Acute Renal Failure secondary to ischemia and associated events such as sepsis and rhabdomyolysis remains a major health problem, about which little has been accomplished therapeutically in the last three decades. Actin cytoskeletal events mediate many of the cellular alterations that in turn result in organ dysfunction. In particular, proximal tubular cell microvillar destruction, resulting in tubular obstruction, reduced proximal ion transport and backleak of solutes between proximal tubular cells, is known to play a major role in reduced glomerular filtration and tubular dysfunction. Therefore, the purpose of this grant is to further delineate proximal tubule cell apical actin alterations as mediated by actin depolymerizing factor (ADF)/cofilin, tropomyosins and myosin Vs. Specifically, we propose that a reduction in ATP results in ADF/cofilin activation and increases in calcium result in apical domain tropomyosin dissociation from the F actin cytoskeleton. These two events then lead to ADF medicated F actin destruction, loss of microvillar integrity, loss of apical membrane through blebbing and internalization. Myosin 1 is affected by both ATP depletion and calcium. We believe its function in mediating apical membrane structure, endocytosis and ion channel function is altered. We will use multi-photon microscopy and adenoviral probes delivered by micropuncture techniques to tubular epithelial cells in vivo to follow intravitally these specific proteins under physiologic conditions, during ischemia and during the reperfusion period. Specific roles for these proteins will be approached using additional proteins known to regulate their function, site directed mutagenesis and the use of truncated constructs. Techniques have been developed to allow for quantitative analysis of the effects observed in vivo. Taken together, we anticipate these data will greatly enhance our understanding of the overall process in vivo as-the ability to follow a differentiated proximal tubular cell utilizing specific molecular approaches has not been previously possible.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK069408-04
Application #
7474766
Study Section
Cellular and Molecular Biology of the Kidney Study Section (CMBK)
Program Officer
Kimmel, Paul
Project Start
2005-08-01
Project End
2010-07-31
Budget Start
2008-08-01
Budget End
2010-07-31
Support Year
4
Fiscal Year
2008
Total Cost
$251,710
Indirect Cost
Name
Indiana University-Purdue University at Indianapolis
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
603007902
City
Indianapolis
State
IN
Country
United States
Zip Code
46202
Molitoris, Bruce A (2017) Rethinking CKD Evaluation: Should We Be Quantifying Basal or Stimulated GFR to Maximize Precision and Sensitivity? Am J Kidney Dis 69:675-683
Molitoris, Bruce A; Reilly, Erinn S (2016) Quantifying Glomerular Filtration Rates in Acute Kidney Injury: A Requirement for Translational Success. Semin Nephrol 36:31-41
Melican, Keira; Sandoval, Ruben M; Kader, Abdul et al. (2011) Uropathogenic Escherichia coli P and Type 1 fimbriae act in synergy in a living host to facilitate renal colonization leading to nephron obstruction. PLoS Pathog 7:e1001298
Basile, David P; Friedrich, Jessica L; Spahic, Jasmina et al. (2011) Impaired endothelial proliferation and mesenchymal transition contribute to vascular rarefaction following acute kidney injury. Am J Physiol Renal Physiol 300:F721-33
Wang, Exing; Sandoval, Ruben M; Campos, Silvia B et al. (2010) Rapid diagnosis and quantification of acute kidney injury using fluorescent ratio-metric determination of glomerular filtration rate in the rat. Am J Physiol Renal Physiol 299:F1048-55
Flynn, Kevin C; Pak, Chi W; Shaw, Alisa E et al. (2009) Growth cone-like waves transport actin and promote axonogenesis and neurite branching. Dev Neurobiol 69:761-79
Molitoris, Bruce A; Sandoval, Ruben M (2009) Techniques to study nephron function: microscopy and imaging. Pflugers Arch 458:203-9
Molitoris, Bruce A; Dagher, Pierre C; Sandoval, Ruben M et al. (2009) siRNA targeted to p53 attenuates ischemic and cisplatin-induced acute kidney injury. J Am Soc Nephrol 20:1754-64
Campos, Silvia B; Ashworth, Sharon L; Wean, Sarah et al. (2009) Cytokine-induced F-actin reorganization in endothelial cells involves RhoA activation. Am J Physiol Renal Physiol 296:F487-95
Weinberg, Joel M; Molitoris, Bruce A (2009) Illuminating mitochondrial function and dysfunction using multiphoton technology. J Am Soc Nephrol 20:1164-6

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