The Effect Of SIRT1 In Pancreatic Islet β-cell Apoptosis And Insulin Resistance

Type 2 diabete has become the epidemic desiease in the whole world, whose morbidity and incidence are rapidly increased both in developed and developing countries. So how to improve insulin resistance andβ-cell function has always been the the hotspot of researchers. The relation of fat metabolism disorder and type 2 diabete has been paid more and more attention. Free fat acid ont only participate ininsulin resistance, but also impareβ-cell insulin secreting and lead toβ-cell apoptosis. In recent years, animal experements studys found that SIRT1 has the positive regulation on the insulin secreting of mammalβ-cell, it also can improve insulin resistance, inhibit many cells apoptosis. However, until now whever SIRT1 can improve lipotoxicity leading toβ-cell apoptosis, whever SIRT1 is related to body mass index, blood glucose, plasma insulin levels, blood fat as well as other factors in newly diagnosed type 2 diabetic mellitus has not been repoted. Baseing the above foundation, we carry out the following research.PartⅠChange of SIRT1 in the effect of FFA on pancreatic INS-1 cell apoptosisObjective To observe the effect of free fatty acid on pancreatic INS-1 cells reactive oxygen species and apoptosis, also to investigate the change of SIRT1 in INS-1 cells in lipotoxicity condition. Method Culivating INS-1 cells, detecting the best time and concentration of FFA on it. Exprement contained control group (bovine serum albumin, containing 16.7 mmol/L glucose) and FFA group (palmitate, containing 16.7 mmol/L glucose), cultivating INS-1 cell in above two groups, after 36 hours detecting SIRT1mRNA, ROS and apoptosis level. Result The best time and concentration of FFA on INS-1 cells is 36 hours and 500μmol/l. SIRT1mRNA levels in FFA group were significantly decreased compared with control group(0.5±0.12 vs 1.02±0.08, P<0.01); ROS level was significantly increased compared with control group(437.88±130.38 vs 136.38±55.23, P<0.01); apoptosis rate was significantly increased compared with control group(35.92±7.81 vs 5.53±1.76, P<0.01). Conclusion FFA can promote ISN-1 cell apoptosis, which function is related to lipotoxicity leading to producton of reactive oxygen species, at the same time, in lipotoxicity condition, the SIRT1mRNA relative expression decreased. The result of study indicated that SIRT1 was related to the apoptosis of ISN-1 cells in lipotoxicity condition.PartⅡRole of SIRT1 anti-apoptosis in pancreatic INS-1 cellObjective Constructing SIRT1-overexpressing plasmid and control plasmid, transfecting INS-1 cell, observing reactive oxygen species and apoptosis level of free fatty acid on SIRT1-overexpressing pancreatic INS-1 cells and control INS-1 cells, also to investigate the mechanism of FFA impairingβcell apoptosis and the mechanism of SIRT1 protecting them. Method We constructed SIRT1-overexpressing plasmid and control plasmid; transfected INS-1 cell by FuGENE HD, cultivated non-transfecting INS-1 cell under BSA and FFA comdition, cultivated transfecting control plasmid INS-1 cell and SIRT1-overexpressing INS-1 cell under FFA comdition, detected ROS and apoptosis level after 36 hours. Result In FFA condition, the ROS level was significantly decreased in SIRT1-overexpressing INS-1 cells group compared with non-transfecting group and control plasmid group (284.80±87.97 vs 458.15±134.94 and 437.45±110.38, P<0.01). Also the apoptosis rate was significantly decreased in SIRT1-overexpressing INS-1 cells group compared with non-transfecting group and control plasmid group (18.72±7.13 vs 36.55±8.16 and 34.42±7.36, P<0.01). The ROS level and apoptosis rate had no difference between the non-transfecting group and control plasmid group (P>0.05). Conclusion FFA can promote ISN-1 cell apoptosis, which function is related to lipotoxicity leading to reactive oxygen species. However, SIRT1-overexpressing INS-1 cells can decrease the production of reactive oxygen species, reduce ISN-1 cell apoptosis in lipotoxicity condition.The result of study indicated that SIRT1 can protect ISN-1 in lipotoxicity condition, which mechanism is related to inhitit the production of reactive oxygen species.PartⅢThe Expression and Significance of SIRT1mRNA in the adipose tissue of newly diagnosed patients with type 2 diabetic mellitusObjective To investigate the expression of SIRT1 and the relationship between SIRT1 levels and body mass index(BMI), waist hip ratio(WHR), blood glucose, plasma insulin levels as well as other factors in newly diagnosed type 2 diabetic mellitus(T2DM). Method Adipose tissue SIRT1mRNA levels were detected by RT-PCR in the patients with T2DM and in the controls. The relationship between SIRT1 levels and BMI, WHR, HOMA-IR, blood lipids, plasma glucose, insulin, HbA1c levels were also analyzed. Result SIRT1 levels in patients with T2DM were significantly decreased compared with controls(1.49±0.47 vs 1.12±0.32, P<0.01); SIRT1 levels correlated negatively with FINS and HOMA-IR(r=-0.421, P<0.01; r=-0.511,P<0.01). Multiple regression analysis showed that HOMA-IR was independently related factor influencing SIRT1 levels. Binomial Logistic regression analysis indicated: Controled sex, age, WHR, BMI, TG, TC, LDL, HDL, SIRT1 levels also correlated negatively with T2DM. Conclusion SIRT1 was negative correlated to fasting insulin , HOMA- insulin resistance index and the occurence of type 2 diabete, The decreasing of SIRT1 was associated with glucose metabolism disorder and insulin resistance. SIRT1 is one of the protecting factors of type 2 diabete.