The complex pathology of diabetic nephropathy in humans

Abstract

This review summarizes the pathomorphological sequences of nephron loss in human diabetic nephropathy (DN). The relevant changes may be derived from two major derangements. First, a failure in the turnover of the glomerular basement membrane (GBM) based on an increased production of GBM components by podocytes and endothelial cells leading to the thickening of the GBM and accumulation of worn-out GBM in the mesangium. This failure may account for the direct pathway to glomerular compaction and sclerosis based on the continuous deposition of undegraded GBM material in the mesangium. Second, an increased leakiness together with an increased propensity of glomerular capillaries to proliferate leads to widespread plasma exudations. Detrimental are those that produce giant insudative spaces within Bowman's capsule, spreading around the entire glomerular circumference and along the glomerulo-tubular junction onto the tubule resulting in tubular obstruction and retroactively to glomerulosclerosis. Tubular atrophy and interstitial fibrosis develop secondarily by transfer of the glomerular damage onto the tubule. Interstitial fibrosis is locally initiated and apparently stimulated by degenerating tubular epithelia. This leads to a focal distribution of interstitial fibrosis and tubular atrophy accompanied by a varying interstitial mononuclear cell infiltration. Spreading of fibrotic areas between intact nephrons, much less to the glomerulus, has not been encountered.

Keywords: diabetic nephropathy; insudative lesions; mesangial matrix expansion; role of endothelial cells; role of podocytes; thickening of the GBM.

Figure 1:

Matrix accumulation in the mesangium. (A–C) Transmission electron micrographs showing the incorporation of GBM portions into the mesangium. (A) The capillary loop is surrounded by a thickened basement membrane (GBM) that forms an infolding with a cleft (asterisk) that narrows toward the bend of the GBM, which merges (joining arrow) with a portion of matrix within the mesangium. A similar situation is seen on the opposite side also indicated by a joining arrow. Note the nucleus (dot) of a mesangial cell. (A1) An enlarged picture of an infolding of the thickened GBM with a narrow cleft. The cytoplasmic extensions of a podocyte are cramped, shedding cytoplasmic material into the cleft (arrows). At the innermost part of the infolding (asterisk) shed podocyte material becomes included into GBM material. (B) Mesangial area filled with matrix that contains randomly distributed cytoplasmic remnants of podocytes identifying the matrix as worn-out undegraded GBM material. (C) A sclerotic nodule bounded by the paramesangial GBM (highlighted in yellow) covered with structurally intact podocytes (arrows) and perfused capillaries (dots). In its peripheral parts it contains mesangial cells typically with many processes, but signs of any synthesizing activity are lacking. Throughout the nodule cytoplasmic remnants of podocytes (asterisks) are encountered, sparse in the very center, dense towards the periphery, allowing the conclusion that the accumulated matrix represents undegraded GBM-material. Biopsies DN. Scale bars: (A) 2.5 µm, (A1) 1 µm, (B) 0.5 µm, (C) 5 µm. (D–F) Schematics showing the sequence of GBM engulfment. The GBM is shown in yellow, a podocyte in pink, a mesangial cell in green and capillaries in red. (D) Normal situation. (E) Thickening of GBM, retraction of podocytes out of the constricted GBM infolding, leaving behind shed cytoplasmic material. (F) Dropping of the innermost portions of the GBM into the mesangium accumulating as mesangial matrix. The inclusions of shed cytoplasmic material from podocytes identify the matrix as undegraded GBM-material. (A–F) Reproduced from [1] with permission.

Figure 2:

Immunofluorescence (IF) data identifying the origin of the accumulated matrix in the mesangium. (A) IF double staining against the α5 chain of collagen IV (green) and synaptopodin (red). Podocytes are cramped in narrow clefts of the GBM infoldings (arrowheads). Displacements of podocyte particles stained in red (arrows) into the mesangium surrounded by diffusely green staining GBM material (α5 chain). (B) IF double staining against the α1 chain of collagen IV (green) and synaptopodin (red). Note the cramped podocytes within the GBM clefts (arrowheads), and the intense staining of the GBM (arrows) and of the mesangial matrix (asterisks). (C) In situ hybridization (ISH) of the α1 chain of collagen IV (red) combined with IF localization of CD31 (green) clearly showing that the mRNA signals (red) are found only in endothelial cells (arrows). (D and D1) IF staining for perlecan. (D) Control, virtually no staining for perlecan in the heathy glomerulus. (D1) Biopsy DN. The GBM is stained throughout the tuft and a faint staining is seen in mesangial areas. Biopsies DN. Bars: (A) 5 µm, (B) 10 µm, (C) 20 µm, (D and D1) 40 µm. (A, C and D) Reproduced from [1] with permission; (B) original. Matrix propagation changes in overall tuft structure. (E) Overview of a glomerular profile with herniation of the tuft to the outside. The GBM and the parietal basement membrane PBM are highlighted in yellow. Note that the transitions between both (arrowheads) are displaced sideways showing that the upper portion of the tuft (star) has come to lie outside of the borders of the glomerulus. This herniated area is devoid of podocytes. It contains the efferent arteriole (dot) and several outgrowing vessels (arrows), some of them with hyaline cuffs. Apart from macula densa (MD) an assembly of structures that may be called JGA cannot be delineated. The rest of the tuft shows segmental sclerosis. (F) Glomerular profile with an extremely widened glomerular entrance delimited by the transition of the GBM into the PBM (marked by arrowheads). Thus, the upper portion of the tuft is herniated to the outside with outgrowth of a bunch of vessels (arrows; all of them with hyaline cuffs) occupying the area of the former JGA. Some of the vessels penetrate in between adjacent tubules. The rest of the tuft shows diffuse to segmental sclerosis. (G) The compacted tuft is largely filled with matrix (asterisks). Podocytes have retracted out of the tuft gathering on its surface (arrows). They are heavily distorted (LC3 and beclin-1 positive; not shown). The vascular pole is marked by a star. (H) In addition to solid matrix, the global sclerotic tuft contains several hyalinotic areas (arrows). Podocytes and parietal epithelial cells have disappeared, Bowman's space is filled with a fibrous matrix (asterisks) thus damage progresses to global glomeruosclerosis. Biopsies DN. All images are light micrographs. (E, G and H) Stained with methylene blue, (F) with PAS. Bars: (E, G and H) 20 µm, (F) 30 µm. (E and F) Reproduced from [2] with permission; (G) reproduced from [3] with permission; (H) original.

