Yuping Zhou1, Xueyou Lv2, Hui Qu2, Kekai Zhao2, Liyun Fu3, Linwen Zhu4, Guoliang Ye2, Junming Guo4. 1. Department of Gastroenterology, The Affiliated Hospital of Medical School of Ningbo University, Ningbo 315020, China. Electronic address: nbuzhouyuping@126.com. 2. Department of Gastroenterology, The Affiliated Hospital of Medical School of Ningbo University, Ningbo 315020, China. 3. Department of Hepatology, Ningbo No. 2 Hospital and the Affiliated Hospital, Medical School of Ningbo University, Ningbo 315010, China. 4. Institute of Biochemistry and Molecular Biology, Medical School of Ningbo University, Ningbo 315211, China; Zhejiang Province Key Laboratory of Pathophysiology Technology Research, Ningbo 315211, China.
Abstract
OBJECTIVE: To screen for circular RNAs (circRNAs) that are associated with the activation of hepatic stellate cell (HSC) by monitoring changes in liver circRNA expression in a model of liver fibrosis. METHODS: The classic mouse model of CCl4-induced liver fibrosis was established and validated by histopathological examination. JS1 cells were activated by TGF-β1 to model HSC activation in vitro. Differentially expressed circRNAs in the fibrotic liver tissues and JS1 cells were determined using circRNA microarray, and some of those circRNAs were verified by RT-qPCR. The target genes of the above circRNAs were then predicted by bioinformatics analysis and summarized into a "circRNA-miRNA-mRNA" network diagram. Constructed plasmid mmu_circ_34116 siRNA was transfected to JS1 cells by Lipo2000, then we detected the expression changes of α-SMA. RESULTS: A total of 10,389 circRNAs were identified by microarray screening, and 69 differentially expressed circRNAs were detected in the fibrotic liver tissues with >2-fold difference in expression level relative to normal liver tissues (P < 0.05); 14 circRNAs were up-regulated and 55 were down-regulated. Five differentially expressed circRNAs in fibrotic liver and JS1 cells were verified by RT-qPCR, while all five showed similar trends with the microarray results in the liver, only 3 circRNAs in the JS1 activation model were consistent with the microarray results while one showed no significant change and one circRNA was not detected. Bioinformatics analysis predicted that the "mmu_circ_34116/miR-22-3P/BMP7" signal axis might be involved in the activation of HSC. Transfection experiment confirmed that the expression of α-SMA is significantly elevated as a result of inhibitory expression of mmu_circ_34116. CONCLUSION: The circRNAs expression profile of liver tissue had changed in fibrosis mouse model, and some of these circRNAs may be associated with HSC activation. For instance, mmu_circ_34116 would inhibit HSC activation.
OBJECTIVE: To screen for circular RNAs (circRNAs) that are associated with the activation of hepatic stellate cell (HSC) by monitoring changes in liver circRNA expression in a model of liver fibrosis. METHODS: The classic mouse model of CCl4-induced liver fibrosis was established and validated by histopathological examination. JS1 cells were activated by TGF-β1 to model HSC activation in vitro. Differentially expressed circRNAs in the fibrotic liver tissues and JS1 cells were determined using circRNA microarray, and some of those circRNAs were verified by RT-qPCR. The target genes of the above circRNAs were then predicted by bioinformatics analysis and summarized into a "circRNA-miRNA-mRNA" network diagram. Constructed plasmid mmu_circ_34116 siRNA was transfected to JS1 cells by Lipo2000, then we detected the expression changes of α-SMA. RESULTS: A total of 10,389 circRNAs were identified by microarray screening, and 69 differentially expressed circRNAs were detected in the fibrotic liver tissues with >2-fold difference in expression level relative to normal liver tissues (P < 0.05); 14 circRNAs were up-regulated and 55 were down-regulated. Five differentially expressed circRNAs in fibrotic liver and JS1 cells were verified by RT-qPCR, while all five showed similar trends with the microarray results in the liver, only 3 circRNAs in the JS1 activation model were consistent with the microarray results while one showed no significant change and one circRNA was not detected. Bioinformatics analysis predicted that the "mmu_circ_34116/miR-22-3P/BMP7" signal axis might be involved in the activation of HSC. Transfection experiment confirmed that the expression of α-SMA is significantly elevated as a result of inhibitory expression of mmu_circ_34116. CONCLUSION: The circRNAs expression profile of liver tissue had changed in fibrosismouse model, and some of these circRNAs may be associated with HSC activation. For instance, mmu_circ_34116 would inhibit HSC activation.