【摘要】：目的:腦膠質(zhì)瘤是中樞神經(jīng)系統(tǒng)最常見的惡性腫瘤。包括神經(jīng)外科手術(shù)、放射治療及化療等多模式治療相結(jié)合,但腦膠質(zhì)瘤患者的預(yù)后仍然不令人滿意。局部復(fù)發(fā)是治療失敗的主要原因。膠質(zhì)瘤細(xì)胞具有侵襲和遷移能力是導(dǎo)致局部復(fù)發(fā)的重要原因。FOXJ2是轉(zhuǎn)錄因子叉頭框(Forkhead box,FOX)蛋白家族成員之一,FOXJ2涉及細(xì)胞周期的調(diào)控以及腫瘤的發(fā)生發(fā)展。已有研究表明FOXJ2表達(dá)的上調(diào)可抑制乳腺癌的遷移和侵襲能力。本研究旨在初步了解FOXJ2在腦膠質(zhì)瘤中的表達(dá)情況及其與膠質(zhì)瘤患者臨床病例因素的關(guān)系,檢測膠質(zhì)瘤細(xì)胞中FOXJ2、E-鈣黏蛋白(E-cadherin)的表達(dá),分析其相互的關(guān)系以及對膠質(zhì)瘤細(xì)胞遷移的影響,探討FOXJ2的異常表達(dá)在膠質(zhì)瘤發(fā)生、發(fā)展過程中的意義。方法:1.從組織學(xué)水平,本研究首先通過Western Blot方法,檢測在兩組人正常腦組織和七組不同級別膠質(zhì)瘤組織中FOXJ2的表達(dá)情況;然后,運(yùn)用免疫組化方法檢測80例膠質(zhì)瘤組織標(biāo)本中FOXJ2和E-cadherin的表達(dá)情況,并采用統(tǒng)計學(xué)方法分析FOXJ2、E-cadherin的表達(dá)情況與膠質(zhì)瘤患者的年齡、性別、腫瘤位置、大小及腫瘤組織學(xué)分級等臨床病理因素之間的相關(guān)性。通過Kaplan-Meier法分析FOXJ2的表達(dá)與膠質(zhì)瘤患者的生存率之間的相關(guān)性;2.從細(xì)胞分子水平,本研究選取U87MG,U373,H4,A172和U251MG等五種膠質(zhì)瘤細(xì)胞株,用Western Blot技術(shù)檢測FOXJ2、E-cadherin和波形蛋白(Vimentin)的表達(dá)。選取U87膠質(zhì)瘤細(xì)胞株,瞬時轉(zhuǎn)染FOXJ2過表達(dá)和干擾質(zhì)粒,通過Western Blot和細(xì)胞免疫熒光檢測膠質(zhì)瘤細(xì)胞E-cadherin和Vimentin的表達(dá);用劃痕實驗和Transwell遷移實驗檢測細(xì)胞遷移能力。明確FOXJ2對膠質(zhì)瘤的遷移的影響。結(jié)果:1.Western Blot及免疫組化結(jié)果均顯示,FOXJ2在膠質(zhì)瘤組織中的表達(dá)隨膠質(zhì)瘤病理級別的增高而降低,提示FOXJ2的表達(dá)強(qiáng)度與膠質(zhì)瘤組織學(xué)分級(P=0.012)之間的關(guān)系有統(tǒng)計學(xué)意義,而與患者的年齡、性別、腫瘤體積大小、位置、是否發(fā)生壞死等臨床病理特征間無差異。Kaplan-Meier生存曲線分析顯示FOXJ2在膠質(zhì)瘤中的低表達(dá)患者的總生存率明顯低于FOXJ2高表達(dá)的患者(P0.01)。2.Western Blot技術(shù)檢測U87,U373,H4,A172,U251膠質(zhì)瘤細(xì)胞株FOXJ2的表達(dá),結(jié)果顯示FOXJ2在遷移能力弱的H4細(xì)胞中表達(dá)較高,而在遷移能力強(qiáng)的U251細(xì)胞中表達(dá)偏低。選取U87和U251細(xì)胞株,檢測上皮間質(zhì)轉(zhuǎn)化(epithelial-mesenchymal transition,EMT)標(biāo)記,觀察到FOXJ2與E-cadherin表達(dá)呈正相關(guān)(r=0.303;P0.01),Vimentin的表達(dá)情況與FOXJ2的表達(dá)情況相反。3.通過對轉(zhuǎn)染過表達(dá)FOXJ2質(zhì)粒的U87膠質(zhì)瘤細(xì)胞,進(jìn)行Western Blot和細(xì)胞免疫熒光檢測,觀察到FOXJ2過表達(dá)可促進(jìn)膠質(zhì)瘤細(xì)胞E-cadherin的表達(dá),抑制Vimentin的表達(dá)。劃痕實驗和Transwell遷移實驗,觀察到FOXJ2過表達(dá)可抑制膠質(zhì)瘤細(xì)胞的遷移。4.通過對轉(zhuǎn)染sh RNA FOXJ2質(zhì)粒的U87膠質(zhì)瘤細(xì)胞,進(jìn)行Western Blot和細(xì)胞免疫熒光檢測,觀察到干擾FOXJ2表達(dá)抑制膠質(zhì)瘤細(xì)胞E-cadherin的表達(dá),促進(jìn)Vimentin的表達(dá)。劃痕實驗和Transwell遷移實驗,觀察到干擾FOXJ2表達(dá)可促進(jìn)膠質(zhì)瘤細(xì)胞的遷移。結(jié)論:1.FOXJ2在膠質(zhì)瘤中的表達(dá)隨病理分級的提高而降低。FOXJ2的低表達(dá)提示膠質(zhì)瘤患者的預(yù)后不良。2.FOXJ2的表達(dá)可影響E-cadherin、Vimentin的表達(dá),影響EMT的進(jìn)程,從而抑制膠質(zhì)瘤細(xì)胞遷移。
[Abstract]:Objective:Glioma is the most common malignant tumor of the central nervous system.It includes neurosurgery,radiotherapy and chemotherapy,but the prognosis of the patients with glioma is still unsatisfactory.Local recurrence is the main cause of treatment failure.The ability of glioma cells to invade and migrate leads to local recurrence. FOXJ2 is a member of the Forkhead box (FOX) protein family. FOXJ2 is involved in cell cycle regulation and tumor development. Upregulation of FOXJ2 expression has been shown to inhibit the migration and invasion of breast cancer. The expression of FOXJ2 and E-cadherin in glioma cells was detected, the relationship between FOXJ2 and E-cadherin and the migration of glioma cells was analyzed, and the significance of the abnormal expression of FOXJ2 in