MicroRNA-103靶向PDCD10抑制前列腺癌的作用及機制研究
發(fā)布時間：2018-08-05 11:27
【摘要】：研究背景和目的:前列腺癌嚴重影響著世界范圍內的男性健康,該疾病在男性癌癥中的患病率已位居第二位,前列腺癌在中老年男性中的發(fā)病率隨年齡的增長快速上升,該疾病難以治療并且致死率也非常高,對人類的健康產生嚴重的威脅。所以目前依據(jù)前列腺癌的發(fā)病機制尋找該疾病的預防以及治療靶點和新方法迫在眉睫。近年來的報道顯示,前列腺癌的發(fā)病機制主要有雄性激素依賴性前列腺癌以及包括原發(fā)性和繼發(fā)性的激素非依賴性前列腺癌,并且指出雄性激素依賴性前列腺癌在該疾病的發(fā)生發(fā)展過程中占據(jù)重要位置。近來的研究發(fā)現(xiàn)微小RNA-103(microRNA-103,miR-103)調控程序性細胞死亡因子10(programmed cell death factor 10,PDCD10)在前列腺細胞中的表達對前列腺癌的發(fā)生發(fā)展具有較大影響,并且在前列腺癌發(fā)病機制中具有非常關鍵的作用。PDCD10能夠調控機體細胞的凋亡過程,對維持機體細胞的健康存活具有非常重要的作用,當PDCD10在前列腺細胞中的表達失調會引起前列腺病變,甚至最終能夠引起前列腺癌變,同時PDCD10在人類細胞中的分布非常廣泛,能夠參與人體多種疾病的生理病理程序,目前已經發(fā)現(xiàn)PDCD10在多種癌癥中發(fā)揮非常重要的調控作用。此外miR-103是一類miRNA,目前已發(fā)現(xiàn)其在直結腸癌等多種癌細胞中調控癌癥的發(fā)生發(fā)展過程。miRNAs一般通過調控與癌癥相關基因的表達,發(fā)揮致癌或者抑癌功能。目前研究發(fā)現(xiàn)miR-103可能通過與靶基因的非編碼區(qū)結合調控目的基因在細胞中的表達,進而影響該基因功能的發(fā)揮,最終對多種癌癥的發(fā)病過程進行調控。采用生物信息學方法(使用miRanda、TargetScan、PicTar等軟件)對miR-103和PDCD10基因的靶向匹配關系進行預測,發(fā)現(xiàn)PDCD10可能是miR-103的潛在靶基因。綜上,miR-103對PDCD10在前列腺細胞中表達的調控與前列腺癌的發(fā)生發(fā)展可能具有緊密聯(lián)系,但是目前對于mi R-103和PDCD10與前列腺癌之間的聯(lián)系以及兩者之間的調控模式尚未有研究,所以相關的重要科學問題急需深入了解及研究。因此,本課題擬研究miR-103及PDCD10在前列腺癌患者癌細胞中的表達變化,并且以兩者之間的表達變化模式為切入點,闡明miR-103及PDCD10在前列腺癌發(fā)病機制中作用,有助于從新的位點了解前列腺癌的發(fā)病機制,為前列腺癌的預防和治療提供新的靶點及理論基礎。研究方法:1.利用Real-time PCR方法檢測miR-103在前列腺癌患者組織和前列腺癌細胞系中的表達變化規(guī)律,并對結果進行統(tǒng)計學分析。2.運用Western blotting以及Real-time PCR方法檢測PDCD10在前列腺癌患者組織和前列腺細胞系中的表達變化規(guī)律,并對結果進行統(tǒng)計學分析。3.體外實驗中,利用MTT實驗、流式細胞實驗、細胞計數(shù)以及細胞轉染等方法檢測miR-103以及PDCD10對前列腺癌細胞的增殖、凋亡以及細胞周期的影響。另外利用酵母雙雜實驗檢測miR-103對PDCD10與MST4互作的影響,利用Western blotting以及Real-time PCR方法檢測miR-103對ERK蛋白磷酸化的影響,利用熒光素酶報告系統(tǒng)分析miR-103與PDCD10的靶向關系。研究結果:1.miR-103在前列腺癌患者組織和前列腺癌系中表達顯著低于正常對照。2.體外實驗中,miR-103在前列腺癌細胞中的表達變化與前列腺癌細胞的增殖率、侵襲呈負相關性,與前列腺癌細胞的凋亡率呈正相關性;miR-103在前列腺癌細胞中超表達時,能夠抑制癌細胞由G1期向S期轉變。3.在前列腺癌患者中,PDCD10在前列腺癌組織和細胞系中的表達量顯著高于正常前列腺內皮細胞。4.在體外實驗中,PDCD10在前列腺癌細胞中的表達變化與前列腺癌細胞的凋亡呈負相關,與前列腺癌細胞的增殖率、侵襲呈正相關;在前列腺細胞中PDCD10蛋白能夠與MST4蛋白互作,并且兩者能夠協(xié)同表達,PDCD10表達上調與ERK蛋白的磷酸化呈現(xiàn)正相關性,同時能夠增加前列腺癌細胞抵御氧化應激對細胞的傷害。5.PDCD10的表達可以受miR-103直接調控,miR-103在前列腺癌細胞中超表達時會抑制PDCD10的表達,并且引起MST4蛋白在前列腺癌細胞中的表達下調,ERK蛋白的磷酸化水平也下降,導致前列腺癌抵御氧化應激的能力下降。研究結論:1.在前列腺癌患者癌細胞中miR-103表達下降,PDCD10的表達水平上升,miR-103表達下降以及PDCD10表達上調會引起前列腺癌細胞持續(xù)增殖,阻礙細胞凋亡。2.在前列腺癌細胞中,miR-103能夠通過直接調控靶基因PDCD10的表達抑制前列腺癌細胞的增殖、侵襲,同時抑制前列腺癌細胞周期由G1期向S期轉變。3.miR-103能夠通過抑制抑制PDCD10的表達,使得MST4蛋白表達下降以及ERK磷酸化水平下降,最終引起前列腺癌細胞抵抗氧化應激刺激的能力下降。4.miR-103靶向調控PDCD10抑制前列腺癌的發(fā)生和進展,為前列腺的治療提供了新的理論依據(jù)和新的靶點。
[Abstract]:Research background and purpose: prostate cancer seriously affects the health of men worldwide. The incidence of the disease is the second in male cancer. The incidence of prostate cancer in middle-aged and old men is rising rapidly with age. The disease is difficult to treat and has a high mortality rate, which produces serious health to human health. So it is imminent to find the prevention and treatment targets and new methods of the disease according to the pathogenesis of prostate cancer. Recent reports have shown that the main pathogenesis of prostate cancer is androgen dependent prostate cancer and primary and secondary hormone non dependent prostate cancer, and the male irritable disease is pointed out. Recent studies have found that the expression of RNA-103 (microRNA-103, miR-103) regulatory programmed cell death factor 10 (programmed cell death factor 10, PDCD10) in prostate cells has a great influence on the development of prostate cancer. The key role of the pathogenesis of prostate cancer is that.PDCD10 can regulate the apoptosis process of the body cells, which plays a very important role in maintaining the healthy survival of the body cells. When the expression of PDCD10 in the prostate cells is dysfunctional, it can cause prostate disease, and even eventually lead to prostate cancer, while PDCD10 is very thin in human beings. The distribution of the cell is very extensive and can participate in the physiological and pathological procedures of various diseases. PDCD10 has been found to play a very important regulatory role in many kinds of cancer. In addition, miR-103 is a kind of miRNA. At