Galacto-oligosaccharide and derivative and application thereof in medicines or health products for improving mitochondrial function and preventing and treating insulin resistance related diseases
阅读说明：本技术 半乳寡糖及衍生物和在作为改善线粒体功能防治胰岛素抵抗相关疾病药物或保健品中的应用 (Galacto-oligosaccharide and derivative and application thereof in medicines or health products for improving mitochondrial function and preventing and treating insulin resistance related diseases ) 是由 于广利 王学良 郝杰杰 蔡超 于 2019-09-25 设计创作，主要内容包括：本发明属于海洋药物领域,具体涉及一种含有D-半乳糖和L-半乳糖的寡糖及其衍生物和在作为改善线粒体功能防治胰岛素抵抗相关疾病药物或保健品中的应用。以含有D-/L-半乳糖的红藻多糖为原料,经物理法、化学法、生物酶法或上述方法的任意组合进行降解,制备获得不同聚合度的半乳寡糖及其衍生物,其分子骨架中含有D-半乳糖和L-半乳糖及其衍生物。本发明产品的原料来源于红藻多糖,具有资源丰富、制备工艺简单、安全性高,靶点明确和易于产业化等优点,用于改善线粒体功能,在防治胰岛素抵抗新药及在降血糖、降血脂和降血压等特医食品的开发领域,具有广阔的应用前景。(The invention belongs to the field of marine medicines, and particularly relates to oligosaccharide containing D-galactose and L-galactose, derivatives thereof and application of the oligosaccharide and the derivatives thereof in medicines or health products for improving mitochondrial functions and preventing and treating insulin resistance-related diseases. The preparation method comprises the steps of taking red algae polysaccharide containing D-/L-galactose as a raw material, and degrading by a physical method, a chemical method, a biological enzyme method or any combination of the methods to prepare galacto-oligosaccharides with different polymerization degrees and derivatives thereof, wherein the molecular skeleton of the galacto-oligosaccharides contains D-galactose, L-galactose and derivatives thereof. The raw materials of the product are derived from the red algae polysaccharide, the product has the advantages of rich resources, simple preparation process, high safety, clear target spot, easy industrialization and the like, is used for improving the function of mitochondria, and has wide application prospect in the development fields of new medicines for preventing and treating insulin resistance and special medical foods for reducing blood sugar, blood fat, blood pressure and the like.)
1. Galacto-oligosaccharide and derivatives thereof, characterized in that the structural general formula of the oligosaccharide is as follows:
wherein R is-H or-SO3Na，n＝0～30；
2. A process for preparing galacto-oligosaccharide and its derivatives as claimed in claim 1, wherein the compounds prepared from red algae polysaccharide rich in D-/L-galactose and its derivatives by one or more of physical degradation, chemical degradation and enzymatic degradation contain both β -1, 3-D-galactose (D-Gal) and α -1, 4-L-galactose (L-Gal) residues or both D-Gal and α -1,4-L-3, 6-lacto (L-AnG) residues; the hydroxyl groups at C6 of the D-Gal and L-Gal sugar residues contain varying degrees of sulfation (Gal6S) modification; the non-reducing end of the prepared oligosaccharide is Gal, Gal6S or AnG, the reducing end is Gal or sugar alcohol (Gal-OH) and sugar acid (Gal-OOH) or AnG sugar alcohol (AnG-OH), or Gal6S and sugar alcohol (Gal6S-OH) and sugar acid (Gal 6S-OOH).
