發(fā)布時(shí)間：2018-06-21 18:53

  本文選題：前列腺癌 + ARRDC3�。� 參考：《南方醫(yī)科大學(xué)》2017年博士論文

【摘要】：研究背景及目的:前列腺癌(prostate cancer,PCa)是常見(jiàn)的泌尿系惡性腫瘤之一,目前我國(guó)的發(fā)病率和病死率呈現(xiàn)逐年上升的趨勢(shì)。前列腺癌根治術(shù)是早期階段的優(yōu)選治療方法,內(nèi)分泌治療可作為腫瘤晚期的主要治療手段。近年來(lái)以阿比特龍(Abiraterone)為代表的內(nèi)分泌治療及分子靶向治療顯現(xiàn)出一定優(yōu)勢(shì),但療效尚未確定。然而幾乎所有患者最終發(fā)展成激素非依賴性前列腺癌,目前尚無(wú)有效的治療方法。既往的研究和我們先前的實(shí)驗(yàn)已經(jīng)證實(shí)了 miR-30d在前列腺癌中表達(dá)上調(diào),并發(fā)揮關(guān)鍵的促癌作用。因此,我們利用基因表達(dá)譜芯片技術(shù)對(duì)過(guò)表達(dá)miR-30d的LNCaP和DU145細(xì)胞株進(jìn)行基因表達(dá)譜分析,獲得一批miR-30d調(diào)控的差異表達(dá)下調(diào)的基因,其中Arrestin domain-containing protein 3(ARRDC3)基因表達(dá)水平明顯下調(diào)。含有Arrestin結(jié)構(gòu)域的蛋白3(ARRDC3)是哺乳動(dòng)物a-抑制蛋白家族的成員,已被確定為腫瘤抑制基因,但其在人類前列腺癌中的功能仍未見(jiàn)報(bào)道。DraheimKM等一項(xiàng)研究顯示ARRDC3直接與磷酸化形式的ITGβ4結(jié)合,導(dǎo)致其內(nèi)化,泛素化并最終降解,抑制乳腺癌的進(jìn)展。本研究旨在確定ARRDC3在前列腺癌中的表達(dá)情況及ARRDC3與前列腺癌的進(jìn)展和預(yù)后的相關(guān)性,探索ARRDC3-ITGβ4通路在前列腺癌中的作用。方法:1.構(gòu)建miR-30d抑制表達(dá)和過(guò)表達(dá)的LNCaP細(xì)胞株,通過(guò)qRT-PCR和蛋白質(zhì)印跡法分別驗(yàn)證miR-30d和ARRDC3之間的表達(dá)相關(guān)性。2.免疫組織化學(xué)分析ARRDC3在臨床前列腺癌組織芯片中的表達(dá)情況。利用統(tǒng)計(jì)學(xué)方法,分析Taylor數(shù)據(jù)庫(kù)中ARRDC3表達(dá)水平與前列腺癌患者的臨床特征和預(yù)后的相關(guān)性,通過(guò)生存曲線分析及Cox回歸分析研究ARRDC3的表達(dá)與總體生存率、無(wú)生化復(fù)發(fā)生存率的關(guān)系及是否可作為預(yù)后的預(yù)測(cè)因子。3.構(gòu)建ARRDC3抑制表達(dá)和過(guò)表達(dá)的DU145和LNCaP細(xì)胞株,細(xì)胞功能實(shí)驗(yàn)分析ARRDC3對(duì)前列腺癌細(xì)胞功能的影響。4.過(guò)表達(dá)DU145細(xì)胞株接種至裸鼠皮下組織構(gòu)建皮下移植瘤模型,用于分析ARRDC3在體內(nèi)對(duì)前列腺癌成瘤的影響。5.免疫組織化學(xué)分析裸鼠皮下腫瘤組織中Ki67,MMP-9和ITGβ4的表達(dá)情況。蛋白質(zhì)印跡法檢測(cè)裸鼠皮下腫瘤細(xì)胞中ITGB4的表達(dá)情況。結(jié)果:1.在過(guò)表達(dá)miR-30d的基因表達(dá)譜分析中,我們獲得一批差異性表達(dá)下調(diào)的基因,其中ARRDC3明顯下調(diào)。qRT-PCR和蛋白質(zhì)印跡法分析結(jié)果顯示ARRDC3和miR-30d存在負(fù)相關(guān)性。2.組織芯片免疫染色分析顯示前列腺癌組織標(biāo)本的ARRDC3低表達(dá)與高Gleason評(píng)分顯著相關(guān)(P = 0.045)。Taylor數(shù)據(jù)庫(kù)分析ARRDC3結(jié)果顯示ARRDC3的下調(diào)表達(dá)與Gleason評(píng)分、術(shù)后發(fā)生轉(zhuǎn)移和生化復(fù)發(fā)呈相關(guān)性(P分別為0.0001,0.01 和 0.02)。3.Kaplan-Meier法和Log rank法分析結(jié)果顯示低表達(dá)ARRDC3與前列腺癌患者不良預(yù)后相關(guān)(P = 0.010),在前列腺癌中作為抑癌基因。COX回歸模型單因素和多因素分析顯示,ARRDC3是前列腺癌患者無(wú)生化復(fù)發(fā)生存的獨(dú)立預(yù)測(cè)因子。4.細(xì)胞功能實(shí)驗(yàn)顯示過(guò)表達(dá)的ARRDC3能降低DU145和LNCaP細(xì)胞株的增殖、遷移和侵襲能力(P0.05),ARRDC3的抑制表達(dá)能增強(qiáng)細(xì)胞的增殖、遷移和侵襲能力(P0.05)。5.動(dòng)物實(shí)驗(yàn)結(jié)果顯示過(guò)表達(dá)ARRDC3能夠顯著抑制裸鼠皮下移植瘤的生長(zhǎng)速度(P0.05)。與對(duì)照組相比,Ki67和MMP-9的免疫組化評(píng)分明顯降低(P0.05),證實(shí)過(guò)表達(dá)ARRDC3抑制腫瘤細(xì)胞的增殖和侵襲。6.蛋白質(zhì)印跡和免疫組織化學(xué)結(jié)果顯示ARRDC3和ITGβ4呈負(fù)相關(guān)性(P0.05)。結(jié)論:1.ARRDC3在前列腺癌中起抑癌作用,抑制前列腺癌進(jìn)展,并作為獨(dú)立預(yù)測(cè)指標(biāo)預(yù)測(cè)術(shù)后生化復(fù)發(fā)和轉(zhuǎn)移的風(fēng)險(xiǎn)。2.ARRDC3可通過(guò)ARRDC3-ITGβ4通路抑制前列腺癌的進(jìn)展,ARRDC3-ITGβ4通路可作為前列腺癌新的治療目標(biāo)。
