本文關鍵詞： KiSS-1 kisspeptin GPR54 復發(fā)性流產　出處：《蘇州大學》2014年碩士論文　論文類型：學位論文

【摘要】：目的：探討KiSS-1基因對妊娠滋養(yǎng)細胞生物學行為的影響及其在復發(fā)性流產中的作用機制。 方法：①免疫組織化學方法檢測kisspeptin、GPR54在復發(fā)性流產患者絨毛及蛻膜組織中表達水平；②構建重組質粒pcDNA3.1（+）-KiSS-1；③將重組質粒pcDNA3.1（+）-KiSS-1轉染至人滋養(yǎng)層腫瘤JAR細胞株，建立單克隆細胞系；④應用RT-PCR及Western Blot方法分別檢測轉染細胞KiSS-1的mRNA及編碼多肽kisspeptin表達；⑤檢測KiSS-1基因及外源性kisspeptin對JAR細胞生長增殖、軟瓊脂克隆形成能力及遷移侵襲能力的影響。 結果：①復發(fā)性流產患者絨毛kisspeptin、GPR54的表達均明顯低于正常妊娠者（P0.05），蛻膜組織kisspeptin的表達明顯低于正常妊娠者（P0.05），蛻膜組織中GPR54的表達在兩組間無明顯差異；②成功構建重組質粒pcDNA3.1（+）-KiSS-1，并經PCR和測序鑒定無誤；③將重組質粒pcDNA3.1（+）-KiSS-1成功轉染至人滋養(yǎng)細胞腫瘤株JAR細胞；④篩選出穩(wěn)定轉染的細胞，并通過RT-PCR和Western Blot鑒定目的基因KiSS-1mRNA和編碼多肽kisspeptin均成陽性表達；⑤KiSS-1基因和kisspeptin對JAR細胞的生長增殖及克隆形成能力無影響，但明顯抑制其遷移和侵襲能力。 結論： Kisspeptin在復發(fā)性流產患者母胎界面絨毛和蛻膜組織的表達均低下，GPR54在復發(fā)性流產患者母胎界面絨毛組織表達低下。KiSS-1基因及其產物kisspeptin能顯著抑制滋養(yǎng)細胞遷移及侵襲能力，，推測KiSS-1基因在妊娠期母胎界面的表達調控對妊娠的維持起重要作用，其表達異常與復發(fā)性流產有關。
[Abstract]:Objective: to investigate the effect of KiSS-1 gene on the biological behavior of gestational trophoblastic cells and its mechanism in recurrent abortion. Methods the expression level of kisspeptinine GPR54 in villi and decidua of recurrent abortion patients was detected by immunohistochemical method. The recombinant plasmid pcDNA3.1 (pDNA3.1- KiSS-1) was constructed and transfected into human trophoblastic tumor JAR cell line. RT-PCR and Western Blot were used to detect the expression of mRNA and kisspeptin, respectively. The effects of KiSS-1 gene and exogenous kisspeptin on the growth and proliferation of JAR cells, the ability of soft Agar clone formation and migration and invasion were detected. Results the expression of GPR54 in chorionic villi of patients with recurrent abortion was significantly lower than that in normal pregnant women (P 0.05), and the expression of kisspeptin in decidua was significantly lower than that in normal pregnancy (P 0.05). There was no significant difference between the two groups in the expression of GPR54 in decidual tissues. The recombinant plasmid pcDNA3.1was successfully transfected into human trophoblastic tumor cell line JAR cell line, and stable transfected cells were screened by PCR and sequencing. By RT-PCR and Western Blot, both the target gene KiSS-1mRNA and the encoding polypeptide kisspeptin were identified as positive expression of p5KiSS-1 gene and kisspeptin, which had no effect on the growth, proliferation and clone formation of JAR cells, but significantly inhibited the migration and invasion of JAR cells. Conclusion: the expression of Kisspeptin in the villi and decidua of the maternal and fetal interface of recurrent abortion patients is low. The expression of GPR54 gene and its product kisspeptin can significantly inhibit the migration and invasion of trophoblast cells in the villus tissues of recurrent abortion patients. It is speculated that the regulation of KiSS-1 gene expression at maternal and fetal interface plays an important role in the maintenance of pregnancy. The abnormal expression of KiSS-1 gene is related to recurrent abortion.
【學位授予單位】：蘇州大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R714.21

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