High-Throughput Sequencing Strategy for miR-146b-regulated circRNA Expression in Hepatic Stellate Cells
Na Cheng, Juhua Xiao, Shanfei Ge, Juntao Li, Jiansheng Huang, Xiaoping Wu, Shouhua Zhang, Tianxin Xiang
Department of Infectious Diseases, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, China (mainland)
Med Sci Monit 2018; 24:8699-8706
This study was designed to detect and analyze miR-146b-mediated circular RNA (circRNA) expression in hepatic stellate cells.
MATERIAL AND METHODS: The experiment was divided into a control group and a siRNA-miR-146b group. The interference efficiency of siRNA-miR-146b was confirmed by real-time quantitative reverse transcription PCR (qRT-PCR) and the cells were collected, and total RNA was collected for high flux sequencing. The miRNA-targeted carcass were predicted. Finally, the expression of 5 circRNAs was verified by qRT-PCR.
RESULTS: miR-146b expression in the siRNA-miR-146b group was significantly lower than that in the control group. The quality of the original sequencing data and the processed data satisfied with the analysis, and the expression of circRNAs was modulated after the reduction of miR-146b. Among them, 18 circRNAs were upregulated, while 77 circRNAs were downregulated in the miR-146b group compared with the control group. The gene prediction showed that hsa_circ1887 was the largest contact point in miRNA and circRNA regulatory networks. qRT-PCR showed that rno-circRNA-469, rno-circRNA-1138, rno-circRNA-2168 and rno-circRAN-1907 were significantly reduced, while circRNA-1984 was significantly promoted in the siRNA-miR-146b group compared with the control group, which were consistent with the measurements by high-throughput sequencing technique.
CONCLUSIONS: miR-146b could regulate the expression of circRNAs in HSCs, which might take part in the formation and development of hepatic fibrosis.
Keywords: High-Throughput Nucleotide Sequencing, MicroRNAs, RNA Cap Analogs