Successful renal transplantation in a child with ANCA-associated microscopic polyangiitis

Crescentic glomerulonephritis (CGN) is a clinicopathologic entity which is characterized by severe renal dysfunction of rapid onset with glomerular crescents. Type III CGN is associated with the absence of glomerular immune complex deposition (pauci-immune) and is associated with antineutrophil cytoplasmic antibody (ANCA). Microscopic polyangiitis and idiopathic pauci-immune necrotizing glomerulonephritis (NCGN) are strongly associated with ANCA directed against myeloperoxidase (anti-MPO). We describe here an unusual pediatric patient with MPO-ANCA-associated rapidly progressive glomerulonephritis (RPGN), emphasizing the management and outcome of the disease.     

Glomerular expression of C-C chemokines in different types of human crescentic glomerulonephritis.

BACKGROUND: Crescentic glomerulonephritis (CGN) presents a rapidly progressive glomerulonephritis clinically, in which macrophages play a crucial role in the pathogenesis. However, the precise molecular mechanism of macrophage recruitment and activation has not been fully elucidated. C-C chemokines, monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha and beta (MIP-1alpha and MIP-1beta), are major chemoattractants for macrophages. We attempted to study the expression of C-C chemokines and their correlation with CD68-positive macrophages in crescentic glomeruli to investigate further their possible roles in crescent formation and progression to fibrosis in different types of human CGN. METHODS: The expression of MCP-1, MIP-1alpha, MIP-1beta and CD68 was detected in glomeruli with different forms of crescents (cellular, fibrocellular and fibrous crescents) by immunohistochemistry in serial sections of renal biopsies taken from 32 patients with biopsy-proven CGN including eight patients with anti-glomerular basement membrane (GBM) disease (type I CGN), 12 patients with immune complex-mediated CGN (type II CGN) and another 12 patients with pauci-immune CGN (type III CGN) enrolled in this study. Eight normal human kidneys were obtained from cadaveric renal transplant donors whose kidneys were technically unsuitable for transplantation, serving as controls. RESULTS: MCP-1, MIP-1alpha, MIP-1beta and CD68 were undetectable in glomeruli of normal kidney. In crescentic biopsies, MCP-1, MIP-1alpha, MIP-1beta and CD68 were detected in fibrocellular crescents and were even more prominent in cellular crescents, but were undetectable in fibrous crescents. Using consecutive sections for staining, it was demonstrated that a high proportion of infiltrating CD68-positive macrophages, mainly localized to the area of the expression of chemokines, were MCP-1, MIP-1alpha and MIP-1beta positive in crescents. Chemokines were expressed mainly by CD68-positive macrophages and parietal epithelial cells in crescents. The number of MCP-1- and MIP-1alpha-positive cells in glomeruli with cellular crescents was positively correlated with the number of CD68-positive cells (r = 0.568 and 0.749, respectively, both P < 0.01). The number of MCP-1- and MIP-1alpha-positive cells and the incidence of Bowman's capsule rupture in glomeruli of patients with type I CGN were higher than those of type II and type III CGN. CONCLUSIONS: These observations suggest that the expressed C-C chemokines, MCP-1, MIP-1alpha and MIP-1beta, may mediate the inflammatory process of crescent formation and progression to fibrosis. The strong correlation of MCP-1 and MIP-1alpha with infiltrating macrophages within glomeruli with cellular crescents suggested that these chemokines might be of particular importance for macrophage recruitment to this site. MCP-1 and MIP-1alpha were correlated to type I CGN with its more severe inflammatory course and worse prognosis. The variance of glomerular expression of C-C chemokines may contribute to the difference in histopathological features and prognosis in these three types of CGN.     

