NDT Advance Access originally published online on March 30, 2006
Nephrology Dialysis Transplantation 2006 21(7):1838-1847; doi:10.1093/ndt/gfl090
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© The Author [2006]. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org
Original Articles: Clinical Nephrology
Quantitative analysis of type IV collagen subchains in the glomerular basement membrane of patients with Alport syndrome with confocal microscopy
Research Institute of Nephrology, Jinling Hospital, Nanjing University School of Medicine, Nanjing, 210002, China
Correspondence and offprint requests to: Zhi-Hong Liu, MD, Prof. of Medicine, Research Institute of Nephrology, Jinling Hospital, Nanjing University School of Medicine, Nanjing 210002, China. Email: zhihong{at}21cn.net
Background. Alport syndrome (AS) is an inherited nephropathy characterized by glomerular basement membrane (GBM) abnormalities due to mutations in the type IV collagen genes. Through immunofluorescence analysis, the absence of 
3(IV), 4(IV) and 5(IV) chains within the GBM has been shown in the majority of AS cases. In some atypical AS cases, however, staining of the GBM with antibodies against the 3(IV), 4(IV) and 5(IV) chains appeared normal. In this study, we studied these atypical AS cases by quantitative analysis of the expression of type IV collagen subchains in GBM.
Methods. Twelve patients diagnosed with AS, yet having normal staining for 3(IV) and 5(IV) chains in the GBM, were recruited. Quantitative analysis of type IV collagen subchains in the GBM was performed using confocal microscopy and immunofluorescence double label techniques.
Results. The absolute amounts of 3(IV), 4(IV) and 5(IV) were significantly lower in AS patients than that in normal subjects, associated with up-regulated expression of type IV collagen in GBM. It was found that eight cases had decreased ratios of 3(IV)/IV, 4(IV)/IV and 5(IV)/IV in the GBM simultaneously; one had reduced levels of 3(IV)/IV and 5(IV)/IV but had a normal level of 4(IV)/IV, and one had reduced 3(IV)/IV with normal 4(IV)/IV and 5(IV)/IV levels. The remaining two patients had normal ratios of 3(IV)/IV, 4(IV)/IV and 5(IV)/IV.
Conclusions. Confocal analysis demonstrated for the first time that the ratios of 3(IV)/IV, 4(IV)/IV and 5(IV)/IV in the GBM decreased in patients with AS, even though routine immunofluorescence staining for (IV) chains appeared normal. This result not only sheds light on the pathogenesis of AS, but also provides an alternative approach to diagnose atypical AS cases.
Keywords: alport syndrome; confocal microscopy; type IV collagen