Protective Effect Of Empagliflozin On Tacrolimus-induced Pancreas Islet Dysfunction And Renal Injury

Objective:New-onset diabetes after transplantation(NODAT)is a serious complication after kidney transplantation.Tacrolimus(TAC)is one of the most commonly used immunosuppressive agent and is associated with an increased incidence of NODAT.This condition is partly related to the direct toxic effect of TAC on pancreas islet cells,and oxidative stress plays a pivotal role in TAC-induced pancreas dysfunction.Empagliflozin(Em)is a inhibitor of sodium-glucose co-transporter type 2(SGLT-2)and exerts its effect by preventing glucose reabsorption in renal tubular cells.Em is recommended in type 2 DM(T2DM)but its use is still undetermined in NODAT.This study aims to evaluate the effect of SGLT-2 inhibitor on TAC-induced DM.First,we tested whether Em has a blood glucose-lowering effect in TAC-induced DM.Second,we evaluated the protective effect of Em on TAC-induced kidney and pancreatic islet injury using an experimental model of TAC-induced DM.Methods:Male Sprague-Dawley rats(Charles River Technology,Seoul,Korea)weighing 200 to 220g were randomized to 6 groups containing 12 rats each and were treated daily with TAC(1.5 mg/kg,subcutaneously[s.c.])or vehicle(VH,olive oil,0.3 mL,s.c.)for 3 weeks.After confirming development of TAC-induced DM,Em(5 and 10 mg/kg,oral gavage)was added for 3 weeks more.Rats were pair-fed and their body weight was monitored daily.After the treatment period,animals were housed individually in metabolic cages(Tecniplast,Gazzada,Italy),and their urine volume and water intake were measured over 24h.Systolic blood pressure(SBP)was recorded in conscious rats by the tail-cuff method with plethysmography.After the treatment period,animals were anesthetized,and blood samples and tissue specimens were obtained.Evaluate the effect of Em on TAC-induced pancreatic islet dysfunctionand renal injury by measuring SCR(Serum creatinine),Clcr(Creatinineclearance),SBP (Systolic blood pressure),SGLT-1(Sodium-glucose co-transporter type 2),SGLT-2,GLUT-1(Glucose transport type 1),GLUT-2(Glucose transport type 2),plasma insulin level,insulin mRNA,GSIS(Glucose-stimulated insulin secretion),pancreatic islet size,TGF-?1(Transforming growth factor beta-1),Col ?(Collagen?),WT1(Wilms tumor),TUNEL(TdT-mediated dUTP nick-endlabeling)positive cell,8-OHdG(8-Hydroxy-2'-deoxyguanosine),Caspase-3(cysteinyl aspartate specific proteinase-3).Results:1.Em treatment decreased body weight compared with the VH and TAC group without Em treatment.TAC increased urine albumin excretion but addition of Em recovered the parameter.There was no significant difference in the electrolyte content,SBP,and trough level of TAC in the whole blood.2.TAC increased SGLT-2,SGLT-1,GLUT-1 expression compared to the VH group in kidney tissues,did not affect the GLUT-2 expression.However,Em decreased SGLT-2,GLUT-1,GLUT-2 expression compared with the TAC group.TAC increased urinary glucose compared to the VH group and Em further increased urinary glucose compared to the TAC group.3.TAC increased the AUCg,and decreased plasma insulin level compared with the VH group,but Em remarkably decreased the AUCg and increased plasma insulin level compared with the TAC group.4.The TAC group showed a smaller islet with lower intensity of insulin staining within islet than the VH group,but Em increased the islet size compared with the TAC group.The insulin mRNA and GSIS in the TAC group were reduced compared with the VH group.However,Em increased insulin mRNA expression and GSIS compared with the TAC group.5.The TAC group demonstrated increased Scr and decreased Clcr compared to the VH group.However,addition of Em to TAC improved renal function compared with the TAC group.6.TAC increased the tubulointerstitial fibrosis(TIF)score accompanied by increased collagen IV and TGF-?1 expression compared with the VH group.Em reduced the TIF score,collagen IV and TGF-?1 expression.7.TAC significantly increased fractional mesangial area,and decreased WT1-positive cell.Em reduced fractional mesangial area and increased WT1-positive cell.TAC caused fusion of foot process but Em preserved podocyte structure.The TAC group increased renin expression compared with the VH group,but Em significantly reduced renin expression compared with the TAC group.8.8-OHdG was much higher in the TAC group than in the VH group in serum,urine,pancreas,and kidney.Em considerably decreased the 8-OHdG level in serum,pancreas,and kidney but increased urine 8-OHdG.The number of TUNEL-positive cells in the TAC group was significantly increased compared with the VH group in the pancreas,and kidney,but the addition of Em decreased TUNEL positive cells in the pancreas and kidney.Caspase-3 expression in the TAC group was significantly increased compared with the VH group in the pancreas and kidney,but the addition of Em decreased caspase-3 expression in pancreas and kidney compared with the TAC group.Conclusion:Em is effective in controlling TAC-induced hyperglycemia and has protective effect on TAC-induced pancreatic islet dysfunction and reanl injury.