Osteopontin expression in human crescentic glomerulonephritis

Osteopontin expression in human crescentic glomerulonephritis. BACKGROUND: Osteopontin is a molecule with diverse biological functions, including cell adhesion, migration, and signaling. The expression of osteopontin has been demonstrated in a number of models of renal injury in association with accumulations of monocyte/macrophages, including recent reports of osteopontin expression in glomerular crescents in a rat model of anti-glomerular basement membrane glomerulonephritis. METHODS: Glomerular expression of osteopontin in biopsies of human crescentic glomerulonephritis (N = 25), IgA nephropathy with crescents (N = 2), and diffuse proliferative lupus glomerulonephropathy with crescents (N = 1) was studied by immunohistochemistry, in situ hybridization, and combined immunohistochemistry/in situ hybridization. Additionally, antibodies to cell-specific phenotypic markers were used to identify cellular components of the glomerular crescent, which express osteopontin protein and mRNA. RESULTS: All of the crescents present in the biopsies studied contained a significant number of cells that expressed osteopontin protein and mRNA, demonstrated by immunohistochemistry and in situ hybridization, respectively. Using replicate tissue sections and combined immunohistochemistry/in situ hybridization, we showed that the majority of the strongly osteopontin-positive cells are monocyte/macrophages. In addition to the very strong and cell-associated localization, a weaker and more diffuse pattern of osteopontin protein and mRNA expression could be seen in a number of crescents. None of the osteopontin mRNA-expressing cells could be identified as parietal epithelial cells, CD3-positive T cells, or alpha-smooth muscle actin-positive myofibroblasts. Interstitial monocyte/macrophages did not express osteopontin, except when located in a periglomerular inflammatory infiltrate. CONCLUSIONS: Macrophages present in the human glomerular crescent express osteopontin protein and mRNA at a high level. This expression supports a role for osteopontin in the formation and progression of the crescentic lesion via chemotactic and signaling properties of the molecule.     

Heparin-binding epidermal growth factor-like growth factor is expressed in the adhesive lesions of experimental focal glomerular sclerosis.

BACKGROUND: In this study, we attempted to determine whether heparin-binding epidermal growth factor-like growth factor (HB-EGF) was up-regulated in two chronic models of proteinuria. METHODS: Chronic passive Heymann nephritis (PHN) and puromycin aminonucleoside (PAN) models were induced in Sprague-Dawley rats. HB-EGF expression was studied by Northern blotting, in situ hybridization, and immunohistochemistry. RESULTS: The chronic PAN model was associated with the development of glomerular lesions of focal glomerular sclerosis (FGS), severe interstitial fibrosis, and renal failure. Lesions of FGS were seen in approximately 80% of glomeruli at all time points, with a slight increase in the number of glomeruli showing extensive adhesion between 40 and 90 days. Northern blots of whole kidney tissue showed a 3- to 5.8-fold increased expression of HB-EGF mRNA in the chronic PAN group. Increased mRNA and protein were localized by in situ hybridization and immunohistochemistry to tubules, glomerular epithelial cells (GECs), and cells of Bowman's capsule. HB-EGF mRNA and protein were strongly expressed by epithelial cells involved in the formation of the lesions of FGS. By contrast, in chronic PHN, there was a small increase in HB-EGF, and the extensive lesions of FGS did not develop despite continued, heavy proteinuria. CONCLUSIONS: These data suggest that HB-EGF may contribute to formation of the lesions of FGS, perhaps through stimulation of abortive mitogenesis in GECs or an adhesive interaction between transmembrane HB-EGF and the exposed glomerular basement membrane.     

Antineutrophil cytoplasmic antibodies (ANCA) in Chinese patients with anti-GBM crescentic glomerulonephritis

OBJECTIVE: To study the prevalence of ANCA and their target antigen in Chinese patients with anti-GBM crescentic glomerulonephritis (CGN), and to evaluate the possible role of ANCA in Chinese anti-GBM CGN patients with coexisting serum ANCA by studying clinicopathologic features of this disease. MATERIAL AND METHODS: Twenty-three sera were collected from 23 renal biopsy-proven anti-GBM CGN patients. According to the standardized procedures, all of the sera were determined by both, indirect immunofluorescence (IIF) ANCA, and enzyme-linked immunosorbent assay (ELISA) MPO-ANCA, PR3-ANCA and BPI-ANCA. The patients were divided into two groups according to serum ANCA positivity (Group A) or negativity (Group B). Thirty-three ANCA-associated pauci-immune CGN patients were regarded as control group (Group C). Their clinicopathologic features were compared to reveal whether ANCA correlated with disease activity. RESULTS: There were 11 (47.8%) cases with positive serum ANCA in 23 anti-GBM glomerulonephritis patients. There were 4/11 MPO-ANCA (one with positive PR3-ANCA and C-ANCA, three with negative IIF-ANCA), 1/11 PR3-ANCA (with positive MPO-ANCA and C-ANCA), 3/11 P-ANCA (with negative ELISA-ANCA) and 5/11 C-ANCA (one with positive PR3-ANCA and MPO-ANCA, and the other four with negative ELISA-ANCA). No BPI-ANCA was detected. No different clinicopathologic features were found between Groups A and B. Both were different from Group C in age, sex ratio, frequence of anuria and ESRD, variety of crescents, glomerular sclerosis, vessel lesion and prognosis. CONCLUSION: Our data demonstrate that ANCA in Chinese patients with anti-GBM CGN is not rare. The major target antigen of ANCA is MPO. ANCA seems not to be correlated with disease activity.     

