Application of rifampicin in preparation of medicine for regulating lipid metabolism

Abstract

The invention discloses an application of rifampicin in preparation of a medicine for regulating lipid metabolism. The invention discloses a new medical application of rifampicin by using a mouse model fed by 3T3-L1 fat cells and high fat diet (HFD). Rifampicin can increase the expression of Sirt6 in vivo and in vitro. In addition, rifampicin effectively blocks differentiation of 3T3-L1 preadipocytes and reduces lipid accumulation by inhibiting lipid generation in adipocytes and enhancing lipid decomposition. In the mechanism level, rifampicin significantly inhibits the expression of adipogenesis genes and proteins, and enhances the expression of genes and proteins of lipolysis, energy consumption and white adipose tissue browning related pathways. In conclusion, data of the invention shows that rifampicin is a potential new treatment strategy for regulating lipid metabolism.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and particularly relates to the application of rifampicin in preparing medicines for regulating lipid metabolism. Background Art

[0002] Adipose tissue plays a vital role in metabolic regulation by regulating the balance of energy storage and consumption at the organ and system levels. Based on its function, adipose tissue can be divided into white adipose tissue (WAT) and brown adipose tissue (BAT): white adipocytes are mainly responsible for storing fat to reserve energy; while brown adipocytes burn fat to produce heat through uncoupling protein 1 (UCP1)-mediated uncoupled respiration. Recent studies have also discovered another type of brown-like adipocyte present in white fat - beige / light brown cells. There is evidence that white adipocytes can transdifferentiate into beige / light brown adipocytes, a process known as "browning." Therefore, promoting WAT browning and activating BAT are considered to be potential strategies for regulating lipid metabolism.

[0003] Sirtuin 6 (Sirt6) is a NAD-binding protein that is primarily localized in the cell nucleus. + Sirt6, a member of the sirtuin family, plays a key role in regulating lipid and glucose metabolism. Studies have shown that overexpression of Sirt6 extends lifespan in mice and protects against high-fat diet (HFD)-induced metabolic stress. Conversely, liver-specific deletion of Sirt6 exacerbates HFD-induced hepatic steatosis / hepatitis, inflammation, insulin resistance, and impaired ketogenesis. Sirt6 overexpression inhibits adipogenesis by downregulating the expression of key lipogenic factors: carbohydrate response element binding protein (ChREBP), stearoyl-CoA desaturase-1 (SCD1), fatty acid binding protein 4 (FABP4), peroxisome proliferator-activated receptor γ (PPARγ), and diacylglycerol acyltransferase 1 (DGAT1). Sirt6 deficiency downregulates the expression of the browning-related gene UCP1, as well as lipolytic genes such as adipose triglyceride lipase ATGL and hormone-sensitive lipase HSL. Therefore, targeted activation of Sirt6 represents a novel therapeutic strategy for regulating lipid metabolism and related metabolic disorders.

[0004] Rifampicin (RFP) is a semisynthetic macrocyclic broad-spectrum antibiotic known for its ability to inhibit bacterial proliferation, but its potential in regulating lipid metabolism has not been explored. Summary of the Invention

[0005] The purpose of the present invention is to provide a new pharmaceutical use of rifampicin or a pharmaceutically acceptable salt thereof.

[0006] The new pharmaceutical use of rifampicin or its pharmaceutically acceptable salt provided by the present invention is its use in preparing products for regulating lipid metabolism.

[0007] Furthermore, the product has at least one of the following effects:

[0008] 1) Increased expression of Sirtuin 6 (Sirt6);

[0009] 2) inhibiting lipid production;

[0010] 3) Promote lipid decomposition;

[0011] 4) Promote browning of fat cells;

[0012] 5) inhibit lipid accumulation in adipose tissue;

[0013] 6) Inhibit adipocyte hypertrophy and abnormal adipocyte differentiation;

[0014] 7) Inhibit the expression of adipogenic genes and proteins;

[0015] 8) Increased expression of lipolytic genes and proteins;

[0016] 9) Reduce serum triglyceride (TG) and non-esterified fatty acid (NEFA) levels;

[0017] 10) Inhibit triglyceride accumulation in 3T3-L1 differentiated cells;

[0018] 11) Inhibit the expression of adipogenic genes and proteins in 3T3-L1 differentiated cells;

[0019] 12) Upregulation of lipolytic protein levels in 3T3-L1 cells differentiated into adipogenic cells;

[0020] 13) Reduce body fat percentage.

[0021] Furthermore, the increasing the expression of Sirtuin 6 (Sirt6) is increasing the expression of Sirtuin 6 (Sirt6) in adipose tissue.

[0022] Furthermore, the adipose tissue includes white adipose tissue (WAT) and brown adipose tissue (BAT).

[0023] Furthermore, the white adipose tissue (WAT) may include inguinal white fat (iWAT), epididymal white fat (eWAT) and perirenal white fat (rWAT).

[0024] Furthermore, the lipogenesis gene is selected from at least one of the following: PPARγ1 / 2, FABP4, Resistin, ChREBP, FAS, SCD1, SREBP1-C, ACC1 / 2, and DGAT1 / 2.

[0025] Furthermore, the lipolytic gene is selected from at least one of the following: ATGL, p-ATGL, HSL and p-HSL.

[0026] The present invention also protects the use of rifampicin or a pharmaceutically acceptable salt thereof in the preparation of a Sirt6 inducer and / or a Sirt6 activator.

[0027] Furthermore, the Sirt6 inducer or Sirt6 activator can target and activate Sirt6 in adipocytes.

[0028] The present invention also protects other uses of rifampicin or its pharmaceutically acceptable salts, which are at least one of the following a)-j):

[0029] a) Use in the preparation of a product for increasing the expression of Sirtuin 6 (Sirt6);

[0030] b) use in the preparation of products that inhibit lipid formation;

[0031] c) application in the preparation of products promoting lipid decomposition;

[0032] d) application in the preparation of products promoting browning of fat cells;

[0033] e) use in the preparation of a product for inhibiting lipid accumulation in adipose tissue;

[0034] f) use in the preparation of products for inhibiting adipocyte hypertrophy and abnormal adipocyte differentiation;

[0035] g) Use in the preparation of products for inhibiting the expression of adipogenic genes and proteins;

[0036] h) use in the preparation of products for increasing the expression of lipolytic genes and proteins;

[0037] i) Use in the preparation of a product for lowering serum triglyceride (TG) and non-esterified fatty acid (NEFA) levels;

[0038] j) Application in the preparation of products for reducing body fat percentage.

