peroxidase-conjugated antisera against x and A light- ..... Gallo G, Feiner HD, Katz LA, Feldman GM, Correa EB, ... Edited by EL Becker, New York, John Wiley.
Glomerular
Capillary
Aneurysms
in Light-Chain
Nephropathy An Ultrastructural Proposal
of
R. SINNIAH, MD, PhD, A. H. COHEN, MD
Pathogenesis
From the Department of Pathology, Harbor-UCLA Medical Center, Torrance, California; the University of California, Los Angeles, School of Medicine, Los Angeles, California; and the Department of Pathology, National University of Singapore, Singapore General Hospital, Singapore
The occurrence of glomerular capillary aneurysms in light-chain nephropathy and a morphologic pattern of development as determined by electron microscopy are described. Renal biopsies from 4 patients with nodular glomerulopathy were evaluated. Light-chain deposition in mesangium and capillary walls was associated with monocyte accumulation in capillaries. Loss ofendothelial cell lining and disruption of mesangial anchoring points ofbasement membranes ensued, resulting in aneurysmal dilatation of the capillaries. The walls collapsed, and the
aneurysms healed as a result of mesangial interposition, a process which, in combination with enlarging nodules, led to thickening and collapse of the walls and narrowing of the lumens. This study, in conjunction with review of photographs from previously published reports, indicates that glomerular aneurysms are a common feature of nodular light-chain glomerulopathy. It also emphasizes additional light-niicroscopic similarities between this glomerulopathy and nodular diabetic glomerulosclerosis. (Am J Pathol 1985, 118:298-305)
SINCE the initial report of 2 cases of multiple myeloma with granular electron-dense deposits containing kappa (xc) light-chains within glomerular mesangial nodules and glomerular and tubular basement membranes, ' many additional cases of light-chain nephropathy have been documented.29 This glomerulopathy was subsequently found to be related to multiple myeloma in less than 50Oo of the cases. Although the predominant lesion was initially emphasized as having a nodular configuration associated with monoclonal x light-chain deposits, other morphologic forms of glomerular injury, including proliferative glomerulonephritis, and deposits of monoclonal A light-chain have been described as constituting the entity of light-chain glomerulopathy.8 There are many light-microscopic similarities between light-chain nodular glomerulopathy and nodular diabetic glomerulosclerosis; as emphasized by many investigators, the composition of the nodules and their ultrastructural appearances are dissimilar and serve as points of diagnostic differentiation.35 Glomerular cap-
illary aneurysms are considered to be an integral feature of nodular diabetic glomerulosclerosis; as postulated by Bloodworth, the nodules form as a consequence of organization of the aneurysmal contents.10 In contrast, glomerular aneurysms in light-chain nephropathy have been described only infrequently,59 and little attention has been directed at their pattern of development. This report, of 4 cases of light-chain nephropathy, examines the relationship between capillary aneurysms and mesangial nodules and traces the morphologic abnormalities leading to the formation and organization of the aneurysms.
Materials and Methods The four biopsies included in this report were referred for evaluation to the renal pathology laboratories at Accepted for publication September 19, 1984. Address reprint requests to Arthur H. Cohen, MD, Department of Pathology, Harbor-UCLA Medical Center, 1000 W. Carson Street, Torrance, CA 90509.
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ANEURYSMS IN LIGHT-CHAIN GLOMERULOPATHY
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Harbor-UCLA Medical Center. They represent all cases of light-chain glomerulopathy studied. The specimens were divided into three portions and processed for light microscopy, electron microscopy, and immunofluorescence as previously described."1 For light microscopy, hematoxylin and eosin, periodic acid-Schiff (PAS), Masson's trichrome, periodic acid-methenamine silver, Congo red, and crystal violet stains were employed. For immunofluorescence, monospecific fluorescein-isothiocyanate conjugated antisera against IgG, IgA, IgM, Clq, albumin, fibrin (Calbiochem-Behring, La Jolla, California), C3, and x and A light-chains (Meloy, Springfield, Virginia) were used; in addition, monospecific peroxidase-conjugated antisera against x and A lightchains (Calbiochem-Behring) were employed on frozen sections. The clinical histories were obtained from the referring physicians and from record review.
