Thymosin-β4 Mediates Hepatic Stellate Cell Activation by Interfering with CircRNA-0067835/miR-155/FoxO3 Signaling Pathway

Abstract

Background/Aims: Hepatic stellate cells (HSCs) are the primary cell type responsible for liver fibrosis. Our study proved that thymosin beta 4 (Tβ4) has anti-fibrogenic effects in HSCs through PI3K/AKT pathway. However, the underlying mechanisms are not fully elucidated. Circular RNAs (circRNAs) play important roles in fine-tuning gene expression and are often deregulated in cancers. However, the expression profile and clinical significance of in liver fibrosis is still unknown. Therefore, we hypothesize that Tβ4 influences circRNAs in liver fibrosis. Methods: Circular RNA microarray was conducted to identify Tβ4-related circRNAs. Pathway analysis and miRNA response elements analysis was conducted to predict the potential roles of differentially expressed circRNAs in liver fibrosis. CCK8 assays and flow cytometric assays were conducted to clarify the role of circRNA in liver fibrosis. Bioinformatics analysis and in vitro experiments were conducted to clarify the mechanism of circRNA-mediated gene regulation in liver fibrosis. Results: A total of 644 differentially expressed circRNAs were identified between the Tβ4-depleted LX-2 cells and the control LX2 cells. The expression of circRNA-0067835 was significantly increased in the Tβ4-depleted LX-2 cells compared with control. Knockdown of circRNA-0067835 observably decreased LX-2 cell proliferation by causing G1 arrest and promoting apoptosis. Bioinformatics online programs predicted that circRNA-0067835 acted as miR-155 sponge to regulate FOXO3a expression, which was validated using luciferase reporter assay. Conclusion: Our experiments showed that circRNA-0067835 regulated liver fibrosis progression by acting as a sponge of miR-155 to promote FOXO3a expression, indicating that circRNA-0067835 may serve as a potential therapeutic target for patients with liver fibrosis.

Regulatory Non-coding RNAs Network in Non-alcoholic Fatty Liver Disease

“…Among these circRNAs, circRNA_34116 was shown to inhibit HSCs activation possibly via miR-22-3p and its targets, BMP7 as predicted by the bioinformatics analysis (Zhou Y. et al, 2018). Another study of HSC inhibition by thymosin beta 4 treatment has identified 644 circRNAs that were differentially expressed and circRNA_0067835 expression was significantly increased in activated HSCs (Zhu L. et al, 2018). It was also shown that circRNA_0067835 acted as a miR-155 sponge and elevated the FOXO3a expression thus promoted HSCs activation and fibrosis (Zhu L. et al, 2018).…”

“…Another study of HSC inhibition by thymosin beta 4 treatment has identified 644 circRNAs that were differentially expressed and circRNA_0067835 expression was significantly increased in activated HSCs (Zhu L. et al, 2018). It was also shown that circRNA_0067835 acted as a miR-155 sponge and elevated the FOXO3a expression thus promoted HSCs activation and fibrosis (Zhu L. et al, 2018). Despite these limited findings of circRNAs roles in liver fibrosis, the evidence of their regulatory interaction with miRNAs and their target mRNAs showed that understanding the molecular regulation and mechanisms underlying liver fibrosis could potentially discover a novel therapeutic target.…”

The Crosstalk of miRNA and Oxidative Stress in the Liver: From Physiology to Pathology and Clinical Implications

“…miR-155 is upregulated in a NASH mouse model induced via high fat diet [98]. iR-155 elevates the Forkhead-Box-Protein O3 (FOXO3a) expression thereby regulating the activation of that pathway, whose proteins are involved in the maintenance of the intercellular redox balance [99]. Additionally, miR-155 regulates lipid metabolism by modulating the protein expression of SREBP-1c and fatty acid synthase (FAS) resulting in increased intracellular lipid accumulation in hepatocytes [100].…”

Circular RNA RSF1 promotes inflammatory and fibrotic phenotypes of irradiated hepatic stellate cell by modulating miR‐146a‐5p

“…LX2 cells were transfected with RAC1 siRNA with or without miR-146a-5p inhibitor and then subjected to 8 Gy X-ray irradiation. (Zhu et al, 2018). It was reported that microRNA-146b regulates HSC activation via targeting of KLF4 (Ge et al, 2016).…”