Figure 6.

Mature MicroRNA Editing and Expression in Normoxic and Ischemic Human Vessels

(A) miR-376a+b-3p, (B) miR-376c-3p, (C) miR-381-3p, and (D) miR-411-5p expression and editing in internal mammary arteries (IMAs, n = 5) and venae saphenae magnae (VSMs, n = 3) cultured in ex vivo control (C) conditions or hypoxia+starvation (H+S) to mimic ischemia. Right before harvest, IMAs were separated manually into the tunica media/intima (media) and the tunica adventitia (adv.), whereas VSMs were left intact. ^#p < 0.01, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, versus its T0 by a paired two-sided Student’s t test. Symbol color shows whether means of either WT or ED expression are compared. (E) miR-376a+b-3p, (F) miR-376c-3p, (G) miR-381-3p, and (H) miR-411-5p expression and editing of the same microRNAs in lower leg vein (LLV) samples from three different patient groups with minimal to end-stage peripheral artery disease (PAD). Normoxic LLV samples (n = 8) from patients with coronary artery disease (CAD) rather than PAD undergoing coronary artery bypass surgery were compared to LLV samples (n = 6) from patients with severe PAD undergoing femoral artery to popliteal artery bypass surgery and to LLV samples (n = 8) from patients with end-stage PAD, undergoing lower limb amputation. All mature microRNA expressions are normalized to U6. (I and J) Relative ADAR1 and ADAR2 mRNA (I) and protein (J) expression in the LLV samples from different patient groups. Expression was normalized to stable household genes RPLP0 or β-actin (for mRNA and protein expression, respectively) and expressed as fold change of the CAD group. See Figure S4 for the western blots used for the ADAR protein quantification. For (E) and (J), all data are presented as mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, versus T0 unless otherwise indicated by one-way ANOVA.