Glomerular Diseases With Organized Deposits

Samy S. Iskandar

Guillermo A. Herrera

INTRODUCTION AND HISTORICAL BACKGROUND

The term glomerular diseases with organized deposits has come into use in the medical literature relatively recently with the gradual increase in number of glomerular diseases defined by recognition at the ultrastructural level of deposits with distinguishing organized substructures.

In the 1970s, consultation of a recently published textbook of renal pathology (1) would have produced only two such entities: amyloidosis and cryoglobulinemia. With the reporting of cases that were initially described as showing Congo red—negative amyloid-like glomerular deposits (2) and later named fibrillary deposits (3), the entity recognized by most renal pathologists as fibrillary glomerulonephritis (FGN) (4) came into existence. About the same time, the much rarer entity of immunotactoid glomerulopathy (ITG) was described (5). This was followed by the description of hereditary collagenofibrotic glomerulopathy (6) and fibronectin glomerulopathy (7). With the increase of these entities arose the need to have an all-encompassing term—hence, the emergence of the term glomerular diseases with organized deposits. The term was first used in Heptinstall’s Pathology of the Kidney in its fifth edition, although the expression “with organized deposits” had been introduced in the renal pathology literature as early as 1967. This chapter focuses primarily on FGN and immunotactoid
glomerulopathy and also discusses other glomerular diseases with organized deposits that are covered in more detail in other chapters (e.g., cryoglobulinemic glomerulonephritis and collagenofibrotic glomerulopathy). In addition, normal extracellular materials that can be confused with organized deposits (e.g., fibrillary collagen and fibrin tactoids) are described. The organized deposits discussed in this chapter are the ones that are seen most often in renal biopsy specimens (8,9,10); however, there are anecdotal reports of organized deposits that do not fit well into these recognized categories (11).

As is the case with most renal biopsies, achieving the correct diagnosis is critical for management and prognostic purposes, but this is particularly true in the case of the topic under discussion where the difference, for example, between AL amyloidosis and FGN means the difference between a condition that can potentially be treated by appropriate chemotherapy or, if necessary, bone marrow ablation and autologous stem cell transplantation and another that currently can only be treated for the most part by measures directed toward control of proteinuria and hypertension.

Because many of the entities listed above are covered more extensively in other chapters of this text, this chapter focuses on FGN, immunotactoid glomerulopathy, fibronectin glomerulopathy, and the accentuation of the mesangial matrix and deposition of precollagen in some sclerosing conditions that enter into the differential diagnosis of in particular FGN. Glomerular diseases with organized deposits can be categorized in a number of ways.

Although for classification purposes it is useful to divide the entities that are the subject of this chapter into those that are Congo red reactive and those that are not (Table 23.1), from the practical point of view, this approach would be helpful only if the Congo red stain was included in the battery of special stains used on a routine basis to evaluate every renal biopsy just like the periodic Schiff (PAS) and the periodic methenamine silver (PAMS/Jones) stains are. However, if that is not the case, then an alternative diagnostic algorithm is needed that is based on the stains that are used by the majority of renal pathologists. That is why we find the algorithm based on silver methenamine staining outlined in Figure 23.1 helpful. An algorithmic approach is a useful first step in making a pathologic diagnosis, but the final diagnosis depends on considering multiple pathologic features that rule in or rule out a specific diagnosis (Table 23.2).

FIBRILLARY GLOMERULONEPHRITIS

Clinical Presentation and Epidemiology

The first case of FGN was described in 1977 (2) in a patient who presented with the nephrotic syndrome and whose biopsy demonstrated immunoglobulin and complement deposits on immunofluorescence microscopy examination and on ultrastructural examination revealed the presence of fibrillary deposits resembling amyloid fibrils in a background of amorphous granular deposits consistent with immune complex-type deposits. All histochemical stains for amyloid were negative. This case was soon followed by a succession of publications that established this as a new diagnostic entity (reviewed in Ref. (12)). Most significant was the small series published by Duffy et al. (3), which used the title “fibrillary renal deposits and nephritis” and established the clinical presentation of the entity, as well as emphasized the need for electron microscopy to avoid missing this diagnosis. Credit for coining the term fibrillary glomerulonephritis goes to Alpers et al. (4). Multiple series have since been published that have further defined the clinical presentation, pathologic features, and prognosis of FGN, and the majority have in particular emphasized the distinction between FGN and the much rarer entity widely known as immunotactoid glomerulopathy (ITGP), although this latter issue was the subject of a heated debate for a number of years (12,13,14,15,16,17,18,19).

