Hypoxia derived exosomes promote the proliferation and metastasis of colorectal cancer through the regulation of HIF-1α/miR-4299/ZBTB4

Abstract

AimsThe biological functions of colorectal cancer (CRC) cell derived exosomes responding to hypoxic microenvironment and its underlying mechanisms remain unclear. Main methodsExtracted exosomes were confirmed. CRC cells were incubated with hypoxic and normoxic exosomes and its biological behavior were analyzed. miRNA microarray were conducted. Cells were incubated with miRNAs mimics, inhibitors, or small interfering RNAs; expression of reporter constructs was measured in luciferase assays. Cells were transfected with Lentivirus vectors containing eGFP-miR-4299 overexpression (or ZBTB4 siRNA expression plasmid) and they were injected into BALB/C nude mice subcutaneously or by tail vein and the growth of xenograft tumors or lung metastasis were measured. The clinical significance of ZBTB4 was measured in tumor tissues and adjacent non-tumor tissues. Key findingsHypoxic exosomes could tranfer to the recipient normoxic cells and promote the cell proliferation and migration. We found several miRNAs were significantly up-regulated in hypoxic exosomes and the expression levels of miR-4299 increased in both hypoxic cells and hypoxic exosomes. We observed that miR-4299 was upregulated in a HIF-1α dependent way. In addition, ectopic expression of miR-4299 promoted the tumor growth and metastasis in vitro and in vivo. ZBTB4, an identified direct target of miR-4299, could abrogate the effect on tumor growth and distant metastasis. The expression of ZBTB4 were decreased in tumor tissues compared with non-tumor colon tissues from patients. SignificanceWe demonstrated that in response to hypoxia, CRC cells had an increased production of exosomes. The hypoxia derived exosomes promote the proliferation and metastasis of colorectal cancer by exporting miR-4299 and modulating its target gene ZBTB4.