Pioglitazone restores the homocysteine-impaired function of endothelial progenitor cells via the inhibition of the protein kinase C/NADPH oxidase pathway

Homocysteine (Hcy) has been shown to impair the migratory and adhesive activity of endothelial progenitor cells (EPCs). As a peroxisome proliferator-activated receptor γ agonist, pioglitazone (PIO) has been predicted to regulate angiogenesis, and cell adhesion, migration and survival. The aim of the present study was to determine whether PIO could inhibit Hcy-induced EPC dysfunctions such as impairments of cell migration and adhesion. EPC migration and adhesion were assayed using 8.0-µm pore size Transwell membranes and fibronectin-coated culture dishes, respectively. Hcy at a concentration of 200 µM was observed to markedly impair cell migration and adhesiveness, and PIO at a concentration of 10 µM attenuated the Hcy-mediated inhibition of EPC migration and adhesion. The mechanism of these effects may be through the inhibition of protein kinase C (PKC) and reactive oxygen species production. The expression levels of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, NADPH oxidase 2 (Nox2) and p67phox, were upregulated by Hcy, with a peak in levels following treatment with a concentration of 200 µM. PIO downregulated the expression levels of Nox2 and p67phox via the PKC signaling pathway. Furthermore, the mechanism of PIO associated with downregulating the p67phox and Nox2 subunits of NADPH oxidase was verified. Thus, PKC and NADPH oxidase may serve a major role in the protective effects of PIO in EPCs under conditions of high Hcy concentrations.