TABLE 23.1 Congo red reactivity of glomerular deposits with ultrastructural organized deposits

Congo red-positive
Amyloidosis composed of >30 precursor proteins among which are the following:
	Immunoglobulin light chain (usually λ light chain but can be κ) (AL amyloid)
	Immunoglobulin heavy chain (AH amyloid)
	Immunoglobulin light and heavy chain (ALH amyloid)
	Amyloid A protein (AA amyloid)
	Apolipoprotein AI and AII
	Apolipoprotein E
	Transthyretin
	Leukocyte chemotactic factor 2 amyloid
	Fibrinogen Aα
	Lysozyme
	Gelsolin
	Cystatin
	β-Microglobulin (no renal involvement but complicates chronic hemodialysis)
Congo red-negative
Cryoglobulinemic glomerulopathy
Fibrillary glomerulonephritis
Immunotactoid glomerulopathy
Fibronectin glomerulopathy
Collagenofibrotic glomerulopathy
Diabetic fibrillosis and other sclerosis with accentuated collagenous fibrils
Fibrin
Glomerular deposits with ultrastructural organized deposits can be divided into Congo red-positive deposits of amyloid of varying composition versus Congo red-negative deposits of other substances.

FGN is found in approximately 1% of native renal biopsies accessioned in most renal biopsy services (12,18). The clinical presentation is not specific. Patients are overwhelmingly Caucasians, even in series where this did not reflect the racial distribution of the overall biopsy population (12,20) with a slight female predominance. Two large series (18,19) give age means and ranges remarkably similar, approximately 57 [28 to 81] years and 53 [19 to 81] years, respectively; however, a case has been described in a 9-year-old girl (21), and we have seen a case in a 14-year-old boy. All patients have proteinuria at the time of biopsy with proteinuria often reaching the nephrotic level and frequently associated with the full nephrotic syndrome. Microscopic hematuria is present in approximately 50% to 60% of cases, and rare cases have episodes of gross hematuria (18,19). Renal insufficiency is
present in 50% of patients at the time of biopsy (16), and 65% are hypertensive (16). In the two most recent and largest series (18,19), associated medical conditions included, respectively, diabetes mellitus (20%, 20%), lymphoproliferative disease (2%, 9%), carcinomas (5%, 14%), and systemic lupus erythematosus (1.6%, 3%). In the series from the Mayo Clinic (19), an additional 15% had miscellaneous autoimmune diseases. Positive serologic tests in the same two series included hepatitis C (17%, 3%), monoclonal immunoglobulin by serum or urine protein electrophoresis (15%, 17%), antinuclear antibodies (16%, 14%), low complement level (2%, 2%), and cryoglobulins (4%, 3%). FGN is a disease confined to the kidney. There are individual case reports indicating involvement of the liver, lungs, and heart (22,23,24); however, careful examination of the published illustrations suggests to us as well as to others (25) that interpretation of these reports warrants caution. An autopsy study appears to have demonstrated the presence of the characteristic fibrils of FGN only in the spleen in addition to the kidney and confirmed that localization by the immunogold labeling method (26).

FIGURE 23.1 Diagnostic algorithm for glomerular mesangial extracellular matrix expansion based on silver methenamine staining. Many but not all processes with organized deposits are included. Rare entities such as fibronectin glomerulopathy and metabolic storage diseases are not included. MIDD, monoclonal immunoglobulin (Ig) deposition disease, including light chain (LC) and heavy chain (HC) deposition disease; GN,glomerulonephritis; IF, immunofluorescence microscopy; IHC, immunohistochemistry; MS, mass spectroscopy.

Pathologic Findings

Gross Pathology

Little is known about the gross pathology of FGN since patients reaching end-stage renal disease (ESRD) are kept alive on dialysis or receive renal transplants such that autopsy experience with the entity is limited. An occasional autopsy case on a patient who died of myocardial infarction close to 5 years after the diagnostic biopsy described atrophic right and left kidneys weighing 100 and 90 g, respectively, with coarse granular external surfaces (26).

