The Mechanism Of Tianmai Xiaokepian Moderating Glucose Metabolism Using Gene Array And MiRNA Array

OBJECTIVEThe incidence of type2diabetes mellitus is increasing rapidly worldwide, reaching8.3percent in developed countries. It is the fourth prior considerable disease which is the fifth main causes of death after carcinoma, AIDS, cardiovascular disease, imposing a major burden on the health. Chromium is an essential mineral that is thought to be necessary for normal glucose homeostasis. Numerous researches give evidence that acarbose and chromium picolinate which is a formulation designed to improve absorption moderate glucose and insulin resistance. The main ingredient of Tianmai Xiaokepian is chromium picolinate. The research is to explore the effects of Tianmai Xiaokepian and acarbose on the weight, blood glucose, lipid metabolism, serum insulin and glucagon-like peptide-1(GLP-1) in diabetic rats and to investigate its possible glucose-lowing mechanism.METHODSSD rats were randomly divided into four groups:low dosage of Tianmai Xiaokepian group (TML, treated with50mg/kg/d Tianmai Xiaokepian, n=8), high dosage of Tianmai Xiaokepian group (TMH, treated with100mg/kg/d Tianmai Xiaokepian, n=8), low dosage of acarbose group (AcarL, treated with30mg/kg/d acarbose, n=8), high dosage of acarbose group (AcarH, treated with60mg/kg/d, n=8), diabetic model group (n=8) and control group(n=8). The fasting blood glucose and weight were tested on week0,2,4,6and8. The OGTT test was done on week7to measure glucose and plasma GLP-1. Serum insulin, TC, TG, HDL-c and LDL-c were detected on week8. Roche NimbleGen gene array experiment was done using skeletal muscle of rats. miRcury LNATM miRNA array experiment was performed using pancrease and ileum of rats. Real time RT-PCR was done to verify the results of array.RESULTSWe found high dosage of Tianmai Xiaokepian could significantly decrease the level of fasting blood glucose (15.4±5.3vs24.1±2.5mmol/L week2,14.5±3.5vs25.3±3.1mmol/L week4,16.3±4.3vs24.8±4.8mmol/L week6,15.3±5.3vs23.6±4.3mmol/L week8), area under curve of blood glucose in oral glucose tolerance test (AUC,35.7±4.6mmol/L vs53.7±3.4mmol/L), serum fasting insulin (15.79±3.75vs31.90±4.68μIU/mL), HOMA-IR index (15.83±4.87vs33.46±8.30), TC (1.27±0.01vs1.44±0.01mmol/L) and TG (0.49±0.04vs0.83±0.05mmol/L), increase HOMA-β (31.85±3.65vs21.92±2.46) and plasma GLP-1at15min after glucose loading (6.7±0.6vs5.7±0.3pmol/L) compared with those of diabetic group (P<0.05or P<0.01). High dosage of acarbose could significantly decrease the level of fasting blood glucose (14.6±4.3mmol/L vs23.8±4.9mmol/L week2,15.4±2.5vs23.0±4.6mmol/L week4,14.7±4.6vs24.6±5.4mmol/L week6,15.3±6.2vs23.7±4.9mmol/L week8). Gene array showed that Tianmai Xiaokepian up-regulated1752gene expression, down-regulated471gene expression. Based on KEGG pathway analysis, we found that the most three significant pathways were "insulin signaling pathway","glycolysis/gluconeogenesis" and "citrate cycle (TCA)". Real time RT-PCR showed the expression of Akt1(fold-change=5.83±0.15) and Irs2(3.62±0.021) increased in high dosage of Tianmai Xiaokepian group; while Fox03(0.24±0.009)、Pck2(0.21±0.017) and Ptpnl (0.34±0.013) reduced. miRNA array showed18miRNAs increased and3miRNAs decreased in high dasage of Tianmai Xiaokepian group. miR-448, let-7b, miR-540, miR-296, miR-880, miR-200a, miR-500, miR-10b, miR-336, miR-30d, miR-208, let-7e, miR-142-5p, miR-874, miR-375, miR-879, miR-501and miR-188were up-regulated, while miR-301b, miR-134and miR-652were down-regulated in TMH group. Real time RT-PCR showed miR-375and miR-30d, which can stimulate insulin secretion in islet significantly increased. We found that miR-151*, miR-10a-5p, miR-205, miR-17-5p, miR-145and miR-664were up-regulated in the AcarH group, while miR-541and miR-135b were down-regulated. Real time PCR verified these results. Our data suggest that acarbose can improve blood glucose in diabetic rats through the MAPK pathway and can down-regulate pro inflammatory factors by activating miR-10a-5p and miR-664in the ileum.CONCLUSIONTianmai Xiaokepian acts the functions of regulating the levels of fasting blood glucose and lipid metabolism, moderating insulin resistance in diabetic rats. Acarbose can reduce the blood glucose in diabetic rats. The glucose-lowing mechanisms may be correlated with insulin signaling pathway, reducing PCK2, Fox03and PTP-1B in skeletal muscle; incresing miR-375and miR-30d to moderate insulin secretion and islet β cell function, increasing let-7b, let7e, miR-375and miR-142-5p to moderate cytokine-cytokine receptor interaction and MAPK signaling pathway in pancrease. Acarbose can improve blood glucose in diabetic rats through the MAPK pathway and can down-regulate proinflammatory factors by activating miR-10a-5p and miR-664in the ileum.