Highly enriched exosomal lncRNA OIP5-AS1 regulates osteosarcoma tumor angiogenesis and autophagy through miR-153 and ATG5.

OBJECTIVE This study aims to investigate the regulatory role of exosome lncRNA OIP5-AS1 in tumor progression and autophagy. METHODS Seventy-three cases of osteosarcoma (OS) tissues and 56 cases of adjacent normal tissues were collected to culture human OS cell line HOS. The exosomes secreted by OS cell line were isolated and collected. Apoptosis and exosome markers were detected by flow cytometry. A nude mouse model of OS was established. The gene expression levels of lncRNA OIP5-AS1, miR-153 and autophagy-related protein 5 (ATG5) were quantified by real-time quantitative PCR (RT-PCR). The binding sites of lncRNA OIP5-AS1 and miR-153 were predicted by Starbase3.0, and the binding sites of miR-153 and ATG5 were predicted by Targetscan7.2. The gene binding sites were verified by luciferase reporter gene detection or RNA immunoprecipitation (RIP). The relative level of protein was tested by Western blot. Transwell was applied to test migration and invasion of OS cells. The angiogenesis of OS cells was tested by tubule formation test. RESULTS The results of RT-PCR showed that lncRNA OIP5-AS1 levels were elevated in OS cells and exosomes secreted by cells. Cell function experiments revealed that the proliferation, migration, and invasion of OS cells were promoted by exosomal lncRNA OIP5-AS1. In exosomes, lncRNA OIP5-AS1 inhibited the expression of LC3-II and Beclin 1 proteins, indicating that exosomal lncRNA OIP5-AS1 inhibited autophagy. According to the results of bioinformatics tools and dual-luciferase reporter (DLR) assay or RNA immunoprecipitation (RIP), miR-153 targeted the 3'-UTR of lncRNA OIP5-AS1 and autophagy-related protein 5 (ATG5). The results of western blot (WB) assay showed that exosomal lncRNA OIP5-AS1 and down-regulated miR-153 led to the enhancement of ATG5 protein expression, while up-regulated miR-153 resulted in the decrease of ATG5 protein expression. ATG5 was negatively correlated with miR-153 and positively correlated with lncRNA OIP5-AS1. The results of tubule formation assay disclosed an increase in the angiogenesis level caused by the exosomal lncRNA OIP5-AS1, which was then reversed by the increase of miR-153 and decrease of ATG5. CONCLUSION Highly enriched exosomal lncRNA OIP5-AS1 can regulate OS tumor angiogenesis and autophagy through miR-153 and ATG5.

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