Significant re-emergence and recirculation of Schmallenberg virus in previously exposed dairy herds in Ireland in 2016.

Schmallenberg virus (SBV) circulation was investigated in 25 previously exposed dairy herds in Ireland in 2016. A population of 1,550 spring-2014-born animals, which had been monitored for SBV infection in 2014 and 2015 as part of a previous SBV surveillance study, were resampled for evidence of SBV infection during 2016. A total of 366 blood samples were collected in the 25 study herds (15 samples per herd) between 3 March 2017 and 10 March 2017 (before the 2017 vector-active season) and analysed for SBV antibodies using a competitive ELISA kit (IDVet). A total of 256 animals tested seropositive, an AP of 69.9% (95% CI: 65.1-74.4) and TP of 77.7% (95% CI: 72.3%-82.8%) when correcting for imperfect test characteristics. These results demonstrate that a new epidemic of SBV circulation occurred in these previously exposed herds in Ireland in 2016.

Schmallenberg virus: Predicting within-herd seroprevalence using bulk-tank milk antibody titres and exploring individual animal antibody titres using empirical distribution functions (EDF).

Schmallenberg virus (SBV) is transmitted by Culicoides spp. biting midges and can cause abortions and congenital malformations in ruminants and milk drop in dairy cattle. Estimating true within-herd seroprevalence is an essential component of efficient and cost-effective SBV surveillance programs. The objectives of this study were: (1) determine the correlation between bulk-tank milk (BTM)-ELISA results and within-herd seroprevalence, (2) evaluate the ability of BTM-ELISA results to predict within-herd seroprevalence and (3) explore the distributions of individual animal serology results using novel statistical methodology. BTM samples (n=24) and blood samples (n=4019) collected from all lactating cows contributing to the BTM in 26 Irish dairy herds (58-444 cows/herd) in 2014 located in a region exposed to SBV in 2012/2013, were analysed for SBV-specific antibodies using IDVet® ELISA kits. The correlation between BTM-ELISA results and within-herd seroprevalence was determined by calculating Pearson's correlation coefficient. Linear regression models were used to assess the ability of BTM-ELISA results to predict within-herd seroprevalence. The distributions of individual animal serology results were explored by determining the empirical distribution functions (EDF) of the individual animal serum ELISA results in each herd. EDFs were compared pairwise across herds, using the Kolmogorov-Smirnov statistical test. Herds with similar BTM-ELISA results, herds with similar within-herd seroprevalence and herds with similar mean-herd serology ELISA results were stratified in order to explore their respective paired-herd EDF comparisons. Statistical significance was set at p<0.05. Twenty-two herds were BTM-ELISA-positive (within-herd seroprevalence 30.6-100%) and two herds were BTM-ELISA-negative (within-herd seroprevalence 10.7 and 16.2%) indicating BTM-ELISA-negative herds can have seropositive animals present. BTM-ELISA results were highly correlated (r=0.807, p<0.0001) with, and predictive of (R2=0.832, p<0.0001) of within-herd seroprevalence. Predictions were most accurate for upper-range BTM-ELISA antibody titres, while they were less accurate at higher and lower antibody titres. This is likely a result of the overall high within-herd seroprevalence. In herds with similar BTM-ELISA results 82% of the paired-herd EDF comparisons were significantly different. In herds with similar within-herd seroprevalence and in herds with similar mean-herd serology ELISA results, 46% and 47% of the paired-herd EDF comparisons were significantly different, respectively. These results demonstrate that BTM antibody titres are highly predictive of within-herd seroprevalence in an SBV exposed region. Furthermore, exploring the serum EDFs revealed that the variation observed in the predicted within-herd seroprevalence in the regression models is likely a result of individual animal variation in serum antibody titres in these herds.

Post-epidemic Schmallenberg virus circulation: parallel bovine serological and Culicoides virological surveillance studies in Ireland.

