My goal is to become an independent investigator in disease-oriented research in the newborn with a specific focus on the regulation of mitochondrial oxidative stress in persistent pulmonary hypertension of the newborn (PPHN). PPHN affects 2-6/1000 live births and is a common cause of cardiopulmonary failure in the newborn. Of these infants, >30% fail medical treatment and need invasive support measures (ECMO). Oxidative stress is strongly implicated in the pathogenesis of PPHN. NADPH oxidases are considered the primary source of superoxide (O2) in the pulmonary endothelium. Mitochondrial oxygen consumption during respiration produc- es influx of O2 in the mitochondria as a byproduct of oxidative phosphorylation. Recent evidence indicates that reactive oxygen species (ROS) produced by mitochondria induce the activation of NADPH oxidases, lead- ing to ROS induced ROS formation. However, the regulation of mitochondrial O2formation remains unknown. Identification of the adaptive mechanisms that minimizes mitochondrial O2formation during exposure to oxy- gen at birth may identify additional therapeutic targets in PPHN. I will begin to achieve this goal by engaging in a Career Development plan that logically allows me to expand my prior skills and build new skills in mouse ge- netics, analysis of protein-protein interactions and identification of novel signaling pathways pertinent to endo- thelial biology. This plan integrates didactic training in genetics, biochemistry and free radical biology with learning of skills from my Mentors and Scientific Advisory Committee at the Medical College of Wisconsin who combined have expertise in endothelial biology, developmental vascular biology, free radical and mitochondrial biology. The Mentorship and Career Development plan are integrated with the proposed research objectives to test the hypothesis that Akt induces a post translational modification of hsp70 and modulates the interactions of hsp70 with two recently identified proteins namely: an Obg like ATPase-1 (OLA1) that facilitates SOD2 im- port, and CHIP, which is a chaperone-associated ubiquitin ligase that targets hsp70 for degradation. Phos- phorylation of hsp70 by Akt promotes the interaction of hsp70 with OLA1 and facilitates the mitochondrial im- port of SOD2 to reduce free O2during postnatal transition.
The first aim seeks to determine the contributions of OLA1 and CHIP to the regulation of mitochondrial redox signaling in PAECs. We will use transgenic mice, in vitro kinetic assays and cell culture to determine the contributions of OLA1 and CHIP to mitochondrial redox signaling in PAECs and identify how OLA1 and CHIP mechanistically regulate mitochondrial SOD2 import and ROS production.
The second aim will determine the mechanistic role of PI3K/Akt signaling pathway in regulat- ing the mitochondrial import of SOD2 and the functional relevance of this mechanism to postnatal adaptation. The successful completion of the proposed studies and training program will lead to future studies investigating preventable and treatment strategies to improve outcomes in PPHN patients, with the goal of reducing the economic and health burden due to PPHN.

Public Health Relevance

Persistent pulmonary hypertension of the newborn (PPHN) has a great public health significance as the leading cause of cardiopulmonary failure in the neonatal intensive care unit. Our proposal aims to identify cellular mechanisms that regulate oxidative stress in vascular cells in order to identify a potential therapeutic target in the management of PPHN. This will enable us to improve overall infants? health as PPHN rates continue to rise among extremely preterm babies whose survival has significantly increased in the United States.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Clinical Investigator Award (CIA) (K08)
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NHLBI Mentored Clinical and Basic Science Review Committee (MCBS)
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Colombini-Hatch, Sandra
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Medical College of Wisconsin
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Teng, Ru-Jeng; Jing, Xigang; Michalkiewicz, Teresa et al. (2017) Attenuation of endoplasmic reticulum stress by caffeine ameliorates hyperoxia-induced lung injury. Am J Physiol Lung Cell Mol Physiol 312:L586-L598
Afolayan, Adeleye J; Alexander, Maxwell; Holme, Rebecca L et al. (2017) Domain Mapping of Heat Shock Protein 70 Reveals That Glutamic Acid 446 and Arginine 447 Are Critical for Regulating Superoxide Dismutase 2 Function. J Biol Chem 292:2369-2378
Sharma, Megha; Afolayan, Adeleye J (2017) Redox Signaling and Persistent Pulmonary Hypertension of the Newborn. Adv Exp Med Biol 967:277-287
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