Pulmonary hypertension (PH) is a devastating cardiopulmonary disorder with significant morbidity and mortality that frequently complicates disorders that affect the veteran patient population. Existing PH therapies are expensive and not optimally effective, indicating that novel approaches to PH treatment are urgently needed. This proposal seeks to define new pathways in PH pathogenesis and develop novel strategies for PH therapy. The proposed studies focus on the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARg) which has been implicated in PH pathogenesis. Reduced PPARg expression caused PH whereas activation of PPARg with existing medications attenuated PH. Unfortunately, these same PPARg ligands also cause significant side effects. Thus to avoid these adverse effects, this proposal examines mechanisms of PPARg downregulation in PH and ways to prevent it as a new and alternative strategy in PH therapy. The investigators will examine PH in mice caused by exposure to chronic hypoxia ? treatment with a vascular endothelial growth factor receptor (VEGF-R) antagonist. Complementary studies will be performed in vitro in human pulmonary artery endothelial or smooth muscle cells (HPAEC or HPASMC) exposed to hypoxia or in HPAEC or HPASMC isolated from control or PH patients. The investigators hypothesize that hypoxia- induced reductions in PPARg in the pulmonary vascular wall promote the expression of downstream mediators that cause vascular cell proliferation, remodeling and PH. Further, it is predicted that preventing or attenuating reductions in PPARg expression and function will reduce proliferative mediator expression and PH. The proposal will address 2 specific aims.
The first aim focuses on transcriptional mechanisms that regulate PPARg expression.
Aim 1 will determine the role of NF-kB in suppressing PPARg promoter activity. Published and preliminary data demonstrate that NF-kB is activated in PH and by exposure to chronic hypoxia. The expression of PPARg and downstream PPARg-regulated proliferative mediators (including NADPH oxidase 4, endothelin-1, and thrombospondin-1) will be measured in the proposed models using quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting. Vascular cell proliferation, vascular remodeling, and PH will be determined with techniques published by the investigative team. Activation of NF-kB will be determined by electrophoretic mobility shift assays. siRNA will be administered to inhibit NF-kB to determine its role in reduced PPARg expression and in the increased expression of proliferative mediators that cause PH.
Aim 2 will focus on the post-transcriptional regulation of PPARg by microRNA-27 (miR-27). Levels of miR-27 will be measured in the models employed in Aim 1 using qRT-PCR, and its role in PH pathobiology will be established using miR-27 antagonists or overexpression. Binding of miR-27 to the PPARg 3'-UTR will be confirmed using luciferase reporter constructs. Intranasal delivery of anti-miR-27 to the lower respiratory tract will be used in vivo to determine if preventing increases in miR-27 can avert reductions in PPARg and attenuate downstream increases in proliferative mediators. The proposed studies will clarify pathogenic mechanisms in PH and define the relative contributions of transcriptional and post-transcriptional pathways to reductions in PPARg. The outcomes will determine if strategies to prevent reductions in pulmonary vascular PPARg expression can provide a novel approach to PH therapy. The proposed strategies might also enhance the therapeutic effects of existing PPARg ligands thereby permitting reductions in dosage and associated side effects. The results of this proposal can thereby define novel and effective therapeutic strategies to regulate programs of gene expression involved in PH pathogenesis.

Public Health Relevance

Pulmonary hypertension, or high blood pressure in the lungs, is a serious and not infrequent complication of many disorders affecting veteran patients including congestive heart failure, chronic obstructive pulmonary disease, interstitial lung disease, obstructive sleep apnea, HIV infection, collagen vascular disease, and thromboembolic disease to name a few. The development of pulmonary hypertension in these conditions significantly increases morbidity and mortality. In addition, the mechanisms that cause pulmonary hypertension remain to be completely defined. These considerations indicate that new and more effective therapies for veteran patients with these disorders could have significant impact on patient outcomes and VA healthcare. The proposed research will further examine how pulmonary hypertension occurs and will explore new approaches for treating pulmonary hypertension.

National Institute of Health (NIH)
Veterans Affairs (VA)
Non-HHS Research Projects (I01)
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Veterans Health Administration
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