[Abstract]:Background and purpose: prostate cancer (PCa) is one of the common malignant tumors of the urinary system. The incidence and fatality rate of our country are increasing year by year. Radical prostatectomy is the preferred treatment method at the early stage, and endocrine therapy can be used as the main treatment for advanced cancer. (Abiraterone) there are some advantages for the representation of endocrine therapy and molecular targeting therapy, but the effect is not yet determined. However, almost all patients eventually develop into hormone non dependent prostate cancer. There is no effective treatment yet. Previous studies and our previous tests have confirmed the expression of miR-30d in prostate cancer. Therefore, we use gene expression chip technology to analyze the gene expression profiles of LNCaP and DU145 cells overexpressing miR-30d, and obtain a number of differentially expressed genes regulated by miR-30d, in which the expression level of Arrestin domain-containing protein 3 (ARRDC3) gene is obviously downregulated. The protein 3 (ARRDC3) of the estin domain is a member of the mammalian a- suppressor family and has been identified as a tumor suppressor, but its function in human prostate cancer has not been reported in a study of.DraheimKM, such as.DraheimKM, which shows that ARRDC3 is directly associated with the form of phosphorylation of ITG beta 4, leading to its internalization, ubiquitination, and final degradation and inhibition of breast cancer. The purpose of this study is to determine the expression of ARRDC3 in prostate cancer and the correlation between ARRDC3 and the progression and prognosis of prostate cancer. The purpose of this study is to explore the role of ARRDC3-ITG beta 4 pathway in prostate cancer. Method: 1. to construct the LNCaP cell lines with miR-30d inhibition and overexpression, and to verify miR-30d and A by qRT-PCR and Western blotting, respectively. Expression correlation between RRDC3 and.2. immunohistochemical analysis of the expression of ARRDC3 in the clinical prostate cancer tissue microarray. The correlation between the expression level of ARRDC3 in the Taylor database and the clinical characteristics and prognosis of the prostate cancer patients was analyzed by statistical method. The expression of ARRDC3 was analyzed by the survival curve analysis and the Cox regression analysis. The relationship between the overall survival rate, the relationship of no biochemical recurrence survival and the predictor of the prognosis.3. construction of ARRDC3 inhibition expression and overexpression of DU145 and LNCaP cell lines, the effect of ARRDC3 on the function of prostate cancer cells by cell function test,.4. overexpressed DU145 cell lines to subcutaneous tissue of nude mice to construct subcutaneous transplantation