Although the clinical relevance of proteinuria, especially albuminuria, has been well documented in chronic kidney disease,1 the quantitative mechanistic significance of different barrier components to albuminuria remains an area of considerable excitement and debate. The glomerular filtration barrier (GFB), formed by fenestrated endothelia with their glycocalyx, podocytes with their interdigitated foot processes and slit diaphragms, the subpodocyte space, and the glomerular basement membrane (GBM), has been long believed to be the major determinant of albuminuria with podocyte foot process fusion being central.2 Multiple genetic, molecular, and morphologic lines of evidence implicate the podocyte and its slit diaphragm as playing a central role in the GFB. However, changing concepts regarding the mechanistic relevance and clinical importance of each of these individual components of the GFB have been advancing at a rapid rate.
The glycocalyx of glomerular endothelial cells has been shown by enzymatic degradation,3 use of vascular endothelial growth factor antibodies to induce glomerular endothelial injury,4,5 and mutations of the laminin β3 gene6 to result in macroalbuminuria without morphologic podocyte injury, although controversy remains.2 Basement membrane charge properties and their molecular constituents have also been reexamined and deleted, respectively, in attempts to understand further their importance. Clearly, laminin β2 mutants and null mice for either laminin β2 or the α3 chain of type IV collagen demonstrate the potential role of the GBM in albuminuria.7–10 In addition, the glomerular endothelial cell and GBM may be able to compensate for changes in the podocyte that lead to foot process fusion and loss of the heparin sulfate glycosaminoglycan and anionic charge.11 The recent discovery of a subpodocyte space has added another dimension to the GFB, although its role in protein filtration remains unknown.12,13
On this background, is it little wonder that a talented investigative team, with considerable experience in the area, used a new technological advancement in scanning electron microscopy in this issue of JASN to produce data that bring into question long-held beliefs regarding the structure and function of the podocyte slit diaphragm.14 They describe the existence of heterogeneous ellipsoidal and circular pores, located in the central region of the slit diaphragm, log normally distributed with a mean diameter far greater than envisioned previously. This was accomplished with careful attention to potential fixation, imaging, and quantitative analysis artifacts, although high-pressure freezing would have been the preferred tissue preparation.
Like many scientific advances challenging previously held beliefs, it is likely to be initially refuted vigorously, which is a healthy and required part of the scientific evaluation process. Pore sizes were quantified and compared between Munich Wistar Fromter (MWF) rats, known to develop albuminuria with age and develop focal glomerular sclerosis, and Wistar rats that do not develop albuminuria. The mean pore sizes of both strains were similar, but MWF rats had a small increase in the very largest pores, which the authors propose to be the mechanism of albuminuria. However, in both rat strains, unique images with high resolution and state-of-the-art quantitative morphometric analysis revealed pore sizes that beg the question, “Is albumin filtered across the glomerulus under normal physiologic conditions in levels previously deemed unrealistic by many investigators in the field?”15 This is especially true given the accumulating evidence from a number of studies using genetic, biochemical, imaging, and molecular approaches and indicating an important role for proximal tubule epithelial cells in albumin reabsorption and reclamation.16–24
The present data could easily be interpreted to substantiate further the role of the proximal tubule under physiologic and pathologic conditions in minimizing albuminuria. The authors invoke endothelial and GBM barriers in series with the podocyte pores to explain the lack of albuminuria under physiologic conditions. However, why do these same glomerular proximal barriers not prevent albuminuria in the MWF rats? This conclusion seems inconsistent with the data. How can they have it both ways?
Questions will also arise regarding the rat strains used for comparisons. Is it appropriate to compare very old MWF rats that have reduced GFR with Wistar rats (age unknown) that have normal serum creatinine? Would more appropriate, insightful, and confirmatory data have been obtained from young MWF rats without proteinuria? This comparison would have allowed the investigators to evaluate pore numbers and size as albuminuria progressed with age and before foot process fusion and decreases in GFR occurred. Do the large pores increase in size and/or number with increasing albuminuria?
In summary, previously held concepts in the complex process of glomerular albumin filtration are being challenged both across the glomerulus and downstream of the glomerulus on the basis of biochemical, molecular, genetic, and methodologic advances. Each of these challenges is met by skepticism, often severe in nature. These investigators are to be congratulated for pushing science forward because far too many reviews have indicated why the past must be correct. Facts are more important than faith when discrepancy exists, and they serve to move the field forward. Undoubtedly, this study will lead to additional studies and the area will be further refined. Discovery is advanced by technological advances and thinking outside the existing box.25
I thank Simon Atkinson, PhD, and Vince Gattone, PhD, for thoughtful insights.
Published online ahead of print. Publication date available at www.jasn.org.
See related article, “Imaging of the Porous Ultrastructure of the Glomerular Epithelial Filtration Slit,” on pages 2081–2089.
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