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目的探讨不同表达水平的钙网蛋白(calreticulin,CRT)与先兆流产的关系。方法 (1)分别应用免疫组织化学法、Western blotting和q PCR检测早期正常妊娠(n=35,均因非意愿妊娠而要求人工流产)及早期先兆流产患者人工流产后的蜕膜和绒毛组织中CRT蛋白及mRNA的表达。(2)分别转染合成并经检测的质粒m Emerald-CRT与CRT-siRNA至BeWo细胞中,检测CRT及Caspase-3的表达。结果免疫组织化学可见CRT阳性信号定位于绒毛两层滋养层细胞的胞质和胞膜上,先兆流产组染色比正常早孕组深、面积大;q PCR与Western blotting检测CRT表达量在先兆流产组明显高于正常早孕组,先兆流产组中mRNA表达量为正常早孕组的1.2倍(P<0.05)。q PCR与Western blotting检测CRT与Caspase-3表达量在质粒(m Emerald-CRT)转染组高于对照组,质粒转染组中CRT及Caspase-3的表达量分别是对照组的3.5倍与2倍(P<0.05),在si-RNA组低于对照组,对照组中CRT及Caspase-3的表达量分别si-RNA组的5倍与2倍。结论早期先兆流产的发生可能与CRT在绒毛和蜕膜组织中的高表达并介导凋亡相关。
Objective To investigate the relationship between different levels of calreticulin (CRT) and threatened abortion. Methods (1) Immunohistochemistry, Western blotting and q-PCR were used to detect the decidual and chorionic tissue after induced abortion in early normal pregnancy (n = 35, both due to unwanted pregnancy) and early threatened abortion CRT protein and mRNA expression. (2) Transfect the synthesized and detected plasmid m Emerald-CRT and CRT-siRNA into BeWo cells respectively to detect the expression of CRT and Caspase-3. Results Immunohistochemistry showed that positive signals of CRT were localized in the cytoplasm and membrane of trophoblast cells in both layers of villi. Threatened abortion group had deeper staining and larger area than normal early pregnancy group. Q PCR and Western blotting detected the expression of CRT in threatened abortion group The expression of mRNA in threatened abortion group was significantly higher than that in normal pregnancy group (1.2%, P <0.05). qPCR and Western blotting showed that the expression of CRT and Caspase-3 in plasmid transfected group was higher than that in control group. The expression of CRT and Caspase-3 in plasmid transfected group were 3.5 times and 2-fold (P <0.05). Compared with the control group, the expression of CRT and Caspase-3 in the si-RNA group was 5-fold and 2-fold higher than that in the si-RNA group. Conclusion The occurrence of early threatened abortion may be related to the high expression of CRT in villus and decidua and the mediation of apoptosis.