[0039] In the above application, "pharmaceutically acceptable salts of rifampicin" refer to salts that, within the scope of sound medical judgment, are suitable for use in contact with the tissues of humans and lower animals without causing excessive toxicity, irritation, allergic reactions, etc., and are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts of rifampicin are well known in the art and include, but are not limited to, sodium salts, potassium salts, hydrochlorides, nitrates, sulfates, bisulfates, phosphates, hydrogenphosphates, methanesulfonates, acetates, and the like.

[0040] In the above applications, the product is a medicine or a pharmaceutical preparation.

[0041] When preparing medicines or pharmaceutical preparations, rifampicin or a pharmaceutically acceptable salt thereof can be used as one of the active ingredients or as the only active ingredient.

[0042] In the above applications, a pharmaceutically acceptable carrier material may be added when preparing the drug.

[0043] The carrier materials include, but are not limited to, water-soluble carrier materials (such as polyethylene glycol, polyvinyl pyrrolidone, organic acids, etc.), poorly soluble carrier materials (such as ethyl cellulose, cholesterol stearate, etc.), and enteric carrier materials (such as cellulose acetate phthalate and carboxymethyl ethyl cellulose, etc.). These materials can be used to produce a variety of dosage forms, including but not limited to tablets, capsules, dripping pills, pills, powders, solutions, suspensions, emulsions, granules, liposomes, buccal tablets, lyophilized powder injections, etc. These can be conventional preparations, sustained-release preparations, controlled-release preparations, and various microparticle delivery systems.

[0044] In order to prepare the unit dosage form into tablets, various carriers known in the art can be widely used. Examples of carriers include diluents and absorbents, such as starch, dextrin, calcium sulfate, lactose, mannitol, sucrose, sodium chloride, glucose, urea, calcium carbonate, kaolin, microcrystalline cellulose, aluminum silicate, etc.; wetting agents and binders, such as water, glycerol, polyethylene glycol, ethanol, propanol, starch slurry, dextrin, syrup, honey, glucose solution, acacia slurry, gelatin slurry, sodium carboxymethylcellulose, shellac, methylcellulose, potassium phosphate, polyvinyl pyrrolidone, etc.; disintegrants , such as dried starch, alginate, agar powder, brown seaweed starch, sodium bicarbonate with citric acid, calcium carbonate, polyoxyethylene, sorbitan fatty acid esters, sodium lauryl sulfate, methylcellulose, ethylcellulose, etc.; disintegration inhibitors, such as sucrose, tristearin, cocoa butter, hydrogenated oil, etc.; absorption enhancers, such as quaternary ammonium salts, sodium lauryl sulfate, etc.; lubricants, such as talc, silicon dioxide, corn starch, stearate, boric acid, liquid paraffin, polyethylene glycol, etc. Tablets can also be further prepared as coated tablets, such as sugar-coated tablets, film-coated tablets, enteric-coated tablets, or bilayer tablets and multilayer tablets. To prepare the unit dosage form into a pill, a wide variety of carriers known in the art can be used. Examples of carriers include diluents and absorbents such as glucose, lactose, starch, cocoa butter, hydrogenated vegetable oils, polyvinylpyrrolidone, kaolin, and talc; binders such as gum arabic, tragacanth, gelatin, ethanol, honey, liquid sugar, rice paste, or flour paste; and disintegrants such as agar powder, dried starch, alginates, sodium lauryl sulfate, methylcellulose, and ethylcellulose. To prepare unit dosage forms as suppositories, a wide variety of carriers known in the art can be used. Examples of carriers include polyethylene glycol, lecithin, cocoa butter, higher alcohols, higher alcohol esters, gelatin, and semi-synthetic glycerides. To prepare unit dosage forms as injectable preparations, such as solutions, emulsions, lyophilized powder injections, and suspensions, all diluents commonly used in the art can be used, including water, ethanol, polyethylene glycol, 1,3-propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene isostearyl alcohol, and polyoxyethylene sorbitan fatty acid esters. In addition, to prepare an isotonic injection, an appropriate amount of sodium chloride, glucose, or glycerol may be added to the injectable formulation. Conventional solubilizers, buffers, and pH adjusters may also be added. Furthermore, colorants, preservatives, fragrances, flavoring agents, sweeteners, or other materials may be added to the pharmaceutical formulation as needed. The above dosage forms can be administered via injection, including subcutaneous, intravenous, intramuscular, and intracavitary injections; respiratory tract administration, such as through the nasal cavity; and mucosal administration.

[0045] When using rifampicin or a pharmaceutically acceptable salt thereof provided by the present invention to regulate lipid metabolism, an effective amount of rifampicin or a pharmaceutically acceptable salt thereof is administered to the subject.

[0046] The dosage and method of use of rifampicin or a pharmaceutically acceptable salt thereof of the present invention depend on many factors, including the patient's age, weight, gender, natural health, nutritional status, activity strength of the compound, time of administration, metabolic rate, severity of the disease, and the subjective judgment of the treating physician.

[0047] In the present invention, the term "effective amount" refers to a dose that can achieve the treatment, prevention, alleviation and / or relief of the diseases or conditions described in the present invention in a subject.

[0048] In the present invention, the term "subject" may refer to a patient or other animal that receives the product of the present invention to treat, prevent, alleviate and / or relieve the disease or condition of the present invention, especially a mammal, such as a human, dog, monkey, cow, horse, etc.

[0049] The present invention uses a mouse model of 3T3-L1 adipocytes and a high-fat diet (HFD) to reveal a new medical use for rifampicin. Rifampicin can increase the expression of Sirt6 in vivo and in vitro. In addition, rifampicin effectively blocks the differentiation of 3T3-L1 preadipocytes and reduces lipid accumulation by inhibiting lipogenesis and enhancing lipolysis in adipocytes. At the mechanistic level, rifampicin significantly inhibits the expression of lipogenesis genes and proteins, and enhances the expression of genes and proteins in pathways related to lipolysis, energy consumption, and browning of white adipose tissue. In summary, rifampicin can be used as a potential new therapeutic strategy for regulating lipid metabolism. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1Rifampicin is a Sirt6 inducer and reduces triglyceride accumulation. (A) Luciferase activity in 3T3-L1 preadipocytes stably expressing the Sirt6 promoter (Sirt6-Luc 3T3-L1) after 18 hours of rifampicin treatment. (B and C) mRNA levels of adipogenic and lipolytic genes during 3T3-L1 differentiation after 3 days (B) and 9 days (C) of rifampicin treatment were assessed by qPCR analysis and presented as mean ± SEM (n = 5-6). (D) Protein levels of adipogenic and lipogenic genes were measured by Western blot analysis after 9 days of rifampicin treatment. (E) Quantified data were normalized to the internal control GAPDH. (F) Oil Red O staining of differentiated 3T3-L1 adipocytes treated with or without rifampicin (50 μM) for 48 hours. Bright-field microscopy images of differentiated cells (scale bar = 200 μm). (G) Relative quantification of triglyceride (TG) content. (H) Levels of lipolytic proteins were measured by Western blot analysis after 48 hours of rifampicin treatment in differentiated 3T3-L1 adipocytes. (I) Quantified data were normalized to the internal control GAPDH. Data are presented as mean ± SEM (n = 3) and statistically analyzed using a two-tailed unpaired t-test. *P < 0.05, **P < 0.01 relative to the control group.