3. The process for preparing galacto-oligosaccharides and derivatives thereof according to claim 2, wherein the galacto-oligosaccharides and derivatives thereof are prepared by the following process:
dissolving agarose in hot water at 60 ℃, preparing 10mg/mL solution by using buffer solution, placing the solution in a water bath kettle at 30 ℃, adding beta-agarase (CAS #37288-57-6) and stirring for degradation for 4 hours, cooling and centrifuging, collecting supernatant, adding 2 times volume of 95% medical ethanol to the solution at 4 ℃ overnight, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, dialyzing and desalting by using a 200Da dialysis bag, performing rotary evaporation, concentrating and freeze drying to obtain a new agaro-oligosaccharide mixture, and further reducing the new agaro-oligosaccharide mixture by using sodium borohydride to obtain new agaro-oligosaccharide alcohol or oxidizing the new agaro-oligosaccharide acid by using a Benedict reagent; or dissolving agarose in hot water at 60 ℃, preparing 10mg/mL solution by using 0.1M dilute hydrochloric acid, stirring and degrading at 80 ℃ for 0.5 hour, cooling, neutralizing by using 2M NaOH aqueous solution, centrifuging, collecting supernatant, adding 2 times volume of 95% medical ethanol at 4 ℃ overnight, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, dialyzing and desalting by using a 200Da dialysis bag, performing rotary evaporation, concentrating, freeze-drying to obtain an oligosaccharide mixture, and further reducing by using sodium borohydride to obtain agaropectin, or oxidizing by using a Benedict reagent to obtain agaropectin; or preparing the sulfur agar into 10mg/mL aqueous solution by using 0.1M dilute sulfuric acid, heating to 60 ℃, stirring and degrading for 1.5 hours, cooling, neutralizing by using 2M NaOH aqueous solution, centrifuging, collecting supernatant, adding 3 times volume of 95% medical ethanol to the mixture at 4 ℃, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, performing dialysis desalination by using a 200Da dialysis bag, performing rotary evaporation, concentration and freeze drying to obtain an oligosaccharide mixture, and then further performing reduction by using sodium borohydride to obtain sulfur agar oligosaccharide alcohol, or oxidizing by using a Benedict reagent to obtain sulfur agar oligosaccharide acid; or preparing the porphyra gel into 10mg/mL aqueous solution by using 0.1M dilute sulfuric acid, heating to 80 ℃, stirring and degrading for 2 hours, cooling, neutralizing by using 2M NaOH aqueous solution, centrifugally collecting supernatant, adding 3 times volume of 95% medical ethanol to the temperature of 4 ℃ overnight, centrifugally collecting supernatant, carrying out rotary evaporation to remove ethanol, carrying out dialysis desalination by using a 200Da dialysis bag, carrying out rotary evaporation, concentration and freeze drying to obtain an oligosaccharide mixture, and further reducing by using sodium borohydride to obtain porphyra gel oligosaccharide alcohol, or oxidizing by using a Benedict reagent to obtain the porphyra gel oligosaccharide acid.
4. The use of the galacto-oligosaccharides and derivatives thereof according to claim 1 as drugs for improving mitochondrial function and preventing and treating insulin resistance-related diseases, wherein the galacto-oligosaccharides and derivatives thereof having these structural characteristics can target mitochondria and regulate and protect the functions thereof, and are used as drugs or health products for preventing and treating insulin resistance-related diseases.
5. Use of galacto-oligosaccharides and derivatives thereof according to claim 4 as drugs for improving mitochondrial function and preventing and treating diseases associated with insulin resistance, characterized in that the galacto-oligosaccharides and derivatives thereof target to mitochondrial binding and modulate its and downstream signaling pathway function.
6. The use of galacto-oligosaccharides and derivatives thereof according to claim 4 as a medicament for improving mitochondrial function and preventing and treating diseases associated with insulin resistance, wherein the galacto-oligosaccharides and derivatives thereof significantly reduce lipid accumulation and are useful for the preparation of medicaments for alleviating insulin resistance, for combating type 2 diabetes, for combating metabolic syndrome, for combating steatohepatitis, for combating hyperlipidaemia, for protecting the liver, for reducing blood glucose or for reducing blood lipid.
7. The use of galacto-oligosaccharides and derivatives thereof according to claim 4 as a medicament for improving mitochondrial function and preventing insulin resistance related diseases, characterized in that the galacto-oligosaccharides and derivatives thereof are used in health products for anti-diabetes, anti-fatty liver, liver protection, blood glucose or blood lipid lowering; or in beverages, beer, dietary supplements, or in combination with other antidiabetic agents, or in combination with hypolipidemic agents; or a compound preparation containing the galacto-oligosaccharide and the derivative thereof; or the derivative prepared by taking the galactooligosaccharide and the derivative thereof as mother nucleus is used in drugs, functional foods or biological products for resisting diabetes, fatty liver, insulin resistance, metabolic syndrome and hyperlipidemia.
