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Upregulation of microRNA-328-3p by hepatitis B virus contributes to THLE-2 cell injury by downregulating FOXO4.

ABSTRACT: BACKGROUND:Hepatitis B virus (HBV) remains a major cause of chronic hepatitis and hepatocellular carcinoma, and miRNAs play important roles in HBV pathogenesis. Our previous study has shown that miR-328-3p is upregulated in HBV-infected patients and serves as a potent predictor for the prognosis of HBV-related liver failure. METHODS:Here, the role of miR-328-3p in modulating cell injury in HBV-infected liver cells THLE-2 was investigated in detail. MiR-328-3p expression was examined using qRT-PCR. The levels of pro-inflammatory cytokines were measured using ELISA. HBV RNA and HBV DNA levels were quantified. The interactions between STAT3 and miR-328-3p promoter as well as miR-328-3p and FOXO4 were analyzed using chromatin immunoprecipitation (CHIP) assay and luciferase reporter assay, respectively. THLE-2 cell injury was evaluated by examining cell viability and apoptosis. RESULTS:HBV promoted expression of miR-328-3p through the STAT3 signal pathway and that increasingly expressed miR-328-3p downregulated its target FOXO4, leading to the promotion of cell injury in HBV-infected liver cells THLE-2. CONCLUSION:These data demonstrate that HBV-STAT3-miR-328-3p-FOXO4 regulation pathway may play an important role in the pathogenesis of HBV infection.

SUBMITTER: Fu X

PROVIDER: S-EPMC7106851 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Upregulation of microRNA-328-3p by hepatitis B virus contributes to THLE-2 cell injury by downregulating FOXO4.

Fu Xiaoyu X Ouyang Yi Y Mo Juan J Li Ronghua R Fu Lei L Peng Shifang S

Journal of translational medicine 20200330 1

<h4>Background</h4>Hepatitis B virus (HBV) remains a major cause of chronic hepatitis and hepatocellular carcinoma, and miRNAs play important roles in HBV pathogenesis. Our previous study has shown that miR-328-3p is upregulated in HBV-infected patients and serves as a potent predictor for the prognosis of HBV-related liver failure.<h4>Methods</h4>Here, the role of miR-328-3p in modulating cell injury in HBV-infected liver cells THLE-2 was investigated in detail. MiR-328-3p expression was examin ...[more]

PMID: 32228643

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Project description:UNLABELLED:Hepatic injuries in hepatitis B virus (HBV) patients are caused by immune responses of the host. In our previous study, microRNA-146a (miR-146a), an innate immunity-related miRNA, and complement factor H (CFH), an important negative regulator of the alternative pathway of complement activation, were differentially expressed in HBV-expressing and HBV-free hepatocytes. Here, the roles of these factors in HBV-related liver inflammation were analyzed in detail. The expression levels of miR-146a and CFH in HBV-expressing hepatocytes were assessed via analyses of hepatocyte cell lines, transgenic mice, adenovirus-infected mice, and HBV-positive human liver samples. The expression level of miR-146a was upregulated in HBV-expressing Huh-7 hepatocytes, HBV-expressing mice, and patients with HBV infection. Further results demonstrated that the HBV X protein (HBx) was responsible for its effects on miR-146a expression through NF-?B-mediated enhancement of miR-146a promoter activity. HBV/HBx also downregulated the expression of CFH mRNA in hepatocyte cell lines and the livers of humans and transgenic mice. Furthermore, overexpression and inhibition of miR-146a in Huh-7 cells downregulated and upregulated CFH mRNA levels, respectively. Luciferase reporter assays demonstrated that miR-146a downregulated CFH mRNA expression in hepatocytes via 3'-untranslated-region (UTR) pairing. The overall effect of this process in vivo is to promote liver inflammation. These results demonstrate that the HBx-miR-146a-CFH-complement activation regulation pathway might play an important role in the immunopathogenesis of chronic HBV infection. These findings have important implications for understanding the immunopathogenesis of chronic hepatitis B and developing effective therapeutic interventions. IMPORTANCE:Hepatitis B virus (HBV) remains an important pathogen and can cause severe liver diseases, including hepatitis, liver cirrhosis, and hepatocellular carcinoma. Although HBV was found in 1966, the molecular mechanisms of pathogenesis are still poorly understood. In the present study, we found that the HBV X protein (HBx) promoted the expression of miR-146a, an innate immunity-related miRNA, through the NF-?B signal pathway and that increasingly expressed miR-146a downregulated its target complement factor H (CFH), an important negative regulator of the complement alternative pathway, leading to the promotion of liver inflammation. We demonstrated that the HBx-miR-146a-CFH-complement activation regulation pathway is potentially an important mechanism of immunopathogenesis caused by chronic HBV infection. Our data provide a novel molecular mechanism of HBV pathogenesis and thus help to understand the correlations between the complement system, an important part of innate immunity, and HBV-associated disease. These findings will also be important to identify potential therapeutic targets for HBV infection.

Project description:BackgroundSorafenib can prolong the survival of patients with advanced hepatocellular carcinoma (HCC). However, drug resistance remains the main obstacle to improving its efficiency. This study aimed to explore the likely molecular mechanism of sorafenib resistance.MethodsDifferentially expressed microRNAs (miRNAs) related to sorafenib response were analyzed with the Limma package in R software. The expression levels of miR-126-3p and sprouty-related EVH1 domain-containing protein 1 (SPRED1) in HCC cells were measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Cell viability and proliferation were detected with Cell Counting Kit-8 (CCK-8), EdU proliferation, and clone formation assays. Transwell assays were performed to measure cell migration and invasion. TargetScan, MicroRNA Target Prediction Database (miRDB), and StarBase v2.0 were used to predict the targets of miR-126-3p. SPRED1 was confirmed as a target gene of miR-126-3p by dual-luciferase reporter assay and Western blotting. Finally, the in vivo anti-tumor effect of LV-miR-126-3p inhibitor combined with sorafenib was evaluated via subcutaneous tumor models.ResultsHCC cells with high expression of miR-126-3p exhibited increased resistance to sorafenib. The results of bioinformatics analysis and the dual-luciferase reporter assay showed that miR-126-3p directly targeted SPRED1. The sensitivity of HCC cells to sorafenib was markedly enhanced by SPRED1 upregulation. Gain- and loss-of function experiments verified that miR-126-3p induced sorafenib resistance in HCC through downregulating SPRED1. Furthermore, the inhibition of miR-126-3p markedly increased the effectiveness of sorafenib against HCC in vivo. Mechanistically, our results suggested that miR-126-3p promoted sorafenib resistance via targeting SPRED1 and activating the ERK signaling pathway.ConclusionsOur study demonstrates that regulating the miR-126-3p/SPRED1 axis might be a promising strategy for enhancing the antitumor effect of sorafenib in the treatment of HCC.

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Upregulation of microRNA-328-3p by hepatitis B virus contributes to THLE-2 cell injury by downregulating FOXO4.

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Upregulation of microRNA-328-3p by hepatitis B virus contributes to THLE-2 cell injury by downregulating FOXO4.

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