Renal histology in Chinese patients with anti-myeloperoxidase autoantibody-positive Wegener's granulomatosis.

BACKGROUND: Proteinase-3 antineutrophil cytoplasmic antibody (PR3-ANCA) was the serological marker for Wegener's granulomatosis (WG), while myeloperoxidase (MPO)-ANCA was the serological marker for microscopic polyangiitis (MPA). However, our previous study suggested that patients with MPO-ANCA positive WG were common in Chinese. This study aimed to analyse the renal histology of patients with MPO-ANCA positive WG. METHODS: Patients in our centre with WG were selected according to both the Chapel Hill Consensus Conference (CHCC) definition and American College of Rheumatology classification criteria. Patients with MPA were selected according to the CHCC definition. The renal histology was compared between patients with MPO-ANCA positive WG and with PR3-ANCA positive WG as well as patients with MPO-ANCA positive MPA. RESULTS: Sixty-one patients with WG had complete renal histological data, 39/61 with positive MPO-ANCA and 22/61 with positive PR3-ANCA. Among patients with crescents in glomeruli, those with MPO-ANCA had fewer cellular crescents and more fibrous crescents than those with PR3-ANCA (P < 0.01 and P < 0.05, respectively). Interstitial fibrosis and tubular atrophy were more prevalent and severe in patients with MPO-ANCA than in those with PR3-ANCA (P < 0.01 and P < 0.05, respectively). Compared with 44 patients with MPO-ANCA positive MPA, patients with MPO-ANCA positive WG had fewer glomeruli with crescents and more normal glomeruli (P < 0.01 and P < 0.01, respectively). CONCLUSION: Patients with MPO-ANCA positive WG are common in Chinese. In renal histology, chronic lesions were more severe and prevalent in patients with MPO-ANCA positive WG than in patients with PR3-ANCA positive WG. Glomerular lesions were less severe and less prevalent in patients with MPO-ANCA positive WG than in those with MPO-ANCA positive MPA.     

Expression of gremlin, a bone morphogenetic protein antagonist, in glomerular crescents of pauci-immune glomerulonephritis.

BACKGROUND: Recent evidence in vitro and in vivo suggests that gremlin, a bone morphogenetic protein antagonist, is participating in tubular epithelial mesenchymal transition (EMT) in diabetic nephropathy as a downstream mediator of TGF-beta. Since EMT also occurs in parietal epithelial glomerular cells (PECs) leading to crescent formation, we hypothesized that gremlin could participate in this process. With this aim we studied its expression in 30 renal biopsies of patients with pauci-immune crescentic nephritis. METHODS: Gremlin was detected by in situ hybridization (ISH) and immunohistochemistry (IMH) and TGF-beta by ISH and Smads by southwestern histochemistry (SWH). Phosphorylated Smad2, CTGF, BMP-7, PCNA, alpha-SMA, synaptopodin, CD-68, and phenotypic markers of PECs (cytokeratin, E-cadherin), were detected by IMH. In cultured human monocytes, gremlin and CTGF induction by TGF-beta was studied by western blot. RESULTS: We observed strong expression of gremlin mRNA and protein in cellular and fibrocellular crescents corresponding to proliferating PECs and monocytes, in co-localization with TGF-beta. A marked over-expression of gremlin was also observed in tubular and infiltrating interstitial cells, correlating with tubulointerstitial fibrosis (r=0.59; P<0.01). A nuclear Smad activation in the same tubular cells, that are expressing TGF-beta and gremlin, was detected. In human cultured monocytes, TGF-beta induced gremlin production while CTGF expression was not detected. CONCLUSION: We postulate that gremlin may play a role in the fibrous process in crescentic nephritis, both in glomerular crescentic and tubular epithelial cells. The co-localization of gremlin and TGF-beta expression found in glomeruli and tubular cells suggest that gremlin may be important in mediating some of the pathological effects of TGF-beta.     

