本文關(guān)鍵詞： 絲裂原活化蛋白激酶 胰島素樣生長(zhǎng)因子 滋養(yǎng)細(xì)胞 信號(hào)轉(zhuǎn)導(dǎo) 細(xì)胞外信號(hào)調(diào)節(jié)蛋白激酶 增殖 分化 侵入　出處：《華中科技大學(xué)》2006年碩士論文　論文類型：學(xué)位論文

【摘要】： 目的:研究MAPK通路信號(hào)在人早孕絨毛組織中的表達(dá),探討IGF-1對(duì)體外培養(yǎng)的人早孕滋養(yǎng)細(xì)胞的調(diào)節(jié)作用以及MAPK通路對(duì)這些作用的影響,闡明胚胎植入過程中滋養(yǎng)細(xì)胞發(fā)揮生理功能的部分分子機(jī)制。 方法:以妊娠5-7周的正常和自然流產(chǎn)絨毛組織為標(biāo)本,應(yīng)用免疫組化、Western Blot、圖像分析等方法,檢測(cè)MAPK通路信號(hào)在人早孕絨毛組織中的表達(dá)部位、表達(dá)水平;以體外培養(yǎng)的人早孕滋養(yǎng)細(xì)胞為標(biāo)本,分別用細(xì)胞ELISA法、MTT法、化學(xué)發(fā)光法以及割傷法觀察IGF-1在滋養(yǎng)細(xì)胞增殖、分化、侵入中的調(diào)節(jié)作用以及MAPK信號(hào)轉(zhuǎn)導(dǎo)通路對(duì)這些作用的影響。 結(jié)果:(1)免疫組化結(jié)果顯示:妊娠5-7周正常組絨毛組織中,磷酸化(活化型)ERK在細(xì)胞滋養(yǎng)細(xì)胞和合體滋養(yǎng)細(xì)胞呈陽(yáng)性或強(qiáng)陽(yáng)性染色,以細(xì)胞漿著色為主,而在自然流產(chǎn)組絨毛組織中,呈弱陽(yáng)性或陰性染色,以合體滋養(yǎng)細(xì)胞胞漿著色為主;磷酸化p-38和磷酸化JNK在正常組絨毛細(xì)胞滋養(yǎng)細(xì)胞和合體滋養(yǎng)細(xì)胞中均呈陽(yáng)性染色;而在自然流產(chǎn)組中,二者在細(xì)胞滋養(yǎng)細(xì)胞呈陽(yáng)性染色,在合體滋養(yǎng)細(xì)胞則呈陰性染色。(2)Western Blot結(jié)果顯示:磷酸化ERK在妊娠5-7周正常組和自然流產(chǎn)組絨毛均有表達(dá),但在正常早孕組中表達(dá)水平明顯高于自然流產(chǎn)組。(3)細(xì)胞ELISA法檢測(cè)滋養(yǎng)細(xì)胞中ERK的相對(duì)激酶活性,結(jié)果發(fā)現(xiàn),不同濃度IGF-1均能激活ERK,且在0.1-100nM范圍內(nèi),隨IGF-1濃度增高,激酶活性增強(qiáng);ERK通路的特異性阻斷劑U0126可抑制滋養(yǎng)細(xì)胞內(nèi)ERK激活。(4)MTT法測(cè)定滋養(yǎng)細(xì)胞的增殖活性,結(jié)果發(fā)現(xiàn),IGF-1可促進(jìn)滋養(yǎng)細(xì)胞增殖,且在0.1-100nM范圍內(nèi),隨IGF-1濃度增高,滋養(yǎng)細(xì)胞增殖活性增強(qiáng);U0126可抑制IGF-1對(duì)滋養(yǎng)細(xì)胞的促增殖作用。(5)化學(xué)發(fā)光法檢測(cè)滋養(yǎng)細(xì)胞HCG含量,結(jié)果表明,IGF-1可促進(jìn)滋養(yǎng)細(xì)胞分泌HCG,且在0.1-100nM范圍內(nèi),隨IGF-1濃度增高,滋養(yǎng)細(xì)胞分泌HCG增加;U0126可抑制滋養(yǎng)細(xì)胞中HCG分泌。(6)割傷法測(cè)定滋養(yǎng)細(xì)胞的遷移活性,結(jié)果表明,IGF-1對(duì)滋養(yǎng)細(xì)胞的遷移無(wú)明顯促進(jìn)作用;而ERK通路阻斷劑U0126可顯著抑制滋養(yǎng)細(xì)胞的遷移。 結(jié)論: (1) MAPK信號(hào)轉(zhuǎn)導(dǎo)通路激活可能是滋養(yǎng)層細(xì)胞增殖、分化及侵入過程中的重要分子機(jī)制。 (2) IGF-1在體外培養(yǎng)的人早孕滋養(yǎng)細(xì)胞中具有促進(jìn)細(xì)胞增殖、分化的作用。 (3) IGF-1促進(jìn)滋養(yǎng)層細(xì)胞增殖、分化的作用可能是通過激活ERK/MAPK信號(hào)通路實(shí)現(xiàn)的。
[Abstract]:Aim: to investigate the expression of MAPK pathway signal in human chorionic villi, and to investigate the regulatory effect of IGF-1 on human trophoblastic cells cultured in vitro and the effect of MAPK pathway on these effects. To elucidate some molecular mechanisms of the physiological function of trophoblast during embryo implantation. Methods: using normal and spontaneous abortion chorionic villi from 5-7 weeks of gestation as specimens, the expression of MAPK pathway signal in human early pregnancy villi was detected by immunohistochemistry and image analysis. The effects of IGF-1 on the proliferation, differentiation and invasion of trophoblastic cells were observed by ELISA assay, chemiluminescence assay and cut wound method. The effects of MAPK signal transduction pathway on these effects were observed. Results Immunohistochemical results