BACKGROUND: Schmallenberg virus (SBV) emerged in northern-Europe in 2011 resulting in an epidemic of ruminant abortions and congenital malformations throughout the continent. In the years following the epidemic there have been reports of SBV overwintering and continued circulation in several European countries. When the population-level of immunity declines in exposed regions, re-introduction of SBV could result in further outbreaks of Schmallenberg disease. The aims of this study were to determine the SBV seroprevalence in previously exposed Irish dairy herds in 2014 and to investigate if SBV continued to circulate in these herds in the three years (2013-2015) following the Irish Schmallenberg epidemic. Whole-herd SBV serosurveillance was conducted in 26 herds before (spring) and following the 2014 vector-season (winter), and following the 2015 vector-season (winter). In spring 2014, 5,531 blood samples were collected from 4,070 cows and 1,461 heifers. In winter 2014, 2,483 blood samples were collected from 1,550 youngstock (8-10 months old) and a subsample (n = 933; 288 cows, 645 heifers) of the seronegative animals identified in the spring. Youngstock were resampled in winter 2015. Culicoides spp. were collected in 10 herds during the 2014 vector-season and analysed for SBV; a total of 138 pools (3,048 Culicoides) from 6 SBV vector species were tested for SBV RNA using real-time PCR. RESULTS: In spring 2014, animal-level seroprevalence was 62.5 % (cows = 84.7 %; heifers = 0.6 %). Within-herd seroprevalence ranged widely from 8.5 %-84.1 % in the 26 herds. In winter 2014, 22 animals (0.9 %; 10 cows, 5 heifers, 7 youngstock) originating in 17 herds (range 1-4 animals/herd) tested seropositive. In winter 2015 all youngstock, including the 7 seropositive animals in winter 2014, tested seronegative suggesting their initial positive result was due to persistence of maternal antibodies. All of the Culicoides pools examined tested negative for SBV-RNA. CONCLUSIONS: SBV appears to have recirculated at a very low level in these herds during 2013 and 2014, while there was no evidence of SBV infection in naïve youngstock during 2015. A large population of naïve animals was identified and may be at risk of infection in future years should SBV re-emerge and recirculate as it has done in continental Europe.

Meaningful engagement of people living with HIV who use drugs: methodology for the design of a Peer Research Associate (PRA) hiring model.

BACKGROUND: Community-based HIV, harm reduction, and addiction research increasingly involve members of affected communities as Peer Research Associates (PRAs)-individuals with common experiences to the participant population (e.g. people who use drugs, people living with HIV [PLHIV]). However, there is a paucity of literature detailing the operationalization of PRA hiring and thus limited understanding regarding how affected communities can be meaningfully involved through low-barrier engagement in paid positions within community-based participatory research (CBPR) projects. We aim to address this gap by describing a low-threshold PRA hiring process. RESULTS: In 2012, the BC Centre for Excellence in HIV/AIDS and the Dr. Peter AIDS Foundation collaborated to develop a mixed-method CBPR project evaluating the effectiveness of the Dr. Peter Centre (DPC)-an integrative HIV care facility in Vancouver, Canada. A primary objective of the study was to assess the impact of DPC services among clients who have a history of illicit drug use. In keeping with CBPR principles, affected populations, community-based organizations, and key stakeholders guided the development and dissemination of a low-barrier PRA hiring process to meaningfully engage affected communities (e.g. PLHIV who have a history of illicit drug use) in all aspects of the research project. The hiring model was implemented in a number of stages, including (1) the establishment of a hiring team; (2) the development and dissemination of the job posting; (3) interviewing applicants; and (4) the selection of participants. The hiring model presented in this paper demonstrates the benefits of hiring vulnerable PLHIV who use drugs as PRAs in community-based research. CONCLUSIONS: The provision of low-barrier access to meaningful research employment described herein attempts to engage affected communities beyond tokenistic involvement in research. Our hiring model was successful at engaging five PRAs over a 2-year period and fostered opportunities for future paid employment or volunteer opportunities through ongoing collaboration between PRAs and a diverse range of stakeholders working in HIV/AIDS and addictions. Additionally, this model has the potential to be used across a range of studies and community-based settings interested in meaningfully engaging communities in all stages of the research process.

