The goal of this research proposal is to advance our understanding of the cellular and metabolic alterations which are responsible for the initiation and recovery from acute renal failure and to obtain new insight which will be applicable in the clinical setting. This proposal is designed to provide new information concerning the effects of alterations in adenine nucleotides metabolism in renal tubular cells. We propose: a) to determine the hierarchy of cellular dysfunction in response to adenine nucleotide depletion and b) to define the role of de novo purine synthesis in the post-ischemic recovery of renal ATP. To evaluate these questions, we will employ an integrated approach in which investigations will be carried out in animal preparations in vivo as well as suspensions of tubular segments in vitro. In vivo stuies will evauate renal function and utilize 31P NMR spectroscopy to determine the pattern of alterations in renal ATP during and after an insult. In vitro studies will evaluate cellular and molecular components of the epithelial cell injury. The cellular aspect will define changes in intracellular calcium, phospholipid profile, and morphological changes. The metabolic components will evaluate adenine nucleotides and breakdown products, and oxygen consumption in tubular suspensions under basal, stimulated, and inhibited conditions. The molecular aspects of these studies will focus on the role of cytoskeletal elements, brush-border proteins, tight-junction protein, Na/K-ATPase, and stress proteins in the restoration of cell integrity. In this manner, we propose to determine the cellular, metabolic, and molecular basis of the morphological and physiologic alterations which occur during altered adenine nucleotide metabolism.
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