8. The use of galacto-oligosaccharides and derivatives thereof according to claim 4 as a medicament for improving mitochondrial function in the prevention and treatment of insulin resistance related diseases, characterized in that the galacto-oligosaccharides and derivatives thereof form a built formulation with metformin, dapagliflozin, canagliflozin or acarbose related clinical drugs.
Technical Field
The invention belongs to the field of marine medicines, and particularly relates to oligosaccharide containing D-galactose and L-galactose, derivatives thereof and application of the oligosaccharide and the derivatives thereof in medicines or health products for improving mitochondrial functions and preventing and treating insulin resistance-related diseases.
Background
Researches show that long-term abnormal blood sugar, blood fat and blood pressure can cause damage to organs of the whole body to further cause the function reduction of the organs, and finally cause diseases such as hyperglycemia, hyperlipidemia, hypertension and the like. Mitochondria are important energy metabolism organelles for cell life activities, for example, dysfunction of mitochondria can lead to generation and development of various metabolic diseases, for example, Shulman proposes that the dysfunction of mitochondria is an important mechanism of insulin resistance, and the idea is supported by a plurality of experimental results. Mitochondrial dysfunction exists in insulin resistance states including obesity,
Research shows that the seaweed polysaccharide and oligosaccharide have the functions of resisting oxidation, reducing blood sugar, reducing blood fat, resisting inflammation, enhancing immunity and the like. A great deal of research is carried out in the field by the team, and agar oligosaccharide (publication No. CN105168232A) is found to have the activity of reducing blood fat and the like, fucoidan sulfate has the activity of inhibiting alpha-glycosidase (publication No. CN103288978A), alginate oligosaccharide and derivatives thereof have the activity of improving insulin resistance and reducing blood sugar (publication No. CN101649004A, publication No. CN101691410A) and the like, but no report that oligosaccharide containing D-galactose and L-galactose and derivatives thereof have the function of improving mitochondria in a targeted manner to prevent and treat insulin resistance related diseases is found. The galactan derived from red algae mainly comprises carrageenan series, agar series and laver gum series, wherein polysaccharide and oligosaccharide of the carrageenan series are both composed of D-galactose and sulfate derivatives thereof (publication No. CN1513880A, publication No. CN101012249A, publication No. CN101279991A), while polysaccharide and oligosaccharide of the agar and laver gum series are composed of D-galactose and L-galactose sugar residues and sulfate derivatives thereof, the difference between agar and laver gum is that the former contains more L-AnG, and the latter contains more 6-sulfuric acid-L-Gal, but the sugar compositions are different in physicochemical properties and biological functions. Although carrageenan and oligosaccharide thereof have antiviral activity as a spray (publication No. CN102516323A, publication No. CN104546895A), oral administration has certain potential safety hazard (Shang Q., Toxicol Let., 2017,279:87-95), and agar, porphyran polysaccharide and oligosaccharide have high safety when oral administration, and are high-quality raw materials for developing medicaments and functional foods. The preparation and application of oligosaccharides are the key points of research and development in recent years because the solubility of galactopolysaccharides is poor and the structural sequence is not clear, so that the quality is difficult to control. In the aspect of agar oligosaccharide preparation technology, mainly acid method, enzyme method and chemical method degradation are adopted, but different methods can obtain oligosaccharides with different structures and activities, for example, acid method degradation can obtain odd agar oligosaccharides (publication number CN1513860A), enzyme method degradation can obtain new agar oligosaccharides (publication number CN 102827899A; publication number CN109576328A), reducing acid degradation can obtain even sugar alcohol (publication number CN100999537A), and free radical degradation can prepare mixed agar oligosaccharides (publication number CN 109400756A); the porphyra gel oligosaccharide and the like can be obtained by acid degradation (LiuY., et al, Mar drugs.2018,16(3) pii: E82) or enzyme degradation (Zhang Y., et al, J.Agr.food chem.2019,67, 9307-9313). The invention further carries out directional reduction and oxidation reaction on various prepared oligosaccharides on the basis of the existing degradation technology to obtain oligosaccharide derivatives with different structures and sequences and sugar alcohol or sugar acid structures at the reducing ends, and experiments prove that the oligosaccharides and the derivatives thereof have the function of remarkably improving the mitochondrial function, and can be used for preparing medicaments for preventing and treating insulin resistance and
Disclosure of Invention
The invention aims to provide an application of galactooligosaccharide and derivatives thereof in medicines for improving mitochondrial function and preventing and treating insulin resistance related diseases, and provides a series of galactooligosaccharides and derivatives thereof obtained from marine red algae polysaccharide, and application of galactooligosaccharides and derivatives thereof in improving mitochondrial function and preventing and treating insulin resistance related diseases.