Chronic humoral rejection of human kidney allografts is associated with MMP-2 accumulation in podocytes and its release in the urine.

Chronic humoral rejection (CHR) is an important cause of late graft failures following kidney transplantation. Overall, the pathophysiology of CHR is poorly understood. Matrix metalloproteinase-2 (MMP-2), a type IV collagenase, has been implicated in chronic kidney disease and allograft rejection in previous studies. We examined the presence of MMP-2 in allograft biopsies and in the urine of kidney transplant recipients with CHR. MMP-2 staining was detected by immunohistochemistry in podocytes for all CHR patients but less frequently in patients with other renal complications. Urinary MMP-2 levels were also significantly higher in CHR patients (median 4942 pg/mL, N = 27) compared to non-CHR patients (median 598 pg/mL, N = 65; p < 0.001). Elevated urinary MMP-2 correlated with higher levels of proteinuria in both CHR and non-CHR patients. Longitudinal analysis indicated that increase in urine MMP-2 coincided with initial diagnosis of CHR as documented by the biopsies. Using an enzymatic assay, we demonstrated that MMP-2 was present in its active form in the urine of patients with CHR. Overall, our findings associate MMP-2 with glomerular injury as well as interstitial fibrosis and tubular atrophy observed in patients with CHR.

Banff 07 classification of renal allograft pathology: updates and future directions.

The 9th Banff Conference on Allograft Pathology was held in La Coruna, Spain on June 23-29, 2007. A total of 235 pathologists, clinicians and scientists met to address unsolved issues in transplantation and adapt the Banff schema for renal allograft rejection in response to emerging data and technologies. The outcome of the consensus discussions on renal pathology is provided in this article. Major updates from the 2007 Banff Conference were: inclusion of peritubular capillaritis grading, C4d scoring, interpretation of C4d deposition without morphological evidence of active rejection, application of the Banff criteria to zero-time and protocol biopsies and introduction of a new scoring for total interstitial inflammation (ti-score). In addition, emerging research data led to the establishment of collaborative working groups addressing issues like isolated 'v' lesion and incorporation of omics-technologies, paving the way for future combination of graft biopsy and molecular parameters within the Banff process.

Banff '05 Meeting Report: differential diagnosis of chronic allograft injury and elimination of chronic allograft nephropathy ('CAN').

The 8th Banff Conference on Allograft Pathology was held in Edmonton, Canada, 15-21 July 2005. Major outcomes included the elimination of the non-specific term "chronic allograft nephropathy" (CAN) from the Banff classification for kidney allograft pathology, and the recognition of the entity of chronic antibody-mediated rejection. Participation of B cells in allograft rejection and genomics markers of rejection were also major subjects addressed by the conference.

The normal kidney filters nephrotic levels of albumin retrieved by proximal tubule cells: retrieval is disrupted in nephrotic states.

The origin of albuminuria remains controversial owing to difficulties in quantifying the actual amount of albumin filtered by the kidney. Here we use fluorescently labeled albumin, together with the powerful technique of intravital 2-photon microscopy to show that renal albumin filtration in non-proteinuric rats is approximately 50 times greater than previously measured and is followed by rapid endocytosis into proximal tubule cells (PTCs). The endocytosed albumin appears to undergo transcytosis in large vesicles (500 nm in diameter), identified by immunogold staining of endogenous albumin by electron microscopy, to the basolateral membrane where the albumin is disgorged back to the peritubular blood supply. In nephrotic rats, the rate of uptake of albumin by the proximal tubule (PT) is decreased. This is consistent with reduced expression of clathrin, megalin, and vacuolar H(+)-ATPase A subunit, proteins that are critical components of the PT endocytotic machinery. These findings strongly support the paradigm-shifting concept that the glomerular filter normally leaks albumin at nephrotic levels. Albuminuria does not occur as this filtered albumin load is avidly bound and retrieved by PTCs. Dysfunction of this retrieval pathway leads to albuminuria. Thus, restoration of the defective endocytotic and processing function of PT epithelial cells might represent an effective strategy to limit urinary albumin loss, at least in some types of nephrotic syndrome.