Results Patient Information The pertinent clinical and laboratory findings are provided in Table 1. Patient 1 has been the subject of an extensive report.9 Only 2 patients were deemed to have multiple myeloma, and the other 2 had slight increases in bone marrow plasma cells of mature appearance. Three patients had an abnormal x light-chain, while one had an abnormal A light-chain.
299
Light Microscopy There was mesangial expansion in a nodular fashion with no appreciable increase in cellularity, although a few glomeruli showed a slight increment in mesangial nuclei. The nodules affected almost all lobules of all glomeruli. Less than 10% of the glomeruli exhibited only mild diffuse expansion of the mesangium. The nodules were strongly PAS-positive and stained in a variable way with silver impregnation. Congo red was negative, there was no birefringence in polarized light, and metachromasia with crystal violet stain was absent. Overlying many of the nodules were aneurysmally dilated capillaries (Figure 1). Aneurysms were not present in glomeruli without nodules. The basement membranes were generally thin; some were wrinkled. A few capillary loops had double contours. Mild to moderate interstitial edema and small focal collections of interstitial lymphocytes were observed. Scattered tubular epithelial cells showed cytoplasmic lipid vacuoles, and a few cells also contained protein reabsorption droplets. Typical myeloma casts with surrounding multinucleated giant cells were absent in Patients 1-3, while they were commonly noted in the biopsy from Patient 4.
Renal Biopsies With the exception of the light-chain class (A, not x) and the presence of myeloma casts in tubules of Patient 4, the changes in all biopsies were similar and will, therefore, be described as a unit, with emphasis solely on glomeruli.
Glomerular Immunohistochemistry Both fluorescein- and peroxidase-conjugated antibodies gave similar results. In Cases 1-3, there was strong staining for x light-chain determinants in all mesangial areas and nodules, and linear staining of the glomerular and tubular basement membranes, while a similar pattern for A light-chain was observed in Case 4 (Figure 2). There was negative staining for other immunoglobulins and complements, except for Patient 1, where IgG, C3, and Clq were present in the same distribution as the x light-chains.9
Table 1 -Summary of Clinical and Laboratory Findings Patient
Age/Sex 71 /F
2
3
4
42/F 67/F
55/M
Presenting complaint Hypertension azotemia Fluid overload anemia Shortness of breath, edema Fatigue, hy-
BUN/Cr (mg/dl)
Urine analysis Protein semiquantitative RBC/HPF
65/2.5
4+
Abnormal light chain
Urine
Serum
3-6
x
x
NA
x
NA
1-2
x
x
(19.4 g/24 hrs) NA/1.6
4+
Bone marrow 6% plasma cells Plasmacytosis
(Nephrotic syndrome) 52.6/2.6
2-3 +
Multiple
myeloma
70/8.0
4+
pertension RBC, red blood cells; HPF, high-power microscopic field; NA,
not available.
0
A
Multiple myeloma
300
SINNIAH AND COHEN
AJP * February 1985
Figure I1-Light-microscopic appearance of a representative glomerulus demonstrating multiple mesangial nodules and capillary aneurysms (arrows).
(DAS
x
270)
Electron Microscopy The prominent changes were observed in the glomerular mesangium by the formation of nodules which contained deposits of finely granular electron-dense material within expanded matrixes (Figure 3). In addition, there were small numbers of fibrils measuring approximately 100-150 A in width and of indefinite length. The basement membranes, particularly in the lamina rara interna, were permeated by a continuous thin
x
fa,
X
{.' 0
F'"
>.e .J.
Figure 3-Low magnification electron micrograph of a typical mesangial nodule composed of matrix impregnated by electron-dense material (arrows). ( x 2210)
to thick, moderately electron-dense, and finely granular substance with the same density as that in the mesangium (Figures 3 and 4). Subendothelial spaces of many capillaries were widened, resulting from accumulation of lucent and loosely flocculent material. In addition,
O_
_s
_
tStR~~~.~k
i..
