發(fā)布時(shí)間：2018-06-25 05:12

  本文選題：鼻咽癌 + RNA干擾　； 參考：《臨床耳鼻咽喉頭頸外科雜志》2017年10期

【摘要】：目的:通過研究VTCN1在鼻咽癌細(xì)胞中的表達(dá)情況,采用慢病毒載體靶向沉默VTCN1基因體外對(duì)人鼻咽癌細(xì)胞的抑制作用。方法:通過RT-PCR技術(shù)和蛋白質(zhì)免疫印跡法,檢測(cè)人鼻咽癌細(xì)胞株CNE1,HONE-1,NP65和HNE2的VTCN1表達(dá)情況,篩選出VTCN1表達(dá)量最多的細(xì)胞株,進(jìn)行后續(xù)的實(shí)驗(yàn)。用脂質(zhì)體lipofectamine2000將沉默VTCN1的慢病毒載體轉(zhuǎn)染高表達(dá)VTCN1的HNE2細(xì)胞,篩選出穩(wěn)定細(xì)胞株;采用RT-PCR技術(shù)和蛋白質(zhì)免疫印跡法檢測(cè)VTCN1ShRNA對(duì)VTCN1基因的靶向沉默效率;ckk-8、基質(zhì)膠侵襲實(shí)驗(yàn)檢測(cè)VTCN1沉默后HNE2細(xì)胞增殖和侵襲的影響;蛋白質(zhì)免疫印跡實(shí)驗(yàn)檢測(cè)VTCN1沉默后,JAK/STAT通路主要蛋白的磷酸化水平的變化。結(jié)果:VTCN1靶向沉默的細(xì)胞VTCN1表達(dá)受到顯著抑制;RT-PCR和蛋白質(zhì)免疫印跡法檢測(cè)發(fā)現(xiàn)穩(wěn)定轉(zhuǎn)染ShRNA后,VTCN1的mRNA表達(dá)水平和蛋白表達(dá)水平均明顯下降(P0.05);轉(zhuǎn)染VTCN1ShRNA后HNE2細(xì)胞的增殖和侵襲能力均明顯降低(P0.05);VTCN1沉默后,JAK、STAT磷酸化水平明顯下降(P0.05)。結(jié)論:VTCN1ShRNA能有效沉默VTCN1的表達(dá),顯著抑制HNE2細(xì)胞的增殖和侵襲,推測(cè)其與下調(diào)JAK/STAT信號(hào)通路蛋白的活化相關(guān)。
[Abstract]:Aim: to investigate the expression of VTCN1 in nasopharyngeal carcinoma cells and to investigate the inhibitory effect of lentivirus vector targeting VTCN1 gene on human nasopharyngeal carcinoma cells in vitro. Methods: the expression of VTCN1 in human nasopharyngeal carcinoma cell line CNE1HONE-1nNP65 and HNE2 was detected by RT-PCR and Western blotting. The lentivirus vector silencing VTCN1 was transfected into HNE2 cells with high expression of VTCN1 by liposome lipofectamine2000, and stable cell lines were selected. RT-PCR and Western blot were used to detect the effect of VTCN1ShRNA on the targeting silencing efficiency of VTCN1 gene and the effect of VTCN1ShRNA on the proliferation and invasion of HNE2 cells after VTCN1 silencing. Western blot assay was used to detect the phosphorylation of major proteins in the JAK / stat pathway after VTCN1 silencing. Results the expression of VTCN1 was significantly inhibited in the cells targeted to VTCN1. RT-PCR and Western blot analysis showed that the mRNA expression level and protein expression level of VTCN1 decreased significantly after stable transfection of ShRNA (P0.05), the proliferation of HNE2 cells and the expression of VTCN1 protein in HNE2 cells were observed after transfection of VTCN1 ShRNA. The invasive ability was significantly decreased (P0.05) and the phosphorylation level of JAKN STAT was significantly decreased after VTCN1 silencing (P0.05). Conclusion the cell proliferation and invasion of HNE2 cells can be significantly inhibited by the inhibition of VTCN1 expression by 1: VTCN1ShRNA, which may be related to the down-regulation of JAK / stat signaling pathway protein activation.
【作者單位】： 武漢大學(xué)中南醫(yī)院耳鼻咽喉頭頸外科;武漢大學(xué)人民醫(yī)院耳鼻咽喉科;
【分類號(hào)】：R762

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