| Objective: Nonalcoholic fatty liver disease(NAFLD)is an important complication of type 2 diabetes mellitus(T2DM),which has become the most common chronic liver disease worldwide.Metabolic associated fatty liver disease “MAFLD” was suggested as a more appropriate overarching term,but there are many issues that require further study and discussion.NAFLD is characterized by lipid accumulation and fatty degeneration of hepatocytes.It is closely related with insulin resistance,oxidative stress,endoplasmic reticulum stress,inflammation and so on.Autophagy can regulate lipid metabolism,insulin resistance,alleviate hepatocyte damage and inflammation through multiple ways.The occurrence,development and outcome of NAFLD are closely related to the down regulation of autophagy.Therefore,upregulating autophagy plays a pivotal role for the prevention and therapy of NAFLD.Sodium-glucose cotransporter 2(SGLT-2)inhibitors are a new class of drugs for T2 DM by inhibiting the reabsorption of glucose in the kidney and promoting the urinary glucose excretion.Several studies with SGLT-2 inhibitors reported a lower of body mass index and fatty liver index in NAFLD.However,the effects of SGLT-2 inhibitors on NAFLD with T2 DM and its mechanism remain unclear.Empagliflozin is a high effective and selective SGLT-2 inhibitor.There is little research on the effects of empagliflozin on hepatic steatosis in T2 DM.We evaluated the beneficial effects and its underlying mechanisms of empagliflozin on T2 DM with NAFLD.Methods: Six-week-old db/db mice was taken as the model of T2 DM with NAFLD.Db/m mice in the same litter was used as a control group(n=6,NC group).Then the db/db mice were randomly divided into 3 groups(n=18): diabetic group(db/db group),empagliflozin group(db/db+Empa group),insulin group(db/db+Ins group).The db/db+Empa group was administered intragastrically with empagliflozin(3.8 mg/kg/d)while NC group and db/db group were given matching normal saline.The weight,blood glucose and urine glucose were measured periodically.After 8 weeks treatment,samples of venous blood were collected for detection of liver function.H&E,Oil Red O,immunohistochemistry and western blotting were used to detect morphological changes,lipid accumulation and autophagy levels,TET2,AMPK,p-AMPK.Human liver cells(HL7702)were used as the cell model(NC group),then 0.5mmol/L palmitic acid(PA)and 25mmol/L high glucose(HG)treated cells as a cell model of steatosis(PA+HG group).Cells were treated with empagliflozin(8μmmol/L,PA+HG+Empa group)for 48 hours and then exposed to autophagy inhibitor 3-methyladenine(3-MA),AICAR(AMP-activated protein kinase activation)or Compound C(AMP-activated protein kinase inhibitor).TET2 si RNA was also transfected into cells.The detecting indexes were same as indexes in vivo.Results: 1.The db/db group showed higher blood glucose levels,body weight,liver weight,ALT,AST.Meanwhile,above all indexes improved obviously in db/db+Empa group.2.H&E and Oil Red O staining showed that lipid accumulation increased obviously in db/db group.However,above all indexes improved in db/db+Empa group.3.Immunohistochemistry and western blotting showed that autophagy level,TET2 and p-AMPK expressions were decreased in db/db group but recoved in db/db+Empa group.But changes above all indexes were insignificantly in db/db+Ins group.4.Oil Red O staining showed that lipid accumulation was increased in PA+HG group.Autophagy level,TET2 and p-AMPK expressions were suppressed in PA+HG group.Above all indexes improved in empagliflozin treatment.5.The effects of AICAR were similar to empagliflozin.But the effects of Compound C were contrast to empagliflozin.6.Depletion of TET2 reversed the effects of empagliflozin.Conclusions: 1.Empagliflozin alleviates nonalcoholic fatty liver disease in type 2 diabetic mice.2.Empagliflozin could increase p-AMPK,TET2 expressions and induce autophagy.3.Empagliflozin could reduce lipid accumulation via autophagy in hepatocytes.4.Empagliflozin could reduce hepatic steatosis through AMPK-TET2-autophagy. |
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