Putative antibody-mediated rejection with C4d deposition in HLA-identical, ABO-compatible renal allografts.

We sought evidence for non-MHC antibody-mediated rejection in renal allografts by a systematic study of rejected HLA-identical sibling renal allografts. Among 162 recipients of HLA-identical, ABO-compatible sibling donor kidneys transplanted at the Massachusetts General Hospital from 1964 to 2005, we identified 15 grafts that were lost from rejection and two additional grafts with reversible acute rejection, which provided 30 samples for study. All samples were stained for C4d by immunofluorescence in frozen tissue (n = 7) or by immunohistochemistry in paraffin embedded tissues (n = 10). We found that two of 17 grafts had positive C4d staining of peritubular capillaries. Histology revealed acute antibody-mediated rejection in one and acute cellular rejection type 1 in the other. Both grafts were matched at HLA-A, B, and C loci and had a nonreactive mixed lymphocyte response. Genotyping and serological analysis were not available. Compared with a published series, C4d+ irreversible rejection was more common in HLA nonidentical than HLA-identical grafts (75% vs 6.7%, respectively, P < .002). We conclude that antibody-mediated rejection, presumably due to non-MHC antigens other than ABO-blood groups does occur, but infrequently. This may account for some of the HLA antibody negative cases that develop antibody-mediated rejection.

Control of antidonor antibody production with tacrolimus and mycophenolate mofetil in renal allograft recipients with chronic rejection.

BACKGROUND: In renal transplantation, chronic rejection is a major cause of late allograft loss. Recent studies indicate that a subset of chronic rejection is associated with anti-HLA donor specific antibodies (DSA) and complement C4d deposition in peritubular capillaries (PTC). Since rescue therapy with tacrolimus and mycophenolate mofetil has been found to limit antidonor B-cell responses in recipients with acute humoral rejection, we sought to determine whether a similar immunosuppressive regimen might be effective in patients with 'chronic humoral rejection'. METHODS: Four renal allograft recipients with 'chronic humoral rejection' were prospectively identified. The diagnosis was based on: (1) progressive rise in serum creatinine over 12 months; (2) typical pathologic features by light microscopy (transplant arteriopathy and glomerulopathy); (3) widespread C4d deposits in PTC by immunofluorescence; (4) detection of 'de novo' DSA at the time of biopsy. Maintenance immunosuppression was CsA, prednisone and azathioprine (n=3) or prednisone and azathioprine (n=1). Rescue therapy with tacrolimus and mycophenolate mofetil was initiated in all patients, 12 hr after cyclosporine and azathioprine discontinuation. RESULTS: At diagnosis, the mean serum creatinine was 3.9 mg/dl (range: 3.3 to 5.4 mg/dl). DSA was an IgG directed against HLA class II (n=3) or class I (n=2), that is one patient had both anti-HLA class I and class II antibodies. Pretreatment antibody titers varied between 1:8 and 1:128. Rescue therapy was associated with a rapid and sustained decrease in antibody titers. In two patients, DSA became undetectable after 9 months and a repeat biopsy performed after 12 months revealed a decrease in C4d deposition in PTC. CONCLUSION: These results suggest that a decrease in DSA production can be induced in renal allograft recipients with 'chronic humoral rejection' by using an immunosuppressive regimen that combines tacrolimus and mycophenolate mofetil. Limitation of antidonor antibody synthesis may be important for the treatment or the prevention of chronic rejection in organ transplantation.

Acute humoral rejection in renal allograft recipients: I. Incidence, serology and clinical characteristics.