Light Microscopy

The common denominator among all the studies including any significant number of cases (3,4,12,13,18,19) is the expansion of the mesangium by eosinophilic, PAS-reactive material (Fig. 23.2) with the exception of one study (18) where the material was described as weakly eosinophilic and weakly PAS reactive. With a Masson trichrome stain, the mesangium stains blue or green, depending on whether one uses an aniline blue- or light green-based stain, with a variable red fuchsinophilic tinge depending on the mixture of the collagenous mesangial matrix and deposits of pathologic material, respectively (Fig. 23.3). The periodic acid methenamine silver stain (PAMS or Jones stain) shows failure of the mesangium to stain uniformly with silver (Fig. 23.4). This should raise concerns about the potential diagnosis of amyloidosis; however, all histochemical stains for amyloid, specifically the most commonly used ones, namely, Congo red and thioflavin T, are negative. An important caveat is that in very rare instances, FGN is Thioflavin T positive.
The above-described mesangial pattern comprised 71% of cases in a large series reported from the Mayo Clinic (19); however, other patterns described (18) include membranoproliferative-like, diffuse proliferative and exudative, segmental necrotizing and crescentic, membranous and diffuse sclerosing patterns. Crescents are reported to be present in between 17% and 31% of cases (12,18,19,20) excluding the category of segmental necrotizing and crescentic glomerulonephritis, which was defined by Nasr et al. (19) as presence of ≥50% glomeruli with segmental tuft necrosis or crescents.

TABLE 23.2 Distinguishing pathologic characteristics of different types of organized glomerular deposits

	Light microscopy	Immunofluorescence microscopy	Electron microscopy	Others
Amyloidosis	Faintly eosinophilic and PAS-negative material. Spicules best appreciated with silver stains with a porcupine (cock’s comb) appearance distinguishable from spikes of membranous glomerulonephritis	Staining for specific subtypes of amyloid. Routine stains may show increased nonspecific background staining.	Feltwork of 7- to 15-nm randomly disposed fibrils	Apple green birefringence with polarized light. Fluorescence with Thioflavin T and S and Congo red
Cryoglobulinemia	Hyaline subendothelial and luminal material strongly PAS positive. MPGN-like pattern. Abundant monocytes	Staining for contributing immunoglobulins and light chain over or around hyaline thrombi	Curved 20- to 30-nm cylindrical and annular structures in subendothelium and lumen. Sometimes deposits amorphous	Circulating cryoglobulins, hypocomplementemia, and rheumatoid factor
Fibrillary glomerulonephritis	Mesangial expansion by eosinophilic and PAS-positive material. Other features highly variable	IgG and C3. Usually both kappa and lambda	Feltwork of randomly disposed 15- to 30-nm fibrils sometimes in background of amorphous immune complex-type material	Absence of congophilic material
Immunotactoid GP	Mesangial and capillary expansion by eosinophilic and PAS-positive material. Often pattern of membranous glomerulonephritis with characteristic spikes. Other cases MPGN-like pattern	IgG, often light chain restricted	Bundles of intersecting 10- to 40-nm microtubules, occasionally up to 90 nm. May be in mesangial, subendothelial, or subepithelial	Paraprotein often identified. Frequent association with lymphoproliferative malignancies
Collagenofibrotic GP	MPGN-like pattern with PAS- and PAMS-negative infiltrate	Usual immunofluorescence panel noncontributory	Frayed collagen bundles with 43- to 65-nm periodicity. Worm and comma shapes where bundles cut transversely	Identification of collagen III by immunofluorescence or other immunohistochemical methods
Fibronectin GP	Accentuation of lobular architecture. Strongly PAS-positive but PAMS-negative material	Usual immunofluorescence panel noncontributory. Positive staining for fibronectin	Some kindreds with 14- to 16-nm fibrils. Other kindreds show only amorphous deposits	Staining for fibronectin by IHC
Collagen deposits in various GPs	Histologic features are those of underlying condition, often with sclerosis	Immunofluorescence pattern of the underlying condition	Collagen recognized by characteristic periodicity and precollagen recognizable by its usual association with mature collagen
Modified from Iskandar SS, Herrera GA. Glomerulopathies with organized deposits. Semin Diagn Pathol 2002;19(3):116-132.

