KiSS-1基因对妊娠滋养细胞生物学行为影响及其在复发性流产中作用机制的研究

发布时间：2018-02-08 16:32

本文关键词： KiSS-1 kisspeptin GPR54 复发性流产　出处：《苏州大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：探讨KiSS-1基因对妊娠滋养细胞生物学行为的影响及其在复发性流产中的作用机制。方法：①免疫组织化学方法检测kisspeptin、GPR54在复发性流产患者绒毛及蜕膜组织中表达水平；②构建重组质粒pcDNA3.1（+）-KiSS-1；③将重组质粒pcDNA3.1（+）-KiSS-1转染至人滋养层肿瘤JAR细胞株，建立单克隆细胞系；④应用RT-PCR及Western Blot方法分别检测转染细胞KiSS-1的mRNA及编码多肽kisspeptin表达；⑤检测KiSS-1基因及外源性kisspeptin对JAR细胞生长增殖、软琼脂克隆形成能力及迁移侵袭能力的影响。结果：①复发性流产患者绒毛kisspeptin、GPR54的表达均明显低于正常妊娠者（P0.05），蜕膜组织kisspeptin的表达明显低于正常妊娠者（P0.05），蜕膜组织中GPR54的表达在两组间无明显差异；②成功构建重组质粒pcDNA3.1（+）-KiSS-1，并经PCR和测序鉴定无误；③将重组质粒pcDNA3.1（+）-KiSS-1成功转染至人滋养细胞肿瘤株JAR细胞；④筛选出稳定转染的细胞，并通过RT-PCR和Western Blot鉴定目的基因KiSS-1mRNA和编码多肽kisspeptin均成阳性表达；⑤KiSS-1基因和kisspeptin对JAR细胞的生长增殖及克隆形成能力无影响，但明显抑制其迁移和侵袭能力。结论： Kisspeptin在复发性流产患者母胎界面绒毛和蜕膜组织的表达均低下，GPR54在复发性流产患者母胎界面绒毛组织表达低下。KiSS-1基因及其产物kisspeptin能显著抑制滋养细胞迁移及侵袭能力，，推测KiSS-1基因在妊娠期母胎界面的表达调控对妊娠的维持起重要作用，其表达异常与复发性流产有关。
[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.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714.21

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