MicroRNA-103靶向PDCD10抑制前列腺癌的作用及机制研究
发布时间：2018-08-05 11:27
【摘要】：研究背景和目的:前列腺癌严重影响着世界范围内的男性健康,该疾病在男性癌症中的患病率已位居第二位,前列腺癌在中老年男性中的发病率随年龄的增长快速上升,该疾病难以治疗并且致死率也非常高,对人类的健康产生严重的威胁。所以目前依据前列腺癌的发病机制寻找该疾病的预防以及治疗靶点和新方法迫在眉睫。近年来的报道显示,前列腺癌的发病机制主要有雄性激素依赖性前列腺癌以及包括原发性和继发性的激素非依赖性前列腺癌,并且指出雄性激素依赖性前列腺癌在该疾病的发生发展过程中占据重要位置。近来的研究发现微小RNA-103(microRNA-103,miR-103)调控程序性细胞死亡因子10(programmed cell death factor 10,PDCD10)在前列腺细胞中的表达对前列腺癌的发生发展具有较大影响,并且在前列腺癌发病机制中具有非常关键的作用。PDCD10能够调控机体细胞的凋亡过程,对维持机体细胞的健康存活具有非常重要的作用,当PDCD10在前列腺细胞中的表达失调会引起前列腺病变,甚至最终能够引起前列腺癌变,同时PDCD10在人类细胞中的分布非常广泛,能够参与人体多种疾病的生理病理程序,目前已经发现PDCD10在多种癌症中发挥非常重要的调控作用。此外miR-103是一类miRNA,目前已发现其在直结肠癌等多种癌细胞中调控癌症的发生发展过程。miRNAs一般通过调控与癌症相关基因的表达,发挥致癌或者抑癌功能。目前研究发现miR-103可能通过与靶基因的非编码区结合调控目的基因在细胞中的表达,进而影响该基因功能的发挥,最终对多种癌症的发病过程进行调控。采用生物信息学方法(使用miRanda、TargetScan、PicTar等软件)对miR-103和PDCD10基因的靶向匹配关系进行预测,发现PDCD10可能是miR-103的潜在靶基因。综上,miR-103对PDCD10在前列腺细胞中表达的调控与前列腺癌的发生发展可能具有紧密联系,但是目前对于mi R-103和PDCD10与前列腺癌之间的联系以及两者之间的调控模式尚未有研究,所以相关的重要科学问题急需深入了解及研究。因此,本课题拟研究miR-103及PDCD10在前列腺癌患者癌细胞中的表达变化,并且以两者之间的表达变化模式为切入点,阐明miR-103及PDCD10在前列腺癌发病机制中作用,有助于从新的位点了解前列腺癌的发病机制,为前列腺癌的预防和治疗提供新的靶点及理论基础。研究方法:1.利用Real-time PCR方法检测miR-103在前列腺癌患者组织和前列腺癌细胞系中的表达变化规律,并对结果进行统计学分析。2.运用Western blotting以及Real-time PCR方法检测PDCD10在前列腺癌患者组织和前列腺细胞系中的表达变化规律,并对结果进行统计学分析。3.体外实验中,利用MTT实验、流式细胞实验、细胞计数以及细胞转染等方法检测miR-103以及PDCD10对前列腺癌细胞的增殖、凋亡以及细胞周期的影响。另外利用酵母双杂实验检测miR-103对PDCD10与MST4互作的影响,利用Western blotting以及Real-time PCR方法检测miR-103对ERK蛋白磷酸化的影响,利用荧光素酶报告系统分析miR-103与PDCD10的靶向关系。研究结果:1.miR-103在前列腺癌患者组织和前列腺癌系中表达显著低于正常对照。2.体外实验中,miR-103在前列腺癌细胞中的表达变化与前列腺癌细胞的增殖率、侵袭呈负相关性,与前列腺癌细胞的凋亡率呈正相关性;miR-103在前列腺癌细胞中超表达时,能够抑制癌细胞由G1期向S期转变。3.在前列腺癌患者中,PDCD10在前列腺癌组织和细胞系中的表达量显著高于正常前列腺内皮细胞。4.在体外实验中,PDCD10在前列腺癌细胞中的表达变化与前列腺癌细胞的凋亡呈负相关,与前列腺癌细胞的增殖率、侵袭呈正相关;在前列腺细胞中PDCD10蛋白能够与MST4蛋白互作,并且两者能够协同表达,PDCD10表达上调与ERK蛋白的磷酸化呈现正相关性,同时能够增加前列腺癌细胞抵御氧化应激对细胞的伤害。5.PDCD10的表达可以受miR-103直接调控,miR-103在前列腺癌细胞中超表达时会抑制PDCD10的表达,并且引起MST4蛋白在前列腺癌细胞中的表达下调,ERK蛋白的磷酸化水平也下降,导致前列腺癌抵御氧化应激的能力下降。研究结论:1.在前列腺癌患者癌细胞中miR-103表达下降,PDCD10的表达水平上升,miR-103表达下降以及PDCD10表达上调会引起前列腺癌细胞持续增殖,阻碍细胞凋亡。2.在前列腺癌细胞中,miR-103能够通过直接调控靶基因PDCD10的表达抑制前列腺癌细胞的增殖、侵袭,同时抑制前列腺癌细胞周期由G1期向S期转变。3.miR-103能够通过抑制抑制PDCD10的表达,使得MST4蛋白表达下降以及ERK磷酸化水平下降,最终引起前列腺癌细胞抵抗氧化应激刺激的能力下降。4.miR-103靶向调控PDCD10抑制前列腺癌的发生和进展,为前列腺的治疗提供了新的理论依据和新的靶点。
[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.
【学位授予单位】：第四军医大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R737.25
，
本文编号：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.
【学位授予单位】：第四军医大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R737.25
，
本文编号：2165656
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