Characterization and differentiation of protein reabsorption granules and punctate IgG in primary podocytopathies

Abstract

Nephrotic syndrome (NS) is a set of symptoms defined by heavy proteinuria and associated with a host of kidney diseases that cause injury to the glomerulus, the filtration apparatus of the kidney. Primary podocytopathies (Px), a group of diseases including minimal change disease (MCD), primary focal segmental sclerosis (pFSGS), and lupus podocytopathy (LP), are the principal cause of idiopathic NS in both children and adults. The hallmark feature of Px is the ultrastructural finding of podocyte foot process effacement (FPE), so current differential diagnosis of Px relies on technically exhaustive electron microscopy (EM) analysis. During routine immunofluorescence (IF) microscopy of many Px cases, we have observed punctate IgG (P-IgG) immunoreactivity in the glomerulus. P-IgG may represent a disease-specific reactivity that could not only provide clues for understanding Px etiology but could also serve as a diagnostic tool. However, we have found that P-IgG may be misinterpreted as protein reabsorption granules (PRGs), a morphological feature seen in proteinuric conditions. We sought to definitively characterize the key characteristics of PRGs and P-IgG in order to differentiate these features. To accomplish this, we reviewed prior IF immunostaining of MCD biopsies to evaluate anti-human IgG and anti-human albumin staining. We conducted additional IF staining on archived MCD biopsies using antibodies against IgG and against the markers of PRGs: albumin, megalin, and cubilin. We found that the P-IgG demonstrates a diffuse, global distribution pattern that is specific to glomerular epithelium and is fine and scattered. Conversely, the PRGs are coarse, clustered, and frequently demonstrate a focal, segmental pattern in the glomeruli and tubules. Co-staining with albumin and megalin revealed that the P-IgG and the PRGs do not colocalize in the tissue. While the cubilin antibody positively stained the tubular epithelium, it did not stain glomeruli. Our results showed that punctate IgG and protein reabsorption granules are morphologically and constitutionally distinct and do not colocalize with each other, indicating that P-IgG is highly likely to represent a distinct process from epithelial protein reabsorption.