【作者】 吴敏；

【导师】 彭依群；

【作者基本信息】 中南大学 ， 内科学， 2012， 硕士

【摘要】 第一章高糖促进小鼠血管平滑肌细胞CTGF的表达目的：本部分探讨高糖对小鼠血管平滑肌细胞(vascular smooth muscle cells, VSMCs)结缔组织生长因子(connective tissue growth factor, CTGF)的表达影响。方法：小鼠VSMCs复苏传代后用正常浓度葡萄糖组(NG组,5.5mmol/L葡萄糖)、甘露醇高渗对照组(5.5mmol/L葡萄糖+19.5mmol/L甘露醇)、高浓度葡萄糖组(HG组,25mmol/L葡萄糖)分别培养小鼠血管平滑肌细胞0、6、12、24、48小时和0、12、24、48、72小时,应用Real-Time PCR和Western Blot观察其CTGF mRNA及蛋白的表达变化。结果：Real-Time PCR显示：与正常浓度葡萄糖组相比,高糖组明显上调VSMCs CTGF mRNA表达,高糖处理后6小时其表达水平达高峰,24小时开始下降,48小时候后回落至基础水平,甘露醇高渗对照组较正常浓度葡萄糖组相比,CTGF mRNA的表达无明显差异。Western Blot示：与正常浓度葡萄糖组相比,高糖处理后12小时CTGF蛋白的表达水平明显增加,24小时达高峰,72小时候回落至基础水平。结论：高糖上调小鼠血管平滑肌细胞结缔组织生长因子的表达。第二章17β雌二醇对高糖诱导的mVSMCs CTGF的表达影响目的：从第一章的研究发现高糖上调mVSMCs CTGF的表达。本部分旨在探讨17p雌二醇对高糖诱导的mVSMCs CTGF的表达影响。方法：分别用培养基含D-葡萄5.5mmol/L,培养基含D-葡萄25mmol/L+终浓度分别为:OnM、0.01nM、1nM,100nM、1000nM17β雌二醇共6组细胞分别培养12小时、24小时,利用Real-Time PCR和Western Blot观察其干预后CTGF mRNA及蛋白质的表达水平。结果：Real-Time PCR结果显示：与正常对照组相比,含不同浓度17p雌二醇的高糖组均促进mVSMCs CTGFmRNA的表达(P<0.05)；与不含17β雌二醇组相比,终浓度分别为0.01nM、1nM、100nM、1000nM17β雌二醇组上调CTGF的表达无显著差异性。Western Blot结果显示：高糖组中,与不含17β雌二醇组相比,0.01nM、1nM、100nM、1000nM17p雌二醇组的CTGF蛋白表达增加,但各浓度组间的区别无明显差异。结论：17β雌二醇促进高糖诱导的mVSMCs CTGF蛋白的表达,但对高糖诱导的CTGFmRNA的表达无明显作用,提示17β雌二醇可能在糖尿病相关的动脉粥样硬化过程中起到一定的促进作用。更多还原

【Abstract】 Chapter one High glucose increases the expression of CTGF in cultured mice vascular smooth muscle cellsObjective:This Part,we investigated the effects of high glucose on the expression of connective tissue growth factor (CTGF) in cultured mice vascular smooth muscle cells (mVSMCs).Methods:mVSMCs were divided into three groups and cultured in three different culture conditions for0、6、12、24、48hours or0、12、24、48、72hours, respectively:normal glucose group (NG group,5.5mm-ol/L Glu), mannitol group (5.5mmol/L Glu+19.5mmol/L mannitol), high glucose group (HG group,25mmol/L Glu), Real-Time PCR and Western blot were used to detect the changes in CTGF mRNA and protein expressions in the cells after the treatment.Results:The expressions of CTGF mRNA were significantly higher in HG group than in NG group (P<0.05). After high glucose treatment, the CTGF mRNA expression increased gradually and reached the peak level at6h, then followed by gradual decrease till recovering the baseline level at48h. However, mannitol group did not differ significantly from NG group in the expressions of CTGF mRNA.The expression level of CTGF protein also increased progressively and reached the peak level at24h, recovering the base-line level at72h.Conclusion:High glucose increases the expression of CTGF in mice vascular smooth muscle cells. Chapter Two Effect of17β estradiol exposure on connective tissue growth factor expression in cultured mice vascular smooth muscle cells by high glucoseObjective:In the above study, we first found high glucose increasesd the expression of CTGF in mice vascular smooth muscle cells. In the present study, we investigated the effect of17(3estradiol exposure on connective tissue growth factor expression in cultured mice vascular smooth muscle cells by high gluclose.Methods:After being cultured and passaged, the VSMCs were cultured in six different culture conditions for12hours or24hours, respectively:normal glucose group (NG group,5.5mmol/L Glu), high glucose group (HG group,25mmol/L Glu) plused different concentrations of17(3estradiol group (End concentration were OnM、0.01nM、1nM、100nM、1000nM), then the expression of CTGF were detected by Real-Time PCR technique and Western blotResults:Real-Time PCR results showed that compared with the normal control group, high glucose groups including different concentrations of17(3estradiol promoted the expression of mVSMCs CTGF mRNA (P<0.05); And compared with group without17(3estradiol,0.01nM、1nM、100nM、1000nM17(3estradiol groups had no significant difference on expression of CTGF.Western Blot results showed that:compared with the normal control group, and the expression level of CTGF were increased in the high glucose groups containing different concentrations of17(3estradiol groups; the protein expression of CTGF were induced by0.01、1、100、1000nM17(3estradiol group compared with without17β estradiol but there was no differences between each group.Conclusion:17β estradiol promote CTGF protein expression induced by high glucose, but not on CTGF mRNA demonstrating that17(3estradiol may play a certain role in the process of diabetes related atherosclerosis.更多还原