1Department of Pediatrics, College of Medicine, Chungbuk National University Cheongju, The Institute of Kidney Disease 2Department of Pediatrics, College of Medicine, Chungbuk National University Cheongju, The Institute of Kidney Disease 3Department of Pediatrics, College of Medicine, Chungbuk National University Cheongju, The Institute of Kidney Disease 4Department of Pediatrics, Yonsei University College of Medicine, Severance Children's Hospital 5Department of Pediatrics, College of Medicine, Chungbuk National University Cheongju, The Institute of Kidney Disease
Received: September 1, 2011; Accepted: October 4, 2011.
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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 ($50{mu}g/mL$) 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 ($50{mu}g/mL$) 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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