The Protective Effect Of Ginsenoside Compound K On Islet β-cells Apoptosis And Its Mechanism In Mice With Type2Diabetes

Diabetes is a severe metabolic disease, and its incidence is increasing year byyear. In patients with diabetes, type2diabetes accounts for more than95%. Apathophysiological change at the early stages of type2diabetes is insulin resistance.As the extension of the course, islet β-cell function gradually diminishes, resulting inthe obvious reduction of the secretion of insulin. At present, type2diabetes drugs areoften accompanied by severe adverse reactions. It is important to developing naturalmedicine of bioactive substances on protection islet β-cells. Therefore, weinvestigated the hypoglycemic effect of ginsenoside Compound K (CK), and its rolein the protection of islet cells.Compound K (CK), a terminal metabolite of protopanaxadiol ginsenosides, isknown to possess anti-diabetic activities but the mechanism for this action is unknown.In addition, an AMP-activated protein kinase (AMPK) is reported to be associatedwith numerous aspects of type2diabetes, including insulin secretion and the functionof pancreatic islets. And the kinase has now become a new therapeutic target for type2diabetes. Activation of AMPK can induce apoptosis in pancreatic β cells in vitro. Inthis study, the snailase was used to transform Ginsenoside Rd to CK under optimizedconditions. And the effect of CK on islet β-cells and AMPK protein was investigatedby using the type2diabetes mice model induced by high-fat diet combined with STZ,as well as the MIN6pancreatic islet β-cells in vitro. This study is divided into twoparts:The first part: the snailase was used to transform Ginsenoside Rd to CK underoptimized conditions. The purity of CK was more than98%. The hypoglycemic effectof CK was investigated by high-fat diet combined intraperitoneal injection of STZ(100mg/kg) induced type2diabetes mice model. The results showed that treatmentof T2D mice with CK (30mg/kg) for4weeks resulted in a significant decrease inFBG, TG, TC and an increase in FINS compared to DM animals. The results of thepancreatic islet HE staining and insulin immunohistochemistry showed that, the damage of islets was weaken and the positive area of average insulin staining wasincreased by the CK treatment. The islet TUNEL staining showed that, model groupapoptosis rate was54±7%, while the normal group and CK group were10.2±5%,39±8%respectively. Western blot analysis showed that, CK inhibited expression ofthe pancreatic tissue AMPK, JNK and mitochondrial apoptosis-related protein(BAX/BCL-2, Cytchrome-c Csapase-3).The second part: the effect of CK on palmitate-induced and hydrogenperoxide-induced MIN6cells damage was investigated. Detection indicators andmethods were as follow: cell viability was determined by MTT assay. The activity ofcaspase-3activity was determined by caspase-3activity kit. Apoptosis wasdetermined by Annexin V/PI double staining kit. ROS was determined byDCHF-DA reactive oxygen kit. The expression of cell protein (AMPK, JNK, GLUT2)was determined by western blot. Intracellular ATP content was determined by HPLC.The results of palmitate-induced model indicated that Palmitate-treatment of MIN6β-cells resulted in reduced cell viability, increased apoptosis and caspase-3activityand activation of AMPK and JNK. CK treatment of MIN6β-cells attenuated thepalmitate-induced reduction in cell viability, apoptosis and caspase-3activity andactivation of AMPK and JNK. The AMPK activator AICAR attenuated theCK-mediated inhibition of palmitate-induced apoptosis, caspase-3activity and JNKactivation. The results of hydrogen peroxide-induced model indicated that CK couldalso reduce caspase-3activity, the rate of apoptosis, and ROS generation. Finally, CKincreased glucose-stimulated insulin secretion in normal MIN6cells, upregualatedGLUT2protein and increased intracellular ATP content.In summary, the following conclusions were drawed:1. CK can improve gluose and lipid disorders, increase insulin secretion and reduceinsulin resistance in type2diabetic mice.2. CK can reduce islet cell apoptosis in type2diabetic mice and palmiticacid-induced apoptosis of MIN6cells through AMPK-JNK-mitochondrial apoptoticpathway.3. CK can reduce hydrogen peroxide-induced MIN6cell apoptosis by reducing ROSgeneration. 4. CK can increase i nsulin secretion by regulating the GLUT2protein in MIN6cells.