本文關(guān)鍵詞：GnT-IVa與黏附分子及肝癌細(xì)胞侵襲和遷移關(guān)系的初探　出處：《哈爾濱工業(yè)大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： GnT-IVa 細(xì)胞粘附分子 信號蛋白 粘附能力

【摘要】：糖,作為繼DNA、RNA和蛋白質(zhì)之后的第四類生物大分子,在生物體中發(fā)揮著極其重要的作用。糖鏈結(jié)構(gòu)的異常與多種腫瘤的形成有關(guān),具有特定結(jié)構(gòu)的糖鏈還可作為腫瘤診斷的生物標(biāo)志物,因此,近年來對催化生成糖鏈結(jié)構(gòu)的糖基轉(zhuǎn)移酶的研究呈快速上升趨勢。有文獻(xiàn)報道,Gn T-III和Gn T-V催化的N-聚糖結(jié)構(gòu)重塑對cadherin和integrin介導(dǎo)的細(xì)胞黏附和遷移起著正向或負(fù)向的調(diào)控作用,但關(guān)于Gn T-IVa對cadherin與integrin作用的研究報道較少。早期研究發(fā)現(xiàn),肝癌組織中N-乙酰氨基葡萄糖轉(zhuǎn)移酶IVa(Gn T-IVa)的表達(dá)比癌旁組織中明顯增強(qiáng),當(dāng)用si RNA對mgat4a基因進(jìn)行干擾沉默時,發(fā)現(xiàn)肝癌細(xì)胞BEl-7402的運(yùn)動能力發(fā)生了變化,這表明Gn T-IVa可能與肝癌的運(yùn)動相關(guān)。在前期研究的基礎(chǔ)上,采用RNAi方法,研究Gn T-IVa基因在BEl-7402細(xì)胞中表達(dá)下調(diào)后,細(xì)胞表面integrinβ1的表達(dá)和糖鏈的變化情況。免疫熒光實(shí)驗和western實(shí)驗結(jié)果表明,Gn T-IVa表達(dá)變化前后,integrinβ1在細(xì)胞中的表達(dá)和定位沒有明顯變化。對integrinβ1的糖鏈進(jìn)行分析,結(jié)果發(fā)現(xiàn)Gn T-IVa表達(dá)下調(diào)后,BEL-7402細(xì)胞中integrinβ1糖鏈發(fā)生了改變。同時,本實(shí)驗采用粘附實(shí)驗及流式細(xì)胞凋亡試驗研究Gn T-IVa表達(dá)下調(diào)所引起的BEL-7402細(xì)胞的生物學(xué)功能改變,結(jié)果發(fā)現(xiàn),Gn T-IVa表達(dá)沉默后,細(xì)胞粘附能力增強(qiáng),但細(xì)胞沒有明顯凋亡的跡象。同時,本實(shí)驗還分析了Gn T-IVa對與細(xì)胞侵襲遷移密切相關(guān)的信號通路中若干信號蛋白的影響,結(jié)果表明,Gn T-IVa表達(dá)下調(diào)導(dǎo)致BEl-7402細(xì)胞中β-catenin表達(dá)增強(qiáng),P-ERK2表達(dá)降低。表明Gn T-IVa可能對細(xì)胞中信號通路產(chǎn)生一定影響。本研究以integrinβ1為切入點(diǎn),初步分析了Gn T-IVa表達(dá)變化引起細(xì)胞運(yùn)動能力改變的分子機(jī)制,為深入研究糖鏈和Gn T-IVa的細(xì)胞生物學(xué)功能提供了新的思路。
[Abstract]:Sugar, as a kind of 4th biological macromolecules after DNA RNA and protein, plays an extremely important role in organism. The abnormal structure of sugar chain is related to the formation of many kinds of tumors. Sugar chain with specific structure can also be used as a biomarker for tumor diagnosis. Therefore, the study of glycosyltransferase catalyzing the formation of sugar chain structure has been increasing rapidly in recent years, which has been reported in literature. Gn-T-III and Gn-T-V catalyzed structural remodeling of N-glycan play a positive or negative role in cell adhesion and migration mediated by cadherin and integrin. However, there are few reports on the effects of Gn T-IVa on cadherin and integrin. The expression of N-acetylglucosamine transferase (IVa(Gn T-IVa) in HCC tissues was significantly higher than that in adjacent tissues, when the mgat4a gene was silenced by si RNA. It was found that the motility of BEl-7402 was changed, which suggested that GnT-IVa might be related to the motility of HCC. Based on the previous studies, RNAi method was used. To study the down-regulation of Gn T-IVa gene expression in BEl-7402 cells. The expression of integrin 尾 1 and the changes of sugar chain on the cell surface. The results of immunofluorescence assay and western assay showed that the expression of GnT-IVa was changed before and after the change of GnT-IVa expression. The expression and localization of integrin 尾 1 did not change significantly. The analysis of the sugar chain of integrin 尾 1 showed that the expression of GnT-IVa was down-regulated. The sugar chain of integrin 尾 1 in BEL-7402 cells was changed. The biological function of BEL-7402 cells induced by down-regulation of GnT-IVa expression was studied by adhesion assay and flow cytometry. After the expression of Gn T-IVa was silenced, cell adhesion was enhanced, but there was no obvious sign of apoptosis. The effects of GnT-IVa on some signal proteins in the signal pathway closely related to cell invasion and migration were also analyzed. The expression of 尾 -catenin in BEl-7402 cells was enhanced by down-regulation of GnT-IVa expression. The decreased expression of P-ERK2 suggests that GnT-IVa may have an effect on the signal pathway in cells. In this study, integrin 尾 1 was used as the starting point. The molecular mechanism of the change of cell motility caused by the change of GnT-IVa expression was preliminarily analyzed, which provided a new idea for the further study of the cellular biological function of glucose chain and GnT-IVa.
【學(xué)位授予單位】：哈爾濱工業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R735.7

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本文編號：1364078