In order to achieve the purpose, the invention adopts the following technical scheme:
a galacto-oligosaccharide and its derivatives, the oligosaccharide has the following structural formula:
wherein R is-H or-SO3Na，n＝0～30；
The preparation method of the galacto-oligosaccharide and the derivative thereof takes red algae polysaccharide rich in D-/L-galactose and the derivative thereof as a raw material, and prepares oligosaccharides and derivatives thereof with different polymerization degrees by one or the combination of more than two degradation methods of physical degradation, chemical degradation and enzymatic degradation, and the prepared compound structure simultaneously contains beta-1, 3-D-galactose (D-Gal) residues and alpha-1, 4-L-galactose (L-Gal) residues or simultaneously contains D-Gal and alpha-1, 4-L-3, 6-diether galactose (L-AnG) residues; the hydroxyl groups at C6 of the D-Gal and L-Gal sugar residues contain varying degrees of sulfation (Gal6S) modification; the non-reducing end of the prepared oligosaccharide is Gal, Gal6S or AnG, the reducing end is Gal or sugar alcohol (Gal-OH) and sugar acid (Gal-OOH) or AnG sugar alcohol (AnG-OH), or Gal6S and sugar alcohol (Gal6S-OH) and sugar acid (Gal 6S-OOH).
The preparation method of the galactooligosaccharide and the derivative thereof adopts the following preparation processes:
dissolving agarose in hot water at 60 ℃, preparing 10mg/mL solution by using buffer solution, placing the solution in a water bath kettle at 30 ℃, adding beta-agarase (CAS #37288-57-6) and stirring for degradation for 4 hours, cooling and centrifuging, collecting supernatant, adding 2 times volume of 95% medical ethanol to the solution at 4 ℃ overnight, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, dialyzing and desalting by using a 200Da dialysis bag, performing rotary evaporation, concentrating and freeze drying to obtain a new agaro-oligosaccharide mixture, and further reducing the new agaro-oligosaccharide mixture by using sodium borohydride to obtain new agaro-oligosaccharide alcohol or oxidizing the new agaro-oligosaccharide acid by using a Benedict reagent; or dissolving agarose in hot water at 60 ℃, preparing 10mg/mL solution by using 0.1M dilute hydrochloric acid, stirring and degrading at 80 ℃ for 0.5 hour, cooling, neutralizing by using 2M NaOH aqueous solution, centrifuging, collecting supernatant, adding 2 times volume of 95% medical ethanol at 4 ℃ overnight, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, dialyzing and desalting by using a 200Da dialysis bag, performing rotary evaporation, concentrating, freeze-drying to obtain an oligosaccharide mixture, and further reducing by using sodium borohydride to obtain agaropectin, or oxidizing by using a Benedict reagent to obtain agaropectin; or preparing the sulfur agar into 10mg/mL aqueous solution by using 0.1M dilute sulfuric acid, heating to 60 ℃, stirring and degrading for 1.5 hours, cooling, neutralizing by using 2M NaOH aqueous solution, centrifuging, collecting supernatant, adding 3 times volume of 95% medical ethanol to the mixture at 4 ℃, centrifuging, collecting supernatant, performing rotary evaporation to remove ethanol, performing dialysis desalination by using a 200Da dialysis bag, performing rotary evaporation, concentration and freeze drying to obtain an oligosaccharide mixture, and then further performing reduction by using sodium borohydride to obtain sulfur agar oligosaccharide alcohol, or oxidizing by using a Benedict reagent to obtain sulfur agar oligosaccharide acid; or preparing the porphyra gel into 10mg/mL aqueous solution by using 0.1M dilute sulfuric acid, heating to 80 ℃, stirring and degrading for 2 hours, cooling, neutralizing by using 2M NaOH aqueous solution, centrifugally collecting supernatant, adding 3 times volume of 95% medical ethanol to the temperature of 4 ℃ overnight, centrifugally collecting supernatant, carrying out rotary evaporation to remove ethanol, carrying out dialysis desalination by using a 200Da dialysis bag, carrying out rotary evaporation, concentration and freeze drying to obtain an oligosaccharide mixture, and further reducing by using sodium borohydride to obtain porphyra gel oligosaccharide alcohol, or oxidizing by using a Benedict reagent to obtain the porphyra gel oligosaccharide acid.