Immunofluorescence Microscopy

The great majority of cases have deposition of polyclonal IgG (Fig. 23.5) along with C3 and κ and λ light chains in a mesangial pattern as well as along the capillary walls. The staining is typically confined to the glomeruli, but we have observed one specimen with staining of arterioles (Fig. 23.5A). Staining for IgA, IgM, and C1q is variable. A small minority of cases stain monotypically for κ light chain (4). The capillary wall deposits can mimic membranous glomerulopathy, in some cases. In rare cases, the capillary wall staining appears linear
and may simulate the appearance of anti-glomerular basement membrane autoantibody disease (27). The appearance of immunofluorescence in FGN is particular to the entity and allows one to anticipate the diagnosis in the great majority of cases even before electron microscopic examination, although of course by definition ultrastructural examination is required to render a confident diagnosis of FGN. Although the signal of staining for IgG is usually strong, it is neither granular nor linear and has been described as smudged (28) (Fig. 23.5). There is a clear predilection for staining with subclass IgG4 first demonstrated in 13 cases studied by Iskandar et al. (12) with only weaker staining in 15% of cases for subclass IgG1. This has generally been confirmed in a subsequent series (18). A single case of IgA-λ restriction has been reported (29). In two cases that appear to be bona fide FGN based on the published electron micrographs, attempts at immunofluorescence staining with antihuman immunoglobulin (Ig) antibodies from multiple commercial suppliers failed to demonstrate the presence of Ig deposits (30).

FIGURE 23.2 Fibrillary glomerulonephritis. A: Representative glomerulus from a case of FGN with extensive mesangial expansion. Note the infiltration of the mesangium by a homogeneous though slightly “moth-eaten” eosinophilic material. (H&E, 400×.) B: Case of FGN displaying a diffuse proliferative pattern and thickening of capillary walls. Note that the infiltrating material is PAS reactive. (PAS stain, 200×.)

FIGURE 23.3 Fibrillary glomerulonephritis. Same case as in Figure 23.2 using Gomori trichrome green stain for collagen, showing the increased extracellular matrices staining green with various degrees of segmental fuchsinophilic tinge. (Gomori trichrome, 200×.)

Electron Microscopy

FGN is largely defined by its ultrastructural appearance, summarized as infiltration of the mesangium and to a variable degree the glomerular basement membrane by randomly disposed fibrils that resemble amyloid fibrils but are about twice as thick and are less densely packed. Whereas the range of amyloid fibril thickness is 7 to 15 nm, that of fibrils in FGN is 15 to 30 nm with a mean of 22.2 nm and a standard
deviation of 7.4 nm (12) (Fig. 23.6). The fibrils are often set in a background of amorphous immune complex-type electron-dense deposits. Occasionally, the fibrillary deposits infiltrate the glomerular basement membrane and produce perpendicular spicular projections similar to those seen in amyloidosis. In rare cases corresponding to a membranous pattern by light microscopy, subepithelial deposits are present with little or no infiltration of the glomerular basement membrane seen in most cases (18,19). Tubular basement membrane deposits are very rarely present (18,19,20,31).

FIGURE 23.4 Fibrillary glomerulonephritis. Periodic acid-Schiff methenamine silver (PAMS/Jones) stain showing markedly reduced staining in the expanded mesangium and the thickened capillary walls. Staining is seen only in the residual GBM and remnants of fibrils of the mesangial matrix. (PAMS stain, 400×.)

FIGURE 23.5 Fibrillary glomerulonephritis. A, B: Glomeruli illustrate the characteristic strong smudgy mesangial and capillary wall staining characteristic of FGN. B: Rare smudgy staining of the hilar arteriole (arrow). (Direct immunofluorescence, fluorescein isothiocyanate-labeled rabbit anti-human IgG, 400×.)

Differential Diagnosis

Although theoretically all entities included in this chapter enter in the differential diagnosis of FGN, the only serious consideration at the ultrastructural level is amyloidosis. As indicated above, amyloidosis is discussed in detail in a separate chapter;
however, this is an appropriate place to mention that of the long list of amyloidogenic proteins provided in Table 23.1, the ones likely by far to involve the kidney are AL amyloidosis and AA amyloidosis although recently cases of leukocyte chemotactic factor 2 amyloid have been reported with increasing frequency (32) in particular in patients of Latino heritage. Leukocyte chemotactic factor 2 amyloid has a striking tendency to involve the interstitial compartment although the glomeruli and extraglomerular blood vessels are not spared. Most other forms of amyloidosis rarely involve the kidneys.

