Mechanism of cystogenesis in nephrotic kidneys: a histopathological study

The nephrotic syndrome (NS) is pathological condition characterized by several components, including proteinuria. In healthy kidneys, proteins are mainly reabsorbed in proximal tubules by receptor-mediated endocytosis [1]. In NS, the most often cause of renal failure is focal segmental glomerulosclerosis (FSGS), which results from interplay of genetic and external factors. Histologically, FSGS is characterized by podocyte depletion, segmental glomerular scaring and glomerular epithelial proliferation [2]. Another type of NS is congenital nephrotic syndrome of the Finnish type (CNF), a glomerular disease caused by mutation of NPHS1 gene that encodes protein nephrin, localized on the slit diaphragm [3]. In CNF, patients show massive proteinuria already in utero[4]. Although histological lesions in CNF are described as un-specific, dilations of proximal tubules with microcyst formation are most often found [5]. Other hereditary conditions are also associated with renal cysts, including ciliopathies - a group of diseases caused by genetic mutations of proteins residing on the primary cilia [6], a specialized cell surface organelle which coordinates cell proliferation, differentiation and apoptosis [7‐9]. Ciliary microtubular organization enables intraflagelar transport (IFT) and transfer of information both in and out of the cell. Faulty cilia signaling in some cases leads to cysts formation [10]. By now, little attention has been paid to primary cilia in CNF, as mutated nephrin has not been associated to cilia function.

During normal kidney development, morphogenesis and function of primary cilia correlated with proper tubular cells proliferation and apoptosis [11], and with periodical appearance of primary cilia on the surfaces of tubular cells between cell cycles [12]. While during post-natal life primary cilia remained on the surfaces of tubular cells [12], they disappeared in rat podocytes already during development [13]. By now, investigations on the cystogenesis in CNF kidneys have not been connected neither to abnormal cell proliferation and apoptosis [4], nor to the primary cilia abnormalities but to the cell dedifferentiation [4].On the other hand, enhanced cell turnover was found in multicystic dysplastic kidneys [14].

In the present study we analyzed the ultrastructural and immunohistochemical characteristics of CNF tubular cells during cystogenesis, and compared them to tubular cells in FSGS and in normal kidneys. We present a novel data on tubular cell pathology in NS kidneys and suggest that the disturbed proliferation and apoptosis, and associated changes of primary cilia structure and function might be involved in cystogenesis of CNF and FSGS kidneys, and possibly influence clinical presentation of NS.

Our ultrastructural and immunohistochemical analysis of CNF and FSGS kidneys disclosed that cystogenesis in proximal tubules was associated with increased cell proliferation, apoptosis and changes of primary cilia on the surfaces of tubular cells. While in the largest cysts of CNF kidneys the primary cilia were completely missing or were short and distorted, in moderately dilated or apparently normal tubules they were 3–8 fold longer than in healthy kidneys. Increased proliferation found in CNF cysts coincided with decreased number of primary cilia, while the increased diameter of proximal tubules or tubular cysts was inversely proportional to proliferation and was accompanied by the reduced height of tubular cells. The same type of changes in tubular cells characterized FSGS nephrotic kidneys during cystogenesis, as well. However, we believe that differences that we found in the course of proliferation between CNF and FSGS kidneys might be attributed to the prenatal appearance of pathological changes in CNF versus their postnatal appearance in FSGS. Both in CNF and FSGS kidneys, deregulations of cell turnover were accompanied by apoptosis of tubular and mesenchymal cells, as also described in human kidney malformations associated with urinary tract obstruction [14].

Experimental studies on kidney primary cilia confirmed association of primary cilia dysfunction and cystogenesis [8]. Thus, deleting of cilia assembly gene IFT20 prevented cilia formation and promoted rapid postnatal cystogenesis [17], while disturbed IFT resulted in a variety of disorders, including polycystic kidney disease [18]. In some cases, disturbed Wnt signaling, which mediates planar cell polarity (PCP), caused renal cystogenesis [19]. Abnormally short or extensively long cilia were found in human juvenile cystic kidney disease [20] and meckel syndrome, as well [21]. Investigations of ciliogenesis during normal human kidney development, described association of primary cilia lengthening with differentiation of tubular cells, apico-basal cell polarity and proper lumen formation. [12]. Similar to described findings in CNF and FSGS kidneys, increased cilium lengthening following ischemia-reperfusion injury characterized distal/collecting tubules segments [22]. Such downstream changes of primary cilia along the affected nephron might represent a compensatory process associated with loss of cilia in the cysts of proximal tubules. We suggest that described alteration of primary cilia number, structure or orientation might diminish the overall quality and quantity of tubular cells signaling, leading to compensatory growth of cilia in distal/collecting tubules segments in effort to increase signaling and preserve function of the damaged nephron. In Ofd1 syndrome, initially normally formed primary cilia disappeared during cystogenesis, suggesting secondary nature of cilia changes [23]. Recent studies on kidney cystogenesis pointed to significant influence of the extracellular milieu on modulation of cilia signaling, which led to deregulation of cell proliferation and cell differentiation [10]. We speculate that the described apoptosis and malfunction of proximal tubular cells during cytogenesis might cause significant deterioration of protein re-absorption in the affected kidneys. Consequently, the urine of CNF and FSGS kidneys becomes overloaded with proteins and therefore milieu for primary cilia signaling becomes further deteriorated.