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Structural analysis of how podocytes detach from the glomerular basement membrane under hypertrophic stress - PubMed

  • ️Wed Jan 01 2014

Structural analysis of how podocytes detach from the glomerular basement membrane under hypertrophic stress

Wilhelm Kriz et al. Front Endocrinol (Lausanne). 2014.

Abstract

Podocytes are lost by detachment from the GBM as viable cells; details are largely unknown. We studied this process in the rat after growth stimulation with FGF-2. Endothelial and mesangial cells responded by hyperplasia, podocytes underwent hypertrophy, but, in the long run, developed various changes that could either be interpreted showing progressing stages in detachment from the GBM or stages leading to a tighter attachment by foot process effacement (FPE). This occurred in microdomains within the same podocyte; thus, features of detachment and of reinforced attachment may simultaneously be found in the same podocyte. (1) Initially, hypertrophied podocytes underwent cell body attenuation and formed large pseudocysts, i.e., expansions of the subpodocyte space. (2) Podocytes entered the process of FPE starting with the retraction of foot processes (FPs) and the replacement of the slit diaphragm by occluding junctions, thereby sealing the filtration slits. Successful completion of this process led to broad attachments of podocyte cell bodies to the GBM. (3) Failure of sealing the slits led to gaps of varying width between retracting FPs facilitating the outflow of the filtrate from the GBM. (4) Since those gaps are frequently overarched by broadened primary processes, the drainage of the filtrate into the Bowman's space may be hindered leading to the formation of small pseudocysts associated with bare areas of GBM. (5) The merging of pseudocysts created a system of communicating chambers through which the filtrate has to pass to reach Bowman's space. Multiple flow resistances in series likely generated an expansile force on podocytes contributing to detachment. (6) Such a situation appears to proceed to complete disconnection generally of a group of podocytes owing to the junctional connections between them. (7) Since such groups of detaching podocytes generally make contact to parietal cells, they start the formation of tuft adhesions to Bowman's capsule.

Keywords: crescents; foot process effacement; podocyte detachment; podocyte hypertrophy; pseudocysts; sealing of filtration slits.

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Figures

Figure 1
Figure 1

Overview and early changes. (A) Light micrograph (LM) of a glomerular profile showing the crucial injuries presented later in more detail in transmission electron micrographs (TEMs) and quantified in Table 1. Hypertrophied podocytes forming large pseudocysts (asterisks), areas of small pseudocysts generally associated with local detachments (enclosed by a hatched circle), detachment of podocytes forming a cluster in Bowman’s space (star), and a large tuft adhesion comprising almost two-thirds of the tuft (clockwise from arrow 1 to arrow 2) are seen. Note that outside the tuft adhesion, the parietal epithelium looks fully normal (two small arrows). (B) The GBM is highlighted in yellow, capillary lumens in green. The cell bodies of podocytes are attenuated (arrows) and pseudocysts (asterisks) have developed underneath them. Note that the foot process pattern is widely preserved including the floors of pseudocysts and, second, the parietal epithelium looks fully normal. Male rats, growth stimulation with FGF-2 for 13 weeks. (A) LM from 1 μm Epon section. Bar: 15 μm. (B) TEM. Bar: 5 μm.

Figure 2
Figure 2

Sealing of the slits may proceed to complete foot process effacement (FPE). The GBM is highlighted in yellow, capillary lumens in green. (A) Early stage of FPE with broadening of FPs and sealing of the slits by occluding junctions (some are labeled by arrows). In the right upper corner part of a capillary, profile with completed FPE (arrowhead) is seen. (B) “Foot processes” have changed into broad plaques (1,2,3,4) that closely stick to the GBM. The sealing of the filtration slits by occluding junctions between them is maintained (arrows). (C) Capillary profile covered by a podocyte that is closely attached to the GBM by “effaced” basal cell portions stuffed with a prominent cytoplasmic mat (arrowheads). At the opposite side, broadened FPs (arrows) are undergoing retraction, whereas at the transition to the mesangial region, FPs appear intact (two small arrows). Pseudocyst formation is also seen (asterisks). (A) and (B) male, (C) female rats, growth stimulation with FGF-2 for 13 weeks. TEMs. Bars: (A) 1 μm, (B) 0.5 μm, (C) 2 μm.

