Mitochondrial Imaging, PTM Analysis & Metabolism Core

ABSTRACT The goal of the Mitochondrial Imaging, PTM analysis, and Metabolism Core (Core B) is to provide a high-quality state-of-the art service to support analyses in all three of the PPG Projects. In this context, Core B will be defined by providing four services. 1) High-resolution imaging and functional characterization of mitochondria. This will include analyses of specific protein redistribution to mitochondria, mitochondrial network remodeling, and markers of mitochondrial function (mitochondrial-derived reactive oxygen species, mitochondrial membrane potential, carnitine metabolites, ATP levels). 2) Bioenergetics: oxygen consumption, rate of glycolysis, fatty acid oxidation, measurement of cellular ATP kinetics, ETC complex analyses. 3) The analysis of protein PTMs in biological samples. Individual proteins will be purified to assay PTMs in vivo. To analyze PTMs, we will utilize a mass spectrometry approach coupled to the detection and quantification of specific sites of modification by tandem mass spectrometry. 4) Metabolomic analyses. Individual metabolites in cultured cells, lung tissue, and plasma from animals and children born with congenital heart disease that result in changes in pulmonary blood flow (PBF). In addition, directional metabolomic flux analysis will be performed to understand the dyanimics of metabolic pathways. Core B will enhance the scientific work for all three projects by assuring that all mitochondrial, PTM, and metabolomic analyses are standardized and performed in a uniform standardized manner, and by highly experienced and trained personnel. Core B is run by experienced investigators who have had many years of experience using these techniques. They have developed or adapted many of these techniques and applied them to perform physiological studies in vitro and in vivo systems. As a central dedicated analytical resource, Core B will ensure these analyses are carried out in a cost-efficient manner.