Date of Award

12-2011

Level of Access Assigned by Author

Campus-Only Thesis

Degree Name

Master of Science (MS)

Department

Biochemistry

Advisor

Sharon Ashworth

Second Committee Member

Robert Gundersen

Third Committee Member

Ian Bricknell

Abstract

Acute kidney injury (AKI) is a growing problem in the United States. The high morbidity and mortality of this disease combined with limited treatment options make AKI an important area of research. Renal proximal tubule and interstitial fibroblast cells respond differently to AKI. Actin cytoskeleton structure in proximal tubule cells is disrupted during an ischemic event (Kellerman and Bogusky, 1992). This change is mediated by cofilin activation and altered tropomyosin localization (Schwartz et al., 1999, Ashworth et al., 2001, Ashworth et al., 2004). While changes in cofilin and tropomyosin have been identified during ischemia, the role of these proteins in recovery is not yet fully understood. The presence of interstitial fibroblasts and extracellular matrix proteins is increased after an ischemic event (Forbes et al., 2000). This increase can lead to a condition known as fibrosis, which leads to end stage renal failure (Qi et al., 2006). The role of cofilin and tropomyosin in this process remains unclear. In this study, we observed increased cofilin activation in proximal tubule and interstitial fibroblast cells with twenty minutes of ATP depletion, a model for renal ischemia. With twenty-four hours of ATP repletion, cofilin activation was decreased in interstitial fibroblast cells but increased in proximal tubule cells. At this time point actin stress fibers and cell-to-cell contacts were disrupted in proximal tubule cells, while tropomyosin bound stress fibers were present in renal interstitial fibroblasts. These findings suggested that the differential response of proximal tubule cells and interstitial fibroblasts to AKI may be a result of differences in cofilin activation and tropomyosin localization in these cells. The death-associated protein kinase (DAPK) family proteins are regulators of the actin cytoskeleton (Bialik and Kimchi, 2006). DAPK1 is activated during recovery from ischemia of the brain, leading to its degradation (Shamloo et al., 2005, Schumacher et al., 2002). DAPK2 expression and localization have not been examined in renal ischemia and may signal to the actin cytoskeleton during or after AKI. These studies demonstrated that with twenty-four hours of recovery from ATP depletion, DAPK2 expression was decreased in proximal tubule cells, but increased in renal interstitial fibroblasts. These differences in DAPK2 expression provided hints to a possible mechanism leading to the observed changes in the actin cytoskeleton during recovery from ATP depletion. Understanding the differential responses of proximal tubule cells and renal interstitial fibroblasts during AKI may lead to better treatment options for this disease. Interruption of signals that lead to proximal tubule disruption and fibroblast activation during and after AKI may reduce the incidence of end stage renal failure that can accompany AKI.

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