Exposure to prolonged
hypoxia can result in pulmonary
vascular remodeling and
pulmonary hypertension.
Hypoxia induces pulmonary vascular smooth muscle cell (PVSMC) proliferation and
vascular remodeling by affecting cell adhesion and migration and secretion of
extracellular matrix proteins. We previously showed that acute
hypoxia decreases
cGMP-dependent protein kinase (PKG) activity in PVSMC and that PKG plays a role in maintaining the differentiated contractile phenotype in normoxia. In this study, we investigated the effect of
hypoxia on PVSMC adhesion and migration and the role of PKG in these functions. Ovine fetal pulmonary artery SMC were incubated in normoxia (Po(2) approximately 100 Torr) or
hypoxia (Po(2) approximately 30-40 Torr) or treated with the PKG inhibitor DT-3 for 24 h in normoxia. To further study the role of PKG in the modulation of adhesion and migration, PVSMC were transiently transfected with a full-length PKG1alpha [PKG-
green fluorescent protein (GFP)] or a dominant-negative construct (G1alphaR-GFP). Cell adhesion to
extracellular matrix proteins was determined, and
integrin-mediated adhesion was assessed by alpha/beta-
integrin-mediated cell adhesion array. Exposure to
hypoxia (24 h) and pharmacological inhibition of PKG1 by DT-3 significantly promoted adhesion mediated by alpha(4)-, beta(1)-, and alpha(5)beta(1)-integrins to
fibronectin,
laminin, and
tenacin and also resulted in increased cell migration. Likewise, inhibition of PKG by expression of a dominant-negative PKG1alpha construct increased cell adhesion and migration, comparable to that induced by
hypoxia. Dynamic actin reorganization associated with
integrin-mediated cell adhesion is partly regulated by the
actin-binding protein cofilin, the (Ser3) phosphorylation of which inhibits its actin-severing activity. We found that increased PKG expression and activity is associated with decreased
cofilin (Ser3) phosphorylation, implying a role for PKG in the modulation of
cofilin activity and actin dynamics. Together, these findings identify cGMP/PKG1 signaling as central to the functional differences between PVSMC exposed to normoxia versus
hypoxia.