The galacto-oligosaccharide and the derivative thereof can be used as medicines for improving mitochondrial function and preventing and treating insulin resistance related diseases, and the galacto-oligosaccharide and the derivative thereof with the structural characteristics can target mitochondria and regulate and protect the functions of the mitochondria and can be used as medicines or health care products for preventing and treating insulin resistance related diseases.
The galacto-oligosaccharide and the derivative thereof can be used as a medicine for improving the mitochondrial function and preventing and treating insulin resistance related diseases, and can be combined with mitochondria in a targeted manner and regulate the functions of the mitochondria and downstream signal paths.
The galacto-oligosaccharide and the derivative thereof can obviously reduce lipid accumulation and are used for preparing medicines for relieving insulin resistance, resisting
The galacto-oligosaccharide and the derivative thereof are applied to medicines for improving mitochondrial function and preventing and treating insulin resistance related diseases, and the galacto-oligosaccharide and the derivative thereof are used for health care products for resisting diabetes, resisting fatty liver, protecting liver and reducing blood sugar or blood fat; or in beverages, beer, dietary supplements, or in combination with other antidiabetic agents, or in combination with hypolipidemic agents; or a compound preparation containing the galacto-oligosaccharide and the derivative thereof; or the derivative prepared by taking the galactooligosaccharide and the derivative thereof as mother nucleus is used in drugs, functional foods or biological products for resisting diabetes, fatty liver, insulin resistance, metabolic syndrome and hyperlipidemia.
The galacto-oligosaccharide and the derivative thereof are applied to the medicines for improving the mitochondrial function and preventing and treating the diseases related to insulin resistance, and the galacto-oligosaccharide and the derivative thereof and the metformin, dapagliflozin, canagliflozin or acarbose related clinical medicines form a compound preparation.
The invention has the advantages and beneficial effects that:
1. the oligosaccharide containing D-and L-galactose residues and the derivative thereof can target mitochondria and relieve insulin resistance by regulating the functions of the mitochondria.
2. The oligosaccharide containing D-and L-galactose residues and the derivative thereof have obvious effect of reducing blood fat, and can be used for preventing and treating fatty liver and hyperlipidemia.
3. The raw materials of the product are derived from marine polysaccharide, and the product has the advantages of rich resources, simple preparation process, good product stability, easy industrialization, safety, effectiveness and the like, is used for improving the functions of mitochondria, and has wide development and application prospects in the development fields of new medicines for preventing and treating insulin resistance and special medical foods for reducing blood sugar, blood fat, blood pressure and the like.
4. The oligosaccharide has the functions of inhibiting hyperglycemia, lipid accumulation, insulin resistance and mitochondrial dysfunction caused by high-fat diet.
