Mechanism Of Scutellarin Regulates Autophagy To Relieve Type 2 Diabetic Liver Injury

Objective:Establish a type 2 diabetes (T2D) rat model to explore whether scutellarin can reduce hepatocyte apoptosis by regulating the autophagy pathway and alleviate liver damage caused by T2D.Method:Feeding 100 SD rats 2 weeks after adaptation of feeding,separation of 10 random normal control group (Con group),given normal diet,the remaining 90 high-sugar high-fat diet 8 weeks.Measured at week 10,all rats fasting glucose,and the rats were given a high-fat high-sugar diet,intraperitoneal injection of small doses of cytotoxic drug pancreatic ? streptozocin (STZ) (35mg/kg) to induced type 2 diabetes model.One week after injection,the fasting blood glucose value was measured,and the blood glucose> 16.7 mmol/L was the successful T2D model rat.63 rats were randomly selected from the rats successfully modeled and divided into 5 groups:T2D model control group (Mod group) 13 rats,scutellarin low-dose group (L group) (25 mg/kg) 13 rats,scutellarin medium-dose group (Group M) (50 mg/kg) 13 rats,scutellarin high-dose group (H Group) (100 mg/kg) 12 rats,metformin positive drug control group (Met group) (120 mg/kg) 12 rats.The drug treatment group was given gavage of scutellarin and metformin,and the saline (10mL/kg) in the Con group and Mod group was gavaged for 8 weeks.During drug treatment,body weight and fasting blood glucose were measured every week.Glucose tolerance test (GTT) and insulin tolerance test (ITT) were performed in the 4th and 8th week of treatment.After 8 weeks of treatment,the apical blood of rats was taken under anesthesia to separate serum for detect the triglyceride (TG);serum liver function indicators:aspartate aminotransferase (AST),alanine alanine aminotransferase (ALT);serum oxidative stress indicators:malondialdehyde (MDA),superoxide dismutase (SOD),glutathione(GSH) ? A part of the rat liver was used for scanning electron microscopy and HE staining to observe the cell structure,and the other part was used for immunohistochemistry (IHC) and Western blotting to detect apoptosis factors(Pro-Caspase3,Cleared-Caspase3) and the expression of autophagy-related proteins(mTOR,Beclinl,LC3 ?/?).Result:1.During the 8 weeks of drug treatment,the fasting blood glucose measured every week was not statistically different between the groups (P> 0.05).2.The blood glucose of GTT and ITT were measured at the 4th and 8th week of drug treatment,and there was no statistically significant difference between the groups (P>0.05).3.Serum triglyceride index:Mod group had higher TG content than Con group;L group and M group had lower TG content than Mod group (P>0.05);H group and Met group had lower TG content than Mod group (P <0.05);Met group had better effect than H group (P <0.05).4.Serum liver function indexes:Mod group had higher AST and ALT than Con group,H group and Met group had significantly lower AST and ALT than Mod group,the difference was statistically significant (P <0.05).5.Serum oxidative stress indicators:MDA in Mod group was higher than Con group,GSH and SOD decreased;MDA in H group and Met group was significantly lower than Mod group,GSH and SOD increased significantly,the difference was statistically significant (P <0.05).6.Scanning electron microscopy:Mod group had more obvious mitochondrial swelling in hepatocytes than Con group,with stronger light transmittance.7.HE staining:the liver cells in the Con group are arranged regularly,and the structure of the liver lobule is clear;the liver cells in the Mod group are arranged irregularly,with a certain degree of atrophy,and the cytoplasmic eosinophilia is enhanced;compared with the Mod group,the cell results change in the L group and the M group Not significant;the H group and Met group had improved liver cell structure and cytoplasmic eosinophilia compared with the Mod group.8.Immunohistochemistry and Western blotting of liver tissue:The expression of Pro-Caspase3,Cleared-Caspase3 and mTOR protein in liver tissue of Mod group was higher than that of Con group (P <0.05);compared with Mod group,the expression of Pro-Caspase3,Cleared-Caspase3 and mTOR protein in L group and M group was decreased (P>0.05);Pro-Caspase3,Cleared-Caspase3,and mTOR in Group H and Met were lower than those in Mod group (P <0.05).The expression of Beclinl protein in liver tissue of Mod group was higher than that of Con group (P <0.05);the expression of Beclinl protein in Group L and M group was higher than that of Mod group,but there was no statistical significance (P>0.05);Group H and Met group were more Mod Group Beclinl protein expression increased (P <0.05).The expression of LC3 I protein in liver tissue of Mod group was higher than that of Con group (P <0.05);compared with Mod grroup (P>0.05) in L group and M group;the expression of LC3 ? protein was lower in H group and Met group than Mod group (P <0.05).Compared with the Con group,the Mod group had a lower LC3:? protein expression (P> 0.05);the L group and M group had a higher LC3? protein expression than the Mod group (P> 0.05);the H group and Met group had a higher LC3 ? protein expression than the Mod group (P<0.05).Compared with the Con group,the Mod group had a lower LC3?/LC3? gray value ratio (P<0.05);the L group compared with the Mod group,the gray value ratio increased;the M group,H group,and Met group had a higher gray value than the Mod group.(P<0.05).Among the above factors,there was no statistically significant difference in protein expression between the H group and the Met group (P> 0.05).Conclusions:The mechanism of scutellarin improving liver injury in type 2 diabetes may be related to the upregulation of autophagy activity leads to inhibition of hepatocyte apoptosis.