Mitochondrial depolarization stimulates vascular repair-relevant functions of CD34 cells via reactive oxygen species-induced nitric oxide generation.

BACKGROUND AND PURPOSE

CD34 haematopoietic stem/progenitor cells have revascularization potential and are now being tested for the treatment of ischaemic vascular diseases in clinical trials. We tested the hypothesis that mitochondrial depolarization stimulates the reparative functions of CD34 cells.

EXPERIMENTAL APPROACH

Peripheral blood was obtained from healthy individuals (n = 63), and mononuclear cells (MNCs) were separated. MNCs were enriched for lineage negative cells, followed by isolation of CD34 cells. Vascular repair-relevant functions of CD34 cells, proliferation and migration, were evaluated in the presence and absence of diazoxide. Mitochondrial membrane potential, ROS and NO levels were evaluated by flow cytometry by using JC-1, mitoSOX and DAF-FM respectively.

KEY RESULTS

Diazoxide stimulated the proliferation and migration of CD34 cells that were comparable to the responses induced by stromal-derived factor-1α (SDF) or VEGF. Effects of diazoxide were blocked by either 5-hydroxydecanoate (5HD), a selective mitochondrial ATP-sensitive potassium channel (mitoK ) inhibitor, or by L-NAME. Diazoxide induced mitochondrial depolarization, and NO and cGMP generation that were 5HD-sensitive. The generation of NO and cGMP by diazoxide was blocked by an endothelial NOS (eNOS)-selective inhibitor, NIO, but not by a neuronal (n)NOS-selective inhibitor, N -propyl-L-arginine (NPA). A Ca chelator, BAPTA, Akt inhibitor, triciribine, or PI3K inhibitor, LY294002, inhibited the NO release induced by diazoxide. Phosphorylation of eNOS at Ser and dephosphorylation at Thr were increased. Diazoxide-induced ROS generation and phosphorylation of eNOS at Ser were reduced by NPA.

CONCLUSION AND IMPLICATIONS

Diazoxide stimulates vascular repair-relevant functions of CD34 cells via the mitoK -dependent release of NO and ROS.

LINKED ARTICLES

This article is part of a themed section on Mitochondrial Pharmacology: Featured Mechanisms and Approaches for Therapy Translation. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.22/issuetoc.