The Role of Mitochondria in Oxygen Sensing of the Ductus Arteriosus and Fetal Pulmonary Arteries
Loading...
Authors
Bentley, Rachel Emily Teresa
Date
2024-08-29
Type
thesis
Language
eng
Keyword
Ductus Arteriosus , Fetal Circulation , Oxygen Sensing , Mitochondria , Transcriptomics , Proteomics
Alternative Title
Abstract
Rationale: Mitochondria underlie the oxygen responses of the ductus arteriosus (DA), adult pulmonary arteries (PA) and other oxygen sensing tissues. However, the fetal PA oxygen response has been little studied, the identity of the DA oxygen sensor remains elusive, and the basis for the opposing oxygen responses of the DA (vasoconstriction) and PA (vasodilation) are unknown. With the transition from fetal (hypoxic) to neonatal (normoxic) life, a failure of the DA to constrict or the PA to dilate in response to oxygen can precipitate patent ductus arteriosus and persistent pulmonary hypertension of the newborn, respectively.
Objectives: 1) determine the molecular changes in human DA smooth muscle cells (DASMC) induced by exposure to postnatal oxygen concentrations, 2) examine the contribution of individual mitochondrial electron transport chain (ETC) subunits to DASMC oxygen response, and 3) explore the proteomic differences between fetal DA and PA that might underlie their opposing oxygen responses.
General Methods: Cell lines were purified with fluorescence activated cell sorting and oxygen response measured with confocal calcium imaging. Human DASMC grown for 96-hours in hypoxia (2.5% oxygen) or normoxia (19.6% oxygen) were used for 3’RNA-sequencing. Silencing RNA was used to selectively knock down ETC subunits, with effects on oxygen responsiveness examined with calcium imaging. The bioenergetic effects of knockdown were examined with micropolarimetry and ETC activity assays. Liquid chromatography with tandem mass spectrometry was used to explore proteomic differences in whole DA versus PA and mitochondrially-enriched DASMC versus PASMC samples.
General Results: 1) Transcriptomic analysis revealed mitochondrial pathway enrichment between hypoxic and normoxic DASMC. Among genes in ETC Complex I, NADH:Ubiquinone oxidoreductase core subunit S2 (NDUFS2) was uniquely downregulated by oxygen. 2) Knockdown of NDUFS2 reduced DASMC acute oxygen responsiveness without bioenergetic inhibition. 3) Rabbit DA and PA have unique proteomes, with differences in structural, proliferative, and metabolic proteins.
Conclusions: This thesis furthers our understanding of the oxygen response of the human DA, demonstrating a key role for NDUFS2 and characterizing the molecular changes in gene transcription caused by oxygen. This first omic comparison between DA and PA identifies mitochondrial-metabolic protein differences that may explain their opposing response to oxygen.
Description
Citation
Publisher
License
Queen's University's Thesis/Dissertation Non-Exclusive License for Deposit to QSpace and Library and Archives Canada
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution-NonCommercial-NoDerivatives 4.0 International
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution-NonCommercial-NoDerivatives 4.0 International