[0051] Figure 2 Effects of rifampicin on 3T3-L1 cell viability. 3T3-L1 cells were treated with various concentrations of rifampicin for 24 hours (A), 48 hours (B), and 72 hours (C). Cell viability was measured using the MTT assay. Data are presented as mean ± SEM. Statistical analysis was performed using one-way ANOVA. **P < 0.01.

[0052] Figure 3 Oral administration of rifampicin prevents high-fat diet (HFD)-induced lipid metabolism abnormalities. C57BL / 6J mice were fed an HFD for 14 weeks and then treated with rifampicin (30 mg / kg, once daily) or vehicle by oral gavage for 6 weeks. An HFD was also administered during drug treatment. (A) Body fat content. (B) Changes in weight of white adipose tissue (WAT) depots, including inguinal WAT (iWAT), epididymal WAT (eWAT), retroperitoneal WAT (rWAT), and brown adipose tissue (BAT) fat pads (n = 8 or 10). (C) H&E staining of iWAT and eWAT depots. (D) Adipocyte size in iWAT and eWAT (n = 6). (E) Frequency distribution of adipocyte size in iWAT and eWAT (n = 6). Data are presented as mean ± SEM. Statistical analysis was performed using a two-tailed unpaired t-test. *P < 0.05, **P < 0.01 relative to the control group.

[0053] Figure 4Plasma analysis was performed on HFD-induced obese mice treated with rifampicin or vehicle. Plasma samples obtained from experimental animals were analyzed using the kit described in the main methods. (A) Plasma triglyceride levels. (B) Plasma non-esterified fatty acid levels. (C) Food intake. Data are presented as mean ± SEM (n = 1).

[0054] =8-10), and statistical analysis was performed using two-tailed unpaired t-test. *P<0.05, **P<0.01 relative to the control group.

[0055] Figure 5 Rifampicin prevents lipid accumulation and promotes browning in adipose tissue of HFD-induced obese mice. (A) mRNA levels of lipogenic and lipolytic genes in iWAT were assessed by qPCR analysis and are presented as mean ± SEM (n = 6-8), normalized to 36b4, and compared with the control group. (B) mRNA levels of lipogenic and lipolytic genes in iWAT were assessed by qPCR analysis and are presented as mean ± SEM (n = 6-8), normalized to 36b4, and compared with the control group. (C) mRNA levels of BAT, mitochondrial, and β-oxidation genes in iWAT were assessed by qPCR analysis and are presented as mean ± SEM (n = 6-8), normalized to 36b4, and compared with the control group. (D) Sirt6, lipogenic, lipolytic, and BAT protein levels were measured in iWAT by Western blot analysis. (E) Quantified data were normalized to the internal control GAPDH. Data are presented as mean ± SEM (n = 3), and statistical analysis was performed using a two-tailed unpaired t-test. *P<0.05, **P<0.01 relative to the control group.

[0056] Figure 6 Rifampicin prevents HFD-induced lipid metabolism abnormalities through the Sirt6-specific pathway. fl / fl ) and adipocyte-specific Sirt6 knockout mice (Sirt6 Adi- / -) were fed an HFD for 4 weeks and then continued on an HFD and received rifampicin (30 mg / kg once daily) or vehicle by oral gavage for 4 weeks. (A) Representative images of mice. (B) Body weight. (C) Body fat content. (D) Changes in weight of white adipose tissue (WAT) depots, including inguinal WAT (iWAT), epididymal WAT (eWAT), retroperitoneal WAT (rWAT), and brown adipose tissue (BAT) fat pads (n = 8 or 10). (E) H&E staining of iWAT, eWAT, and BAT depots. (F) Adipocyte size of iWAT, eWAT, and BAT (n = 6). (G) Frequency distribution of adipocyte size of iWAT, eWAT, and BAT (n = 6). Data are expressed as mean ± SEM. Statistical analysis was performed using two-way ANOVA. *, comparison within the same genotype; #, comparison between two genotypes. * or #, P < 0.05; ** or ##, P < 0.01.

[0057] Figure 7 To investigate the effect of Sirt6 on HFD-fed rats treated with rifampicin or vehicle fl / fl and Sirt6 Adi- / - Plasma analysis of mice. Plasma samples obtained from experimental animals were analyzed using the kits described in the main methods. (A) Plasma triglyceride levels. (B) Plasma non-esterified fatty acid levels. Data are presented as mean ± SEM (n = 8-10) and statistically analyzed using two-way ANOVA. *, comparison within the same genotype; #, comparison between two genotypes. * or #, P < 0.05; ** or ##, P < 0.01

[0058] Figure 8 Rifampicin alleviates lipid accumulation and promotes browning in adipose tissue through a Sirt6-dependent mechanism. (A) mRNA levels of lipogenic, (B) adipogenic, (C) lipolytic and BAT genes, and (D) mitochondrial and β-oxidation genes in iWAT were assessed by qPCR analysis and presented as mean ± SEM (n = 6-8), normalized to 36b4, and compared with the control group. (E) Protein levels of lipogenic, adipogenic, lipolytic, and BAT genes were measured in iWAT by Western blot analysis. (F) Quantified data were normalized to the internal control GAPDH. Data are presented as mean ± SEM (n = 3) and statistically analyzed using two-way ANOVA. *, comparison within the same genotype; #, comparison between two genotypes. * or #, P < 0.05; ** or ##, P < 0.01.

