Figure 5.

Transferred exosomal miRs regulate biological properties of recipient CPCs. (A) CPCs were transfected with a luciferase reporter vector containing either miR-126 or miR-210 recognition sequence or a control luciferase vector lacking the sequence. CPCs were then given EC^HIF-CM in the presence or absence of exosomes as indicated. CPCs were then cultured for another 24 hours and luciferase activity was determined and expressed as fold-decrease of cells transfected with the luciferase reporter containing specific miR recognition sequence over the vector lacking the recognition sequence for each culture condition. ^*P<0.05 (N=4) vs. cells transfected with vector lacking miR recognition sequence. (B) Representative immunoblots and densitometry quantification of indicated proteins in CPCs grown in either normal conditions, or supplemented with EC^GFP-Exo, EC^HIF-Exo, or EC^{HIF/miR126-KO}-exosomes. ^*P<0.05 vs. untreated CPCs; ^#P<0.05 vs. EC^HIF (N=4). (C) Expression of angiogenic genes in CPCs grown in each culture condition as indicated was determined by qPCR. ^*P<0.05 vs. untreated; ^#P<0.05 vs. EC^GFP-Exo; ^!P<0.05 vs. EC^HIF-Exo (N=8). (D) Expression of Iscu, a validated miR-210 target gene, was determined in CPCs supplemented with either EC^GFP-Exo or EC^HIF-Exo. ^*P<0.05 (N=4) vs. CPCs supplemented with EC^GFP-Exo. CPCs were cultured in conditions as specified for 24 hours. (E) Oxygen consumption and (F) non-mitochondrial respiration were then determined and expressed as fold-change compared to untreated control cells. Dimethyloxalylglycine (DMOG), a known HIF-1 activator was used as positive control for reduced oxygen consumption. ^*P<0.05 vs. untreated controls (N=6).