Figure 3
Figure 3

Sequence of podocyte disconnection from the GBM. The GBM is highlighted in yellow, capillary lumens in green, and spaces considered as pseodocysts connected to bare GBM in yellow-orange. Individual podocytes are highlighted in different shades of violet. (A) Early stage of podocyte detachment from the GBM. A sealing of filtration slits by occluding junctions replacing slit membranes was only partially successful (arrowheads). At many other sites, retraction of FPs has left behind circumscribed areas of bare GBM (arrows) that start to merge (red dots). (B) Advanced stage of podocyte detachment from the GBM. The dome-shaped spaces (asterisks) above bare areas of GBM have enlarged bulging toward the urinary space. Merging of these spaces leads to a communicating system of pseudocysts (arrows). (C) The pseudocysts (asterisks) increase in size by merging and bulging; they are related to large stretches of bare GBM and overarched by broadened and attenuated primary processes. (D) The final detachment of a podocyte is frequently characterized by the merging of innumerous interconnected (red arrows) pseudocysts that start over bare GBM (red dots) and drain into Bowman’s space (not seen in this section). Other portions of this podocyte may closely stick to the GBM with completed FPE (black arrows). Male rats, growth stimulation with FGF-2 for 13 weeks. TEMs. Bars: (A) 1 μm, (B) 2 μm, (C) 1 μm, (D) 2.5 μm.

Figure 4
Figure 4

Clusters of detaching podocytes in Bowman’s space. Capillary lumens are highlighted in green, the GBM in yellow. In B and C cell nuclei of podocytes are shown in violet, the PBM in light brown. (A) A group of 6 podocytes in the process of detachment. Only podocytes 1 to 4 still have contact with the GBM, whereas 5 and 6 are attached to the podocytes beneath by junctions (red circles). Within this cluster of podocytes, a communicating system of pseudocysts is seen that starts at bare areas of GBM (red dots). It may readily be suggested that the filtrate entering through bare GBM has to pass these spaces to reach Bowman’s space; openings into Bowman’s space are not hit in this section. Note that additional detaching podocytes (colored in various shades of violet) are gathered in another cluster. (B) Cluster of some 10 detaching or detached podocytes forming a cell bridge between the tuft and Bowman’s capsule. The unlabeled cell within the parietal epithelium cannot be assigned by any plausible criterion to either parietal cells or to podocytes. The innermost cells are still partially attached to the GBM (arrows). The cells are interconnected to each other and still contain abundant pseudocysts that form a continuous chamber system starting from bare GBM (red dots) extending to the outermost layer. Note that the walls of many pseudocysts are undergoing shedding (asterisks). (C) Podocytes after almost complete detachment from the GBM are assembled as a cluster in Bowman’s space interconnected to each other by cell contacts (red circles). The inner row of podocytes shows assemblies of small pseudocysts, the outer row appears as rounded cells. The outermost cells have contacts to Bowman’s capsule (arrows). Male rats, growth stimulation with FGF-2 for 13 weeks. TEMs. Bars: (A) 5 μm, (B) 4 μm, (C) 2 μm.

Figure 5
Figure 5

Clustering of podocytes in Bowman’s space is associated with shrinkage of the mesangio-capillary area. The GBM is highlighted in yellow, capillary lumens in green, the PBM and cell nuclei of parietal cells in brown, podocyte cell nuclei in violet. Glomerular profile with a tuft that is broadly connected to Bowman’s capsule by a large cellular crescent that surrounds the tuft from three sides. The mesangio-capillary area (delimited by the GBM) is small compared to the area occupied by podocytes. Note that podocyte cell bodies (visualized by their cell nuclei) have almost completely disappeared from central tuft areas being all contained within the crescent. Most of the podocytes have retained pseudocysts. The cluster of podocytes on top of the tuft does not show any contacts to the tuft nor to the PBM, whereas the major part of the crescent displays extensive contacts to the GBM as well as to Bowman’s capsule. Nuclei of cells, which by no plausible criterion can be assigned as podocytes or parietal cells are uncolored. Note that the GBM is heavily wrinkled at several sites and the mesangial area appears to be collapsed with disappearance of capillaries at several loci (asterisks). Male rat, growth stimulation with FGF-2 for 13 weeks. TEM. Bar: 10 μm.

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