MicroRNA-140-5p Suppresses The PD-L1 Level In Exosomes Derived From Hepatocellular Carcinoma Cells, Thereby Attenuating The Chemoresistance, Autophagy And Stemness Induced By Sorafenib In Hepatocellular Carcinoma

Abstract

Background: Sorafenib (SO), inhibitor of receptor tyrosine kinase, possesses anti-tumor ability in hepatocellular carcinoma (HCC) by anti-angiogenic effects. However, patients with advanced HCC are resistant to SO. Dysregulated micro-RNA (miRNA) has been shown to be associated with the drug resistance of tumors. Little is known about the effect of miRNA-140-5p on SO-resistance of HCC.Methods: The human HCC cell line Hep3B and Huh-7 were used in this study. The miRNA microarray was used to sequence the transcripts of SO-chemoresistance HCC tissues and SO-chemosensitive tissues. Then, the HCC tissue chip in the GEO database was also analyzed to screen the target gene. Flow cytometry, in situ hybridisation (ISH), immunohistochemistry (IHC), Western blot, cell proliferation assay, transmission electron microscopy (TEM), real-time PCR analyses (qRT-PCR), and pull-down assay were used to explore the effects of miRNA-140-5p and programmed death ligand-1 (PD-L1) on the chemoresistance, autophagy, and stemness of SO-resistant HCC cells treated with SO. In vivo, the SO-resistant HCC cell lines treated with miRNA-140-5p overexpression were injected subcutaneously into mice to construct HCC subcutaneous xenograft tumor models.Results: Based on miRNA microarray, GEO database, and bioinformatics analysis, it was found that miRNA-140-5p and PD-L1 are the core molecules in the SO resistance regulatory network in HCC, and lower miRNA-140-5p and higher PD-L1 expression levels were observed in SO-resistant HCC tissues and cells. IHC results showed that compared with HCC tissues with high miRNA-140-5p expression, PD-L1 protein expression levels in HCC tissues with low miRNA-140-5p was significantly higher (P<0.01). Moreover, compared with SO-sensitive tissues, the levels of PD-L1 protein expression in SO-resistance tissues were significantly higher (P<0.01). In in vitro and in vivo experiments, the introduction of miRNA-140-5p can decreased PD-L1 expression. Mechanically, we found that the Hep3B/SO and Huh-7/SO cells pretreated with miRNA-140-5p inhibitor or exosomes containing PD-L1 have higher autophagy activity, higher stemness, and lower apoptosis rate, which could be abrogated by co-treating cells with anti-PD-L1 antibody or miRNA-140-5p mimic. Conclusions: MiRNA-140-5p can suppress the PD-L1 expression in HCC cells-derived exosomes, thereby attenuating the chemoresistance, autophagy and stemness induced by SO in HCC.