for 6 days until differentiation and maturity. MDPAO1 (100 μM) was added to the differentiated and mature primary brown adipocytes for 24 hours, and the expression levels of mitochondrial oxidative phosphorylation (OXPHOS) proteins CI-NDUF88, CII-SDHB, CIII-UQCRC2, CIV-MTCO2, and CV-ATP5A were detected.

[0091] The results are as follows Figure 3As shown, differentiated mature primary brown adipocytes were used as the research object, and MDPAO1 was added to the cells for 24 hours. After MDPAO1 treatment, the expression of mitochondrial oxidative phosphorylation proteins CI-NDUF88, CII-SDHB, CIII-UQCRC2, CIV-MTCO2, and CV-ATP5A increased.

[0092] 4. MDPAO1 reduces mitochondrial autophagy

[0093] Primary brown adipocytes were used as the research object, and the growth medium was prepared: DMEM / 12 medium, 20% fetal bovine serum, 1% double antibody (penicillin, streptomycin);

[0094] Induction medium: DMEM / 12 medium, 10% fetal bovine serum, 1% double antibody (penicillin, streptomycin), 16.5 μg / mL insulin, 0.25 μM dexamethasone, 0.5 mM IBMX, 2 μM rosiglitazone;

[0095] Differentiation medium: DMEM / 12 medium, 10% fetal bovine serum, 1% double antibody (penicillin, streptomycin), 1 μg / mL insulin, 10 nM T3.

[0096] Primary brown adipocytes were seeded in 6-well plates at 2 × 10 6 cells at 37°C and 5% CO 2 When the incubator culture reached 100% cell density, the induction medium was replaced and cultured for 72 hours. After 72 hours, the differentiation medium was replaced and cultured for 6 days until differentiation and maturity. MDPAO1 (100 μM) was added to the differentiated and mature primary brown adipocytes for 24 hours, and mt-Keima was observed under a confocal fluorescence microscope.

[0097] Results from Figure 4 As shown, differentiated and mature primary brown adipocytes were used as the research object, and MDPAO1 was added to the cells for 24 hours. After MDPAO1 treatment, the fluorescence signal excited by the 561nm (red light) channel weakened, indicating that the degree of mitochondrial autophagy was reduced.

[0098] The above-described embodiments only express several implementation methods of the present application, which is convenient for understanding the technical solution of the present application in detail, but it cannot be understood as a limitation on the scope of protection of the patent application. It should be pointed out that for ordinary technicians in this field, without departing from the concept of the present application, several deformations and improvements can be made, which all belong to the protection scope of the present application. In addition, it should be understood that after reading the above-mentioned teaching content of the present application, the technicians in this field can make various changes or modifications to the present application, and the equivalent forms obtained also fall within the protection scope of the present application. It should also be understood that the technical solutions obtained by the technicians in this field through logical analysis, reasoning or limited experiments on the basis of the technical solutions provided in the present application are all within the protection scope of the claims attached to the present application. Therefore, the protection scope of the patent of the present application shall be based on the content of the attached claims, and the description and drawings can be used to explain the content of the claims.

Claims

1. Use of a breast milk-derived polypeptide in the preparation of a drug for preventing and treating childhood obesity, characterized in that: Its amino acid sequence is shown in SEQ ID NO.1; The concentration of the breast milk-derived polypeptide at which biological activity is exerted is 8 mg / kg to 12 mg / kg.

2. The use according to claim 1, characterized in that: The childhood obesity includes secondary obesity.

3. The use according to any one of claims 1 to 2, characterized in that: The dosage form of the drug includes one or more of tablets, capsules, granules, oral solutions and injections.

4. The use according to any one of claims 1 to 2, characterized in that: The subject of the drug is a mammal.

5. The use according to any one of claims 1 to 2, characterized in that: The subjects of the drug are primates.

6. The use according to claim 5, characterized in that: The subject of the drug is a human.

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