Figure 2-Glomeruli stained with fluorescein-labeled anti-x antiserum; light chain is present in the mesangial nodules and basement membranes of capillaries, Bowman's capsules, and tubules. (x 160)
Figure 4-Portion of glomerular capillary wall demonstrating a continuous layer of granular dense material (arrows) internal to the lamina densa. The endothelial cell is partially detached (arrowhead). (x 7225)
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.. i
301
;:S.,NxX't~~~~~~~~le,,
Figure 5-Early lesion in the evolution of aneurysm formation. There is marked widening and lucency of the subendothelial zone. Note the intact endothelial cell cytoplasm. (x 5000) Inset-Higher magnification of 5. The subendothelial space contains pale material, fine fibrils (arrow), and fibrin. There is a very thin basement membrane immediately beneath the endothelial cells. Note the small portion offluffy interposed mesangial cell (double arrow) directly apposed to a small portion of original basement membrane. (x 6000)
in areas adjacent to the mesangium, there were portions of interposed mesangial cells which were closely apposed and possibly adherent to segments of basement membranes; in capillary aneurysms, the remainder of the basement membranes were prominently "ballooned" from the original contour of the capillary wall (Figure 5). In some of these capillaries, monocytes accumulated in the widened subendothelial spaces (Figures 6 and 7); in several, endothelial cells were partially or completely detached (Figures 4 and 6). In these capillaries, two additional abnormalities were present. Portions of basement membranes had small single (Figures 6 and 7) or multiple ruptures or discontinuities (Figure 8), often
close to the leukocytes (Figures 6 and 7). Furthermore, there was more prominent dilatation of the capillaries, especially where basement membranes were not "attached" to the mesangial cells. This combination of changes led to the observed aneurysms. The aneurysms were, therefore, partially lined by endothelium; they contained cytoplasmic fragments, erythrocytes, fibrin, and leukocytes. The less dilated capillaries were the sites of a greater degree of mesangial interposition and, therefore, more "anchoring" points, while the more dilated ones had fewer anchoring points or, at the extreme, lacked even any mesangial anchors except at their
peripheries (Figure 9).
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Figure 6-Portion of capillary wall with interposed mesangial cell (M) and monocyte (arrow) in widened subendothelial space. Note the defect (D) in the basement membrane, the residual anchor points (arrowheads), and the lack of endothelial cell. (x 3500)
In what are interpreted as the older and healing lesions of the capillary aneurysms, there was also evidence of mesangial interposition, which was initially partial, and later progressed to circumferential involvement. Mesangial matrix and cells were adherent at multiple points along the basement membranes, a process which resulted in a convoluted capillary wall (Figure 10). Nevertheless, this restored the continuity and substance of the capillary wall, and the new lumen was lined completely by endothelial cytoplasm. There was also deposition of new mesangial matrix-basement membrane material in the capillaries (Figure 10), corresponding to the "double contours" described above in the light microscopy section. Further progression of this process, especially that of matrix production, led to considerable capillary wall thickening. It is likely that simultaneous enlargement of the nodules, in concert with the capillary wall changes, resulted in marked narrowing of capillary lumens (Figure 10).
ment further concerning significance, pathogenesis, or ultrastructural features. Gipstein et al,9 in a detailed case report of Patient 1, also pointed out the presence of glomerular aneurysms. As the result of our experience with that case, we carefully studied subsequent biopsies submitted to our laboratory from patients with light-chain nephropathy. Our review of published lightmicroscopic photographs of this glomerulopathy has disclosed the presence of undescribed aneurysms in several instances,2'4 7'8 12.13 suggesting the frequency to be greater than previously suspected. We postulate that the aneurysms arise as a consequence of mesangial and basement-membrane injury from the deposited light-chains, in concert with accumulation of and infiltration by monocytes. The exact stimulus for monocyte involvement in this injury is not known, but it is possible that these cells secrete various enzymes, including proteases, which disrupt both peripheral basement membranes and possibly the anchor points between basement membranes and the peripheral portions of the mesangium to which they are attached. If this postulate is true, then the pattern of injury leading to glomerular aneurysms in nodular diabetic glomerulosclerosis, as suggested by Bloodworth,10 and in light-chain glomerulopathy would be different. As envisioned by Bloodworth, the diabetic aneurysms form first, the nodules representing "organization" of the aneurysmal contents. In that disorder, it is suggested that there is disruption of mesangial anchoring points as the primary event in aneurysm formation; in contrast, our observations suggest that the
