S Rongxin1, L Pengfei, S Li, J Xiaochen, H Yihe. 1. Department of Orthopedics, Xiangya Hospital, Central South University, Changsha, China. dempseywell@163.com.
Abstract
OBJECTIVE: To study the effects of miR-340-5p on the proliferation of osteosarcoma (OS) U2OS cells. PATIENTS AND METHODS: miR-340-5p expression was measured by quantitative Real-time Polymerase Chain Reaction (qRT-PCR) in cancer tissues and paracancerous tissues from OS patients, and in OS cell line U2OS and normal osteoblast cell line hFOB1.19. The dual luciferase reporter vector was constructed to verify whether Signal transducers and activators of transcription 3 (STAT3) were a potential target gene of miR-340-5p. MiR-340-5p mimics/inhibitor was transfected into U2OS cells using liposomes, and transfection results were verified by qRT-PCR. Cell Counting Kit-8 (CCK-8) and Annexin V/PI were used to analyze the proliferation and apoptosis of transfected cells, respectively. Western Blot was used to detect STAT3 protein expression and Wnt/β-catenin pathway-related proteins after transfection. U2OS cells were injected into nude mice to observe the effects of over expression of miR-340-5p on tumor growth in vivo. RESULTS: miR-340-5p gene expression in OS tissues and U2OS cells was significantly lower than that in paracancerous tissues and hFOB1.19 cells. miR-340-5p directly interacted with the 3'-untranslated region (3'-UTR) of STAT3 gene and negatively regulated its expression. The increase of miR-340-5p expression could significantly inhibit U2OS proliferation in vitro and induce apoptosis, and vice versa. STAT3, β-catenin, c-Myc, TCF-4, CyclinD1, and ROCK1 protein expression in U2OS cells was significantly decreased after miR-340-5p over-expression, and vice versa. MiR-340-5p over-expression in nude mice significantly decreased tumor size and weight. CONCLUSIONS: Low expression of miR-340-5p in OS and U2OS cells could inhibit the course of OS by negatively regulating Wnt/β-catenin signaling pathway through targeting STAT3 gene. Therefore, miR-340-5p might be a potential biomarker and target for the diagnosis and treatment of OS.
OBJECTIVE: To study the effects of miR-340-5p on the proliferation of osteosarcoma (OS) U2OS cells. PATIENTS AND METHODS: miR-340-5p expression was measured by quantitative Real-time Polymerase Chain Reaction (qRT-PCR) in cancer tissues and paracancerous tissues from OS patients, and in OS cell line U2OS and normal osteoblast cell line hFOB1.19. The dual luciferase reporter vector was constructed to verify whether Signal transducers and activators of transcription 3 (STAT3) were a potential target gene of miR-340-5p. MiR-340-5p mimics/inhibitor was transfected into U2OS cells using liposomes, and transfection results were verified by qRT-PCR. Cell Counting Kit-8 (CCK-8) and Annexin V/PI were used to analyze the proliferation and apoptosis of transfected cells, respectively. Western Blot was used to detect STAT3 protein expression and Wnt/β-catenin pathway-related proteins after transfection. U2OS cells were injected into nude mice to observe the effects of over expression of miR-340-5p on tumor growth in vivo. RESULTS:miR-340-5p gene expression in OS tissues and U2OS cells was significantly lower than that in paracancerous tissues and hFOB1.19 cells. miR-340-5p directly interacted with the 3'-untranslated region (3'-UTR) of STAT3 gene and negatively regulated its expression. The increase of miR-340-5p expression could significantly inhibit U2OS proliferation in vitro and induce apoptosis, and vice versa. STAT3, β-catenin, c-Myc, TCF-4, CyclinD1, and ROCK1 protein expression in U2OS cells was significantly decreased after miR-340-5p over-expression, and vice versa. MiR-340-5p over-expression in nude mice significantly decreased tumor size and weight. CONCLUSIONS: Low expression of miR-340-5p in OS and U2OS cells could inhibit the course of OS by negatively regulating Wnt/β-catenin signaling pathway through targeting STAT3 gene. Therefore, miR-340-5p might be a potential biomarker and target for the diagnosis and treatment of OS.