present, it has been found that.MiRNAs regulates and regulates the progression of cancer in a variety of cancer cells such as colon cancer. The expression of cancer related genes exerts carcinogenic or tumor suppressor functions. At present, it is found that miR-103 may regulate the expression of target genes in cells by combining the non coding regions of the target genes, and then affect the function of the gene, and ultimately regulate the pathogenesis of various cancers. Bioinformatics (using miRanda, Ta) The target matching relationship between miR-103 and PDCD10 genes is predicted by rgetScan, PicTar and other software. It is found that PDCD10 may be a potential target gene for miR-103. To sum up, the regulation of miR-103 on the expression of PDCD10 in prostate cells may be closely related to the development of prostate cancer, but it is currently available for MI R-103 and PDCD10 and prostate cancer. The relationship between the two and the mode of regulation between the two has not been studied, so the important scientific questions need to be deeply understood and studied. Therefore, this topic intends to study the changes in the expression of miR-103 and PDCD10 in the cancer cells of the patients with prostate cancer, and to clarify that miR-103 and PDCD10 are in the forefront. The role of adenocarcinoma pathogenesis is helpful to understand the pathogenesis of prostate cancer from new sites and provide new targets and theoretical basis for the prevention and treatment of prostate cancer. Research methods: 1. the expression of miR-103 in the tissues of prostate cancer patients and prostate cancer cell lines was detected by Real-time PCR method, and the results were added to the results. Statistical analysis.2. used Western blotting and Real-time PCR to detect the changes in the expression of PDCD10 in the tissues and the prostate cell lines of the prostate cancer patients, and the results were statistically analyzed in.3. in vitro, and miR-103 and P were detected by MTT experiment, flow cytometry, cell count and cell transfection. The effect of DCD10 on the proliferation, apoptosis and cell cycle of prostate cancer cells. In addition, the effect of miR-103 on the interaction between PDCD10 and MST4 was detected by yeast double heterozygosity. The effect of miR-103 on the phosphorylation of ERK protein was detected by Western blotting and Real-time PCR method. The target direction of miR-103 and PDCD10 was analyzed by the fluorescent enzyme report system. The results: the expression of 1.miR-103 in the tissue and prostate cancer of the prostate cancer patients was significantly lower than that in the normal control.2. in vitro. The expression of miR-103 in the prostate cancer cells was negatively correlated with the proliferation rate and invasion of the prostate cancer cells, and was positively correlated with the apoptosis rate of the prostate cancer cells; miR-103 was in the prostate cancer. The expression of PDCD10 in prostate cancer patients was significantly higher than that of normal prostate cancer cells and cell lines. The expression of PDCD10 in prostate cancer tissues and cell lines was significantly higher than that of normal prostate endothelial cells (.4.) in vitro. The expression of PDCD10 in prostate cancer cells was negatively correlated with the apoptosis of prostate cancer cells, and the expression of.3. in prostate cancer cells was negatively correlated with the apoptosis of prostate cancer cells. The proliferation rate and invasion of prostate cancer cells are positively correlated. In prostate cells, PDCD10 protein can interact with MST4 protein, and both can be co expressed. The up regulation of PDCD10 expression is positively correlated with the phosphorylation of ERK protein, and can also increase the expression of.5.PDCD10 in prostate cancer cells against oxidative stress. Under the direct regulation of miR-103, the overexpression of miR-103 in prostate cancer cells inhibits the expression of PDCD10, and causes the expression of MST4 protein in the