Modulation of pulmonary vascular smooth muscle cell phenotype in hypoxia: role of cGMP-dependent protein kinase.
Chronic hypoxia triggers pulmonary vascular remodeling, which is associated with a modulation of the vascular smooth muscle cell (SMC) phenotype from a contractile, differentiated to a synthetic, dedifferentiated state. We previously reported that acute hypoxia represses cGMP-dependent protein kinase (PKG) expression in ovine fetal pulmonary venous SMCs (FPVSMCs). Therefore, we tested if altered expression of PKG could explain SMC phenotype modulation after exposure to hypoxia. Hypoxia-induced reduction in PKG protein expression strongly correlated with the repressed expression of SMC phenotype markers, myosin heavy chain (MHC), calponin, vimentin, alpha-smooth muscle actin (alphaSMA), and thrombospondin (TSP), indicating that hypoxic exposure of SMC induced phenotype modulation to dedifferentiated state, and PKG may be involved in SMC phenotype modulation. PKG-specific small interfering RNA (siRNA) transfection in FPVSMCs significantly attenuated calponin, vimentin, and MHC expression, with no effect on alphaSMA and TSP. Treatment with 30 microM Drosophila Antennapedia (DT-3), a membrane-permeable peptide inhibitor of PKG, attenuated the expression of TSP, MHC, alphaSMA, vimentin, and calponin. The results from PKG siRNA and DT-3 studies indicate that hypoxia-induced reduction in protein expression was also similarly impacted by PKG inhibition. Overexpression of PKG in FPVSMCs by transfection with a full-length PKG construct tagged with green fluorescent fusion protein (PKG-GFP) reversed the effect of hypoxia on the expression of SMC phenotype marker proteins. These results suggest that PKG could be one of the determinants for the expression of SMC phenotype marker proteins and may be involved in the maintenance of the differentiated phenotype in pulmonary vascular SMCs in hypoxia.
Zhou
Weilin
W
Division of Neonatology, Harbor-UCLA Medical Center, Los Angeles Biomedical Research Institute at Harbor-UCLA, 1124 West Carson St., Torrance, CA 90502, USA. wzhou@labiomed.org
Dasgupta
Chiranjib
C
Negash
Sewite
S
Raj
J Usha
JU
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D017869:Cyclic GMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation / Q000502:physiology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D049452:Green Fluorescent Proteins / Q000235:genetics; D000860:Hypoxia / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000201:enzymology*; D010641:Phenotype; D011247:Pregnancy; D011667:Pulmonary Veins / Q000166:cytology / Q000201:enzymology*; D034741:RNA, Small Interfering; D012756:Sheep; D014162:Transfection
D015415:Biomarkers; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins; D017869:Cyclic GMP-Dependent Protein Kinases