Background: Recent studies have demonstrated that exosomes from diabetic animals/cells have detrimental effects on the post-injury vascular repair. Here, we tested the hypothesis that systemic exosome inhibition in vivo improves blood flow recovery in ischemic hindlimbs of diabetic db/db following surgical ischemia.
Methods and Results: Exosomes were isolated from plasma of non-diabetic db/+ and diabetic db/db mice by standard ultracentrifugation method. Unilateral hindlimb ischemia surgery was conducted by ligation of left femoral artery in 12-week old, male db/+ and db/db mice. Exosome inhibitor GW-4869 (GW, 2 μg/g body weight) was administrated by intraperitoneal injection every other day for 4 weeks starting from one week before the hindlimb ischemia (HLI) surgery. HLI mice injected with vehicle served as controls. Mice were divided into four groups: 1) db/+ + vehicle; 2) db/db+ vehicle; 3) db/+ GW; 4) db/db + GW. We found that systemic administration of GW decreased necrosis and loss of toe/toenail, improved blood flow; enhanced capillary/arterial density determined with CD31 and SMA-α staining; increased skeletal muscle weight and cell survival and enhanced satellite cell marker-PAX-7 expression; and decreased fibrosis in the muscles of HLI of db/db mice 21 days post-ligation. Furthermore, our in vitro experiments also revealed that plasma exosomes from db/db mice significantly impaired human cardiac microvascular endothelial cells (HCMVECs) tube formation and migration. Mechanistically, diabetic plasma exosomes decreased H3K4me3 (active epigenetic mark of transcription) hepatocyte growth factor (HGF) and heme oxygenase 1 (HO-1), whereas increased H3K27me3 (suppressor mark of transcription).
Conclusions: Inhibition of circulating exosome formation/secretion by GW increased ischemic limb repair in diabetic db/db, at least partially, via regulating of histone methylation, HGF and HO-1 signaling pathways. Our findings suggest that therapeutic targeting of systemic dysfunctional exosomes could represent a new avenue for the prevention and treatment of ischemic tissue injury in diabetic patients.