Publication Only: Acute myeloid leukemia - Biology & translational research
Leukemic cells could stimulate neovascularization in bone marrow and this angiogenesis has been linked to adverse prognosis in AML. Exosomes could accelerate angiogenesis and promote tumor progression by harboring microRNAs (miRNAs). So the exosomal miRNAs may become new targets for AML treatment. High p62 expression is related to poor prognosis of AML, however, the regulation of angiogenensis by p62 is remain largely unknown.
The aim of this study was to investigate the effect of p62 on miRNAs expression profile in exosomes derived from AML and the effect of exosomes derived from AML cells on angiogenesis.
Electron microscopy and Western blot(WB) were used to confirm the exosomes derived from U937 cells, the controls and the cells knockdown p62. The Exiqon v19.0 microRNA MicroArray was used for profiling of miRNAs in exosomes derived from our specimens. The miRCURY™ Hy3™/Hy5™ Power labeling kit was used for miRNA labelling. Then the Hy3™-labeled samples were hybridized on the miRCURYTM LNA Array (v.19.0). The gene ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) databases were used to predict the biological functions and potential mechanisms of the differentially expressed miRNAs in AML exosomes. Quantitative real-time polymerase chain reaction(qRT-PCR) was used to confirm the results of two downregulated microRNAs of exosomes. Then endothelial cell tube formation assays were used to investigate the effect of the angiogenesis of HUVECs which added exosomes derived from cells.
We demonstrated that 2080 miRNAs were expressed in exosomes derived from U937 cells with knockdown p62 and the controls, of which 215 miRNAs were upregulated and 208 miRNAs were downregulated in U937 cells with knockdown p62(Fold Change≥2.0, P < 0.05). The results revealed miR-3064-3p and miR-339-5p were downregulated in U937 cells with knockdown p62 by qRT-PCR and consistent with microarray profile. GO pathway analysis indicated that the highest enrichment was intercellular part by miRNA. There were 1460 biological processes in downregulated transcripts and there were 19 pathways related with vesicle and 1515 pathways in upregulated transcripts and 8 pathways about vesicle by BP analysis. MF analysis inducted protein binding, transcription regulator activity and DNA binding transcription factor activity were enriched (P < 0.05). Pathway analysis indicated that 84 pathways corresponded to upregulated transcripts and 55 pathways corresponded to downregulated transcripts (P < 0.05). Then we found that exosomes deviated from U937 cells can promote the angiogenesis of HUVECs
Our data suggested that miRNA in exosomes may play an important role in the pathogenesis of AML, and knockdown p62 may treat AML by the miRNAs in exosomes as reducing angiogenesis.