the occurrence and development of glioma was explored. Methods: 1. From the histological level, this study was first carried out by W. Estn Blot method was used to detect the expression of FOXJ2 in normal brain tissues of two groups and seven groups of glioma tissues of different grades. Immunohistochemical method was used to detect the expression of FOXJ2 and E-cadherin in 80 glioma tissues, and statistical method was used to analyze the expression of FOXJ2 and E-cadherin and the age of glioma patients. Kaplan-Meier method was used to analyze the correlation between FOXJ 2 expression and survival rate of glioma patients. 2. At the cellular and molecular level, five glioma cell lines, including U87MG, U373, H4, A172 and U251MG, were selected for Western Blot assay. The expression of FOXJ2, E-cadherin and Vimentin was detected. U87 glioma cell line was transfected with FOXJ2 overexpression and interfering plasmid. The expression of E-cadherin and Vimentin was detected by Western Blot and immunofluorescence assay. The migration ability of glioma cells was detected by scratch test and Transwell migration test. Western Blot and immunohistochemical results showed that FOXJ2 expression in glioma tissues decreased with the increase of pathological grade, suggesting that FOXJ2 expression intensity and histological grade of glioma (P = 0.012) were statistically significant, but with the age, sex, tumor size and location of patients. Kaplan-Meier survival curve analysis showed that the overall survival rate of patients with low FOXJ2 expression in gliomas was significantly lower than that of patients with high FOXJ2 expression (P 0.01). 2 Western Blot assay showed that the expression of FOXJ2 in U87, U373, H4, A172, U251 gliomas was significantly lower than that of patients with high FOXJ2 expression (P 0.01). U87 and U251 cell lines were selected to detect epithelial-mesenchymal transition (EMT) markers. FOXJ2 was positively correlated with E-cadherin expression (r = 0.303; P 0.01). Vimentin expression was contrary to FOXJ2 expression. Western Blot and immunofluorescence assay were performed on U87 glioma cells transfected with FOXJ2 plasmid. Overexpression of FOXJ2 could promote the expression of E-cadherin and inhibit the expression of Vimentin. Scratch test and Transwell migration test showed that over-expression of FOXJ2 could inhibit the migration of glioma cells. Western Blot and immunofluorescence were used to detect the expression of E-cadherin and Vimentin in U87 glioma cells infected with SH RNA FOXJ2 plasmid. Scratch test and Transwell migration test showed that interfering with FOXJ2 expression could promote the migration of glioma cells. The expression of FOXJ 2 decreased with the increase of pathological grade. The low expression of FOXJ 2 suggested that the prognosis of glioma patients was poor. 2. The expression of FOXJ 2 could affect the expression of E-cadherin and Vimentin, affect the process of EMT and inhibit the migration of glioma cells.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R739.41

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