Jun Chen1, Ze-Bing Huang1, Cheng-Jin Liao1, Xing-Wang Hu1, Sha-Ling Li1, Min Qi1, Xue-Gong Fan1, Yan Huang2. 1. Hunan Key Laboratory of Viral Hepatitis, Department of Infectious Disease, Xiangya Hospital, Central South University, Changsha 410008, Hunan Province, P.R.China. 2. Hunan Key Laboratory of Viral Hepatitis, Department of Infectious Disease, Xiangya Hospital, Central South University, Changsha 410008, Hunan Province, P.R.China. Electronic address: drhyan@csu.edu.cn.
Abstract
PURPOSE: Our study aimed to study the role of lncRNA TP73-AS1/miR-539/MMP-8 axis in modulating M2 macrophage polarization in hepatocellular carcinoma (HCC). METHODS: The gene expression levels of TP73-AS1, miR-539 and MMP-8 were modified by transfection with the overexpression or knockdown vectors. The patient survival rate was analyzed using Kaplan-Meier method. The levels of TP73-AS1, miR-539, MMP-8 and M1/2 macrophage polarization markers were analyzed by qRT-PCR, western blot, and flow cytometry. The release of TGF-β1 in the supernatant was determined by ELISA assay. The interaction between TP73-AS1, miR-539 and MMP-8 was analyzed by bioinformatics analysis and dual-luciferase reporter assays. Mouse xenograft model was further established to examine the therapeutic effects of the TP73-AS1 knockdown and miR-539 overexpression in vivo. RESULTS: We found TP73-AS1 and MMP-8 upregulation, and miR-539 downregulation in HCC tissues and cell lines. Lower TP73-AS1 and MMP-8 expressions and higher miR-539 expression were associated with higher survival rate of patients. M2-macrophage markers CD206, Arg-1 and CD163 were significantly upregulated in the tumor tissues. TP73-AS1 negatively and directly regulated miR-539 and knockdown of TP73-AS1 inhibited MMP-8 expression and M2 macrophage polarization. Also, overexpression of miR-539 suppressed M2 macrophage polarization by negatively regulating MMP-8. Furthermore, knockdown of MMP-8 also restrained M2 macrophage polarization via inhibiting TGF-β1 signaling. We also found knockdown of TP73-AS1 or overexpression of miR-539 inhibited HCC tumor growth and M2 macrophage infiltration in vivo. CONCLUSION: Our study demonstrated lncRNA TP73-AS1 negatively regulated miR-539 to promote MMP-8 expression, which activated TGF-β1 signaling to induce M2 macrophage polarization in HCC.
PURPOSE: Our study aimed to study the role of lncRNA TP73-AS1/miR-539/MMP-8 axis in modulating M2 macrophage polarization in hepatocellular carcinoma (HCC). METHODS: The gene expression levels of TP73-AS1, miR-539 and MMP-8 were modified by transfection with the overexpression or knockdown vectors. The patient survival rate was analyzed using Kaplan-Meier method. The levels of TP73-AS1, miR-539, MMP-8 and M1/2 macrophage polarization markers were analyzed by qRT-PCR, western blot, and flow cytometry. The release of TGF-β1 in the supernatant was determined by ELISA assay. The interaction between TP73-AS1, miR-539 and MMP-8 was analyzed by bioinformatics analysis and dual-luciferase reporter assays. Mouse xenograft model was further established to examine the therapeutic effects of the TP73-AS1 knockdown and miR-539 overexpression in vivo. RESULTS: We found TP73-AS1 and MMP-8 upregulation, and miR-539 downregulation in HCC tissues and cell lines. Lower TP73-AS1 and MMP-8 expressions and higher miR-539 expression were associated with higher survival rate of patients. M2-macrophage markers CD206, Arg-1 and CD163 were significantly upregulated in the tumor tissues. TP73-AS1 negatively and directly regulated miR-539 and knockdown of TP73-AS1 inhibited MMP-8 expression and M2 macrophage polarization. Also, overexpression of miR-539 suppressed M2 macrophage polarization by negatively regulating MMP-8. Furthermore, knockdown of MMP-8 also restrained M2 macrophage polarization via inhibiting TGF-β1 signaling. We also found knockdown of TP73-AS1 or overexpression of miR-539 inhibited HCC tumor growth and M2 macrophage infiltration in vivo. CONCLUSION: Our study demonstrated lncRNA TP73-AS1 negatively regulated miR-539 to promote MMP-8 expression, which activated TGF-β1 signaling to induce M2 macrophage polarization in HCC.