Pleiotrophin-induced endothelial cell migration is regulated by xanthine oxidase-mediated generation of reactive oxygen species

Pleiotrophin (PTN) is a heparin-binding growth factor that induces cell migration through binding to its receptor protein tyrosine phosphatase beta/zeta (RPTP beta/zeta) and integrin alpha v beta 3 (alpha(v)beta(3)). In the present work, we studied the effect of PIN on the generation of reactive oxygen species (ROS) in human endothelial cells and the involvement of ROS in PTN-induced cell migration. Exogenous PTN significantly increased ROS levels in a concentration and time-dependent manner in both human endothelial and prostate cancer cells, while knockdown of endogenous PTN expression in prostate cancer cells significantly down-regulated ROS production. Suppression of RPTP beta/zeta through genetic and pharmacological approaches, or inhibition of c-src kinase activity abolished PTN-induced ROS generation. A synthetic peptide that blocks PTN-alpha(v)beta(3) interaction abolished PIN-induced ROS generation, suggesting that alpha(v)beta(3) is also involved. The latter was confirmed in CHO cells that do not express beta(3) or over-express wild-type beta(3) or mutant beta(3)-3Y773F/Y785F. PIN increased ROS generation in cells expressing wild-type beta(3) but not in cells not expressing or expressing mutant 133. Phosphoinositide 3-kinase (PI3K) or Erk1/2 inhibition suppressed PIN-induced ROS production, suggesting that ROS production lays down-stream of PI3K or Erk1/2 activation by PIN. Finally, ROS scavenging and xanthine oxidase inhibition completely abolished both PIN-induced ROS generation and cell migration, while NADPH oxidase inhibition had no effect. Collectively, these data suggest that xanthine oxidase-mediated ROS production is required for PIN-induced cell migration through the cell membrane functional complex of alpha(v)beta(3) and RPTPf3g and activation of c-src, PI3K and ERK1/2 kinases. (C) 2015 Elsevier Inc. All rights reserved.