Yi-Chia Lin1, Ji-Fan Lin2, Te-Fu Tsai1, Kuang-Yu Chou1, Hung-En Chen3, Thomas I-Sheng Hwang4. 1. Department of Urology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan. 2. Central Laboratory, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan. 3. Department of Urology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan. 4. Department of Urology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan; School of Medicine, Taipei Medical University, Taipei, Taiwan. Electronic address: M001009@ms.skh.org.tw.
Abstract
BACKGROUND: Prostate cancer (PCa) is a leading cause of cancer-related death in men, which emphasizes the need for novel therapeutic approaches. Targeting microRNA (miRNA) has been considered as a therapeutic strategy against cancers. Human miR-204-5p potentially targeting BCL2 has been reported to be downregulated in various cancers. We hypothesized that miR-204-5p overexpression induces cancer cell apoptosis by repressing BCL2 expression. METHODS: A vector harboring mature miR-204-5p was constructed and delivered into human PCa cells. The expression level of miR-204-5p was determined by miRNA quantitative polymerase chain reaction (QPCR). Luciferase reporter assays were performed to verify the function of mature miR-204-5p and its direct binding to BCL2 transcripts. The expression levels of BCL-2 messenger RNA (mRNA) and protein samples were measured by QPCR and Western blot, respectively. Cell viability was detected by WST-1 assays. Induction of apoptosis was determined by increased levels of cleavage caspase 3 and caspase 3/7 activity. RESULTS: The expression levels of miR-204-5p were downregulated in PCa cells compared with normal prostate epithelial cells. Transfection of pSM-204 resulted in up to 6.2-fold higher expression of miR-204-5p when compared with pSM control. The mRNA levels of several potential target genes of miR-204-5p were decreased in pSM-204-transfected PC3 and Rv1 cells. BCL2 mRNA and protein expression decreased in miR-204-5p-transfected cells, which led to cytochrome C release from mitochondria. It subsequently increased cleaved caspase 3 and caspase 3/7 activities and reduced cell viability. Cotransfection of a reporter vector harboring the BCL2 3'-untranslated region to compete with endogenous transcripts partially rescued miR-204-5p-induced apoptosis. CONCLUSION: Human miR-204-5p targets BCL2 in PCa cells. Restoration of miR-204-5p in PCa could therefore be considered as a novel strategy by targeting antiapoptotic BCL2.
BACKGROUND:Prostate cancer (PCa) is a leading cause of cancer-related death in men, which emphasizes the need for novel therapeutic approaches. Targeting microRNA (miRNA) has been considered as a therapeutic strategy against cancers. HumanmiR-204-5p potentially targeting BCL2 has been reported to be downregulated in various cancers. We hypothesized that miR-204-5p overexpression induces cancer cell apoptosis by repressing BCL2 expression. METHODS: A vector harboring mature miR-204-5p was constructed and delivered into human PCa cells. The expression level of miR-204-5p was determined by miRNA quantitative polymerase chain reaction (QPCR). Luciferase reporter assays were performed to verify the function of mature miR-204-5p and its direct binding to BCL2 transcripts. The expression levels of BCL-2 messenger RNA (mRNA) and protein samples were measured by QPCR and Western blot, respectively. Cell viability was detected by WST-1 assays. Induction of apoptosis was determined by increased levels of cleavage caspase 3 and caspase 3/7 activity. RESULTS: The expression levels of miR-204-5p were downregulated in PCa cells compared with normal prostate epithelial cells. Transfection of pSM-204 resulted in up to 6.2-fold higher expression of miR-204-5p when compared with pSM control. The mRNA levels of several potential target genes of miR-204-5p were decreased in pSM-204-transfected PC3 and Rv1 cells. BCL2 mRNA and protein expression decreased in miR-204-5p-transfected cells, which led to cytochrome C release from mitochondria. It subsequently increased cleaved caspase 3 and caspase 3/7 activities and reduced cell viability. Cotransfection of a reporter vector harboring the BCL2 3'-untranslated region to compete with endogenous transcripts partially rescued miR-204-5p-induced apoptosis. CONCLUSION:HumanmiR-204-5p targets BCL2 in PCa cells. Restoration of miR-204-5p in PCa could therefore be considered as a novel strategy by targeting antiapoptotic BCL2.
Authors: Asmahan A El Ezzi; Vladyslav G Boyko; Monika T Baker; Wissam R Zaidan; Kalim M Hraiki; Mohammad A El Saidi; Ruhul H Kuddus Journal: Asian Pac J Cancer Prev Date: 2017-01-01
Authors: Aleksandra Butrym; Piotr Łacina; Kazimierz Kuliczkowski; Katarzyna Bogunia-Kubik; Grzegorz Mazur Journal: BMC Cancer Date: 2018-01-30 Impact factor: 4.430