The mechanism of lncRNA‐CRNDE in regulating tumour‐associated macrophage M2 polarization and promoting tumour angiogenesis

Abstract

M2 macrophages can promote liver cancer metastasis by promoting tumour angiogenesis; however, the mechanism underlying macrophage polarization has not been completely revealed. In this study, we mainly explored the mechanism underlying long non‐coding RNA‐CRNDE (lncRNA‐CRNDE) in regulating M2 macrophage polarization and promoting liver cancer angiogenesis. The expression of CRNDE was up‐regulated or down‐regulated in THP‐1 cells (CRNDE‐/‐‐THP‐1 cells and pcDNA3.1‐CRNDE‐THP‐1). THP‐1 cells were co‐cultured with liver cancer cell line H22, and M2 polarization was induced in THP‐1 by IL‐4/13 to simulate tumour‐induced macrophage polarization. As a result, after CRNDE overexpression, THP‐1 cell viability was up‐regulated, the expression of M2 membrane marker CD163 was up‐regulated, and the proportion of F4/80 + CD163+ cells was also up‐regulated. ELISA assay showed that the expression of M2 markers (including TGF‐β1 and IL‐10) and chemokines (including CCl22 and CCL22) was up‐regulated, and the expression of key signals (including STAT6, JAK‐1, p‐AKT1, and Arg‐1) was also up‐regulated, which were significantly different compared with the control group (Con). In addition, the intervention effect of CRNDE on THP‐1 was consistent between co‐culture with H22 cells and IL‐4/13 induction assay. The induced M2 THP‐1 cells were co‐cultured with HUVEC. As a result, THP‐1 cells with CRNDE overexpression can promote the migration and angiogenesis of HUVEC cells in vitro and simultaneously up‐regulate the expression of Notch1, Dll4 and VEGFR2, indicating that THP‐1 M2 polarization induced by CRNDE could further promote angiogenesis. The H22 cell tumour‐bearing mouse model was constructed, followed by injection of CRNDE anti‐oligosense nucleotides and overexpression plasmids to interfere CRNDE expression in tumour‐bearing tissues. Consequently, down‐regulation of CRNDE could down‐regulate tumour volume, simultaneously down‐regulate the expression of CD163 and CD31 in tissues, decrease the expression of key proteins (including JAK‐1, STAT‐6, p‐STAT6 and p‐AKT1), and down‐regulate the expression of key angiogenesis‐related proteins (including VEGF, Notch1, Dll4 and VEGFR2). In this study, we found that CENDE could indirectly regulate tumour angiogenesis by promoting M2 polarization of macrophages, which is also one of the mechanisms of microenvironmental immune regulation in liver cancer.