[0059] Figure 9Sirt6 is required for rifampicin to increase adipocyte lipolysis via ATGL. (AD) 3T3-L1 cells were infected with lenti-shSirt6 and then differentiated into adipocytes in the presence of a hormone cocktail (n = 6). (A) Oil Red O staining of differentiated 3T3-L1 adipocytes treated with or without rifampicin (50 μM). (B) Relative quantification of triglyceride (TG) content. (C) Analysis of free fatty acids (FFA) or (D) glycerol released in the presence of isoproterenol (10 μM), atglistatin (20 μM), and / or rifampicin for 2 hours. (E and F) Activation of Sirt6 from adipocytes ^fl / fl^ and *Sirt6 Adi- / - Primary WAT adipocytes were isolated from iWAT tissue of mice and then differentiated into adipocytes (n = 6). (E) Oil Red O staining of differentiated primary adipocytes treated with or without rifampicin (50 μM). (F) Relative quantification of TG content. (G and H) Sirt6 expression was measured in the presence of isoproterenol (10 μM), atglistatin (20 μM), and / or rifampicin (50 μM). ^fl / fl^ and *Sirt6 Adi- / - Lipolysis in mouse iWAT tissue for 2 hours (n = 6). (G) Analysis of FFA or (H) glycerol release over 2 hours. Data are presented as mean ± SEM. Statistical analysis was performed using two-way ANOVA. *, comparison within the same genotype; #, comparison between two genotypes. * or #, P < 0.05; ** or ##, P < 0.01.

[0060] Figure 10 Sirt6-deficient adipocytes inhibit lipolysis by ATGL. Lipolysis was performed for 2 hours in the presence of isoproterenol (10 μM) and / or atglistatin (20 μM) and / or rifampicin (50 μM). Basal free fatty acid (FFA) (A) or glycerol (B) release was measured. Basal FFA (C) or glycerol (D) release in the presence of isoproterenol (10 μM) and / or atglistatin (20 μM) and / or rifampicin (50 μM) was analyzed. Data are expressed as mean ± SEM (n = 6) and statistically analyzed using two-way ANOVA. DETAILED DESCRIPTION

[0061] The present invention will be further described in detail below in conjunction with specific embodiments. The examples provided are only for illustrating the present invention and are not intended to limit the scope of the present invention. The examples provided below can serve as a guide for further improvements by those skilled in the art and are not intended to limit the present invention in any way.

[0062] Unless otherwise specified, the experimental methods in the following examples are conventional methods and were performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials and reagents used in the following examples, unless otherwise specified, were all commercially available.

[0063] The abbreviations used in the present invention are as follows: ACC, acetyl-CoA carboxylase; ATGL, adipose triglyceride lipase; BAT, brown adipose tissue; Cytc, mitochondrial gene cytochrome c; eWAT, epididymal white adipose tissue; FABP4, fatty acid binding protein 4; FAS, fatty acid synthase; HFD, high-fat diet; HSL, hormone-sensitive lipase; iWAT, inguinal white adipose tissue; PGC1α, peroxisome proliferator-activated receptor gamma coactivator 1α; PPARγ, peroxisome proliferator-activated receptor gamma; Prdm16, PR domain-containing protein 16; rWAT, retroperitoneal white adipose tissue; UCP1, uncoupling protein 1; WAT, white adipose tissue.

[0064] Example 1: Pharmacodynamics of Rifampicin in Regulating Lipid Metabolism

[0065] 1. Materials and Methods

[0066] 1. Mice and Diet

[0067] The Institute of Laboratory Animals, Chinese Academy of Medical Sciences and Peking Union Medical College, successfully constructed Sirt6 gene conditional knockout (floxed) mice (Sirt6 fl / fl There are four transcripts of the Sirt6 gene. Based on its gene structure, the exon 3 to exon 8 region of transcript Sirt6-201 (ENSMUST00000042923.8) is recommended as the knockout region. This region contains 811bp of coding sequence. Knocking out this region will lead to loss of protein function. To obtain mice with Sirt6 knockout specific to adipocytes, floxed mice were mated with adipoq-Cre mice (JAX strain number: 028020) to produce a control group (Sirt6 fl / fl ) mice and mice with adipocyte-specific Sirt6 knockout (Sirt6 Adi- / -). C57BL / 6J mice used in some experiments were purchased from Beijing Huafukang Biotechnology Co., Ltd. Because estrogen regulates abdominal fat deposition and changes the biological characteristics of adipocytes, only male mice were used in all experiments. All mice received humane care and were kept in an environment with a temperature controlled at 20-22°C and a 12-hour light / dark cycle, with free access to drinking water and food. Male mice were used throughout the experiment and were kept in a specific pathogen-free (SPF) environment certified by the Institute of Medical Biotechnology, Peking Union Medical College. All animal experiments were approved by the Institutional Animal Care and Use Committee (Approval No.: IMB-20240216D506). Unless otherwise stated, all mice were fasted for 5-6 hours before euthanasia. The high-fat diet (HFD) containing 60% fat calories used in the experiment was purchased from Research Diets (Product No.: D12492).

[0068] 2. Chemicals

[0069] Rifampicin (RIF, catalog number #R3501), dexamethasone (DEX, catalog number #D4902), bovine serum albumin (BSA, catalog number #A3803), fetal bovine serum (FBS, catalog number F8687), dimethyl sulfoxide (DMSO, catalog number #D2438), 3-isobutyl-1-methylxanthine (IBMX, catalog number #I5879), insulin (catalog number #I9278), isoproterenol (catalog number #I5627), and atglistatin (catalog number #sml1075) were purchased from Sigma.

[0070] 3. Body composition analysis

[0071] Body fat content was measured by EchoMR02-025V (EchoMRI, NIUMAG, Suzhou, China).

[0072] 4. Cell culture

[0073] 3T3-L1 cells (ATCC, No. CL-173) were grown in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum and 1% penicillin and streptomycin.

[0074] After the cells were infected with the knockdown Sirt6 lentivirus (HYKY-220930015-DLV Shanghai Heyuan Biotechnology Co., Ltd.), a cell line with stable knockdown of Sirt6 was obtained by puromycin resistance screening. The stably knocked-down cell line was induced to differentiate. On day 1, the cells were induced to differentiate in DMEM containing 10% FBS, 0.5mM IBMX, 1μM DEX and 1.7μM insulin. On day 3, the differentiation medium was replaced with maintenance medium containing 10% FBS, 1μM DEX and 1.7μM insulin. The maintenance medium was replaced every 2 days until the cells were used for the experiment.

