CircNFIX promotes stemness and evasion of phagocytosis in glioma stem cells by sponging miR-449a to upregulate the expression of CD47

Abstract

Background

Cancer stem cells play important roles in tumorigenesis and progression of glioma. Circular RNA nuclear factor I X (circNFIX, hsa_circ_0049658) has been found to facilitate the development of glioma. The regulation of circNFIX in glioma stem cells (GSCs) has never been studied.

Methods

RNA levels of circNFIX, microRNA-449a (miR-449a) and cluster of differentiation 47 (CD47) were detected through reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The protein analysis was performed by western blot. Cell proliferation and self-renewal abilities were assessed by EdU assay and tumor sphere formation assay. The macrophage-mediated phagocytosis and cytotoxicity were examined by flow cytometry and lactate dehydrogenase (LDH) assay, respectively. The target analysis was performed using dual-luciferase reporter assay, RNA immunoprecipitation (RIP) and RNA pull-down assays. In vivo assay was conducted through GSCs model in mice.

Results

The levels of circNFIX and CD47 were upregulated in glioma tissues and GSCs. Silencing circNFIX or CD47 suppressed proliferation and self-renewal abilities but enhanced phagocytosis and cytotoxicity in GSCs. CircNFIX induced the positive regulation of CD47 by exerting the sponge effect on miR-449a in GSCs. The regulatory role of circNFIX was achieved by targeting miR-449a in GSCs. CircNFIX downregulation also repressed tumorigenesis of glioma and maintenance of stem cells by the miR-449a/CD47 axis in GSCs model in vivo.

Conclusion

The evidence suggested that circNFIX acted as a miR-449a sponge to regulate the CD47 level, thereby promoting the features of glioma stem cells and the evasion of phagocytosis.