Up-regulation of extracellular matrix proteoglycans and collagen type I in human crescentic glomerulonephritis

BACKGROUND: The pathogenesis of crescentic glomerulonephritis (CGN) involves cellular migration and proliferation in the urinary space, frequently followed by fibrous organization. Extracellular matrix proteoglycans (PGs) may regulate these events via effects on cellular migration, interactions with growth factors, including transforming growth factor-beta (TGF-beta), and control of collagen fibrillogenesis. The expression of PG in human CGN is unknown. METHODS: Renal tissues from 18 patients with CGN were examined immunohistochemically for versican, decorin, biglycan and collagen type I, and were compared with morphologically normal tissues from six tumor nephrectomies. Synthesis of decorin, biglycan, and procollagen type I mRNAs was evaluated by in situ hybridization. RESULTS: Versican was strongly expressed in cellular crescents and periglomerular areas, whereas decorin and biglycan accumulated in collagen type I-enriched regions, including fibrocellular and fibrous crescents, and interstitial fibrosis. PG and collagen type I accumulation colocalized with myofibroblasts in crescents, periglomerular areas, and interstitium. CONCLUSIONS: The temporal and spatial patterns of expression demonstrated in this study provide evidence to support pathogenic roles for PG in the evolution of CGN. Based on known biological properties of this molecule, versican may facilitate migration of cells in developing crescents. Decorin and biglycan may contribute to progression of CGN, perhaps via interactions with collagen type I in the remodeled extracellular matrix.     

Expression of the chemokine monocyte chemoattractant protein-1 and its receptor chemokine receptor 2 in human crescentic glomerulonephritis

Crescents are morphologic manifestations of severe glomerular injury. Several chemokines and their receptors have been demonstrated to be involved in animal models of crescentic glomerulonephritis (cGN) and are potential targets for therapeutic interventions. Therefore, the expression of monocyte chemoattractant protein-1 (MCP-1), its receptor chemokine receptor 2B (CCR2B), and CCR5 in human cGN was studied. MCP-1 and CCR2B mRNA expression was evaluated, by in situ hybridization, in serial sections of 23 renal biopsies from patients with cGN. T cells, macrophages, and CCR5-expressing cells were examined by immunohistochemical analysis. MCP-1 mRNA was expressed by cells in crescents, parietal epithelium, and tubular epithelium, as well as by infiltrating leukocytes in the tubulointerstitium. The expression of CCR2B mRNA was observed in cells in glomeruli and crescents and in infiltrating leukocytes in the tubulointerstitium. CCR2B mRNA expression could not be clearly localized to intrinsic renal cells; evidence that most of the CCR2B-expressing cells were leukocytes is provided. CD3-positive T cells formed the major part of the interstitial cell infiltrates but were rare within the glomerular tufts. CD68-positive macrophages constituted a major population of infiltrating cells in crescents and contributed significantly to the interstitial infiltrates. The number of glomerular macrophages was associated with the number of MCP-1- and CCR2B-positive glomerular cells. Expression of CCR2B was significantly correlated with interstitial CD3-positive T cells. CCR5 expression was restricted to infiltrating leukocytes and was correlated quantitatively and by localization with interstitial CD3-positive T cells and CD68-positive macrophages. These first morphologic data on the distribution of CCR2-positive cells in human cGN suggest differential effects of chemokines and their receptors on the distribution of infiltrating leukocytes in different compartments of the kidney.     

ANCA-associated glomerulonephritis/systemic vasculitis in childhood: clinical features–outcome

Background

Antineutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis and systemic vasculitis (AAGNV) is uncommon in childhood.

Methods

This is a retrospective study of AAGNV cases diagnosed over a 13-year period in a tertiary pediatric nephrology department.

Results

Thirteen cases of AAGNV were identified: seven Wegener granulomatosis (WG) and six microscopic polyangiitis (MPA). Acute renal failure/nephrotic range proteinuria (NRP) was found in 77 % of the patients (4 with WG, all with MPA). Eleven (85 %) patients showed necrotizing glomerulonephritis (NGN), with ≥50 % crescents identified in nine patients (69 %) (4 with WG, 5 with MPA). Treatment with methylprednisolone, cyclophosphamide and plasma exchange resulted in extra-renal remission and antibody reduction in all patients and renal function improvement/stabilization in 77 % of the patients. Three patients, all without oliguria at presentation and few sclerotic lesions, had normal renal function at follow-up. Chronic kidney disease (CKD) stages 2 and 3–4 were observed in four (WG) and three (MPA) patients, respectively. Three patients (23 %) developed end stage renal disease: two were MPA patients with severe presentation (markedly impaired glomerular filtration rate, oliguria, NRP, crescentic NGN, glomerular sclerosis) and one was a WG patient with extensive interstitial fibrosis/tubular atrophy.