[0075] 5. Isolation and differentiation of stromal vascular fraction into adipocytes

[0076] From Sirt6 fl / fl and Sirt6 Adi- / - Primary preadipocytes were isolated from iWAT of mice and differentiated into mature adipocytes after some modifications. Briefly, the tissue was cut into small pieces, digested in KRBH buffer containing type I collagenase at 37°C with shaking for 1 hour, and the stromal vascular fraction (SVF) was separated from mature adipocytes by centrifugation. The SVF containing preadipocytes was precipitated, washed and resuspended in growth medium (DMEM / F-12 supplemented with 20% FBS, 1% antibiotics / antimycotics), filtered and plated. The growth medium was changed every other day. To induce differentiation, on day 1, cells were induced in induction medium (DMEM / F-12 supplemented with 10% FBS, 1% penicillin and streptomycin, 1 μg / mL insulin, 200 μM indomethacin, 500 μM IBMX, 1 μM dexamethasone and 5 μM rosiglitazone). On day 3, the induction medium was removed and differentiation medium (DMEM / F-12 supplemented with 10% FBS, 1% penicillin and streptomycin, 5 μg / mL insulin and 1 μM rosiglitazone) was added. The differentiation medium was changed every two days for 6 days until the cells were considered to be fully differentiated adipocytes.

[0077] 6. Fat decomposition assay

[0078] Lipolysis in WAT was measured as follows: Approximately 50-60 mg of inguinal fat pads or 3T3-L1 cells were incubated in 2% fatty acid-free BSA with or without 10 μM isoproterenol and / or 20 μM atglistatin at 37°C for 2 hours. After inactivation at 65°C for 10 minutes, glycerol release was measured using a Sigma kit (catalog #MAK117-1KT), and free fatty acids (FFAs) were measured using Leadman's reagent (catalog #NE7001). Basal lipolysis rates (0 h) and induced lipolysis rates (2 h) were normalized to protein content.

[0079] 7. RNA Extraction and Quantitative Real-Time PCR

[0080] Cells were washed with PBS and collected in Trizol reagent (QIAGEN, catalog #79306). Approximately 50 mg of tissue was homogenized in Trizol reagent. cDNA was synthesized from total RNA using the HiScript III RT SuperMix for qPCR kit (Vazyme, catalog #R323-01) according to the manufacturer's instructions. cDNA was diluted and Used on 480 Instrument II (Roche) mRNA levels were quantified by quantitative real-time PCR (qRT-PCR) using 480 SYBR Green I Master (Roche, Cat. #04707516001). Primer sequences are described in Table 1. Relative mRNA levels were quantified and normalized to Rplp0 (36b4).

[0081] Table 1 Primer sequences for QRT-PCR

[0082]

[0083] 8. Western Blot Analysis

[0084] Western blot analysis was performed using total tissue or cell lysates. Antibodies against hormone-sensitive lipase (HSL) (catalog #4107), phosphorylated HSL (p-HSL) (catalog #45804), adipose tissue triglyceride lipase (ATGL) (catalog #2439), and uncoupling protein 1 (UCP1) (catalog #14670) were purchased from Cell Signaling Technology (Boston, MA, USA). Antibody against phosphorylated adipose tissue triglyceride lipase (ATGL) (catalog #ab135093) was purchased from Abcam (Cambridge, UK). Anti-GAPDH antibody (catalog #ap0063, Bioworld) was purchased from Bioworld Technology (MN, USA). Anti-tubulin antibody was used at a dilution of 1:5000, and other primary antibodies were used at a dilution of 1:1000. Western blot images were obtained using ChemiDoc. TM Images were acquired using the MP Imaging System (Bio-Rad), and immunoblots were quantified using ImageJ.

[0085] 9. Histology, adipocyte size and number

[0086] WAT or BAT were fixed in 10% formalin and then embedded in optimal cutting temperature compound or paraffin. Hematoxylin and eosin (H&E) staining was performed using standard techniques. Adipocyte size and number were quantified using ImageJ.

[0087] 10. Oil Red O staining

[0088] After 9 and 14 days of differentiation of 3T3-L1 cells or primary adipocytes from preadipocytes, respectively, the differentiated adipocytes were washed twice with PBS and fixed in 10% formalin for 1 hour. They were then stained with Oil Red O working solution for 1 hour. Prior to microscopic analysis, the cells were washed four times with distilled water. The Oil Red O stain was eluted with 100% isopropanol (v / v) and quantified by measuring absorbance at 200 nm.

[0089] 11. In vitro promoter analysis

[0090] For in vitro promoter analysis, the Sirt6 promoter (-2000 to 24) (SEQ ID No. 1) was cloned into the H26561pSLenti-CMV-Luc2-3xFLAG-PGK-Puro-WPRE lentiviral vector between the MluI and EcoRI restriction sites to replace the CMV promoter, constructing the pSLenti-SIRT6Promoter-Luc2-3xFLAG-PGK-Puro-WPRE lentiviral vector (HYKY-220929064-DLV, H27138, Shanghai Heyuan Biotechnology Co., Ltd.). The shuttle vector (pSLenti-SIRT6Promoter-Luc2-3xFLAG-PGK-Puro-WPRE) was co-transfected with the packaging plasmids H1 (expressing gag / pol and the regulatory gene Rev) and H2 (expressing the envelope protein VSVG) into 293T cells, and the cell supernatant was collected and the virus was purified and collected by ultracentrifugation. qPCR was used to detect the number of exogenous DNA copies in the genome of 293T cells infected with the virus, and the titer was calculated to be 2.75E+8 TU / ml. The packaged lentivirus was used to infect 3T3L1 precursor cells, and 3T3-L1 preadipocyte clones stably overexpressing the Sirt6 promoter were selected by puromycin resistance. Cell clones were seeded at 50,000 cells / well in 96-well plates and cultured in 100 μL of DMEM medium for 12 hours. Cells were then treated with the indicated concentrations of rifampicin or vehicle (0.1% DMSO). After incubation for 18 hours, cells were washed with PBS (137 mM NaCl, 2.7 mM KCl, 4.3 mM Na2HPO4, 1.4 mM KH2PO4, pH 7.3), and luciferase activity was measured using a luciferase assay system (Promega, Madison, WI, USA) and expressed as relative luminescence units (RLUs).

[0091] 12. Cell Viability Analysis

[0092] 3T3-L1 cells were seeded at 5,000 cells per well in 96-well plates and treated with rifampicin (at concentrations of 1, 2, 5, 10, 20, 50, 100, and 200 μM) for 24, 48, and 72 hours. Cell viability was analyzed by the MTT assay.