BACKGROUND: Acute rejection (AR) associated with de novo production of donor-specific antibodies (DSA) is a clinicopathological entity that carries a poor prognosis (acute humoral rejection, AHR). The aim of this study was to determine the incidence and clinical characteristics of AHR in renal allograft recipients, and to further analyze the antibodies involved. METHODS: During a 4-year period, 232 renal transplants (Tx) were performed at our institution. Assays for DSA included T and B cell cytotoxic and/or flow cytometric cross-matches and cytotoxic antibody screens (PRA). C4d complement staining was performed on frozen biopsy tissue. RESULTS: A total of 81 patients (35%) suffered at least one episode of AR within the first 3 months: 51 had steroid-insensitive AR whereas the remaining 30 had steroid-sensitive AR. No DSA were found in patients with steroid-sensitive AR. In contrast, circulating DSA were found in 19/51 patients (37%) with steroid-insensitive AR, and widespread C4d deposits in peritubular capillaries were present in 18 of these 19 (95%). In at least three cases, antibodies were against donor HLA class II antigens. DSA were not found in the remaining 32 patients but C4d staining was positive in 2 of 32. The DSA/C4d positive (n=18) and DSA/C4d negative (n=30) groups differed in pre-Tx PRA levels, percentage of re-Tx patients, refractoriness to antilymphocyte therapy, and outcome. Plasmapheresis and tacrolimus-mycophenolate mofetil rescue reversed rejection in 9 of 10 recipients with refractory AHR. CONCLUSION: More than one-third of the patients with steroid-insensitive AR had evidence of AHR, often resistant to antilymphocyte therapy. Most cases (95%) with DSA at the time of rejection had widespread C4d deposits in peritubular capillaries, suggesting a pathogenic role of the circulating alloantibody. Combined DSA testing and C4d staining provides a useful approach for the early diagnosis of AHR, a condition that often necessitates a more intensive therapeutic rescue regimen.

Circulating thyroglobulin transcytosed by thyroid cells in complexed with secretory components of its endocytic receptor megalin.

After its endocytosis from the colloid, some thyroglobulin (Tg) is transcytosed intact across thyrocytes, accounting in part for its presence in the circulation. We previously showed that megalin (gp330), an endocytic Tg receptor, mediates apical to basolateral Tg transcytosis. Here we investigated whether a portion of megalin remains combined with Tg after its transcytosis, using studies with cultured thyroid cells and in vivo observations. FRTL-5 cells, a rat thyroid cell line, cultured on filters in dual chambers form tight junctions and exhibit features of polarity, with expression of megalin exclusively on the upper (apical) surface. After the addition of unlabeled Tg to the upper chamber and incubation at 37 C, some Tg was transcytosed intact across FRTL-5 cells into the lower chamber. Two antimegalin ectodomain antibodies precipitated transcytosed Tg in fluids collected from the lower chamber. After the addition of Tg to surface-biotinylated FRTL-5 cells, an anti-Tg antibody and the two antimegalin ectodomain antibodies precipitated high molecular mass biotinylated material in fluids collected from the lower chamber, corresponding to much of the megalin ectodomain, as well as smaller amounts of lower molecular mass material. The results indicate that Tg transcytosed across FRTL-5 cells remains complexed with megalin ectodomain components, which we refer to as megalin secretory components. In aminotriazole-treated rats, which develop increased megalin-mediated Tg transcytosis, antimegalin antibodies precipitated some of the Tg in the serum. Tg was also precipitated by antimegalin antibodies in sera from patients with Graves' disease, in which we found increased megalin expression on the apical surface of thyrocytes. In contrast, in thyroidectomized patients with metastatic papillary thyroid carcinoma, in whom Tg is directly secreted by neoplastic thyroid cells into the circulation rather than transcytosed, serum Tg was not precipitated by antimegalin antibodies. The detection of Tg-megalin complexes may help identify the source of serum Tg in patients with thyroid diseases.

