Effect Of Jin Lida On Glucagon In Type1Diabetic Rats And Its Mechanism Research

Diabetes represents a common metabolic disorders, which chronic high bloodglucose as the main features of it.With further research, there are always new discoveriesto perfect and complement its pathogenesis. The traditional view that insulin deficiencyand/or dysfunction is the main pathogenesis of diabetes. In recent years, the role ofglucagon,as a hormone secreted by the pancreaticαcell, in the development of diabetescaused more and more people’s attention.Recent studies show that the phenomenon ofglucagon elevated can occur starting from the abnormal glucose tolerance patients.Bothin type1or type2diabetes are present abnormal basic or postprandial glucagonsecretion, leading to an increase in hepatic glucose output, is an important factor causingelevated blood glucose in diabetic patients.Study found that the sensibility of α cells to the stimulation of hyperglycemia isdecreased in diabetes,hyperglycemia can not effectively inhibit the secretion of glucagon,suggesting that high glucose on the islet α cells with glucose toxicity. NF-κB is atranscription factor within the cells, as the primary targets of hyperglycemia, which canpromote the activation of TNF-α, IL-1β, IL-6and other inflammatory cytokines, causingdamage and apoptosis.Another study found that the activation of NF-κB can increaseiNOS expression,increased NO levels,decreased inhibition of hyperglycemia on glucagonsecretion, as a reason of glucagon increased.the theory of traditional Chinese medicineholds that diabetes associated with "spleen"(including pancreatic concept) to transportdisorders. Jin lida based on the theory of from the "spleen", ginseng, dogwood, berberineand Kudzu as the main ingredient, from the benefits temper, nourishing spleen yin,temperature Spleen yang and other aspects to benefit Spleen, aimed at from the wholeangle to improve islet function.Previous studies have demonstrated that Jin lida caneffectively improve the function of pancreatic β cells and, therefore, this study will focuson the mechanism of its protective effect on pancreatic islet α cells.This study, with different doses of Jin lida,Jin lida+TXL, metformin, saxagliptinintervention in diabetic rats, focusing on the function of Jin lida to glucagon in diabeticrats.By HE staining of pancreatic tissue, glucagon immunohistochemical staining toobserve its effect on α cells.Explore the possible mechanism of protection of pancreaticislet cells by detecting levels of inflammatory factors. The experiment was divided intotwo parts, the first part is observe the affect of Jin lida on glucagon,glucagon-likepeptide-1and glucose on type1diabetic rats, and by HE staining of pancreatic, glucagon immunohistochemistry staining to observed the protective effect on pancreatic cells. Thesecond part, by detecting pancreatic tissue NF-κB mRNA levels and inflammatoryfactors to discussed the possible mechanism of Jin lida protective effect on islet cells.Objective:To observe the effect of different doses of Jin lida on glucagon,pancreatic tissueNF-κB, inflammatory factors in diabetic rat, explore its effects and the mechanisms onisletαcell function,to further clarify the therapy theory of Jin lida.Methods:Sprague-Dawley (SD) rats, injection of STZ (60mg/kg) to preparation of diabetic ratmodel. All rats were randomly divided into normal control group, diabetes model group,Jin lida low-dose group, Jin lida medium-dose group, Jin lida+TXL group, Jin lidahigh-dose group, metformin and saxagliptin group, each group consecutive gavage for8weeks.Part I: monitoring of blood glucose and weight after administration of each group,at the end of eight weeks,detection the fasting blood glucose (FBG), fasting C-peptide byradioimmunoassay;use enzyme-linked immunosorbent assay to test glycosylatedhemoglobin(HbA1c),fasting glucagon, fasting glucagon-like peptide-1; and use pancreastissue HE staining and glucagon immunohistochemical to comparative study. The secondpart: using real-time PCR detection of pancreatic tissue glucagon mRNA, NF-κB mRNA,I-κB mRNA, enzyme-linked immunosorbent assay to detection the level of TNF-α, IL-1β,IL-6.Results:1.General: After successfully induced diabetic rats, blood glucose increased inmodel group, appeared significantly more food, polydipsia, polyuria phenomenon,listlessness, decreased activity, coat dull, slow weight gain. Normal control rats were ingood general condition, shiny hair, normal eating, normal drinking, without obviousabnormalities, faster weight gain. The performance of different drug intervention groupwith varying degrees of improvement.2. Hematology: Compared with normal control group, the level of FBG, HbA1c,glucagon in model group were significantly increased(P <0.01);the level of C-peptide,GLP-1were decreased significantly (P <0.01). Compared with the model group,thelevel of FBG, in Jin lida+TXL group, metformin group were decreased (P <0.05); thelevel of C-peptide, in Jin lida+TXL group, metformin group were elevated (P <0.05);compared with the model group, the level of HbA1c,,in Jin lida medium-dose group, Jinlida+TXL group, Jin lida high-dose group，metformin group were decreased (P <0.01); compared with the model group,the level of glucagon, in Jin lida low-dose group,medium-dose group, high dose group, Jin lida+TXL group, metformin group,saxagliptin group were decreased,(P <0.01); and compared with model group,the level ofglucagon-like peptide-1in Jin lida low-dose group, medium-dose group, high dosegroup, Jin lida+TXL group, metformin group, saxagliptin group were elevated,(P<0.05).3. HE staining of pancreatic islet: normal control group showed that the number ofislets is rich, cell mass is round or oval, boundaries clear, cell morphology complete, cellpulp is rich, round nuclei stained centrally located. Rat islets of diabetic model groupsignificantly reduced the number of the volume marked atrophy, irregular, ill-defined,larger islet cell swelling, increased cytoplasmic vacuoles, lighter colored, nuclearcondensation, eccentric distribution, partial deletion. Compared with diabetic modelgroup, the number of islets in Jin lida low-dose group, middle dose group,Jin lida+TXLgroup, and saxagliptin group, were certainly improved, in the high-dose Jin lida groupand metformin group improved significantly.4. Immunohistochemistry showed that in normal control group α cells mainly in theperipheral parts of the islets; compared with normal control group,the amount of modelgroup α cells were in a growing trend;in different drug intervention groupglucagon-positive cells compared with diabetic model group were not obvious decline.5. RT-PCR to detect the gene expression in rat pancreatic tissue: Compared withnormal control group, the expression of diabetic model group glucagon mRNA wasincreased (P <0.01), the expression of NF-κB mRNA increased (P <0.01), I-κB mRNAexpression was increased (P <0.01); compared with diabetic model group,the expressionof glucagon mRNA, NF-κB mRNA and I-κB mRNA in Jin lida low-dose group, middle-dose group, high-dose group, Jin lida+TXL group, metformin group, saxagliptin groupwere decreased (P <0.05).6. Enzyme-linked immunosorbent assay (ELISA) Results: Compared with normalcontrol group, the levels of TNF-a, IL-1β, IL-6in diabetic group were elevated (P <0.01);Compared with diabetic model group,the level of IL-1β, IL-6in Jin lida low-dose group,middle-dose group, high-dose group,Jin lida+TXL group, metformin group, saxagliptingroup were decreased (P <0.05), the level of TNF-a in Jin lida middle-dose group, highdosec Jin lida group, Jin lida+TXL group, metformin group, saxagliptin group weredecreased (P <0.05). Conclusion:Jin lida can effectively reduce glucagon levels in diabetic rats, possibly by reducinginflammation factor levels of the pancreas tissue to improve theα-cell function.