prostate cancer cells to decrease, and the level of phosphorylation of ERK protein also decreases, which leads to the decline in the ability of prostate cancer to resist oxidative stress. The conclusion: 1. in the cancer cells of prostate cancer patients, miR-103 The expression level of PDCD10 increased, the expression level of miR-103 and the up-regulated expression of PDCD10 may cause the proliferation of prostate cancer cells and inhibit the apoptosis of.2. in prostate cancer cells. MiR-103 can inhibit the proliferation, invasion and inhibition of prostate cancer cell cycle by direct regulation of the expression of target gene PDCD10. The transition from G1 to S phase.3.miR-103 can inhibit the expression of PDCD10, decrease the expression of MST4 protein and decrease the level of ERK phosphorylation, and eventually lead to the decline in the ability of prostate cancer cells to resist oxidative stress stimulation..4.miR-103 targeted regulation of PDCD10 to inhibit the occurrence and development of prostate cancer and provide the prostate treatment. New theoretical basis and new targets.
【學位授予單位】：第四軍醫(yī)大學
【學位級別】：博士
【學位授予年份】：2017
【分類號】：R737.25
，
本文編號：2165656
[Abstract]:Research background and purpose: prostate cancer seriously affects the health of men worldwide. The incidence of the disease is the second in male cancer. The incidence of prostate cancer in middle-aged and old men is rising rapidly with age. The disease is difficult to treat and has a high mortality rate, which produces serious health to human health. So it is imminent to find the prevention and treatment targets and new methods of the disease according to the pathogenesis of prostate cancer. Recent reports have shown that the main pathogenesis of prostate cancer is androgen dependent prostate cancer and primary and secondary hormone non dependent prostate cancer, and the male irritable disease is pointed out. Recent studies have found that the expression of RNA-103 (microRNA-103, miR-103) regulatory programmed cell death factor 10 (programmed cell death factor 10, PDCD10) in prostate cells has a great influence on the development of prostate cancer. The key role of the pathogenesis of prostate cancer is that.PDCD10 can regulate the apoptosis process of the body cells, which plays a very important role in maintaining the healthy survival of the body cells. When the expression of PDCD10 in the prostate cells is dysfunctional, it can cause prostate disease, and even eventually lead to prostate cancer, while PDCD10 is very thin in human beings. The distribution of the cell is very extensive and can participate in the physiological and pathological procedures of various diseases. PDCD10 has been found to play a very important regulatory role in many kinds of cancer. In addition, miR-103 is a kind of miRNA. At present, it has been found that.MiRNAs regulates and regulates the progression of cancer in a variety of cancer cells such as colon cancer. The expression of cancer related genes exerts carcinogenic or tumor suppressor functions. At present, it is found that miR-103 may regulate the expression of target genes in cells by combining the non coding regions of the target genes, and then affect the function of the gene, and ultimately regulate the pathogenesis of various cancers. Bioinformatics (using miRanda, Ta) The target matching relationship between miR-103 and PDCD10 genes is predicted by rgetScan, PicTar and other software. It is found that PDCD10 may be a potential target gene for miR-103. To sum up, the regulation of miR-103 on the expression of PDCD10 in prostate cells may be closely related to the development of prostate cancer, but it is currently available for MI R-103 and PDCD10 and prostate cancer. The relationship between the two and the mode of regulation between the two has not been studied, so the important scientific questions need to be deeply understood and studied. Therefore, this topic intends to study the changes in the expression of miR-103 and PDCD10 in the cancer cells of the patients with