Chronic kidney disease (CKD) is a growing public health problem that affects 10% of Americans, increases morbidity and mortality, and imposes a huge economic burden. Persistent renal tubular injury is an important component of tubulointerstitial fibrosis (TIF), the common feature of progressive CKD of any etiology. Growth factors such as transforming growth factor-? (TGF??) are important determinants of how tubula epithelia respond to injury. Although excessive TGF-? signaling promotes TIF, blocking TGF-??using genetic tools results in increased tubular damage after mouse models of CKD, suggesting that TGF-??signaling has protective effects as well. TGF-??mediates many differet cellular effects including G1 arrest of cell cycle progression and epithelial de-differentiation. Tis proposal tests the hypothesis that TGF???signaling in the proximal tubule promotes survival ater chronic renal injury by inducing cell cycle arrest and epithelial de- differentiation. The first ai investigates how TGF-? alters cell cycle progression and how these changes affect epithelial survival using murine models of injury and pharmacologic inducers of G1 arrest. To test this, we will use genetically modified mice that lack the TGF?? receptor (T?RII) specifically in the proximal tubule and induce chronic kidney injury by either angiotensin/ uninephrectomy or aristolochic acid. We anticipate that mice lacking T?RII have impaired G1 arrest and reduced epithelial survival after injury and that this improves with administration of palbociclib, an FDA-approved inducer of G0/G1 cell cycle arrest. Proximal tubule cells with and without T?RII in vitro will be used to understand the signaling pathways through which TGF-? induces cell cycle arrest.
The second aim explores mechanisms whereby TGF????lters epithelial differentiation nd how these changes impact survival. The role of epithelial differentiation in chronic kidney injury will be defined using mice with epithelial-specific deletion of Snail, a key transcription factor downstream of TGF-?? Proximal tubule cells with and without T?RII will be transfected with siRNA and constructs in vitro to test the hypothesis that Snail mediates TGF-?-dependent de-differentiation.
The third aim defines how the Wnt/?-catenin pathway, important in renal injury, interacts with TGF-? to affect epithelial survival. Genetic models and cell culture techniques wil be used to test the hypothesis that TGF-? increases canonical Wnt/??catenin signaling to improve epithelial survival through increased de-differentiation and G0/G1 cell cycle arrest. This proposal uses innovative genetic approaches and a new strategy for analyzing injury in a quantitative and location-specific way. These studies will provide valuable insights into the roles of cell cycle and epithelial plasticity in the epithelial response to CKD, focusing on the proximal tubule which is the target of acute kidney injury and a mediator of CKD. However, information about cell cycle, epithelial plasticity and response to chronic injury has implications beyond the kidney as most organ dysfunction results from chronic epithelial injury.
Chronic kidney disease is a growing public health problem which affects 10% of Americans. The proposed studies will shed valuable insight into how tubular epithelia, the target of injury, respond to chronic renal injury using genetically modified mouse models, cell culture techniques, and an FDA-approved pharmacologic agent. These studies have the potential to broaden our understanding of how epithelia respond to chronic injury in other organs with important therapeutic implications.
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