| ObjectiveColonic motility disorder of diabetes is a common complication of diabetes, seriously affects the quality of patients'life, its pathogenesis is not fully understood, may be related to autonomic nervous system, Cajal interstitial cells[1]and smooth muscle disease, high blood sugar abnormalities and other factors[2.3]. Recent studies have found that apoptosis play an important role in the DM and its complications[4]. In this study, we intervent diabetic rats by insulin, detect the smooth muscle cells of Bax, Bcl-2, caspase-3 expression in smooth muscle by using real-time fluorescence quantitative PCR method,study the effect that smooth muscle apoptosis in the mechanism of diabetes colon motility disorder and Insulin, glucose effecting on the mechanism of colonic smooth muscle,from the mRNA levels.ContentsHealthy male SD rats were randomly divided into normal group and the model group.Rats in the model were injected streptozocin (STZ) 40mg/kg one time from tail vein, after seventy-two hours the rats who were measured tail vein blood glucose≥16.7mmol/L and can maintain more than one week were the successful models. Successful model of rats were randomly divided into DM6 week group (C group), DM10 weeks group (D group), insulin intervention in the first 0 week and killed 6 weeks group (E group), insulin 0 Week intervention and killed 10 weeks group (F group), insulin intervention in the first 6 weeks and 10 weeks to kill group (G group). the normal group was divided into normal 6w group (A group), normal 10-week group (B group).The rats of E, F group since the day after the successful model were injected protamine zinc insulin 16U/kg subcutaneously, The rats of G group who were kept six weeks were injected. A group, C group, E group were sacrificed at week 6, and the remaining group were sacrificed at week 10. The changes in the smooth muscle of colon were examined by HE dyeing. The expression levels of Bax and Bcl-2 in the colon smooth muscle cells among the two groups were determined using real-time PCR.Methods 1.Experimental methods included:the establishment of diabetes model; paraffin-embedded technology, HE staining; Extraction of total RNA;real time quantitative PCR,2.Statistic methods:All analysis were performed with SPSS 11.5 software package.Normal distribution of the data was tested using kolmogorov-Smirnov nonparametric test.Groups comparions were performed with one way analysis of variance (post levene test).comparion of any two group among the groups were performed with SNK-q test. Difference were considered statistically significent at P＜0.05.Results1.DM model successfully replicated by STZ. We detect that blood glucose after the model were> 16.7mmol/1, the serum level of fasting insulin decreased, gastrointestinal transit rate slowed down.Diabetes colon motility disorder model reproduced successfully.2.In contrast to he normal control group,The smooth muscle of colon in DM group became thinner,the expression of bax and caspase-3 were higher and the expression of bcl-2 was lower (P<0.05) and the changes were more obviously with the progression of DM.3.In contrast to DM group, the smooth muscle of colon in insulin intervention group became thinner,the number of muscle cells reduced. Bcl-2 expression increased, Bax, Caspase3 expression is reduced, (p<0.05). Early insulin intervention group, Bcl-2, Bax, Caspase3 change more obvious (p<0.05). Conclusions. Since Bcl-2, Bax, Caspase3 is the key to apoptosis, so the experimental results show that apoptosis of smooth muscle cells is one of the pathogenesis of diabetic colonic motility disorders,that insulin has anti-apoptotic function. |
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