The Effects Of LncRNA MALAT1in Bladder Urothelial Carcinoma And The Molecular Mechanism

Part One:The Expression Patterns and Functions of MALAT1in Bladder Urothelial Carcinoma[Purpose] Currently, lots of experiments confirmed lncRNA MALAT1overexpression in a variety of cancers, and regulation of cell proliferation, apoptosis, invasion, migration in vitro. The purpose of this study is to investigate the expression patterns of incRNA MALAT1and its function in bladder urothelial carcinoma.[Methods]Real-time PCR were used to detect the expression levels of MALAT1in tumor tissues and paired normal tissues in a total of45patients with bladder cancer. Bladder urothelial carcinoma EJ and5637cells were transfected with MALAT1siRNA(si-MALATl) or negative control siRNA(si-NC). MTT assay were used to determine cell proliferation changes. Flowcytometry assay were used to analyzed apoptosis and cell cycle.[Results] Compared with normal urothelium, MALAT1was overexpressed in bladder urothelial carcinoma. The MALAT1expression levels were greater in high-grade carcinoma than in low-grade carcinoma (P=0.0109); The MALAT1expression levels were greater in invasive carcinoma than in noninvasive carcinoma (P=0.0258); he MALAT1expression levels were greater in metastasis carcinoma than in no metastasis carcinoma (P=0.032). After transfected with si-MALATl, Cell proliferation inhibition, increased apoptosis, and arrest of G0/G1cell cycle were observed in both EJ and5637cells.[Conclusions]lncRNA MALAT1is overexpressed in bladder urothelial carcinoma, and associates with the grade, stage and metastasis. lncRNA MALAT1promotescell proliferation and inhibits apoptosis, playing an oncogenic role in bladder urothelial carcinoma. Part Two:The Mechanism of Oncogenic Effects of MALAT1in Bladder Urothelial Carcinoma[Purpose] EZH2was confirmed in several experiments are an oncogene, closely related to bladder cancer, having a plurality of tumor-associated downstream genes. This experiment is designed to study the expression patterns of EZH2in bladder cancer, and the interaction between MALAT1and EZH2.[Methods]Real-time PCR were used to detect the expression levels of EZH2in tumor tissues and paired normal tissues in a total of45patients with bladder cancer. RNA-IP were used to analyzed the interaction between MALAT1and EZH2protein. Bladder urothelial carcinoma EJ and5637cells were transfected with si-MALATl or si-NC. EZH2, Bcl2and c-myc expression levels were determined by immunoblotting and Real-time PCR.[Results] Compared with normal urothelium, EZH2was overexpressed in bladder urothelial carcinoma. The EZH2expression levels were greater in high-grade carcinoma than in low-grade carcinoma (P=0.0182); The EZH2expression levels were greater in invasive carcinoma than in noninvasive carcinoma (P=0.0206). The EZH2expression levels were positively correlated with MALAT1in bladder urothelial carcinoma(r=0.5242, P=0.0002). MALAT1binded EZH2protein in EJ and5637cells. After transfected with si-MALAT1, EZH2, Bcl2and c-myc protein levels were down-regulated in both EJ and5637cells, and Bcl2and c-myc mRNA expression were also down-regulated.[Conclusions] EZH2is overexpressed in bladder urothelial carcinoma, and associates with the grade and stage. The EZH2expression level is positively correlated with MALAT1in bladder cancer. MALAT1binds EZH2protein and regulates its downstream genes. Part Three:miR-101Inhibits the Expression of MALAT1and its Role in Bladder Urothelial Carcinoma[Purpose] miR-101has been reported down-regulated in a variety of cancers. EZH2is one of the target genes of miR-101. This study was to investigate the interaction between miR-101and MALAT1and its effects in bladder urothelial carcinoma.[Methods]Real-time PCR were used to detect the expression levels of mrR-101in tumor tissues and paired normal tissues in a total of45patients with bladder cancer. Bladder urothelial carcinoma EJ and5637cells were transfected with anti-miR-101or plasmid. MTT assay were used to analyze cell proliferation changes. EZH2protein levels were determined by immunoblotting.[Results] Compared with normal urothelium, miR-101was down-regulated in bladder urothelial carcinoma. The miR-101expression levels were negative correlated with MALAT1in bladder urothelial carcinoma(r=-0.5335, P=0.0002). The expression levels of MALATl were suppressed after the overexpression of miR-101, and was upregulated after silencing miR-101. Silencing miR-101promoted cell proliferation of EJ and5637cells, while transfected with si-MALAT1could suppress this trend. Silencing miR-101upregulated the expression of EZH2protein in EJ and5637cells, while transfected with si-MALAT1could suppress this trend.[Conclusions]. miR-101is down-regulated in bladder urothelial carcinoma, and its expression level is negative correlated with MALAT1in bladder cancer. miR-101regulates the expression of MALAT1in bladder cancer cells. miR-101inhibits cell proliferation through MALAT1partly. MALAT1suppresses the inhibition effects of miR-101on EZH2partially.