Exosomes Derived From RM-1 Cells Promote The Recruitment Of Mdscs Into Tumor Microenvironment By Up-regulating CXCL12/CXCR4 RM-1

Objective To investigate the expression of chemokine(C-X-C motif)receptor 4(CXCR4)of chemokine ligand 12(CXCL12)wether could be upregulated by exosomes derived from prostate cancer to promote the migration of MDSCs into the tumor microenvironment and verify the potential mechanism.Methods Exosomes in the supernatant of RM-1cells were collected by ultracentrifugation.The typical morphology of exosomes was observed by electron microscopy,and the biomakers of exosomes were identified by Western blot.Flow cytometry was used to detect the proportion of MDSCs in tumor tissues at different time point,and the ratio of MDSCs in bone marrow and spleen after injected exosomes into normal mice,and the proportion of MDSCs in tumor tissues after injected with PBS?exosomes?exosomes + AMD3100(the inhibitor of CXCR4)?AMD3100.Migration ability of MDSCs to the lower chamber processed by the four methods as above was detected by Transwell chemotaxis aasay.Expression of CXCR4 in MDSCs after co-culture with exosomes was detected by WB.Flow cytometry was used to detect the expression of CXCR4 of MDSCs in bone marrow and spleen after injection exosomes into mice.After treated with PBS?exosomes?exosomes + C29(inhibitor of TLR2)?C29,the expression of TLR2?p65?Pp65?CXCR4 in MDSCs were detected by WB.Results Electron microscopy and WB confirmed that the extract was exosomes.The proportion of MDSCs in spleen and tumor tissues increased with tumor-bearing time.The proportion of MDSCs in bone marrow ?spleen and tumor tissues was significantly increased compared with the control group(p<0.05)after injected with exosomes,and the migration effect of MDSCs mediated by exosomes in tumor tissues could be significantly suppressed by AMD3100,the result of transwell assay was similar with that.Both in vivo and in vitro experiments showed that the expression of CXCR4 in MDSCs after treated with exosomes was significantly increased compared with the control group(p <0.05).Western Blot showed that CXCR4's expression in MDSCs was remarkably increased when MDSCs co-cultured with exosomes,meanwhile the NF-kB's activation and TLR2's expression in MDSCs were up-regulated obviously(p <0.05),In contrast,TLR2,Pp65 and CXCR4's expression were decreased markedly after inhibited TLR2 by C29(p <0.05).Conclusion prostate cancer-derived exosomes could up-regulate CXCR4 expression in MDSCs through TLR2/NF-KB signaling pathway,and promote MDSC migration into tumor microenvironment by CXCR4-CXCL12 axis.