Conclusions

Severe renal involvement was more common in children with MPA than WG. Treatment with methylprednisolone, cyclophosphamide and plasma exchange induced extra-renal remission/serological response and renal function improvement/stabilization. Markedly decreased GFR, oliguria, NRP, and chronic glomerular lesions at presentation were predictors of poor outcome.     

Intrarenal synthesis of IL-6 in IgA nephropathy

Summary: Recent in vitro studies have shown the synthesis of interleukin-6 (IL-6) in glomerular mesangial and epithelial cells, and suggested the involvement of IL-6 in mesangial proliferative glomerulonephritis. However, the expression site of IL-6 mRNA in renal tissue of IgA nephropathy (IgAN), the most common chronic mesangial proliferative glomerulonephritis, remains obscure. to localize IL-6 mRNA in renal biopsy specimens of IgAN, we used nonradioactive in situ hybridization (ISH) developed in our laboratory, sensitive in detecting individual cells positive for a specific mRNA. In some sections, periodic acid-Schiff staining was performed after ISH in order to identify the topographical relation between IL-6 mRNA positive cells and glomerular basement membrane and mesangial area. In situ hybridization for IL-6 mRNA and immunohistochemistry for CD3 and CD68, markers for lymphocytes and monocytes, respectively, were also performed on serial sections to examine the contribution of infiltrated mononuclear cells to cells positive for IL-6 mRNA in glomeruli. Glomerular resident cells, including glomerular mesangial and epithelial cells and cells of Bowman's capsule, as well as tubular epithelial cells and infiltrated mononuclear cells expressed IL-6 mRNA. We also compared the localization of IL-6 mRNA and protein and showed different distribution between the gene product and protein. the expression of IL-6 mRNA correlated with the degree of mesangial cell proliferation and tubulointerstitial changes. Our results indicate that IL-6 is synthesized in renal tissues of IgAN and suggest that the increased IL-6 expression may be important in the pathogenesis of IgAN.     

Expression of basic fibroblast growth factor and its receptor in the progression of rat crescentic glomerulonephritis

Summary: The aim of this study was to examine the relationship of basic fibroblast growth factor (FGF-2) and its receptor (FGF-R) to tubular proliferation, interstitial fibrosis, glomerular cell proliferation and crescent formation in the development of anti glomerular basement membrane (GBM) glomerulonephrids in the rat. Using new microwave-based histochemistry techniques, weak constitutive expression of mRNA and protein for FGF-2 and FGF-R was evident in tubules and glomeruli of normal rat kidney. Double and triple staining with the proliferating cell nuclear antigen (PCNA) demonstrated that in disease there was focal up-regulation of FGF-2 and FGF-R mRNA and protein expression within proliferating tubules and fibroblast-like cells in areas of interstitial fibrosis. In contrast, tubules in non-inflamed areas of kidney showed no change in FGF-2 and FGF-R expression or cellular proliferation. In addition, many FGF-2⁺PCNA⁺ and FGF-R⁺PCNA⁺ cells, probably mesangial cells and podocytes, were evident within glomerular segmental proliferative lesions. of particular interest was the observation that many epithelial cells within cellular crescents, inculding proliferating (PCNA⁺) epithelial cells, strongly expressed both FGF-2 and FGF-R. Furthermore, there was strong FGF-2 and FGF-R expression by proliferating fibroblast-like cells within fibrocellular crescents suggesting that FGF-2 is involved in the formation and fibrotic progression of glomerular crescents. In conclusion, this study provides evidence that increased expression of FGF-2 and its receptor may participate in the proliferative/fibrotic response in progressive crescentic glomerulonephritis.     

Renal, major histocompatibility complex antigens and cellular components in rapidly progressive glomerulonephritis identified by monoclonal antibodies

Identification of crescent-forming cells in rapidly progressive glomerulonephritis (RPGN) is very difficult, and controversial results on the participation of different epithelia as well as of monocytes have been reported. In the present study different monoclonal antibodies were used to analyze cellular infiltrates of crescents and the interstitium as well as the distribution of well-defined renal antigens and major histocompatibility complex (MHC) encoded antigens along the human nephron in cryostat sections of renal biopsies from patients with RPGN. The results demonstrate that monocytes/macrophages infiltrate Bowman's space and that cellular components of crescents present with phenotypes of parietal glomerular and proximal tubular cells. T lymphocytes are significantly found in glomeruli and also in interstitium with predominance for CD4+ lymphocytes. Reduction of MHC class-II antigens within diseased glomeruli correlates with changes in renal antigen expression. Tubular cells, however, often presented an abnormal expression of MHC class-II antigens. Differences of renal and MHC-encoded antigen expression may be due to rapid regeneration episodes of renal parenchymal cells in RPGN.     

