Prolonged exposure to decreased oxygen tension, as occurs with many pulmonary diseases, results in pulmonary hypertension, significantly worsening prognosis. The mechanism underlying the pathogenesis of this process remains unknown. Pulmonary arterial smooth muscle cell (PASMC) contraction associated with chronic hypoxia (CH) may be caused by elevated intracellular Ca2+ concentration ([Ca2+]i). In PASMCs, depolarization is observed with CH, fueling speculation that [Ca2+]i is increased due to activation of voltage-gated Ca2+ channels or enhanced Na+/Ca2+ exchange. The endothelium-derived constricting factor, endothelin-1 (ET-1), may contribute to the pathogenesis of CHPH, since ET-1 increases [Ca2+]i. Following exposure to CH, the ET-1-induced rise in [Ca2+]i is reduced but contraction is maintained, suggesting activation of Ca2+-independent contractile pathways. This may be due to ET-1-induced activation of tyrosine kinases (TK). Hypoxic induction of ET-1 occurs via activation of the transcription factor, HIF-1, in systemic endothelium and in mice partially deficient for HIF-1, CH-induced pulmonary hypertension is markedly reduced. Therefore, we hypothesize induction of HIF-1 is an initiating step in the development of pulmonary hypertension, leading to elevated ET-1 levels. ET-1 then diffuses to PASMCs, activating three contractile mechanisms. First, ET-1 decreases voltage-gated K+ channel expression, leading to depolarization-driven activation of Na+/Ca2+ exchange and elevation of resting [Ca2+]i. Second, ET-1 causes TK-mediated Ca2+ influx through L-type Ca2+ channels. Both of these mechanisms increase phosphorylation of myosin light chains (MLCs). Finally, ET-1 causes changes in Ca2+-sensitivity of the contractile apparatus via TK-mediated regulation of actin binding proteins. This final step allows actin to interact with the phosphorylated MLCs generated in steps 1 and 2, and results in contraction. To test these hypotheses, we will use a combination of techniques in our model of hypoxic pulmonary hypertension, including isometric tension recording in arterial segments, Northern and Western blot analysis, whole-cell patch-clamp and microfluorescence measurements, to accomplish the following Specific Aims: 1) confirm that HIF-1 regulates hypoxic induction of ET-1 in the pulmonary vasculature and identify the cell type(s) involved; 2) determine the mechanisms responsible for the CH-induced increase in resting [Ca2+]i and 3) determine the mechanisms by which ET-1 causes contraction during CH.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL067191-01A2
Application #
6617750
Study Section
Lung Biology and Pathology Study Section (LBPA)
Program Officer
Gail, Dorothy
Project Start
2003-08-08
Project End
2007-07-31
Budget Start
2003-08-08
Budget End
2004-07-31
Support Year
1
Fiscal Year
2003
Total Cost
$286,125
Indirect Cost
Name
Johns Hopkins University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
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Shimoda, Larissa A; Laurie, Steven S (2013) Vascular remodeling in pulmonary hypertension. J Mol Med (Berl) 91:297-309
Pisarcik, Sarah; Maylor, Julie; Lu, Wenju et al. (2013) Activation of hypoxia-inducible factor-1 in pulmonary arterial smooth muscle cells by endothelin-1. Am J Physiol Lung Cell Mol Physiol 304:L549-61
Wang, Jian; Shimoda, Larissa A; Sylvester, J T (2012) Ca2+ responses of pulmonary arterial myocytes to acute hypoxia require release from ryanodine and inositol trisphosphate receptors in sarcoplasmic reticulum. Am J Physiol Lung Cell Mol Physiol 303:L161-8
Abud, Edsel M; Maylor, Julie; Undem, Clark et al. (2012) Digoxin inhibits development of hypoxic pulmonary hypertension in mice. Proc Natl Acad Sci U S A 109:1239-44
Luke, Trevor; Maylor, Julie; Undem, Clark et al. (2012) Kinase-dependent activation of voltage-gated Ca2+ channels by ET-1 in pulmonary arterial myocytes during chronic hypoxia. Am J Physiol Lung Cell Mol Physiol 302:L1128-39
Sylvester, J T; Shimoda, Larissa A; Aaronson, Philip I et al. (2012) Hypoxic pulmonary vasoconstriction. Physiol Rev 92:367-520
Shimoda, Larissa A; Polak, Jan (2011) Hypoxia. 4. Hypoxia and ion channel function. Am J Physiol Cell Physiol 300:C951-67
Weigand, Letitia; Shimoda, Larissa A; Sylvester, J T (2011) Enhancement of myofilament calcium sensitivity by acute hypoxia in rat distal pulmonary arteries. Am J Physiol Lung Cell Mol Physiol 301:L380-7

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