[0093] 13. Blood biochemistry

[0094] For serum indicators, the levels of triglycerides (catalog number #TG8160) and non-esterified fatty acids (NEFA) (catalog number #NE7001) were measured according to the instructions of each test kit (Leedman, Beijing, China).

[0095] 14. Statistical analysis

[0096] Data are presented as mean ± standard error (SEM). Statistical significance was analyzed by unpaired Student's t-test or ANOVA (with post hoc Bonferroni test for multiple comparisons) using Prism software (GraphPad, CA). The number of samples per group ranged from 6 to 10. A value of P < 0.05 was designated as statistically significant. A single symbol (# or *) indicates P < 0.05, and two symbols indicate P < 0.01.

[0097] 2. Results

[0098] 1. Sirt6 inducer rifampicin inhibits triglyceride accumulation in 3T3-L1 cells

[0099] We cloned the Sirt6 promoter into the pGL4-Luc luciferase reporter vector and screened 3T3-L1 preadipocyte clones stably overexpressing the Sirt6 promoter to identify compounds that activate Sirt6 transcription. Among the candidate compounds, rifampicin significantly increased Sirt6 promoter activity. A dose-response curve for rifampicin-activated luciferase activity was determined in Sirt6-Luc 3T3-L1 cells, with an EC50 value of 17.38 μM, reaching a maximum of 822% at a concentration of 50 μM ( Figure 1 A). We incubated 3T3-L1 cells with rifampicin at concentrations ranging from 1 to 200 μM for 24, 48, and 72 hours and measured cytotoxicity using the MTT assay. The data showed that incubation of 3T3-L1 cells with rifampicin up to 100 μM for 24, 48, or 72 hours had no cytotoxic effect ( Figure 2 ). Therefore, 50 μM rifampicin was used in subsequent experiments.

[0100] To determine whether rifampicin affects lipid accumulation in 3T3-L1 cells, we incubated differentiated 3T3-L1 cells with rifampicin (50 μM). The results showed that rifampicin significantly increased Sirt6 expression ( Figure 1 C), and inhibited the expression of genes involved in adipogenesis (C / EBPα, PPARγ2) and lipogenesis (ChREBP, FAS, SCD1) after 3 and 9 days of incubation ( Figure 1 B and C). After 9 days of incubation with rifampicin, the protein levels of C / EBPα, PPARγ2, and ChREBP were also significantly decreased ( Figure 1 D and E). In addition, rifampicin inhibited the formation of lipid droplets and reduced lipid content in mature 3T3-L1 cells ( Figure 1F and G). According to previous studies, activation of Sirt6 enhances lipolysis of stored triglycerides (TG) in white and brown adipocytes. Therefore, we explored whether rifampicin regulates adipocyte lipolysis in mature 3T3-L1 cells. Rifampicin significantly upregulated Sirt6, p-ATGL (ser 406 phosphorylation) and p-HSL (ser 660 and Serine 563 The protein level of phosphorylated Figure 1 H and I). These results indicate that rifampicin inhibits adipogenesis and enhances lipolysis in 3T3-L1 adipocytes.

[0101] 2. Rifampicin improves lipid metabolism abnormalities and increases energy expenditure in HFD-induced mice

[0102] Next, we investigated whether rifampicin plays a role in regulating lipid metabolism in vivo. C57BL / 6 mice were randomly divided into two groups (control group and RFP group). Both groups of mice were fed HFD for 14 weeks. Then, the RFP group was gavaged with rifampicin (30 mg / kg) every day, and the control group was gavaged with an equal amount of 0.5% sodium carboxymethylcellulose every day for 6 weeks. HFD was also fed during drug treatment, and fat content was measured in the last week. We found that rifampicin reduced body fat percentage ( Figure 3 A).

[0103] We then examined whether rifampicin reduces body fat percentage by reducing WAT weight.After mice were sacrificed, adipose tissue from the inguinal, epididymal, retroperitoneal, and dorsal scapular regions was isolated and weighed.

[0104] In rifampicin-treated mice, the weights of inguinal WAT (iWAT), epididymal WAT (eWAT), retroperitoneal WAT (rWAT), and BAT were decreased ( Figure 3 B). Increased adipose tissue mass can result from either hypertrophy (increase in size of existing adipocytes) or hyperplasia (formation of new adipocytes through differentiation of resident precursor cells (called preadipocytes). Therefore, we further measured adipocyte size in adipose tissue of each group of mice. Hematoxylin and eosin (H&E) staining showed that adipocyte size was smaller in epididymal and inguinal adipose tissue of rifampicin-treated mice compared with that of control mice ( Figure 3 C). The adipocyte area of iWAT and eWAT in mice given rifampicin ( Figure 3 D) and cell size ( Figure 3 E) was significantly reduced. The data showed that rifampicin treatment reduced the conversion of preadipocytes to mature adipocytes and the subsequent adipocyte hypertrophy and proliferation. In addition, rifampicin reduced serum TG and non-esterified fatty acid (NEFA) levels ( Figure 4 A and B).

[0105] 3. Rifampicin prevents lipid accumulation in adipose tissue of HFD-fed mice and induces WAT browning

[0106] To understand the mechanism by which rifampicin regulates lipid metabolism, we further examined the expression of lipogenic and lipolytic genes.

[0107] Twenty C57BL / 6 mice were randomly divided into two groups (control and RFP) after 14 weeks of HFD feeding. The RFP group was then gavaged daily with rifampicin (30 mg / kg), while the control group was gavaged daily with an equivalent volume of 0.5% sodium carboxymethylcellulose. The HFD feeding continued. After 6 weeks, the mice were fasted for 6 hours and then sacrificed, with adipose tissue collected from various parts. Inguinal adipose tissue from both groups was lysed with TRIZOL, and mRNA was extracted. The mRNA was reverse-transcribed into cDNA using a kit (Nanjing Novozymes). Gene expression changes were detected using a QCPR instrument (Roche 480II).