tumor model Type, used to analyze the effect of ARRDC3 on the tumor formation of prostate cancer in the body.5. immunohistochemical analysis of the expression of Ki67, MMP-9 and ITG beta 4 in subcutaneous tumor tissues of nude mice. The expression of ITGB4 in subcutaneous tumor cells of nude mice was detected by Western blot. Results: 1. in the analysis of gene expression profiles of over expressed miR-30d, we obtained a group of differences. Down-regulation of heterosexual genes, in which ARRDC3 was obviously down regulated by.QRT-PCR and the results of Western blot analysis showed that there was a negative correlation between ARRDC3 and miR-30d,.2. tissue chip immunostaining analysis showed that the low ARRDC3 expression of prostate cancer tissue specimens was significantly correlated with the high Gleason score (P = 0.045).Taylor database analysis ARRDC3 results showed ARR. Down regulated expression of DC3, Gleason score, postoperative metastasis and biochemical recurrence (P respectively 0.0001,0.01 and 0.02).3.Kaplan-Meier and Log rank analysis results showed that low expression ARRDC3 was associated with poor prognosis of prostate cancer patients (P = 0.010). In prostate cancer as a single factor and multiple cause of the tumor suppressor gene.COX regression model ARRDC3 is an independent predictor of the survival of the prostate cancer patients without biochemical recurrence..4. cell function experiments show that the overexpressed ARRDC3 can reduce the proliferation, migration and invasion ability (P0.05) of DU145 and LNCaP cell lines, and the inhibition of ARRDC3 can enhance cell proliferation, migration and invasion (P0.05).5. animal experiment results The expression of ARRDC3 could significantly inhibit the growth rate of subcutaneous xenografts in nude mice (P0.05). Compared with the control group, the immunohistochemical score of Ki67 and MMP-9 was significantly lower (P0.05). It was confirmed that the overexpression of ARRDC3 inhibited the proliferation of tumor cells and the invasion of.6. protein blots and immunohistochemical results showed that ARRDC3 and ITG beta 4 were negatively correlated (P0.05). Conclusion: 1.ARRDC3 plays an inhibitory role in prostate cancer and inhibits the progress of prostate cancer and is an independent predictor for predicting the risk of postoperative biochemical recurrence and metastasis..2.ARRDC3 can inhibit the progression of prostate cancer through the ARRDC3-ITG beta 4 pathway. The ARRDC3-ITG beta 4 pathway can be used as a new therapeutic target for prostate cancer.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R737.25

【相似文獻(xiàn)】

相關(guān)博士學(xué)位論文 前1條

1 鄭煜;ARRDC3及ARRDC3-ITGβ4通路在人類前列腺癌中的表達(dá)及其作用的研究[D];南方醫(yī)科大學(xué);2017年

，

本文編號(hào)：2049677