5. The invention adopts HepG2 insulin resistance cell model induced and constructed by sodium Palmitate (PA) to evaluate the prepared series of oligosaccharides with the related functions of relieving insulin resistance and the like. Research results show that the oligosaccharide containing D-galactose residues and L-galactose residues and derivatives thereof can be combined with mitochondria in a targeted manner to regulate the functions of the oligosaccharide, and has the effects of obviously reducing triglyceride and cholesterol, increasing glucose consumption, improving oxidative stress state and obviously increasing insulin sensitivity, thereby improving insulin resistance and treating diabetes, fatty liver and hyperlipidemia.
Drawings
FIG. 1 shows high resolution mass spectra and structural formulas of neoagarotetraose (a), sugar alcohol (b) and sugar acid (c). In the figure, the abscissa m/z represents the mass-to-charge ratio, and the ordinate Relative Absundance represents the Relative Abundance.
Fig. 2 is a diagram of the localization of the sulfoagaro-oligosaccharide mixture to mitochondria. In the figure, λex578nm represents the excitation wavelength of 578nm, λex488nm represents the excitation wavelength of 488nm, SAOs represents the fluorescein isothiocyanate labeled sulfoagaro oligosaccharide treatment group, and Control represents the blank Control group.
FIG. 3 is a graph of SAOs promoting mitochondrial proliferation of HepG2 cells. SAOs stands for sulfoagaro oligosaccharides. P<0.05, compared to the FFA-free BSA treated group; # P<0.05, compared to the PA treatment group; @ P<0.05, SAOs high dose group compared to SAOs low dose group. Wherein, the picture (a) is a protein immunoblotting picture of SIRT1, SIRT1 represents
FIG. 4 is a diagram of the mitochondrial function of SAOs in regulating insulin resistance in HepG2 cells. P <0.05, compared to FFA-free BSA treated group; # P <0.05, compared to PA treated group; @ P <0.05, SAOs high dose group compared to SAOs low dose group. Wherein, the picture (a) is a cell mitochondrial Complex activity picture, the abscissa Complex I, Complex III and Complex IV respectively represent mitochondrial complexes I, III and IV, and the ordinate Complex enzymic activity represents the corresponding mitochondrial Complex activity; (b) the graph is a plot of oxygen consumption by respiration of cells with the abscissa Time representing Time (minutes) and the ordinate Fluorescence intensity (a.u.) representing Fluorescence intensity; (c) the figure is an intracellular ADP/ATP graph, the ADP/ATP ratio represents the content ratio of adenosine diphosphate to adenosine triphosphate, the ordinate Fold change represents the change of the ADP/ATP ratio of each group compared with a control group, PA (0.2mM) represents the table 0.2mM sodium palmitate, and SAOs (mu M) represents the treatment of different concentrations of SAOs; (d) the graph is a content Ratio graph of JC-1 fluorescent probe polymer and monomer, the ordinate Ratio of JC-1 polymer/monomer represents the content Ratio of the fluorescent probe JC-1 polymer to the fluorescent probe monomer, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (mu M) represents different concentrations of SAOs treatment.
FIG. 5 is a graph of SAOs improving oxidative stress status of insulin resistant HepG2 cells. P <0.05, compared to FFA-free BSA treated group; # P <0.05, compared to PA treated group; @ P <0.05, SAOs high dose group compared to SAOs low dose group. Wherein, the graph (a) is a graph of the Content of various oxidation indexes, the abscissa SOD (U/mg), CAT (U/mg) and MDA (nmol/mg) respectively represent superoxide dismutase, catalase and malonaldehyde, and the ordinate Content represents the Content of various oxidation indexes; (b) FIG. is a graph showing ROS active oxygen content, Control represents a blank Control group, Model represents a 0.2mM sodium palmitate-treated group, SAOs-L represents a 0.2mM sodium palmitate-treated group and a 10. mu.M SAOs-treated group, and SAOs-H represents a 0.2mM sodium palmitate-treated group and a 50. mu.M SAOs-treated group; (c) the graph shows the phosphorylation level of JNK protein, JNK represents c-Jun amino-terminal kinase, pJNK represents phosphorylated c-Jun amino-terminal kinase, beta-actin represents beta-actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (mu M) represents different concentrations of SAOs treatment; (d) the graph shows phosphorylation levels of c-Jun protein, wherein c-Jun is a nuclear proto-oncogene, p-c-Jun represents a phosphorylated c-Jun protein, β -actin represents β -actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (. mu.M) represents treatments with different concentrations of SAOs.
