Effect Of Potato Resistant Starch On Insulin Resistance In HepG2 Cells

Objective:In this study,we cultured Escherichia coli to simulate intestinal digestion and fermentation of potato resistant starch（RS）,and used its end product short chain fatty acid（SCFA）to affect the insulin resistance HepG2 cell model.Preliminary exploration of possible molecular biological mechanisms for improving insulin resistance cell models,providing experimental basis for further development and utilization of potato RS to improve insulin resistance.Methods:（1）The potato RS was fermented by Escherichia coli,and the content of acetic acid,propionic acid and butyric acid in the product SCFA was determined by liquid chromatography tandem mass spectrometry.（2）Detect the effect of different concentrations of SCFA on the activity of HepG2 cells using CCK-8 method.（3）Establish a model of insulin resistance in HepG2 cells induced by high glucose and high insulin culture medium,and determine the optimal insulin concentration for establishing a model of insulin resistance in HepG2 cells.（4）The glucose consumption of HepG2 cells in insulin resistance model was measured by glucose hexokinase method.（5）Real time fluorescence quantitative PCR was used to detect the expression of GLUT2,GLUT4,AMPK,PI3K,and Akt m RNA in insulin resistant HepG2 cells.（6）Western blot（WB）was used to detect the expression of GLUT2,GLUT4,AMPK,PI3K,and Akt proteins in short chain fatty acid treated HepG2 insulin resistant cells.Results:（1）The content of short chain fatty acids（acetic acid,propionic acid,butyric acid）in potato RS products produces the least butyric acid and the most propionic acid.（2）The optimal modeling condition for HepG2 cell insulin resistance model is insulin concentration of 1×10^-6mol/L,glucose concentration 30mmol/L.（3）When the SCFA concentration is 1×Glucose consumption is highest at 10^-6mol/L.（4）Compared with the normal control group,the m RNA expression in HepG2 insulin resistant cells in the model group decreased significantly in each group（P<0.01）;Compared with the model group,the high concentration group showed high expression of AMPK,PI3K,and Akt m RNA,with a statistically significant difference（P<0.05）;The low concentration group showed high expression of GLUT4,and the difference was statistically significant（P<0.05）;The GLUT2m RNA expression was high in the medium concentration group,and the difference was statistically significant（P<0.05）.（5）Compared with the normal control group,the protein expression of each group in HepG2 insulin resistant cells in the model group decreased significantly（P<0.05）;Compared with the model group,the expression of GLUT2,GLUT4,AMPK,PI3K,and Akt proteins increased in the high concentration SCFA group.Conclusion:（1）（1）Potato RS fermentation can produce various SCFA.（2）SCFA can upregulate the protein expression of AMPK,PI3K,Akt,and GLUT2,GLUT4.By enhancing AMPK and PI3K/Akt signal transduction,SCFA acts on various substrate receptor molecules such as GLUT2 and GLUT4,inducing the expression of GLUT2 and GLUT4,further improving glucose transport efficiency,reducing glucose content,improving insulin resistance in HepG2 cells,and effectively controlling blood sugar.