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Dissection of novel pathways leading to podocyte dysfunction and proteinuria

Wang, Dan (2010) Dissection of novel pathways leading to podocyte dysfunction and proteinuria. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Podocytes are highly differentiated glomerular epithelial cells that play an essential role in the establishment of the glomerular filtration barrier, a structural apparatus that selectively restricts the filtration of different macromolecules in the blood stream on the basis of their sizes, shape and charge. Podocyte dysfunction, one of the major causes of proteinuria, is of pathogenetic and prognostic significance in human glomerular disease. My study is focused on the investigation of novel pathways leading to podocyte dysfunction and proteinuria.In the first part, immunoblotting and quantitative reverse transcriptase PCR (RT-PCR) were used to demonstrate that LIM and senescent cell antigen-like domains 1 (PINCH1) is induced and undergoes nuclear translocation in podocytes after transforming growth factor, beta 1 (TGF-beta1) treatment. Bioinformatics analysis revealed the putative nuclear export signal/nuclear localization signal (NES/NLS) at the PINCH1 C-terminus which is required for its nuclear translocation. Immunoprecipitation and GST pull-down assay identified the interaction between PINCH1 and Wilms tumor 1 (WT1) which led to suppression of the WT1-mediated podocalyxin gene expression. In vivo, PINCH1 also underwent nuclear translocation and interacted with WT1 after TGF-beta1 stimulation. Our data identifies nuclear transcription factor WT1 as a novel binding partner for PINCH1, and provides novel insight into the mechanism of podocyte dysfunction under pathological conditions. In the second part, RT-PCR results revealed that treatment with TGF-beta1 induced gene expression of several wingless-type MMTV integration site family members (Wnts), predominantly Wnt1, and activated beta-catenin in mouse podocytes. Wnt antagonist Dickkopf-1 (DKK1) blocked TGF-beta1-induced beta-catenin activation and preserved nephrin expression. In vivo, ectopic expression of constitutively active TGF-beta1 induced Wnt1 expression, activated glomerular beta-catenin, upregulated its downstream target genes, and led to podocyte injury and proteinuria. Consistently, concomitant expression of DKK1 gene abolished beta-catenin activation in mouse glomeruli, inhibited TGF-beta1-triggered Wnt/beta-catenin target genes, and ameliorated proteinuria. These results establish a role for Wnt/beta-catenin signaling in the pathogenesis of podocyte injury and also suggest that this signaling pathway could be exploited as a therapeutic target for the treatment of proteinuric kidney diseases.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Wang, Dandaw35@pitt.eduDAW35
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairZarnegar, Rezarezazar@pitt.eduREZAZAR
Committee MemberWu, Carycarywu@pitt.eduCARYWU
Committee MemberStolz, Donna Bdstolz@pitt.eduDSTOLZ
Committee MemberWeisz, Ora Aweisz@pitt.eduWEISZ
Committee MemberMonga, Satdarshan Pal Singhsmonga@pitt.eduSMONGA
Committee MemberLiu, Youhualiuy@upmc.eduYHLIU
Date: 29 November 2010
Date Type: Completion
Defense Date: 22 November 2010
Approval Date: 29 November 2010
Submission Date: 29 November 2010
Access Restriction: 5 year -- Restrict access to University of Pittsburgh for a period of 5 years.
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Cellular and Molecular Pathology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: beta-catenin; DKK; pdocyte; PINCH1; TGF-beta1; Wnt
Other ID: http://etd.library.pitt.edu/ETD/available/etd-11292010-153331/, etd-11292010-153331
Date Deposited: 10 Nov 2011 20:06
Last Modified: 19 Dec 2016 14:37
URI: http://d-scholarship.pitt.edu/id/eprint/9863

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