FIG. 6 is a graph of SAOs increasing insulin resistance to insulin sensitivity in HepG2 cells. P < 0.05; NS, nonstationary significant difference indicates no statistically significant difference. Wherein, the ordinate glucose consumption form of control represents the change in glucose consumption of each group compared to the control group, PA (0.2mM) represents 0.2mM sodium palmitate, SAOs (. mu.M) represents the treatment with different concentrations of SAOs, and Insulin (1. mu.M) represents the treatment with 1. mu.M added Insulin.
FIG. 7 is a graph of SAOs activating the HepG2 cell insulin signaling pathway and regulating sugar metabolism. P <0.05, compared to FFA-freeBSA treated group; # P <0.05, compared to PA treated group; @ P <0.05, SAOs high dose group compared to SAOs low dose group. Wherein, the graph (a) is an IRS-1 protein phosphorylation level graph, IRS-1 represents insulin receptor substrate 1, pIRS-1 represents phosphorylated insulin receptor substrate 1, beta-actin represents beta-actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (mu M) represents different concentrations of SAOs treatment; (b) the figure is a graph of the phosphorylation levels of AKT protein, AKT representing protein kinase B, pAKT representing phosphorylated protein kinase B, β -actin representing β -actin, PA (0.2mM) representing 0.2mM sodium palmitate, SAOs (μ M) representing treatments with different concentrations of SAOs; (c) the graph is a GSK-3 beta protein phosphorylation level graph, GSK-3 beta represents glycogen synthase kinase-3, pGSK-3 beta represents phosphorylated glycogen synthase kinase-3, beta-actin represents beta-actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (mu M) represents different concentrations of SAOs treatment; (d) the figure is a graph of GS protein phosphorylation levels, GS for glycogen synthase, pGS for phosphorylated glycogen synthase, β -actin for β -actin, PA (0.2mM) for 0.2mM sodium palmitate, and SAOs (. mu.M) for different concentrations of SAOs treatment.
FIG. 8 is a graph of SAOs decreasing lipid synthesis and accumulation in insulin resistant HepG2 cells. P <0.05, compared to FFA-freeBSA treated group; # P <0.05, compared to PA treated group; @ P <0.05, SAOs high dose group compared to SAOs low dose group. Wherein, the picture (a) is a picture of protein phosphorylation level of HMGCR, the HMGCR represents hydroxymethyl glutaryl coenzyme A reductase, the pHMGCR represents phosphorylated hydroxymethyl glutaryl coenzyme A reductase, the beta-actin represents beta-actin, the PA (0.2mM) represents 0.2mM sodium palmitate, and the SAOs (mu M) represents SAOs treatment with different concentrations; (b) FIG. is a graph showing the phosphorylation levels of ACC protein, ACC represents acetyl-CoA carboxylase, pACC represents phosphorylated acetyl-CoA carboxylase, β -actin represents β -actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (. mu.M) represents treatments with different concentrations of SAOs; (c) the figure shows the expression levels of the protein precursor SREBP-1C and the mature body, SREBP-1C represents the sterol regulatory element binding protein-1C, beta-actin represents beta-actin, PA (0.2mM) represents 0.2mM sodium palmitate, and SAOs (mu M) represents the treatment of different concentrations of SAOs; (d) the graph shows the intracellular TC and TG contents, TC represents total cholesterol, TG represents triglyceride, and Content (μ g/mg protein) represents the TC and TG contents per mg cellular protein.
Detailed Description
The technical solution of the present invention will be further described with reference to specific examples.
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