FIGURE 23.6 Fibrillary glomerulonephritis and amyloidosis. A: Electron micrograph of FGN deposit illustrating the randomly disposed nonbranching fibrils resembling amyloid fibrils but noticeably thicker. B: Electron micrograph of amyloidosis to contrast the thickness of the fibrils with those of fibrillary glomerulonephritis. (Uranyl acetate and lead citrate, 25,000×.)

Both amyloidosis and FGN do not pick up the silver component in the PAMS or Jones silver stain, and according to one group (18), the H&E stain and the PAS stain show weakly eosinophilic and equally weakly PAS-reactive mesangial material. This would make FGN indistinguishable from amyloidosis by light microscopy using routine stains. However, this has not been our experience or that of other investigators who have found the mesangium in FGN to be eosinophilic and PAS reactive (19). The immunofluorescence findings of amyloidosis depend on the type of amyloidogenic protein involved; although even in cases of light chain-related amyloidosis, the immunofluorescence findings are not always reliable and can even be misleading if not corroborated by additional laboratory results. Although it is reasonable to resort to immunohistochemistry to determine the type of amyloidosis, when in doubt, a reliable tool is mass spectrometry performed on laser-dissected tissue (33).

At the ultrastructural level, both FGN and amyloidosis are characterized by randomly disposed fibrils, which in the case of amyloidosis range from 7 to 15 nm in thickness, whereas in the case of FGN, the thickness of the fibrils is roughly double that ranges at 15 to 30 nm (Fig. 23.6), making the results of histochemical stains for amyloidosis such as Congo red, thioflavin T, and immunofluorescence microscopy for immunoglobulin and complement critical in distinguishing between the two entities when the fibrils’ thickness in a particular case is at the borderline between that of amyloidosis and FGN. Therefore, even if the block has been cut through and would not allow for additional sections to be obtained to be stained with Congo red, one of the sections already stained with H&E or PAS ought to be destained and restained with Congo red to ascertain the diagnosis, since one defining feature of FGN is its Congo red negativity despite the ultrastructural resemblance to amyloidosis.

Another pitfall in the diagnosis of FGN is encountered when, often in a background of focal segmental glomerulosclerosis or diabetic glomerulosclerosis, the fibrillary nature of the mesangial matrix becomes particularly prominent for unclear reasons (34,35,36), needless to say, the Congo red stain if obtained is negative. Distinction from FGN usually relies on the negative immunofluorescence studies. At the ultrastructural level, the fibrils in most cases tend to be curvilinear and run together in serpentine bundles that intersect, although we have seen cases where the fibrils appeared to be disposed randomly, adding to the diagnostic confusion. Awareness of the existence of such cases and correlation with the underlying light microscopic features and immunofluorescence findings should assist in resolving problems that they may pose. This is discussed in further detail and illustrated in a separate section below on altered mesangial matrix.

The most important differential diagnostic consideration is the entity to be discussed next, namely, immunotactoid glomerulopathy (ITG). Not as much because of the difficulty of distinguishing between the two entities but rather because of the extensive debate that has been going on for over a decade between those who believe that there is no need to distinguish between these two entities merely on morphologic grounds and the majority of renal pathologists and nephrologists who agree that these are two entities that are distinguishable on both morphologic and clinical grounds.

Pathogenesis

The pathogenesis of FGN is not well understood. However, based on the study of Iskandar et al. where they found that of 13 cases stained for subclasses of IgG all stained strongly for IgG4 and only 15% of cases in addition exhibited weak staining for IgG1, it appears that even when the IgG component of deposits in FGN is polyclonal, they demonstrate a degree of homogeneity that may be one of the factors that along with the concentrating microenvironment provided by the filtration function in the glomerulus contribute to the complexes forming fibrillary structures. It has been pointed out that a similar profile is seen in membranous glomerulopathy, which is not associated with formation of fibrillary structures. However, the dominance of IgG4 in membranous glomerulopathy is not as pronounced as in FGN. Although IgG4 is incapable of activating the complement cascade via the classical pathway, aggregates of IgG4 and IgG4-containing immune complexes can activate the alternate pathway of complement. A study using immunogold labeling demonstrated that the immunogoldlabeled antibodies to IgG, C3, and amyloid P component, but not those similarly labeled antibodies to fibronectin, fibrillin, or collagen IV, localized along the fibrils of FGN, suggesting that amyloid P component may play a role in fibrillogenesis but not any of the other proteins (37).