prostate cancer, and to clarify that miR-103 and PDCD10 are in the forefront. The role of adenocarcinoma pathogenesis is helpful to understand the pathogenesis of prostate cancer from new sites and provide new targets and theoretical basis for the prevention and treatment of prostate cancer. Research methods: 1. the expression of miR-103 in the tissues of prostate cancer patients and prostate cancer cell lines was detected by Real-time PCR method, and the results were added to the results. Statistical analysis.2. used Western blotting and Real-time PCR to detect the changes in the expression of PDCD10 in the tissues and the prostate cell lines of the prostate cancer patients, and the results were statistically analyzed in.3. in vitro, and miR-103 and P were detected by MTT experiment, flow cytometry, cell count and cell transfection. The effect of DCD10 on the proliferation, apoptosis and cell cycle of prostate cancer cells. In addition, the effect of miR-103 on the interaction between PDCD10 and MST4 was detected by yeast double heterozygosity. The effect of miR-103 on the phosphorylation of ERK protein was detected by Western blotting and Real-time PCR method. The target direction of miR-103 and PDCD10 was analyzed by the fluorescent enzyme report system. The results: the expression of 1.miR-103 in the tissue and prostate cancer of the prostate cancer patients was significantly lower than that in the normal control.2. in vitro. The expression of miR-103 in the prostate cancer cells was negatively correlated with the proliferation rate and invasion of the prostate cancer cells, and was positively correlated with the apoptosis rate of the prostate cancer cells; miR-103 was in the prostate cancer. The expression of PDCD10 in prostate cancer patients was significantly higher than that of normal prostate cancer cells and cell lines. The expression of PDCD10 in prostate cancer tissues and cell lines was significantly higher than that of normal prostate endothelial cells (.4.) in vitro. The expression of PDCD10 in prostate cancer cells was negatively correlated with the apoptosis of prostate cancer cells, and the expression of.3. in prostate cancer cells was negatively correlated with the apoptosis of prostate cancer cells. The proliferation rate and invasion of prostate cancer cells are positively correlated. In prostate cells, PDCD10 protein can interact with MST4 protein, and both can be co expressed. The up regulation of PDCD10 expression is positively correlated with the phosphorylation of ERK protein, and can also increase the expression of.5.PDCD10 in prostate cancer cells against oxidative stress. Under the direct regulation of miR-103, the overexpression of miR-103 in prostate cancer cells inhibits the expression of PDCD10, and causes the expression of MST4 protein in the prostate cancer cells to decrease, and the level of phosphorylation of ERK protein also decreases, which leads to the decline in the ability of prostate cancer to resist oxidative stress. The conclusion: 1. in the cancer cells of prostate cancer patients, miR-103 The expression level of PDCD10 increased, the expression level of miR-103 and the up-regulated expression of PDCD10 may cause the proliferation of prostate cancer cells and inhibit the apoptosis of.2. in prostate cancer cells. MiR-103 can inhibit the proliferation, invasion and inhibition of prostate cancer cell cycle by direct regulation of the expression of target gene PDCD10. The transition from G1 to S phase.3.miR-103 can inhibit the expression of PDCD10, decrease the expression of MST4 protein and decrease the level of ERK phosphorylation, and eventually lead to the decline in the ability of prostate cancer cells to resist oxidative stress stimulation..4.miR-103 targeted regulation of PDCD10 to inhibit the occurrence and development of prostate cancer and provide the prostate treatment. New theoretical basis and new targets.
【學位授予單位】：第四軍醫(yī)大學
【學位級別】：博士
【學位授予年份】：2017
【分類號】：R737.25
，
本文編號：2165656
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