Exosomes secreted under hypoxia enhance stemness in Ewing’s sarcoma

Hypoxic Ewing’s sarcoma cells release exosomes that promote sphere formation, a stem-like phenotype, in EWS cells by enhancing survival.

Analysis of the hypoxic exosomal miRNA cargo identified a HIF-1α regulated miRNA, miR-210, as a potential mediator of sphere formation in cells exposed to hypoxic exosomes. The knockdown of HIF-1α in hypoxic EWS cells led to decreased exosomal miR-210 levels and reduced the capacity of hypoxic exosomes to form spheres. Inhibition of miR-210 in hypoxic spheres attenuated sphere formation and overexpression of miR-210 in normoxic spheres significantly enhanced the number of EWS spheres.

Together, the findings suggest that hypoxic exosomes promote stemness in EWS cells by delivering enriched miR-210 that is capable of down-regulating apoptotic pathways, resulting in the survival of cells with increased sphere formation.

Dr. Shantaram S. Joshi from The University of Nebraska Medical Center said, “Ewing’s sarcoma (EWS) is an aggressive and highly malignant bone tumor that develops in children and adolescents.”

HIF -1α has been demonstrated to regulate tumor formation and stem cell survival in hypoxic cancer cells by inhibiting apoptosis. Emerging evidence indicates intercellular communication between tumor cells in hypoxic and normoxic regions contributes to functional differences associated with hypoxic tumors. Other reports demonstrated that shCD99 EWS-derived exosomes could transfer enriched miR-34a to recipient EWS cells and stimulate neural differentiation while in another study, EWS-derived exosomes carrying EZH2 mRNA could be delivered intact to mesenchymal stem cells.

EWS HypoxicEXO carry specific hypoxia regulated miRNA.

EWS HypoxicEXO carry specific hypoxia regulated miRNA.

(A) Representative electropherogram profile of total RNA in normoxic and hypoxic SK-ES-1 cells and exosomes. Ribosomal subunits 28S and 18S were observed in cellular RNA profiles and were absent in exosomal RNA. Unsupervised cluster analysis of SK-ES-1 miRNAs in (B) normoxic and hypoxic cells and (C) NormoxicEXO and HypoxicEXO. Each column represents a biological replicate and every row denotes an individual miRNA. The Z-score color scale indicates high expression represented by red ranging to low expression denoted by green. Volcano plot of miRNA differentially expressed (≥ 1.5 fold threshold) in SK-ES-1 (D) normoxic compared to hypoxic cells and (E) HypoxicEXO compared to NormoxicEXO. Significance (-log10 p-value) was plotted versus fold change (log2 fold change) along the y and x axis, respectively. (F) Validation of hypoxia regulated miRNAs miR-210 A673 and SK-ES-1 NormoxicEXO and HypoxicEXO using qRT-PCR (Mean ± SEM, n = 3, *P ≤ 0.05). miR-16 was used as a control to normalize miRNA expression.

Studies in other cancer models investigating the role of hypoxic exosomes have provided insight into how hypoxic tumors can secrete exosomes that propagate an aggressive phenotype in cells outside the hypoxic niche. Exosomes released from hypoxic prostate cancer cells enhanced sphere formation in normoxic cells, but the authors were unable to elucidate a mechanism describing how hypoxic exosomes promote stemness in normoxic cells.

The Joshi Research Team concluded that this study describes a mechanism whereby EWS cells under hypoxic conditions release exosomes that enhance stemness in EWS cells. Ongoing studies are investigating the role of HIF-1α on regulating EWS stemness and together, the authors future aim is to investigate how HIF-1α selectively modulates the packaging of miRNAs into HypoxicEXO, and validate additional miRNAs that promote aggressive hypoxic phenotypes.

Source – Eurekalert

Kling MJ, Chaturvedi NK, Kesherwani V, Coulter DW, McGuire TR, J. Sharp JG, Joshi SS. (2020) Exosomes secreted under hypoxia enhance stemness in Ewing’s sarcoma through miR-210 delivery. Oncotarget 11:3633-3645. [article]

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