[0108] In inguinal adipocytes, rifampicin significantly reduced the expression of adipogenic genes (PPARγ1 / 2, FABP4, Resistin) and lipogenic genes (ChREBP, FAS, SCD1, SREBP1-C, ACC1 / 2, DGAT1 / 2) ( Figure 5 A and B). Consistent with the changes in mRNA levels, the protein levels of adipocyte differentiation and adipogenesis markers were also significantly decreased by rifampicin ( Figure 5 B, D, and E). In contrast, rifampicin increased the expression of Sirt6 and the phosphorylation levels of the lipolytic proteins ATGL and HSL ( Figure 5 D and E). WAT is crucial for triglyceride storage, and browning of white adipocytes has a strong ability to increase lipid metabolism. We analyzed the expression of BAT, mitochondrial, and β-oxidation marker genes. Rifampicin significantly increased the mRNA levels of BAT genes (UCP1, Prdm16), mitochondrial genes (Cytc), and β-oxidation genes (HSP70, Mcad, Cpt1α). Figure 5 C). In addition, rifampicin promoted the UCP1 protein level in iWAT ( Figure 5 D and E). In conclusion, our data indicate that rifampicin promotes Sirt6 expression, adipocyte lipolysis, and iWAT browning, and inhibits TG accumulation, adipocyte hypertrophy, and abnormal adipocyte differentiation.

[0109] 4. Sirt6 KO adipocytes abolished the protective effect of rifampicin against HFD-induced lipid metabolism abnormalities

[0110] To further test our hypothesis that the effect of rifampicin on lipid regulation depends on the Sirt6 signaling pathway, we administered rifampicin to 10 Sirt6 fl / fl mice and 10 Sirt6 Adi- / - Mice were fed HFD for 4 weeks and then Sirt6 fl / fl Mice were randomly divided into two groups: one group (Sirt6 fl / fl +RFP group) were given rifampicin (30 mg / kg, once a day) by gavage, and the other group (Sirt6 fl / fl control group) were gavaged with a vehicle (0.5% sodium carboxymethylcellulose (CMC-Na)) for 4 weeks; similarly, Sirt6 Adi- / - The mice were randomly divided into two groups. One group (Sirt6 Adi- / - +RFP group) were given rifampicin (30 mg / kg, once a day) by gavage, and the other group (Sirt6 Adi- / - The control group) was given a vehicle (0.5% sodium carboxymethylcellulose (CMC-Na)) by gavage for 4 weeks. Adi- / - Rifampicin administration did not reduce Sirt6 expression in HFD-fed mice compared to control mice. Adi- / - The body weight and fat content of mice ( Figure 6 AC). Sirt6 in HFD-fed Adi- / - In mice, no beneficial effects of rifampicin were observed in reducing the weight of iWAT, eWAT, rWAT, and BAT depots ( Figure 6 D) Histological staining showed that rifampicin induced adipocyte size reduction in HFD-fed Sirt6 Adi- / - There is also very little in iWAT, eWAT and BAT of mice ( Figure 6 EG). In addition, rifampicin did not alter Sirt6 expression in HFD-fed mice. Adi- / - Serum TG and NEFA levels in mice ( Figure 7 A and B). These results demonstrate that Sirt6 is essential for rifampicin to regulate lipid metabolism.

[0111] 5. Fat-specific Sirt6 KO abolishes the effects of rifampicin on adipogenesis and lipolysis

[0112] We further examined the expression of lipogenic and lipolytic genes in iWAT. fl / fl and Sirt6 Adi- / - The inguinal fat tissue of the mice in the treatment group and the control group (a total of 4 groups) was lysed, and mRNA was extracted and reverse transcribed into cDNA using a kit (Nanjing Novozymes). The gene expression changes were detected by QCPR instrument (Roche480II). Adi- / -In mice, the expression of adipogenic genes (PPARγ2, FABP4) and lipogenic genes (ChREBP, FAS, SCD1, SREBP1-C, ACC1 / 2, DGAT1 / 2) was not affected by rifampicin ( Figure 8 A and B). Rifampicin also did not increase Sirt6 in HFD-fed mice Adi- / - mRNA levels of lipolysis genes (ATGL, HSL), BAT (UCP1, Prdm16), mitochondrial (Cytc) and β-oxidation (HSP70, Mcad, Cpt1α) marker genes in mice ( Figure 8 C and D). Consistent with the changes in gene expression, Sirt6 Adi- / - In mice, no beneficial effects of rifampicin were observed on protein levels of markers of adipocyte differentiation (PPARγ), adipogenesis (ChREBP), lipolysis (ATGL), and BAT (UCP1) ( Figure 8 E and F). Thus, these data suggest that rifampicin not only inhibits adipogenesis but also promotes adipocyte lipolysis and browning through a Sirt6-dependent mechanism.

[0113] Sirt6-deficient adipocytes abolish rifampicin-induced lipolysis by inhibiting ATGL

[0114] ATGL (adipose triglyceride lipase) and HSL (hormone-sensitive lipase) hydrolyze triglycerides and diglycerides into FFA and glycerol, respectively, and are the first and second steps in lipolysis in adipocytes. To better understand how rifampicin regulates lipolysis, we knocked down Sirt6 in preadipocyte 3T3L-1 cells, which can be differentiated into mature adipocytes in the presence of a hormone cocktail containing insulin and dexamethasone (DEX). Sirt6 knockdown and control differentiated 3T3-L1 cells were treated with or without rifampicin (50 μM) for 48 hours. Oil red staining showed that rifampicin could not inhibit the formation of lipid droplets in Sirt6 knockdown 3T3-L1 cells ( Figure 9 A and B). Isoproterenol, a β1 / β2 adrenergic receptor agonist, significantly stimulated the release of FFA and glycerol in control 3T3-L1 cells. Atglistatin, an ATGL inhibitor, significantly inhibited the release of FFA and glycerol in control 3T3-L1 cells. Rifampicin did not increase FFA in Sirt6 knockdown 3T3-L1 cells. Figure 9 C) and glycerol ( Figure 9 Under basal, unstimulated conditions, there was no change in FFA or glycerol release ( Figure 10 A and B).

[0115] We also investigated the effect of rifampicin on the Sirt6-derived Adi- / - Regulation of lipolysis in mouse primary iWAT adipocytes differentiated into adipogenic adipocytes by Sirt6 fl / fl and Sirt6 Adi- / - The inguinal white tissue of mice was hydrolyzed and separated into primary preadipocytes, which were induced to differentiate into adipocytes, and then stained with oil red and the triglyceride content was measured (see Materials and Methods 5, 10). Our data showed that in the absence of Sirt6, rifampicin did not inhibit lipid droplet formation ( Figure 9 E and F). Figure 9 As shown in G and H, rifampicin did not increase FFA and glycerol release in Sirt6-deficient iWAT adipocytes. Under basal, unstimulated conditions, there was no difference in FFA or glycerol release between the two genotypes ( Figure 10 C and 10D ). Taken together, our data suggest that rifampicin enhances lipolysis and reduces lipid accumulation via the Sirt6-ATGL pathway in adipocytes.