Figure 3:

Formation of tuft adhesions and spreading of insudative changes. (A–D) Schematics. (A) Normal situation. Podocytes (shown in blue) are sitting on the GBM (shown in violet) that covers three capillaries (shown in red). Parietal epithelial cells (PECs) are shown in green. The PBM is shown in brown. (B) Local expansion of the tuft leads to contacts of podocytes to PECs. (C) The junctional contact of podocytes to PECs results in a gap in both epithelial layers followed by a gap in the GBM. Podocytes extend on the PBM. A glomerular capillary deprived of the GBM delivers an exudate into the space between the parietal epithelium and its basement membrane (PBM). PECs produce a new basement membrane (dashed in pale brown). (D) The exudate spreads within BC circumferentially around the glomerulus and, via the glomerulo-tubular junction, onto the tubule separating the tubular epithelium from its basement membrane (TBM). (E–H) Selected originals. (E) Due to matrix accumulations displaced perfused capillaries (asterisks) lead to contact sites of podocytes with the parietal epithelium (arrows). (F) CD31-stained glomerular capillaries that have penetrated into BC and have come to lie inside the PBM. (G) Early adhesion with two capillaries (arrows) penetrating into BC became to lie inside the PBM. Between the PBM and the parietal epithelium (that has produced a thin new basement membrane) a fluid-filled space has developed (asterisk). (H) Tuft adhesion (star) with outgrowth of a glomerular capillary (arrow) into a fluid filled space (insudative space) within BC that is bordered inside by the parietal epithelium (arrowhead), outside by the PBM and, in continuation, by the TBM. The beginning portion of the tubule (T) is atrophied, deprived of its basement membrane and included into the fluid-filled space (asterisk), as well as a further capillary (dot). Biopsies DN. (E–H) LMs, (E) stained with methylene blue, (F) immune-stained with CD31, (G and H) stained with PAS. Bars: (E and H) 10 µm, (F and G) 20 µm. (A–D and F–H) Reproduced from [3] with permission; (E) original.

Figure 4:

Tubulo-interstitial changes. (A) Tubulo-interstitial changes develop exclusively by transfer from the glomerulus. Shown here is the encroachment of the insudative changes in BC onto the tubule. Within BC the parietal epithelium (arrowhead) has been separated from its basement membrane (PBM) by plasma exudation from outgrown capillaries. The resulting insudative spaces within BC spread around the entire glomerular circumference. Via the glomerulo-tubular junction they extend (arrow) onto the tubule separating the tubular epithelium from its basement membrane (TBM); the tubular epithelium undergoes degeneration (dots) leading to obstruction. This process extends to subsequent tubular coils. Finally, wrinkled remnants of the TBM surrounded by proliferating interstitial tissue (circle) are left. Note, the starting point of this development, the original adhesion of the tuft to BC (star) is still seen. (B) Tubulointerstitial area with intermingled damaged (asterisks) and intact tubules (open circles). No tubular profile can be seen that displays intact and injured portions. No signs of a damage encroachment from damaged to intact tubules is encountered. The degenerating group of tubules in the upper right corner is surrounded by proliferating interstitial tissue. (C) Profile of a proximal tubule with a structurally healthy epithelium. This intact tubule is fully surrounded by fibrotic tissue with degenerated tubular profiles (_). Proximal tubule with a structurally healthy epithelium. This intact tubule is embedded in fibrotic tissue (asterisks) that contains also degenerated tubular profiles (arrows). Biopsies DN. (A and B) LMs stained with PAS, (C) is a transmission electron micrograph. Bars: (A and C) 20 µm, (B) 50 µm. (A–C) Reproduced from [3] with permission.