本文選題：神經(jīng)膠質(zhì)瘤細(xì)胞　切入點(diǎn)：人巨細(xì)胞病毒　出處：《青島大學(xué)》2017年碩士論文

【摘要】：人巨細(xì)胞病毒(Human cytomegalovirus,HCMV)屬于β亞科皰疹病毒,在人群中感染非常普遍,一系列的研究顯示HCMV與神經(jīng)膠質(zhì)瘤細(xì)胞的發(fā)病機(jī)制關(guān)系密切。在腦組織中,細(xì)胞之間傳遞信息主要依靠于細(xì)胞縫隙連接通訊(gap juncion intercelluar communication,GJIC),細(xì)胞縫隙連接通訊是機(jī)體中傳導(dǎo)信號最直接的通訊方式,另外,神經(jīng)膠質(zhì)瘤細(xì)胞之間的GJIC被預(yù)測在膠質(zhì)瘤侵襲中起到關(guān)鍵性的作用,并且,腦膠質(zhì)瘤的侵潤性是臨床中治療的難點(diǎn)�？p隙連接基因及蛋白的表達(dá)和功能異常與多種神經(jīng)系統(tǒng)疾病相關(guān),此方面的研究在未來將是新的方向和重點(diǎn)。本研究擬探討HCMV感染對神經(jīng)膠質(zhì)瘤細(xì)胞縫隙連接基因及蛋白的影響,并深入了解縫隙連接蛋白在腫瘤病理機(jī)制中的作用。分別培養(yǎng)并使用HCMV ADl69株(MOI=1)感染原代神經(jīng)膠質(zhì)瘤細(xì)胞和U251細(xì)胞,使用相差顯微鏡觀察細(xì)胞的形態(tài)學(xué)變化,觀察結(jié)果顯示原代神經(jīng)膠質(zhì)瘤細(xì)胞及U251細(xì)胞經(jīng)HCMV感染后逐漸變大變圓。檢測HCMV感染原代神經(jīng)膠質(zhì)瘤細(xì)胞及U251細(xì)胞后縫隙連接基因GJA8和GJB1的表達(dá),結(jié)果顯示縫隙連接基因GJA8和GJB1在原代神經(jīng)膠質(zhì)瘤細(xì)胞及U251細(xì)胞中均可表達(dá)。Real-time PCR檢測原代神經(jīng)膠質(zhì)瘤細(xì)胞及U251細(xì)胞經(jīng)HCMV感染后縫隙連接基因GJA8和GJB1的表達(dá),結(jié)果表明原代神經(jīng)膠質(zhì)瘤細(xì)胞經(jīng)HCMV感染0h、12h、24h、48h后,GJB1的相對表達(dá)水平下降(p㩳0.05),GJA8在24h后相對表達(dá)水平下降(p㩳0.05),在24h之前無明顯變化(p㧐0.05);U251細(xì)胞經(jīng)HCMV感染0h、12h、24h、48h后,GJA8和GJB1的相對表達(dá)量均是下降趨勢(p㩳0.05)。再使用Western-Blot檢測原代神經(jīng)膠質(zhì)瘤細(xì)胞及U251細(xì)胞經(jīng)HCMV感染后縫隙連接蛋白Cx50及Cx32的表達(dá),結(jié)果表明,原代神經(jīng)膠質(zhì)瘤細(xì)胞經(jīng)HCMV感染0h、12h、24h、48h后,Cx32的相對表達(dá)量減少(p㩳0.05),Cx50的相對表達(dá)量在24h之前無明顯變化(p㧐0.05),24h后相對表達(dá)量出現(xiàn)減少趨勢(p㩳0.05);U251細(xì)胞經(jīng)HCMV感染0h、12h、24h、48h后,Cx50和Cx32的相對表達(dá)量均是下降趨勢(p㩳0.05)。使用細(xì)胞劃痕法檢測HCMV感染對U251細(xì)胞的遷移能力的影響,結(jié)果顯示經(jīng)HCMV感染的U251細(xì)胞的侵襲轉(zhuǎn)移能力增強(qiáng)。以上這些實(shí)驗(yàn)結(jié)果表明原代神經(jīng)膠質(zhì)瘤細(xì)胞和U251細(xì)胞經(jīng)HCMV感染后,縫隙連接基因及蛋白的表達(dá)量有所差異,此研究可為縫隙連接基因及蛋白的表達(dá)與腫瘤的惡性程度的相關(guān)性提供一定的理論基礎(chǔ)。
[Abstract]:Human cytomegalovirus (human cytomegalovirus) belongs to the 尾 subfamily herpes virus and is very common in the population. A series of studies have shown that HCMV is closely related to the pathogenesis of glioma cells. The transmission of information between cells mainly depends on cell gap junction communication gap juncion intercelluar communication, gap junction communication is the most direct mode of communication in the body, in addition, GJIC between glioma cells is predicted to play a key role in glioma invasion, and, Invasion of glioma is a difficult point in clinical treatment. The expression and function of gap junction gene and protein are related to many nervous system diseases. This study aims to explore the effects of HCMV infection on gap junction genes and proteins in glioma cells. In addition, the role of gap junction protein in the pathological mechanism of tumor was studied. The primary glioma cells and U251 cells were infected with HCMV ADl69 strain, and the morphologic changes of the cells were observed by phase contrast microscope. The results showed that the primary glioma cells and U251 cells became large and round after HCMV infection. The expression of gap junction gene GJA8 and GJB1 in primary glioma cells and U251 cells infected with HCMV was detected. The results showed that gap junction gene GJA8 and GJB1 could be expressed in primary glioma cells and U251 cells. Real-time PCR was used to detect the expression of gap junction genes GJA8 and GJB1 in primary glioma cells and U251 cells after HCMV infection. The results showed that the relative expression level of GJB1 in primary glioma cells was decreased after infection with HCMV for 12 h or 24 h and 48 h later. The relative expression level of GJA8 decreased after 24 hours. 0. 05%, no significant change before 24 hours. The relative expression of GJA8 and GJB1 in the cells infected with HCMV for 48 h were decreased. Western-Blot was used to detect the expression of gap junction protein Cx50 and Cx32 in primary glioma cells and U251 cells infected by HCMV. The results showed that the relative expression of Cx32 in primary glioma cells was decreased after HCMV infection for 24 h or 48 h after HCMV infection. The relative expression of Cx50 was not changed significantly before 24 hours. The relative expression level decreased after 24 hours. The relative expression of Cx50 and Cx32 in the cells infected with HCMV for 24 h and 48 h after HCMV infection showed a decreasing trend. The effect of HCMV infection on the migration ability of U251 cells was detected by cell scratch method. The results showed that the invasion and metastasis ability of U251 cells infected with HCMV was enhanced. These results indicated that the primary glioma cells and U251 cells were infected by HCMV. The expression of gap junction genes and proteins is different. This study may provide a theoretical basis for the correlation between the expression of gap junction genes and proteins and the malignancy of tumors.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R739.4

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