The number of CD10-positive glomerular epithelial cells reflects renal prognosis in IgA nephropathy patients

BACKGROUND: The glomerular epithelial cells play an important role in glomerular filtration of the kidney. The disruption of these cells contributes to the development of glomerulosclerosis. The present study was performed to elucidate whether loss of the glomerular epithelial cells is associated with renal injury in patients with IgA nephropathy. PATIENTS AND METHODS: Thirty renal biopsy specimens from IgA nephropathy, 12 from minor glomerular abnormalities and 5 from normal controls were observed. The specimens from IgA nephropathy were divided into 2 groups: Group IgA-1, including 11 patients who had received a follow-up renal biopsy because of deterioration of renal function, and Group IgA-2, consisting of the remaining 19 patients without follow-up biopsy. Immunohistochemistry was performed using a monoclonal antibody against CD10 antigen that appears on mature epithelial cells of glomeruli. RESULTS: The average number of CD10-positive glomerular epithelial cells (GECs) was significantly lower in IgA nephropathy than in either minor glomerular abnormalities or the normal controls. In IgA nephropathy, there were significant correlations of the GECs with renal functions. The GECs were reduced along with the progression of histopathological damage. In group IgA-1, the GECs were significantly reduced at the second biopsy compared with the first biopsy, and significantly fewer in group IgA-1 than in group IgA-2 at the first biopsy. The GECs showed a significant correlation with renal prognosis during the follow-up period. CONCLUSIONS: The reduction of GECs was associated with renal dysfunction, histopathological damage and renal prognosis. The GECs may be a useful predictor of renal prognosis in IgA nephropathy.     

Fas on renal parenchymal cells does not promote autoimmune nephritis in MRL mice

BACKGROUND: Although Fas on pancreatic islets promotes autoimmune diabetes in mice, the role of Fas expression on kidney parenchymal cells during autoimmune disease is unknown. METHODS: To determine whether Fas on renal parenchymal cells promotes autoimmune renal destruction, we compared apoptosis and pathology in Fas-intact and Fas-deficient kidneys in an autoimmune milieu. For this purpose, we transplanted single, normal kidneys from MRL-++ (Fas-intact) mice (3 months of age) into age-matched, congenic MRL-Faslpr (Fas-deficient) recipients after removal of nephritic kidneys. These Fas-intact kidneys were compared with Fas-deficient nephritic kidneys. RESULTS: There is a progressive increase of FasL on kidney-infiltrating cells and Fas and FasL on renal parenchymal cells in MRL-++ kidneys during engraftment (0, 2, 4-6, and 8 weeks). By comparison, we detected an increase in FasL in MRL-Faslpr kidneys (3 to 5 months of age), whereas Fas was not detectable. The engagement of T cells bearing FasL with Fas expressing tubular epithelial cells (TECs) induced TEC apoptosis in vitro. However, apoptosis and pathology were similar in kidneys (MRL-++, 8 weeks postengraftment vs. MRL-Faslpr, 5 months) with equivalent amounts of FasL-infiltrating cells or FasL TECs, regardless of Fas on renal parenchymal cells. CONCLUSION: The expression of Fas on renal parenchymal cells does not increase apoptosis or promote renal disease in MRL-++ mice. We speculate that the autoimmune milieu evokes mechanisms that mask, counter, or pre-empt Fas-FasL-initiated apoptosis in MRL kidneys.     

Renal expression of ICAM-1 and VCAM-1 in ANCA-associated glomerulonephritis--are there differences among serologic subgroups?

