Ling Yang1, Ning Wei2, Li Wang1, Xiang Wang1, Qing-Huai Liu3. 1. Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing, Jiangsu, 210029, China. 2. Department of Child Healthcare, Obstetrics and Gynecology Hospital Affiliated to Nanjing Medical University, Nanjing, China. 3. Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing, Jiangsu, 210029, China. liuqinghuainjmu@126.com.
Abstract
PURPOSE: Retinoblastoma (Rb) is the most common intraocular tumor in children. MicroRNAs (miRNAs) play a crucial role in gene regulation and cell growth/apoptosis/differentiation. The current study aimed to investigate the role of miR-498 in Rb. METHODS: Quantitative real-time polymerase chain reaction (QRT-PCR) was used to test mRNA level of miR-498. http://www.targetscan.org and http://www.microrna.org were applied to predict target of miR-498. Dual-luciferase reporter assay was applied to investigate if miR-498 targeted cell cycle progression 1 (CCPG1). Western blot (WB) was carried out to assess CCPG1 protein levels. 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay was used to evaluate cell proliferation. Annexin-V Fluorescein (FITC) was adopted to explore cell apoptosis. RESULTS: In Y79 cells, miR-498 was higher than in normal ARPE-19 cells. MiR-498 could recognize CCPG1-3' untranslated region (UTR). CCPG1 protein level was remarkably decreased when overexpressed miR-498, nevertheless, significantly increased when inhibiting miR-498. Y79 cells that were transfected with miR-498 mimics manifested notable cell apoptosis down-regulation and cell proliferation promotion; whereas, those transfected with miR-498 inhibitor displayed significant cell apoptosis up-regulation and cell proliferation inhibition compared with control group. CONCLUSION: Taken together, miR-498 promotes cell proliferation and inhibits cell apoptosis in Rb by directly targeting CCPG1.
PURPOSE:Retinoblastoma (Rb) is the most common intraocular tumor in children. MicroRNAs (miRNAs) play a crucial role in gene regulation and cell growth/apoptosis/differentiation. The current study aimed to investigate the role of miR-498 in Rb. METHODS: Quantitative real-time polymerase chain reaction (QRT-PCR) was used to test mRNA level of miR-498. http://www.targetscan.org and http://www.microrna.org were applied to predict target of miR-498. Dual-luciferase reporter assay was applied to investigate if miR-498 targeted cell cycle progression 1 (CCPG1). Western blot (WB) was carried out to assess CCPG1 protein levels. 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay was used to evaluate cell proliferation. Annexin-V Fluorescein (FITC) was adopted to explore cell apoptosis. RESULTS: In Y79 cells, miR-498 was higher than in normal ARPE-19 cells. MiR-498 could recognize CCPG1-3' untranslated region (UTR). CCPG1 protein level was remarkably decreased when overexpressed miR-498, nevertheless, significantly increased when inhibiting miR-498. Y79 cells that were transfected with miR-498 mimics manifested notable cell apoptosis down-regulation and cell proliferation promotion; whereas, those transfected with miR-498 inhibitor displayed significant cell apoptosis up-regulation and cell proliferation inhibition compared with control group. CONCLUSION: Taken together, miR-498 promotes cell proliferation and inhibits cell apoptosis in Rb by directly targeting CCPG1.
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