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J Korean Soc Pediatr Nephrol. 2011 Oct;15(2):138-145. Korean. In Vitro.
Choi JY , Ahn EM , Park HY , Shin JI , Ha TS .
Department of Pediatrics, College of Medicine, Chungbuk National University, Cheongju, Korea. tsha@chungbuk.ac.kr
The Institute of Kidney Disease, Department of Pediatrics, Yonsei University College of Medicine, Severance Children's Hospital, Seoul, Korea.
Abstract

PURPOSE: To test whether the expression of beta-catenin, a component of podocyte as a filtration molecule, would be altered by puromycin aminonucleoside (PAN) in the cultured podocyte in vitro. METHODS: We cultured rat glomerular epithelial cells (GEpC) with various concentrations of PAN and examined the distribution of beta-catenin by confocal microscope and measured the change of beta-catenin expression by Western blotting and reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: We found that beta-catenin relocalized from peripheral cytoplasm to inner cytoplasm, therefore, intercellular separations were seen in confluently cultured cells by high concentrations of PAN in immunofluorescence views. In Western blotting of GEpC, PAN (50 microg/mL) decreased beta-catenin expression by 34.9% at 24 hrs and 34.3% at 48 hrs, compared to those in without PAN condition (P<0.05). In RT-PCR, high concentrations (50 microg/mL) of PAN also decreased beta-catenin mRNA expression similar to protein suppression by 25.4% at 24 hrs and 51.8% at 48 hrs (P<0.05). CONCLUSION: Exposure of podocytes to PAN in vitro relocates beta-catenin internally and reduces beta-catenin mRNA and protein expression, which could explain the development of proteinuria in experimental PAN-induced nephropathy.

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