[0116] In this study, we utilized a Sirt6-specific promoter screening approach and adipocyte-specific Sirt6 knockout mice to demonstrate the potential therapeutic role of rifampicin in regulating lipid metabolism. Through virtual screening and in vitro activity assessment, we identified rifampicin as a potent Sirt6 activator and demonstrated its ability to inhibit adipocyte differentiation and lipid accumulation in 3T3-L1 cells. Adipogenic genes such as ChREBP, Fasn, and SREBP1-C are upregulated during adipocyte differentiation. Rifampicin inhibited the expression of the adipogenic gene PPARγ and the adipogenic gene ChREBP, leading to reduced triglyceride accumulation in 3T3-L1 cells. Furthermore, rifampicin reduced triglyceride content by upregulating the levels of lipolytic proteins (ATGL, p-ATGL, HSL, and p-HSL) in mature 3T3-L1 cells. Our studies reveal that rifampicin inhibits preadipocyte differentiation and triglyceride accumulation in 3T3-L1 cells by altering the expression of adipogenic and lipolytic genes.

[0117] We investigated the role of rifampicin in regulating lipid metabolism and its in vivo mechanisms. Oral administration of rifampicin resulted in a decrease in WAT fat depot weight and adipocyte size in subcutaneous WAT (iWAT) and visceral WAT (eWAT and rWAT) of HFD-fed obese mice. However, rifampicin failed to inhibit Sirt6 expression in HFD-fed obese mice. Adi- / -Increased body fat percentage and fat accumulation in mice. ChREBP is highly expressed in the liver and adipose tissue and plays a key role in regulating the expression of lipogenic genes, including acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), and stearoyl-CoA desaturase 1 (SCD1). In addition, ChREBP has been reported to be involved in lipogenesis in adipocytes. Previous studies have shown that in C57BL / 6J mice, hepatic Sirt6 deficiency reduces lipogenesis by inhibiting ChREBP. Our study found that rifampicin reduced the mRNA and protein levels of ChREBP in iWAT and 3T3-L1 cells of HFD-fed mice. Rifampicin-inhibited ChREBP in Sirt6 Adi- / - Sirt6 is eliminated in mice, confirming that Sirt6 is essential for rifampicin to reduce lipid accumulation through downregulation of ChREBP.

[0118] Lipolysis is the process by which triglycerides (TG) are broken down, releasing glycerol and FFA for energy expenditure. ATGL catalyzes the initial and rate-limiting step of lipolysis in adipocytes. Our studies further demonstrated that rifampicin increases ATGL activity by upregulating Sirt6 expression and promotes lipolysis through ATGL. Increased lipolysis in adipocytes may enhance FFA release and lipid accumulation in organs. However, our results showed that rifampicin also reduced plasma TG and NEFA levels in HFD-fed obese mice. Furthermore, we found that rifampicin-increased lipolysis was accompanied by WAT browning, as rifampicin increased the expression of a BAT marker (UCP1) in both in vitro and in vivo studies. BAT plays a key role in regulating TG levels and promoting plasma lipid clearance in rodents. Furthermore, WAT browning can promote UCP1-related mitochondrial and β-oxidation gene activation.

[0119] In conclusion, our studies demonstrate that the Sirt6 inducer rifampicin modulates lipid metabolism both in vitro and in vivo.

[0120] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be practiced over a wide range of parameters, concentrations, and conditions without departing from the spirit and scope of the present invention and without unnecessary experimentation. Although specific embodiments have been given herein, it should be understood that further modifications may be made to the present invention. In summary, this application is intended to encompass any variations, uses, or improvements to the present invention, including those made by conventional techniques known in the art that depart from the scope of the present invention. Applications of the essential features may be made within the scope of the following claims.

Claims

1. Use of rifampicin or a pharmaceutically acceptable salt thereof in the preparation of a product for regulating lipid metabolism.

2. The use according to claim 1, characterized in that: The product has at least one of the following effects: 1) Increase the expression of Sirt6; 2) inhibiting lipid production; 3) Promote lipid decomposition; 4) Promote browning of fat cells; 5) inhibit lipid accumulation in adipose tissue; 6) Inhibit adipocyte hypertrophy and abnormal adipocyte differentiation; 7) Inhibit the expression of adipogenic genes and proteins; 8) Increased expression of lipolytic genes and proteins; 9) Reduce serum triglyceride and non-esterified fatty acid levels; 10) Inhibit triglyceride accumulation in 3T3-L1 differentiated cells; 11) Inhibit the expression of adipogenic genes and proteins in 3T3-L1 differentiated cells; 12) Upregulation of lipolytic protein levels in 3T3-L1 cells differentiated into adipogenic cells; 13) Reduce body fat percentage.

3. The use according to claim 2, characterized in that: The increasing the expression of Sirt6 is to increase the expression of Sirt6 in adipose tissue.

4. The use according to claim 2, characterized in that: The adipose tissue includes white adipose tissue and brown adipose tissue; Furthermore, the white adipose tissue includes inguinal white fat, epididymal white fat and perirenal white fat.

5. The use according to claim 2, characterized in that: The lipogenesis gene is selected from at least one of the following: PPARγ1 / 2, FABP4, Resistin, ChREBP, FAS, SCD1, SREBP1-C, ACC1 / 2, and DGAT1 / 2.

6. The use according to claim 2, characterized in that: The lipolytic gene is selected from at least one of the following: ATGL, p-ATGL, HSL and p-HSL.

7. Use of rifampicin or a pharmaceutically acceptable salt thereof in the preparation of a Sirt6 inducer and / or a Sirt6 activator.

8. The use according to claim 7, characterized in that: The Sirt6 inducer or Sirt6 activator targets and activates Sirt6 in adipocytes.

9. Use of rifampicin or a pharmaceutically acceptable salt thereof, which is at least one of the following a) to j): a) Use in the preparation of a product for increasing the expression of Sirt6; b) use in the preparation of products that inhibit lipid formation; c) application in the preparation of products promoting lipid decomposition; d) application in the preparation of products promoting browning of fat cells; e) use in the preparation of a product for inhibiting lipid accumulation in adipose tissue; f) use in the preparation of products for inhibiting adipocyte hypertrophy and abnormal adipocyte differentiation; g) Use in the preparation of products for inhibiting the expression of adipogenic genes and proteins; h) use in the preparation of products for increasing the expression of lipolytic genes and proteins; i) use in the preparation of a product for reducing serum triglyceride and non-esterified fatty acid levels; j) Application in the preparation of products for reducing body fat percentage.

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