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Updated: Jun 26, 2025
A Mouse Model of Chronic Liver Fibrosis for the Study of Biliary Atresia
Published on: February 3, 2023
Hsa_circ_0009096/miR-370-3p modulates hepatic stellate cell proliferation and fibrosis during biliary atresia
Zhouguang Wu1, Bin Wang1, Siqi Chen1
1Department of General Surgery, Shenzhen Children's Hospital, Shenzhen, China.
Insights
Circular RNA hsa_circ_0009096 promotes hepatic stellate cell proliferation and hepatic fibrosis in biliary atresia by sponging miR-370-3p. This study reveals a novel regulatory mechanism in liver fibrosis.
Area of Science:
- Hepatology
- Molecular Biology
- Cell Biology
Background:
- Hepatic stellate cell (HSC) activation and fibrosis are central to biliary atresia (BA) pathogenesis.
- The molecular mechanisms underlying HSC activation and fibrosis in BA remain poorly understood.
Purpose of the Study:
- To investigate the role of circRNA hsa_circ_0009096 in regulating HSC proliferation and hepatic fibrosis.
- To elucidate the molecular mechanism by which hsa_circ_0009096 influences BA.
Main Methods:
- Established a cellular hepatic fibrosis model using LX-2 cells treated with transforming growth factor β (TGF-β1).
- Assessed hsa_circ_0009096 stability using RNaseR and actinomycin D assays.
- Quantified expression of hsa_circ_0009096, miR-370-3p, and target genes via reverse transcription-qPCR.
- Validated direct binding of hsa_circ_0009096 to miR-370-3p using dual luciferase reporter assay.
- Analyzed cell cycle progression, apoptosis, and protein levels of α-SMA, COL1A1, and TGFBR2 using flow cytometry, immunocytochemistry, and western blotting.
Main Results:
- Hsa_circ_0009096 demonstrated significant stability and its expression increased upon TGF-β1 treatment in LX-2 cells.
- Hsa_circ_0009096 directly binds to miR-370-3p, suppressing its activity and promoting TGFBR2 expression.
- Knockdown of hsa_circ_0009096 inhibited HSC proliferation, promoted apoptosis, and reduced fibrotic markers (α-SMA, COL1A1) in TGF-β1-treated cells.
- Inhibition of miR-370-3p reversed the effects of hsa_circ_0009096 knockdown on cell cycle, apoptosis, and fibrotic markers.
Conclusions:
- Hsa_circ_0009096 promotes HSC proliferation and hepatic fibrosis in BA pathogenesis.
- This promotion is mediated by the sponging of miR-370-3p, leading to accelerated TGFBR2 expression.
- Hsa_circ_0009096 represents a potential therapeutic target for biliary atresia.
Background:
Hepatic stellate cell (HSC) activation and hepatic fibrosis mediated biliary atresia (BA) development, but the underlying molecular mechanisms are poorly understood. This study aimed to investigate the roles of circRNA hsa_circ_0009096 in the regulation of HSC proliferation and hepatic fibrosis.
Methods:
A cellular hepatic fibrosis model was established by treating LX-2 cells with transforming growth factor β (TGF-β1). RNaseR and actinomycin D assays were performed to detect hsa_circ_0009096 stability. Expression of hsa_circ_0009096, miR-370-3p, and target genes was detected using reverse transcription-qPCR. Direct binding of hsa_circ_0009096 to miR-370-3p was validated using dual luciferase reporter assay. Cell cycle progression and apoptosis of LX-2 cells were assessed using flow cytometry. The alpha-smooth muscle actin (α-SMA), collagen 1A1 (COL1A1), and TGF beta receptor 2 (TGFBR2) protein levels in LX-2 cells were analyzed using immunocytochemistry and western blotting.
Results:
Hsa_circ_0009096 exhibited more resistance to RNase R and actinomycinD digestion than UTRN mRNA. Hsa_circ_0009096 expression increased significantly in LX-2 cells treated with TGF-β1, accompanied by elevated α-SMA and COL1A1 expression. Hsa_circ_0009096 siRNAs effectively promoted miR-370-3p and suppressed TGFBR2 expression in LX-2 cells, mediated by direct association of hsa_circ_0009096 with miR-370-3p. Hsa_circ_0009096 siRNA interfered with the cell cycle progression, promoted apoptosis, and reduced α-SMA and COL1A1 expression in LX-2 cells treated with TGF-β1. MiR-370-3p inhibitors mitigated the alterations in cell cycle progression, apoptosis, and α-SMA, COL1A1, and TGFBR2 expression in LX-2 cells caused by hsa_circ_0009096 siRNA. In conclusion, hsa_circ_0009096 promoted HSC proliferation and hepatic fibrosis during BA pathogenesis by accelerating TGFBR2 expression by sponging miR-370-3p.
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