gw9662 and Hypertension--Pulmonary

It has been reported that sodium hydrosulfide (NaHS) stimulated high stretch induced-atrial natriuretic peptide (ANP) secretion via ATP sensitive potassium (K

Topics: 2-Methoxyestradiol; Anilides; Animals; Atrial Natriuretic Factor; Bosentan; Gene Expression Regulation; Glyburide; Heart Atria; Hydrogen Sulfide; Hypertension, Pulmonary; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; KATP Channels; Male; Monocrotaline; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type III; Organ Culture Techniques; Oxygen; Pinacidil; Potassium Channel Blockers; PPAR gamma; Rats; Rats, Sprague-Dawley; Signal Transduction; Sulfides

Pulmonary hypertension (PH) is a progressive disorder that causes significant morbidity and mortality despite existing therapies. PH pathogenesis is characterized by metabolic derangements that increase pulmonary artery smooth muscle cell (PASMC) proliferation and vascular remodeling. PH-associated decreases in peroxisome proliferator-activated receptor γ (PPARγ) stimulate PASMC proliferation, and PPARγ in coordination with PPARγ coactivator 1α (PGC1α) regulates mitochondrial gene expression and biogenesis. To further examine the impact of decreases in PPARγ expression on human PASMC (HPASMC) mitochondrial function, we hypothesized that depletion of either PPARγ or PGC1α perturbs mitochondrial structure and function to stimulate PASMC proliferation. To test this hypothesis, HPASMCs were exposed to hypoxia and treated pharmacologically with the PPARγ antagonist GW9662 or with siRNA against PPARγ or PGC1α for 72 hours. HPASMC proliferation (cell counting), target mRNA levels (qRT-PCR), target protein levels (Western blotting), mitochondria-derived H

Topics: Anilides; Animals; Cell Hypoxia; Cell Proliferation; Cells, Cultured; Humans; Hypertension, Pulmonary; Mice, Inbred C57BL; Mitochondria, Muscle; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; PPAR gamma; Pulmonary Artery; RNA Interference

Peroxisome proliferator-activated receptor type gamma (PPARgamma) is a subgroup of the PPAR transcription factor family. Recent studies indicate that loss of PPARgamma is associated with the development of pulmonary hypertension (PH). We hypothesized that the endothelial dysfunction associated with PPARgamma inhibition may play an important role in the disease process by altering cellular gene expression and signaling cascades. We utilized microarray analysis to determine if PPARgamma inhibition induced changes in gene expression in pulmonary arterial endothelial cells (PAEC). We identified 100 genes and expressed sequence tags (ESTs) that were upregulated by >1.5-fold and 21 genes and ESTs that were downregulated by >1.3-fold (P < 0.05) by PPARgamma inhibition. The upregulated genes can be broadly classified into four functional groups: cell cycle, angiogenesis, ubiquitin system, and zinc finger proteins. The genes with the highest fold change in expression: hyaluronan-mediated motility receptor (HMMR), VEGF receptor 2 (Flk-1), endothelial PAS domain protein 1 (EPAS1), basic fibroblast growth factor (FGF-2), and caveolin-1 in PAEC were validated by real time RT-PCR. We further validated the upregulation of HMMR, Flk-1, FGF2, and caveolin-1 by Western blot analysis. In keeping with the microarray results, PPARgamma inhibition led to re-entry of cell cycle at G(1)/S phase and cyclin C upregulation. PPARgamma inhibition also exacerbated VEGF-induced endothelial barrier disruption. Finally we confirmed the downregulation of PPARgamma and the upregulation of HMMR, Flk-1, FGF2, and Cav-1 proteins in the peripheral lung tissues of an ovine model of PH. In conclusion, we have identified an array of endothelial genes modulated by attenuated PPARgamma signaling that may play important roles in the development of PH.

Topics: Anilides; Animals; Arteries; Blood-Air Barrier; Cattle; Cell Adhesion; Cell Cycle; Cell Proliferation; Disease Models, Animal; Endothelial Cells; Gene Expression Profiling; Gene Expression Regulation; Humans; Hypertension, Pulmonary; Lung; Neovascularization, Physiologic; PPAR gamma; Reproducibility of Results; Reverse Transcriptase Polymerase Chain Reaction; Sheep; Ubiquitin; Vascular Endothelial Growth Factor A; Zinc Fingers