亚砷酸钠对大鼠胰岛β细胞p53、mdm2和Kras基因和蛋白表达的影响

发布时间：2018-04-28 21:07

  本文选题：亚砷酸钠 + 大鼠胰岛β细胞　； 参考：《大连医科大学》2013年硕士论文

【摘要】：目的:砷（As）是一种广泛存在于地壳中的类金属元素，主要以有机砷和无机砷两种形式存在，其中无机砷的毒性更强。人类接触砷最主要的途径是环境暴露，如接触被砷污染的水源。近年有研究显示，长期居住在富砷地区的人群糖尿病高发，提示砷可能对胰岛细胞具有毒性作用。有研究证实砷可引起胰岛细胞凋亡和DNA损伤。但关于砷对胰岛细胞癌基因和抑癌基因表达及其编码的蛋白质的影响尚不明确。本研究探讨了砷对胰岛细胞p53、mdm2和Kras等基因和蛋白表达的影响。 方法：选定大鼠胰岛β细胞（INS-1）。用MTT法检测亚砷酸钠对INS-1的毒性作用。用荧光实时定量PCR法检测亚砷酸钠对大鼠胰岛β细胞野生型p53（Tp53）、mdm2和Kras基因表达的影响。用Western blot免疫印迹检测亚砷酸钠对大鼠胰岛β细胞内突变型p53蛋白和mdm2蛋白表达的影响。 结果：（1）亚砷酸钠对大鼠胰岛细胞的毒性作用：亚砷酸钠（0、2.5、5、10、20、40、80、160μM）在37℃条件下，分别作用于INS-1细胞48h和72h后， OD值随浓度的递增而降低，，细胞存活率随着亚砷酸钠剂量的增大而降低，计算IC50分别为5.66μM、2.78μM；（2）亚砷酸钠对大鼠胰岛细胞Tp53、mdm2和Kras基因表达的影响：48h染毒各组Tp53（野生型p53肿瘤抑制基因）表达呈下降趋势， mdm2、Kras基因表达增高；72h染毒中、低剂量组Tp53肿瘤抑制基因表达下降更加明显，中、高剂量组Kras基因表达率升高更加明显；（3）亚砷酸钠对大鼠胰岛细胞突变型p53蛋白和mdm2蛋白质表达的影响：48h染毒后中、高剂量组突变型p53蛋白表达明显增加，低剂量组突变型p53蛋白表达无明显变化；72h染毒各组突变型p53蛋白表达均增加；而48h染毒和72h染毒均为中、高剂量组mdm2蛋白表达增加；72h染毒组与48h染毒组相比，中、高剂量染毒的p53蛋白表达量增加的差别显著。 结论：亚砷酸钠作用于INS-1可造成细胞内抑癌基因Tp53表达率降低，且致癌基因mdm2和Kras的表达率增加；砷可导致INS-1细胞Tp53向突变型p53的转变。
[Abstract]:Objective: as is a kind of metalloid element in the crust, which mainly exists in organic arsenic and inorganic arsenic, among which inorganic arsenic is more toxic. The main route of human exposure to arsenic is environmental exposure, such as exposure to water contaminated by arsenic. Recent studies have shown that there is a high incidence of diabetes in people living in arsenic-rich areas for a long time, suggesting that arsenic may have toxic effects on islet cells. Studies have shown that arsenic can induce apoptosis and DNA damage in islet cells. However, the effects of arsenic on the expression of oncogenes and tumor suppressor genes in islet cells and the proteins encoded by them are unclear. In this study, we investigated the effects of arsenic on the expression of p53 mdm2 and Kras genes and proteins in islet cells. Methods: rat islet 尾 cells were selected. The toxicity of sodium arsenite to INS-1 was detected by MTT method. The effect of sodium arsenite on the expression of wild-type p53Tp53mdm2 and Kras in rat islet 尾 cells was detected by real-time fluorescence quantitative PCR. The effect of sodium arsenite on the expression of mutant p53 and mdm2 protein in rat islet 尾 cells was detected by Western blot Western blot. Results the toxic effect of sodium arsenite on islet cells of rats: sodium arsenite at 37 鈩

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