showed that phosphorylated ERK was positive or strongly positive in cytotrophoblast and syncytiotrophoblast in normal villi of 5-7 weeks gestation, mainly cytoplasm. In the villi of spontaneous abortion group, weak positive or negative staining, mainly cytoplasmic staining of syncytiotrophoblast, phosphorylated p-38 and phosphorylated JNK were positive staining in trophoblast and syncytiotrophoblast of normal group. In spontaneous abortion group, both of them were positive in cytotrophoblast and negative staining in syncytiotrophoblast. Western Blot results showed that phosphorylated ERK was expressed in villi of normal group and spontaneous abortion group at 5-7 weeks of gestation. But the expression level in normal early pregnancy group was significantly higher than that in spontaneous abortion group. The activity of ERK relative kinase in trophoblastic cells was detected by ELISA method. The results showed that IGF-1 at different concentrations could activate ERKs and increase with the concentration of IGF-1 in the range of 0.1-100nM. U0126, a specific inhibitor of kinase-enhanced ERK pathway, inhibited ERK activation in trophoblastic cells. The proliferative activity of trophoblastic cells was determined by MTT assay. The results showed that IGF-1 could promote the proliferation of trophoblastic cells and increase with the concentration of IGF-1 in the range of 0.1 ~ 100nM. The enhancement of trophoblast proliferation activity (U0126) inhibited the proliferation of trophoblastic cells by IGF-1. The chemiluminescence method was used to detect the content of HCG in trophoblast. The results showed that the HCG level of trophoblastic cells increased with the concentration of IGF-1 in the range of 0.1 ~ 100nM. The increased secretion of HCG by trophoblastic cells inhibited the secretion of HCG in trophoblastic cells by slit method. The results showed that HCG 1 did not promote the migration of trophoblastic cells. ERK pathway blocker U0126 significantly inhibited trophoblast migration. Conclusion:. 1) Activation of MAPK signal transduction pathway may be an important molecular mechanism of trophoblast cell proliferation, differentiation and invasion. 2) IGF-1 can promote the proliferation and differentiation of human early pregnancy trophoblastic cells in vitro. 3) IGF-1 promotes the proliferation of trophoblast cells, and the differentiation of trophoblast cells may be mediated by activation of ERK/MAPK signaling pathway.
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2006
【分類號(hào)】：R321

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相關(guān)期刊論文 前1條

1 李蕾,李尚為,曹澤毅,彭芝蘭,韓字研;胰島素樣生長(zhǎng)因子系統(tǒng)在人早孕絨毛滋養(yǎng)層細(xì)胞中的表達(dá)[J];華西醫(yī)科大學(xué)學(xué)報(bào);2002年02期

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本文編號(hào)：1553406