Complement activation in acute humoral renal allograft rejection: diagnostic significance of C4d deposits in peritubular capillaries.

The distinction between acute humoral rejection (AHR) and acute cellular rejection (ACR) in renal allografts is therapeutically important, but pathologically difficult. Since AHR is probably mediated by antibodies to the donor endothelium that activate the classical complement pathway, it was hypothesized that peritubular capillary C4d deposition might distinguish this group. Renal biopsies (n = 16) from 10 patients with AHR who had acute graft dysfunction, neutrophils in peritubular capillaries, and a concurrent positive cross-match were stained for C4d by immunofluorescence. Control biopsies for comparison showed ACR (n = 14), cyclosporin A toxicity (n = 6), or no abnormality (n = 4). Peribiopsy sera were tested for anti-donor HLA antibody. C4d deposited prominently and diffusely in the peritubular capillaries in all AHR biopsies (16 of 16). IgM and/or C3 were also present in 19 and 44%, respectively. With two-color immunofluorescence, C4d was localized in basement membranes (type IV collagen+) and in the endothelium (Ulex europaeus agglutinin-I+). In ACR, no more than trace C4d was found in peritubular capillaries (P < 0.0001 versus AHR), and no patient had anti-donor HLA antibodies (0 of 8); 27% had neutrophils in peritubular capillaries. One of six biopsies with cyclosporin A toxicity had similar C4d deposits, and circulating anti-donor class I antibody was detected. Grafts with AHR were lost (40%) more often than those with ACR (0%; P < 0.02). C4d in peritubular capillary walls distinguishes AHR from ACR, is more specific and sensitive than traditional criteria, and is a potentially valuable adjunct in the diagnosis of graft dysfunction.

Comparison of anti-GBM antibodies in sera with or without ANCA.

An appreciable percentage of patients with serum anti-glomerular basement membrane (anti-GBM) antibodies also have antineutrophil cytoplasmic antibodies (ANCA), against either myeloperoxidase (MPO-ANCA), or proteinase 3 (PR3-ANCA). In sera without ANCA, the anti-GBM antibodies have been shown to react mainly with the noncollagenous domain (NC1) of Type IV collagen, and especially with its alpha 3 chain, alpha 3(IV)NC1. In most sera, the antibodies can be partially blocked by a monoclonal antibody (Mab17) against alpha 3(IV)NC1, suggesting that a limited region is recognized. Although there is evidence that some anti-GBM antibodies that coexist with ANCA react with alpha 3(IV)NC1, extensive analysis of the specificity of such anti-GBM antibodies has not been reported. In the study presented here, sera were analyzed from 332 patients tested both for anti-GBM antibodies and ANCA (MPO or PR3-ANCA) and found to have one or more positive tests. Of the 100 sera with anti-GBM antibodies, 38 also had ANCA-25 with MPO-ANCA (66%), 12 with PR3-ANCA (32%), and one with both (2%). Of the 232 sera with ANCA only, 153 had MPO-ANCA (66%), 75 had PR3-ANCA (32%), and four had both (2%). Sera was also analyzed from 259 other patients who had positive ANCA tests and were not tested for anti-GBM antibodies: 138 had MPO-ANCA (54%), and 121 had PR3-ANCA (46%). The relative frequencies of MPO or PR3-ANCA in patients with coexisting anti-GBM antibodies did not differ significantly from those in all patients with ANCA (P = 0.35). Seventeen sera with anti-GBM antibodies only and 16 sera with anti-GBM antibodies plus ANCA were selected for further studies to compare the specificity of anti-GBM antibodies in sera with or without ANCA. Using enzyme-linked immunosorbent assays (ELISA), all sera in both groups were found to react with the NC1 domain (as a hexamer) of bovine Type IV collagen and with alpha 3 (IV)NC1 monomers. Furthermore, all but six sera also reacted with one or more of the alpha 1, 2, and 4 (IV)NC1 monomers, generally with considerably lower titers. Reactivity to alpha 3(IV)NC1 was partially blocked by Mab17, with comparable degrees of inhibition in both groups. Western blot analysis with the human NC1 domains revealed no differences in reactivity between the two groups. Thus, differences in antigen specificities of anti-GBM antibodies in sera with or without ANCA were not detected. The anti-GBM response in both situations is hypothesized to be driven by the same immunogen, which is probably derived from NC1 domains of endogenous Type IV collagen.

