Abstract

Delta-9-tetrahydrocannabinol (THC), the major bioactive component of marijuana, has been shown to induce functional MDSCs in vivo. Here, we studied the involvement of miRNA in this process. CD11b+Gr-1+ MDSCs were purified from peritoneal exudates of mice administered with THC and used for genome-wide miRNA profiling. Expression of CD31 and Ki67 confirmed that the THC-MDSC were immature and proliferating. THC-induced MDSCs exhibited distinct miRNA expression signature relative to various myeloid cells and BM precursors. We identified 13 differentially expressed (>2 fold) miRNA in THC-MDSC relative to control BM precursors. In silico target prediction for these miRNA and pathway analysis using multiple bioinformatics tools revealed significant overrepresentation of Gene Ontology clusters within hematopoiesis, myeloid cell differentiation and regulation categories. IGF1 signaling involved in cell growth and proliferation, and myeloid differentiation pathways were among the most significantly enriched canonical pathways. Among the differentially expressed, miRNA-690 was highly overexpressed in THC-MDSC (~16 fold). Transcription factor C/EBPα was identified as a potential functional target of miR-690. Supporting this, C/EBPα expression was attenuated in THC-MDSC as compared to BM precursors, and exhibited an inverse relation with miR-690. miR-690 knockdown using PNA-antagomiR was able to unblock and significantly increase C/EBPα expression establishing the functional link. Further, CD11b+Ly6G+Ly6C+ and CD11b+Ly6G‒Ly6C+ purified subtypes showed high levels of miR-690 with attenuated C/EBPα expression. Moreover, EL-4 tumor-elicited MDSC showed increased miR-690 expression. In conclusion, miRNA are significantly altered during the generation of functional MDSC from BM. Select miRNA such as mir-690 targeting genes involved in myeloid expansion and differentiation likely play crucial roles in this process and therefore in cannabinoid-induced immunosuppression.