A pitfall of glomerular sieving: Profibrotic and matrix proteins derive from the Bowman's capsule and not the glomerular tuft in rats with renovascular hypertension

Abstract

Background. The glomeruli in the non-clipped kidney of rats with 2-kidney, 1-clip hypertension are a classical model for studying the mechanisms of glomerular injury. Methods. In the present study, we compared the glomerular expression of PAI-1 and collagen I α1 mRNA from glomeruli isolated by the classic technique of sieving with the recently developed technique of tissue laser microdissection. For quantification of mRNA from both methods, real-time PCR was used. Results. Real-time PCR revealed a 9.0 ± 1.3- and a 7.1 ± 0.2-fold induction of PAI-1 and collagen I α 1, respectively, in the glomeruli from hypertensive rats isolated by sieving. However, in situ hybridization and microdissection revealed that expression of both mRNAs was mainly from the Bowman's capsule and not from the glomerular tuft (10.7 ± 1.3- and 7.2 ± 0.6-fold higher induction in whole glomeruli compared with tuft alone). Conclusion. This emphasizes that studies focusing on processes in the mesangium, endothelial cells or podocytes should not rely on glomeruli obtained by sieving. Rather, a technique like the laser microdissection or in situ hybridization should be applied which allows the clear separation of different glomerular and periglomerular compartments. © The Author [2007]. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.