There is a fundamental gap in our understanding of the mechanisms that determine functional impairment in acute kidney injury (AKI). Continued existence of this gap presents an important clinical problem because therapeutic interventions to prevent, treat or hasten recovery in AKI have not and cannot be fully realized until this gap is filled. The long term goal is to understand the cellular and molecular regulatory functions of the protein p53 in the kidney under health and disease and to develop novel and specific therapeutic interventions targeting these mechanisms. The objective of this application, which is a step toward attainment of the long term goal, is to selectively determine how p53 regulates basal kidney metabolism and modulates inflammation during AKI. The central hypothesis of this application is that p53 is an important modulator of glucose metabolism in the kidney, as well as the innate immune response during AKI. This hypothesis has been formulated on the basis of existing literature and strong preliminary data from the applicants'laboratories. The rationale fo the proposed research is that once it is determined how p53 regulates kidney metabolism and inflammatory responses during AKI, it then permits the strategic modulation of p53 as a new and innovative pharmacological approach towards preventing and treating AKI. The central hypothesis will be tested by pursuing two specific aims: 1) Delineate, by way of advanced imaging and biochemical techniques, the capacity to shift kidney glucose metabolism towards a protective glycolytic phenotype through inhibition of p53;and 2) Determine the relative roles of tubular and leukocyte p53 in regulating the inflammatory response during AKI. Under the first aim, in vivo inhibition of p53 will be accomplished by pharmacologic inhibition (pifithrin alpha), siRNA silencing techniques, and targeted genetic deletion. Shifts in glycolysis and oxidative phosphorylation at the cellular level will be determined with intravital multiphoton microscopy of the kidney in living animals by employing fluorescent probes specific for the two major metabolic pathways and global shifts in kidney metabolism will be quantified with PET scanning techniques. These studies will be complemented by techniques to measure changes in key metabolic enzymes known to be modulated by p53. The protective capacity of this metabolic shift will be examined in models of AKI. Under the second aim, similar strategies will be used to inhibit p53 in vivo. Flow cytometry coupled with advanced molecular and cell biological techniques will be used to define alterations in the inflammatory response during AKI. These studies will be complemented by chimeric mouse models to further discern the mechanisms involved. The approach is innovative, because our approach to manipulate p53 signaling in order to modify kidney metabolism and innate immunity represents a new and substantial departure from the status quo. The proposed research is significant, because it is the next step in a continuum of research that is expected to lead to the development of more specific and targeted therapeutic interventions aimed at p53 that will prevent and limit subsequent dysfunction in AKI.
The proposed research is relevant to public health because the discovery of pathophysiological mechanisms modulating epithelial cell metabolism and inflammation in acute kidney injury is ultimately expected to provide novel and rationale targets for therapeutic intervention in this deadly disease. Thus the proposed research is relevant to the part of the NIH's mission that pertains to developing knowledge that will help to reduce the burden of human disease and improve people's health and quality of life.
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