Discussion This study has described the occurrence and traced the morphogenesis of capillary aneurysms in glomeruli with nodular light-chain nephropathy. Our experience is in contrast to the many reported cases of this glomerulopathy, for with two notable exceptions,5'9 there has been no mention of capillary aneurysms. Seymour and coworkers5 briefly described microaneurysms in two patients with nodular glomerulopathy, but did not com-
Figure 7-Another capillary with a monocyte directly apposed to the basement membrane. There is a small early defect in the basement membrane
(arrow). (x 8900)
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303
Figure 8-Multiple disruptions of basement membrane. (x 8105)
nodules are present prior to disruption of mesangial anchoring points, for aneurysms were not observed in glomeruli with nodules. Immunoelectron-microscopic studies have indicated the electron-dense material in the glomerular basement membrane to be abnormal light-chain deposits.6 The deposits expand the mesangial matrical channels, appear to increase mesangial cell activity, and perhaps also result in moderate increase in mesangial matrix. The widened mesangium shows admixture of fine fibrils and granular deposits. The fibrils appear to be intrinsic to the mesangial matrix and are more prominent in certain forms of glomerular injury. There is a close similarity of these fibrils in normal and diseased glomeruli.'4 Despite the marked widening of the mesangium, collagen fibrils were not detected, unlike other types of chronic glomerulonephritis with mesangial immune complex deposits, such as IgA nephropathy,15 and unlike the observations of Schubert and Adam, who described long-spacing collagen in the nodules. 12 The abnormal mesangial deposits probably influence the metabolism of the cells, and stimulate increased production of mesangial matrix.15 Abnormalities of the peripheral glomerular capillaries include endothelial cell detachment and the accumulation of fluffy and fibrillary material and new basement membrane in the subendothelial space. These changes are similar to thrombotic microangiopathy,16 chronic transplant rejection,1I and radiation nephritis,18 entities in which the capillary loops also show detachment of endothelial cells, thinning of capillary walls, and aneurysms. Common to all of these entities is mesangiolysis,19 a feature which was not present in the glomeruli studied by us. Instead, we found that the capillary wall changes were due to light-chain deposits within the basement membranes; with participation of blood monocytes producing further injury, there was disruption of anchor points between the capillary loops of a lobule where the peripheral capillary basement membrane is attached to the mesangium. Rupture of
several anchors appeared to produce a microaneurysm composed of several capillary loops and to lead to additional dilatation of the aneurysm. These alterations are similar to the glomerular abnormalities induced by ...
..
'..
Figure 9-An aneurysmally dilated capillary containing distorted erythrocytes and tactoids of fibrin. Only remnants of endothelial cells are still partially attached to the basement membrane (arrow). Note the portion of the mesangial cell also adherent to the basement membrane (double arrows).
(x 2750)
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Figure 10-"Healed" aneurysm overlying mesangial nodule. The original basement membrane is wrinkled; the capillary wall is greatly thickened with mesangial interposition and extensive mesangial matrix beneath the endothelial cell lining an extremely narrowed capillary lumen (L). (x 4285) InsetHigher magnification of "healed" capillary wall lesion. Note the interposed mesangial cell (asterisk), abundant matrix (MM), and narrow lumen (L). (x 9325)
administration of habu snake venom, in which microaneurysms result in 24 hours.20 The ability of light-chain deposits to cause vascular wall weakening and aneurysm formation is suggested by the finding of an intracerebral aneurysm with dense eosinophilic deposits in the outer media in a case of x light-chain disease.7 Rouleaux formation or sludging probably does not contribute to this disorder, although intravascular coagulation has been proposed as a factor in the production of renal lesions in multiple myeloma.21 In this regard, capillary loops in nodular light-chain nephropathy show evidence of decreased vascularity; there is fragmentation of red blood cells and aggregation of platelets. Whether sluggish blood flow is a result of endothelial damage and aneurysmal dilatation or a cause of these alterations is not known. The microaneurysms appear to organize by several processes, including direct extension of mesangial nodules into the lumens and the formation of mesangial bridges. Dendritic mesangial projections acting as bridges between the mesangium and peripheral loops subdivide the capillaries into branches. Mesangial bridges also occur in the chronic phase of Masugi nephritis, apparently representing an irreversible stage in this glomerulonephritis,22 and in diabetes mellitus.23
The bridges are irreversible lesions ultimately leading to segmental obliteration of glomerular capillaries. This study suggests that light-chain deposits have a direct and simultaneous effect on the mesangium and peripheral glomerular capillaries and act with participation of monocytes in the genesis of glomerular capillary microaneurysms. The formation of these aneurysms is a common feature in nodular glomerulo-
sclerosis of light-chain nephropathy.
