A Podocyte-Based Automated Screening Assay Identifies Protective Small Molecules.

Autor:	Lee HW; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Khan SQ; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Faridi MH; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Wei C; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Tardi NJ; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Altintas MM; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Elshabrawy HA; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Mangos S; Department of Medicine, Rush University Medical Center, Chicago, Illinois;, Quick KL; Cellular Imaging and Analysis, PerkinElmer Life Sciences, Waltham, Massachusetts; and., Sever S; Department of Medicine, Harvard Medical School, Division of Nephrology, Massachusetts General Hospital, Charlestown, Massachusetts., Reiser J; Department of Medicine, Rush University Medical Center, Chicago, Illinois; jochen_reiser@rush.edu vineet_gupta@rush.edu., Gupta V; Department of Medicine, Rush University Medical Center, Chicago, Illinois; jochen_reiser@rush.edu vineet_gupta@rush.edu.
Jazyk:	angličtina
Zdroj:	Journal of the American Society of Nephrology : JASN [J Am Soc Nephrol] 2015 Nov; Vol. 26 (11), pp. 2741-52. Date of Electronic Publication: 2015 Apr 09.
DOI:	10.1681/ASN.2014090859
Abstrakt:	Podocyte injury and loss mark an early step in the pathogenesis of various glomerular diseases, making these cells excellent targets for therapeutics. However, cell-based high-throughput screening assays for the rational development of podocyte-directed therapeutics are currently lacking. Here, we describe a novel high-content screening-based phenotypic assay that analyzes thousands of podocytes per assay condition in 96-well plates to quantitatively measure dose-dependent changes in multiple cellular features. Our assay consistently produced a Z' value >0.44, making it suitable for compound screening. On screening with >2100 pharmacologically active agents, we identified 24 small molecules that protected podocytes against injury in vitro (1% hit rate). Among the identified hits, we confirmed an β1-integrin agonist, pyrintegrin, as a podocyte-protective agent. Treatment with pyrintegrin prevented damage-induced decreases in F-actin stress fibers, focal adhesions, and active β1-integrin levels in cultured cells. In vivo, administration of pyrintegrin protected mice from LPS-induced podocyte foot process effacement and proteinuria. Analysis of the murine glomeruli showed that LPS administration reduced the levels of active β1 integrin in the podocytes, which was prevented by cotreatment with pyrintegrin. In rats, pyrintegrin reduced peak proteinuria caused by puromycin aminonucleoside-induced nephropathy. Our findings identify pyrintegrin as a potential therapeutic candidate and show the use of podocyte-based screening assays for identifying novel therapeutics for proteinuric kidney diseases. (Copyright © 2015 by the American Society of Nephrology.)
Databáze:	MEDLINE
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