BACKGROUND: Antineutrophil cytoplasmic autoantibodies (ANCA) play a role in the expression of adhesion molecules. Differences in ANCA test results in ANCA-associated vasculitis may provide differences in their renal expression. PATIENTS AND METHODS: We assessed the renal expression of ICAM-1 and VCAM-1 with monoclonal antibodies in 19 patients with ANCA-vasculitis: 7 microscopic polyangiitis, 5 Wegener's granulomatosis, 4 renal-limited vasculitis and 3 Churg-Strauss disease. Immunofluorescence and ELISA for myeloperoxidase (MPO) and proteinase 3 (PR3) were performed for ANCA-testing. 10 normal renal tissues were used as controls. RESULTS: The ANCA staining pattern was perinuclear in 14 patients, with MPO-ANCA 31 - 220 EU/ml, and cytoplasmic in 5, with PR3-ANCA 37 - 144 EU/ml. Abnormal tubular expression of ICAM-1 and VCAM-1 was seen in more than 80% of biopsies and abnormal expression of VCAM-1 in glomerular tuft was seen in 60%. Glomerular tuft stains of ++ or +++ for VCAM-1 were observed in 10% of renal biopsies from MPO-ANCA-GN patients, but in 60% of biopsies from PR3-ANCA-GN patients (Fi = 8.538, p = 0.03). IN CONCLUSION: De novo expression of VCAM-1 on glomerular tuft suggests that the endothelial cells play a role in ANCA-GN. De novo glomerular expression of VCAM-1 is associated more with ANCA directed against PR3 than with ANCA directed against MPO. Upregulated glomerular expression of VCAM-1 may reflect a higher histological activity in patients with PR3-ANCA, and supports the existence of specific immune activation mechanisms in the different serologic subgroups in ANCA-GN. The de novo tubular expression of ICAM-1 and VCAM-1 suggests that the epithelial cells may participate in adhesive interactions in ANCA-GN.     

ANCA in haemodialysis patients

The prevalence of positive ANCA as well as the prevalence ofPR-3 and MPO antibodies were examined in a cross-sectional sampleof 1277 haemodialysis patients from 16 German haemodialysiscentres. We found 32 patients positive for c-ANCA (median titre1:40; range 1:20–1:320) and 65 for p-ANCA (1:80; 1:20–1:1280).Twenty-two percent of the c-ANCA-positive and 31% of the p-ANCA-positivepatients had PR-3 and MPO antibodies by ELISA respectively.Clinical evidence of vasculitis was found in 11 of 32 c-ANCA-positiveand 19 of 65 p-ANCA-positive patients. Of the 11 c-ANCA-positive,four had a known diagnosis of Wegener's granulo-matosis (WG);WG was recognized after the test in a further five patientsand two had renal limited RPGN. Of the 19 p-ANCA-positive patients,three had a clinical diagnosis of microscopic polyarteritis(MP), MP was newly diagnosed in a further 12, WG in one andrenal limited RPGN in three. The patients had not received cyclophosphamide(the diagnosis had been non-specified ‘systemic disease’).Thus false-positive ANCA, as defined by absence of vasculitis,was found in 5% of dialysis patients versus 0% in patients withpreterminal renal failure {n=152) or blood donors (n=150). Patientswith vasculitis tended to have higher c-ANCA and p-ANCA titresrespectively, but there was a considerable overlap. Titres werenot higher in patients symptomatic at the time of examination(6 of 11 c-ANCA and 10 of 19 p-ANCA), but PR-3 and MPO ELISAwere positive in all but two. Thus c-ANCA were false positive(i.e. not associated with clinical vasculitis) in 66% and p-ANCAin 71%. In some patients ANCA was positive, but ELISA negative. We conclude that (i) in dialysis patients c-ANCA and p-ANCAare frequently present in the absence of vasculitis; (ii) specificitycan be improved, but sensitivity is lost, by using PR-3 andMPO ELISA respectively; (iii) serology permits the recognitionof WG or MP in cases where the diagnosis has been missed. Itis recommended that all patients with unknown primary renaldisease admitted to renal replacement therapy be examined usingboth ANCA-IF and PR-3/MPO ELISA.     

Renal expression of matrix metalloproteinases in human ANCA-associated glomerulonephritis.