References 1. Antonovych T, Lin C, Parrish E, Mostofi K: Light-chain deposits in multiple myeloma (Abstr). Seventh Annual Meeting, American Society of Nephrology, 1973 2. Randall RE, Williamson WC, Mullinax F, Tung MY, Still WJS: Manifestations of systemic light-chain deposition. Am J Med 1976, 60:293-299 3. Herf S, Pohl SL, Sturgill B, Bolton WK: An evaluation of diabetic and pseudodiabetic glomerulosclerosis. Am J Med 1979, 66:1040-1045 4. Gallo G, Feiner HD, Katz LA, Feldman GM, Correa EB, Chuba JV, Buxbaum JN: Nodular glomerulopathy as-
sociated with nonamyloidotic kappa light-chain deposits and excess immunoglobulin light-chain synthesis. Am J Pathol 1980, 99:621-644 5. Seymour AE, Thompson AJ, Smith PS, Woodroffe AJ, Clarkson AR: Kappa light-chain glomerulosclerosis in multiple myeloma. Am J Pathol 1980, 101:557-573
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6. Tubbs RR, Gephardt GN, McMahon JT, Hall PM, Valenzuela R, Vidt DG: Light-chain nephropathy. Am J Med 1981, 78:263-269 7. Colvin R: Case records of the Massachusetts General Hospital. N Eng J Med 1981, 304:33-43 8. Ganeval D, Mignon F, Preud'homme JL, Noel LH, MorelMaroger L, Droz D, Brouet JC, Mery JPH, Grunfeld J-P: Visceral deposition of monoclonal light-chains and immunoglobulins: A study of renal and immunopathologic abnormalities. Adv Nephrol 1982, 11:25-63 9. Gipstein RM, Cohen AH, Adams DA, Adams T, Grabie MT: Kappe light-chain nephropathy without evidence of myeloma cells. Am J Nephrol 1982, 2:276-281 10. Bloodworth JMB Jr: A re-evaluation of diabetic glomerulosclerosis 50 years after the discovery of insulin. Human Pathol 1980, 9:439-453 11. Cohen AH, Border WA: Myeloma kidney: An immunomorphogenetic study of renal biopsies. Lab Invest 1980, 42:248-256 12. Schubert GE, Adam A: Glomerular nodules and longspacing collagen in kidneys of patients with multiple myeloma. J Clin Pathol 1974, 27:800-805 13. Morel-Maroger L, Verroust P, Preud'homme JL: Glomerular lesions in plasma cell dyscrasias, in Pathology of Glomerular Disease. Edited by S Rosen. New York, Churchill Livingstone, 1983, p. 207-224 14. Hsu H-C, Churg J: Glomerular microfibrils in renal dis-
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ease: A comparative electron microscopic study. Kidney Int 1979, 16:497-504 Sinniah R, Churg J: Effect of IgA deposits on the glomerular mesangium in Berger's disease. Ultrastruct Pathol 1983, 4:9-22 Goldstein MH, Churg J, Strauss L, Gribetz D: Hemolytic-uremic syndrome. Nephron 1979, 23:263-272 Hsu H-C, Suzuki Y, Churg J, Grishman E: Ultrastructure of transplant glomerulopathy. Histopathology 1980, 4:351-367 Churg J, Madrazo A: Radiation nephritis. Seminars in Nephrology. Edited by EL Becker, New York, John Wiley & Sons, 1977, pp. 83-96 Morita T, Churg J: Mesangiolysis. Kidney Int 1983, 24:1-9 Bradfield JWB, Cattell V, Smith J: The mesangial cell in glomerulosclerosis. II. Mesangial proliferation caused by habu snake venom in the rat. Lab Invest 1977, 36:487-492 Ward AM, Preston FE: The kidney and intravascular coagulation in myelomatosis. Br Med J 1974, 2:529-531 Kondo P, Shigamatsu H, Okabayashi A: Cellular aspects of rabbit Masugi nephritis: IJI. Mesangial changes. Lab Invest 1976, 34:363-371 Watanabe T, Emoto K: Mesangial bridges: A mode of progression in diabetic glomerulosclerosis. Acta Pathol Jpn 1981, 31:1045-1051