Treatment and Prognosis

The treatment of FGN has so far consisted into administration of steroids with or without cytotoxic drugs, which has not been effective with the disease progressing within 2 to 4 years from diagnosis to ESRD in approximately 50% of cases. The only study that has tried to correlate histologic pattern with prognosis found a negative correlation between the presence of crescents and tubulointerstitial damage and outcome (18). There was also a correlation between the different patterns observed in that study and the risk of progression as well as the rate of progression to ESRD. Transplantation is a valid option since although the fibrils recur in ≥50% of grafts, the influence on the outcome of the grafts is minimal (38,39). Rituximab showed encouraging effects on proteinuria in three patients (40).

IMMUNOTACTOID GLOMERULOPATHY

Clinical Presentation and Epidemiology

The origin of the term immunotactoid glomerulopathy goes back to 1980, when Schwartz and Lewis (5) described the case of a middle-aged man presenting with the nephrotic syndrome whose biopsy revealed mesangial and capillary wall deposits of IgG3-κ and ultrastructural examination showed mesangial and subepithelial microtubular deposits with a diameter of approximately 27 nm. The authors therefore speculated that the organized microtubules were the result of crystallization of
the immunoglobulins in a manner analogous to tactoid formation in the case of sickle cell disease.

Immunotactoid glomerulopathy is 10-fold rarer than FGN such that in most large referral biopsy centers, its frequency amounts to about 0.1% of native biopsies accessioned. Virtually all cases described have been in Caucasians, the gender distribution is equal, and the incidence is highest in the elderly with most cases reported in patients greater than 50 years of age. In one of the largest series (20), the age of incidence was significantly more advanced (62 years ± 2) compared to that of cases of FGN (50 years ± 2). Its clinical presentation is similar to that of FGN with one very important feature that, in addition to the ultrastructural appearance, justifies in the view of most pathologists and nephrologists the distinction between the two entities, which is its high association with a serum or urine paraprotein and/or an overt lymphoproliferative disorder, frequently a chronic lymphocytic leukemia (CLL) (17,18,20). As an example, 6 of the 14 cases in the series of Bridoux et al. (17) had CLL, 1 had a small lymphocytic B-cell lymphoma, and 3 had a monoclonal gammopathy of uncertain significance (MGUS).

FIGURE 23.7 Monoclonal IgG lambda immunotactoid glomerulopathy with a membranoproliferative pattern with mesangial matrix expansion and hypercellularity, thick capillary walls, and slight lobulation A: H&E stain; B: PAS stain; C: Masson trichrome stain; D: Immunofluorescence microscopy showing capillary wall and mesangial staining for lambda light chains. Staining for IgG was similar, and staining for kappa light chains was negative. (400×.) (Courtesy of Carlos Andres Jimenez Guerrero and Adil Mohamed Hussein Gasim, Chapel Hill, NC.)

Pathologic Findings

Gross Findings

As in the case of FGN, there is no experience with autopsy examination of cases that have not been modified by dialysis or transplantation.

Light Microscopy

Most series describe cases with membranoproliferative-like features (Fig. 23.7) or features of atypical membranous glomerulopathy (17,18,20). Depending on the abundance of the
infiltrating microtubular material, the glomeruli may show focal segmental hyalinosis, and sometimes, intraluminal hyaline thrombi are present, representing aggregates of microtubules. None of the 6 cases reported by Fogo et al. (20) and Rosenstock et al. (18) demonstrated crescents, 2 of the 14 cases reported by Bridoux et al. (17) had crescents, and none of the 16 cases reported by the Mayo Clinic group had crescents (41). The trichrome-stained sections present an appearance similar to that described in connection with FGN. The sections stained with Jones stain are described as showing a moth-eaten appearance because of failure of the mesangium to pick up the silver stain. Of course, the stains for amyloid are negative.

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