BACKGROUND: Expression of matrix metalloproteinases (MMPs) by infiltrating and intrinsic renal cells is increased in inflammatory conditions, and may correlate with disease activity of glomerulonephritis. We analysed renal expression of MMPs, tissue inhibitor of metalloproteinase-1 (TIMP-1) and markers of neutrophil and monocyte infiltration in renal biopsies of patients with active anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis. METHODS: Immunohistochemical expression of MMP-2, -3, -9, TIMP-1, the neutrophil- and monocyte-derived MMP activators cathepsin G, neutrophil elastase and myeloperoxidase (MPO), and the monocyte marker CD14 was determined in renal biopsies of active proteinase 3 (PR3)-ANCA (n = 7) and MPO-ANCA (n = 6) associated glomerulonephritis, and in normal renal tissue (n = 4). Double labelling experiments of MMPs and TIMP-1 were performed with MPO and CD68, labelling neutrophils and macrophages. RESULTS: MMP-2-, MMP-3-, MMP-9- and TIMP-1-positive cells were detected in ANCA-associated glomerulonephritis in glomeruli with active inflammation (cellular crescents or fibrinoid necrosis), only occasionally in normal appearing glomeruli, and not in sclerotic glomeruli and positive cells were found in the tubulo-interstitium. MMPs and TIMP-1 were expressed predominantly by MPO-and CD68-positive cells. In normal renal tissue, no expression was detected, with the exception of weak mesangial staining for MMP-2. In ANCA-associated glomerulonephritis, glomerular MMP-2, -9 and TIMP-1 correlated with glomerular cathepsin G expression, while the number of MMP-9-expressing cells per glomerulus correlated with the percentage of crescentic glomeruli. Tubulo-interstitial expression of MMPs correlated with all markers of neutrophil and monocyte infiltration, and interstitial MMP-9 and TIMP-1 expression correlated with renal function at the time of renal biopsy. CONCLUSIONS: Expression of glomerular and interstitial MMP-2, -3, -9 and TIMP-1 is increased in active ANCA-associated glomerulonephritis and correlates with inflammatory activity.     

Expression of 90-kDa heat-shock protein within cellular crescents in human diseased kidneys

Summary: We previously reported the induction of 90 kDa heat-shock protein (HSP90) in rat kidneys with cisplatin-induced acute tubular injury, gentamicin-induced acute renal failure and ischaemia-induced acuterenal failure. In the present study, we examined the expression of HSP90 in normal and diseased human kidneys. the 90-kDa heat-shock protein (HSP90) from mouse brains was purified, and a specific antibody against the protein in a rabbit was produced. This antibody cross-reacted with human renal HSP90 on immunoblot analysis. Using this antibody we observed the intrarenal immunohistochemical localization of HSP90 in both normal and various human diseased kidneys. We also examined the differences between the distributions of macrophages and HSP90 in cellular crescents. In the normal kidney, HSP90 was present in glomerular podocytes, Bowman's epithelia and epithelial cells from the distal tubules to the collecting duct. In diseased kidneys HSP90 was markedly expressed in the cytoplasm of proliferative cells within cellular crescents, but not in fibrocellular crescents. the localization of HSP90 in crescents did not coincide with that of macrophages. These results demonstrated that HSP90 is induced in the cytoplasm of cellular crescents. Heat-shock protein 90 may be expressed partly in response to some factors produced by macrophages.     

HIV-1 induces renal epithelial dedifferentiation in a transgenic model of HIV-associated nephropathy

BACKGROUND: Human immunodeficiency virus-associated nephropathy (HIVAN) is the most common cause of renal failure in HIV-1-seropositive patients. Recent studies using an HIV-1 transgenic mouse model have demonstrated that expression of HIV-1 in the kidney is required for the development of HIVAN. What has remained unclear, however, is the renal cell type responsible for pathogenesis and the essential pathological process. METHODS: To address these issues, we used a transgenic murine model of HIVAN. We identified the cell types in kidney in which HIV transgene expression occurs using in situ hybridization. We evaluated evidence of proliferation by immunocytochemical analysis using an antibody to Ki-67 and cell type-specific markers, including WT-1, synaptopodin, Na+,K+-ATPase, adducin, and desmin. TUNEL assay was used to evaluate apoptosis. RESULTS: We found that glomerular and tubular epithelial cells express the HIV-1 transgene early in the disease process when renal architecture is well preserved. Transgene expression is lost, however, in tubular epithelial cells when they lose their differentiated cuboidal phenotype. In glomerular epithelial cells, dedifferentiation occurs with reduced expression of WT-1 and synaptopodin, in association with activation of desmin expression. Tubular microcysts also form with mislocalization of Na+,K+-ATPase expression to the lateral and apical cellular membranes. CONCLUSIONS: These studies support the hypothesis that the glomerular and renal epithelial cells are the primary targets of HIV-1 pathogenesis in the kidney. The essential pathologic process is dysregulation of the epithelial cell cycle with increased proliferation, apoptosis, cellular dedifferentiation, and altered cellular polarity.