Pulmonary vascular resistance (PVR) in the fetus undergoes a dramatic decrease at birth to facilitate gas exchange during postnatal life. Exposure to higher oxygen tension at birth plays a major role in this adaptation. Endothelial nitric oxide synthase (eNOS) regulates the transition in PVR and is stimulated directly by oxygen and indirectly by an increase in oxidative phosphorylation and ATP release. Oxidative phosphorylation also generates superoxide (O27-) as a byproduct. However, the mechanisms that regulate mitochondrial oxidative stress in the fetal pulmonary arteries during this transition remain unclear. Failure of PVR to decrease at birth leads to persistent pulmonary hypertension of the newborn (PPHN), a condition associated with increased oxidative stress in pulmonary arteries. Identification of the sources of oxidative stress is critical to the development of specific therapies to correct the vascular dysfunction in PPHN. Studies in fetal lamb pulmonary artery endothelial cells (PAEC) demonstrated that exposure to ATP, a NOS agonist, or transition to postnatal oxygen tension stimulate the association of eNOS with the mitochondrial outer membrane protein, porin. NOS agonists increase mitochondrial NO release in fetal PAEC. We propose to investigate the novel hypothesis that targeting of eNOS to the mitochondrial outer membrane regulates the rate of oxidative phosphorylation and O27- generation in PAEC by targeted NO release during birth- related transition. The broad specific aims of the proposed studies are to (1) Investigate whether eNOS - mitochondrial interactions regulate the mitochondrial O27- during birth related transition in PAEC and to define the mechanism of this targeting and (2) determine whether eNOS-mitochondrial interactions regulate the endothelial functions critical to postnatal adaptation: synthesis of NO, PGI2 and endothelin-1 and postnatal pulmonary vasodilation. Studies will be done in PAEC and intact fetal lambs. Cell biology tools will be developed to (A) target eNOS to specific sub-cellular compartments in fetal PAEC and (B) to inhibit the interactions of eNOS with the client proteins involved in mitochondrial targeting - hsp90 and porin. The proposed studies will investigate a new regulatory mechanism for the mitochondrial oxidative stress during oxygen exposure at birth. Identification of this new source of oxidative stress in PAEC may lead to more specific therapies, such as mitochondrial targeted anti- oxidants to restore pulmonary vasodilation and oxygenation in PPHN.

Public Health Relevance

Exposure of the fetal lung to oxygen at birth mediates the increase in pulmonary blood flow which is necessary to establish gas exchange during postnatal life. Oxygen increases the metabolism in mitochondria which in turn can increase free radical production. The proposed studies will investigate a new mechanism that regulates the formation of free radicals during this adaptation.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Small Research Grants (R03)
Project #
5R03HD065841-02
Application #
8118777
Study Section
Pediatrics Subcommittee (CHHD)
Program Officer
Raju, Tonse N
Project Start
2010-07-20
Project End
2013-06-30
Budget Start
2011-07-01
Budget End
2013-06-30
Support Year
2
Fiscal Year
2011
Total Cost
$73,440
Indirect Cost
Name
Medical College of Wisconsin
Department
Pediatrics
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Afolayan, Adeleye J; Eis, Annie; Alexander, Maxwell et al. (2016) Decreased endothelial nitric oxide synthase expression and function contribute to impaired mitochondrial biogenesis and oxidative stress in fetal lambs with persistent pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 310:L40-9
Teng, Ru-Jeng; Wu, Tzong-Jin; Afolayan, Adeleye J et al. (2016) Nitrotyrosine impairs mitochondrial function in fetal lamb pulmonary artery endothelial cells. Am J Physiol Cell Physiol 310:C80-8
Mahajan, Chaitali N; Afolayan, Adeleye J; Eis, Annie et al. (2015) Altered prostanoid metabolism contributes to impaired angiogenesis in persistent pulmonary hypertension in a fetal lamb model. Pediatr Res 77:455-62
Konduri, Girija G; Afolayan, Adeleye J; Eis, Annie et al. (2015) Interaction of endothelial nitric oxide synthase with mitochondria regulates oxidative stress and function in fetal pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol 309:L1009-17
Teng, Ru-Jeng; Rana, Ujala; Afolayan, Adeleye J et al. (2014) Nogo-B receptor modulates angiogenesis response of pulmonary artery endothelial cells through eNOS coupling. Am J Respir Cell Mol Biol 51:169-77
Afolayan, Adeleye J; Teng, Ru-Jeng; Eis, Annie et al. (2014) Inducible HSP70 regulates superoxide dismutase-2 and mitochondrial oxidative stress in the endothelial cells from developing lungs. Am J Physiol Lung Cell Mol Physiol 306:L351-60
Teng, Ru-Jeng; Du, Jianhai; Afolayan, Adeleye J et al. (2013) AMP kinase activation improves angiogenesis in pulmonary artery endothelial cells with in utero pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 304:L29-42
Afolayan, Adeleye J; Eis, Annie; Teng, Ru-Jeng et al. (2012) Decreases in manganese superoxide dismutase expression and activity contribute to oxidative stress in persistent pulmonary hypertension of the newborn. Am J Physiol Lung Cell Mol Physiol 303:L870-9
Teng, Ru-Jeng; Du, Jianhai; Welak, Scott et al. (2012) Cross talk between NADPH oxidase and autophagy in pulmonary artery endothelial cells with intrauterine persistent pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 302:L651-63
Caldwell, Ray A; Grove, Diane E; Houck, Scott A et al. (2011) Increased folding and channel activity of a rare cystic fibrosis mutant with CFTR modulators. Am J Physiol Lung Cell Mol Physiol 301:L346-52

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