Li-Li Zhang1, Yan-Jun Guo1, Chun-Na Zhao1, Jian-Yun Gao2. 1. Department of Gastroenterology, Qiqihaer Medical College Affiliated training hospital Daqing Oil Field General Hospital, Daqing City, Heilongjiang Province, China. 2. Department of Gastroenterology, Qiqihaer Medical College Affiliated training hospital Daqing Oil Field General Hospital, Daqing City, Heilongjiang Province, China. Electronic address: zhanglili020@126.com.
Abstract
OBJECTIVE: To explore the role of miR-214 in the progression of hepatocellular carcinoma (HCC) and its inhibitory mechanisms in depressing the signaling pathway of β-catenin, this study was conducted. METHODS: We ectopically expressed miR-214 in HepG2 cells to obtain cell lines Lv-miR-214-HepG2 and their control Lv-control-HepG2. Differences between the two cell lines were compared in cell growth, proliferation, colony forming ability and cell cycles. RT-PCR method was applied for the quantification of β-catenin mRNA expression. Western-blot method was applied for the determination of the protein level of β-catenin and their downstream targets (ie. Cyclin D1, c-Myc and TCF-1). The effect of miR-214 on cells was further explored through RNA interference and restoring miR-214 expression. RESULTS: In comparison with negative (Lv-control-HepG2) and blank (HepG2) control, a significant inhibition of cell growth and proliferation caused by miR-214 was observed after 48∼72h of cell culture experiments (P<0.05). The miR-214 treatment resulted in a colony forming efficiency of (23.28±3.26)%, which was significantly lower than that of negative control [(51.31±3.97)%] (P<0.05). According to FCM results, the experimental group, compared with control, showed a higher proportion of cells in G0/G1 phase [(70.32±3.12)%] but a lower proportion in S phase [(18.42±2.90)%] (P<0.05). The MTT assay demonstrated a significant inhibition of the proliferation and β-catenin expression of HCC cells compared with control (P<0.05), while no significant difference was observed after HCC cells being transfected with β-catenin overexpression plasmid (P>0.05). By comparing to the RT-PCR and Western-blot results of control, the miR-214 treatment led to a slightly decrease in the β-catenin mRNA expression (P>0.05), but an extremely inhibition in the protein level of β-catenin and its downstream targets Cyclin D1, c-Myc, and TCF-1 (P<0.05). CONCLUSIONS: miR-214 functions as a suppressor during the progression of HCC, and its inhibitory role was achieved by down-regulating β-catenin signaling pathway.
OBJECTIVE: To explore the role of miR-214 in the progression of hepatocellular carcinoma (HCC) and its inhibitory mechanisms in depressing the signaling pathway of β-catenin, this study was conducted. METHODS: We ectopically expressed miR-214 in HepG2 cells to obtain cell lines Lv-miR-214-HepG2 and their control Lv-control-HepG2. Differences between the two cell lines were compared in cell growth, proliferation, colony forming ability and cell cycles. RT-PCR method was applied for the quantification of β-catenin mRNA expression. Western-blot method was applied for the determination of the protein level of β-catenin and their downstream targets (ie. Cyclin D1, c-Myc and TCF-1). The effect of miR-214 on cells was further explored through RNA interference and restoring miR-214 expression. RESULTS: In comparison with negative (Lv-control-HepG2) and blank (HepG2) control, a significant inhibition of cell growth and proliferation caused by miR-214 was observed after 48∼72h of cell culture experiments (P<0.05). The miR-214 treatment resulted in a colony forming efficiency of (23.28±3.26)%, which was significantly lower than that of negative control [(51.31±3.97)%] (P<0.05). According to FCM results, the experimental group, compared with control, showed a higher proportion of cells in G0/G1 phase [(70.32±3.12)%] but a lower proportion in S phase [(18.42±2.90)%] (P<0.05). The MTT assay demonstrated a significant inhibition of the proliferation and β-catenin expression of HCC cells compared with control (P<0.05), while no significant difference was observed after HCC cells being transfected with β-catenin overexpression plasmid (P>0.05). By comparing to the RT-PCR and Western-blot results of control, the miR-214 treatment led to a slightly decrease in the β-catenin mRNA expression (P>0.05), but an extremely inhibition in the protein level of β-catenin and its downstream targets Cyclin D1, c-Myc, and TCF-1 (P<0.05). CONCLUSIONS:miR-214 functions as a suppressor during the progression of HCC, and its inhibitory role was achieved by down-regulating β-catenin signaling pathway.
Authors: G Y He; J L Hu; L Zhou; X H Zhu; S N Xin; D Zhang; G F Lu; W T Liao; Y Q Ding; L Liang Journal: Br J Cancer Date: 2016-11-03 Impact factor: 7.640