The syndrome of lung hemorrhage and nephritis is usually an ANCA-associated condition.

BACKGROUND: In the absence of evidence of arteritis or Wegener's granulomatosis, the syndrome of lung hemorrhage and nephritis has been commonly associated with anti-glomerular basement membrane (GBM) antibodies. However, it has been increasingly recognized that many cases are associated with antineutrophil cytoplasmic antibodies (ANCAs). OBJECTIVE: To review available clinical and pathologic findings to determine the diseases accounting for lung hemorrhage and nephritis. METHODS: We studied the records of 750 patients from whom serum samples were sent to our laboratory for anti-GBM antibody assays between 1981 and 1993 and found 88 patients with evidence of lung hemorrhage and nephritis. Serum samples were retested, using current methods, for anti-GBM antibodies (against noncollagenous 1 domain of the alpha 3 chain of type IV collagen) and for antibodies to proteinase 3 and myeloperoxidase--the two types of ANCA of diagnostic value. RESULTS: Of 88 patients with evidence of lung hemorrhage and nephritis, 48 had ANCAs, six had anti-GBM antibodies, and seven had both. In 48 patients with ANCAs, the pathologic findings that accounted for the pulmonary renal syndrome were pauci-immune necrotizing and crescentic glomerulonephritis and pulmonary capillaritis. Only eight had convincing evidence (during life) of Wegener's granulomatosis and only one other had documented arteritis. In 27 patients without ANCAs or anti-GBM antibodies, a variety of unrelated renal and pulmonary diseases were found. CONCLUSIONS: The largest group of patients who present with the syndrome of lung hemorrhage and nephritis have ANCAs and not anti-GMB antibodies. Appropriate tests for antibodies to proteinase 3, antibodies to myeloperoxidase, and anti-GBM antibodies provide reliable guides for making a diagnosis in patients with this pulmonary renal syndrome.

Effect of CTLA-4 chimeric protein on rat autoimmune anti-glomerular basement membrane glomerulonephritis.

The interaction of the T cell receptor with the antigen/major histocompatibility class II complex is insufficient to induce optimal T cell activation. Co-stimulatory signals, including those provided by CD28/CTLA-4 on T cells and B7 molecules (B7-1, -2, and -3) on antigen-presenting cells, are also required. CD28-B7 interactions can be blocked by a soluble human CTLA-4 chimeric protein (CTLA4Ig). We tested the effect of administration of CTLA4Ig on experimental anti-glomerular basement membrane (GBM) autoimmune glomerulonephritis in Wistar-Kyoto rats induced by immunization with bovine GBM. The disease is characterized by development of antibody to the alpha 3 chain of type IV collagen (Goodpasture's antigen), deposition of rat IgG in GBM, infiltration of the kidney by T cells and macrophages, severe crescent formation and renal failure leading to death in 5-6 weeks. Animals injected with human CTLA4Ig from day 0 to day 14 or to day 35 had reduced disease severity. Beneficial effects were observed even when injections were begun after the onset of glomerulonephritis on day 14. However, the rats developed antibody to the human CTLA4Ig, associated with reduction in levels of circulating CTLA4Ig. The results provide evidence for CD28/CTLA-4 signaling in rat autoimmune glomerulonephritis, and suggest that more effective inhibition of B7-dependent T cell activation, such as might be achieved with homologous CTLA4Ig, could be useful in the treatment of autoimmune diseases.

Antibodies to intercellular adhesion molecule 1/lymphocyte function-associated antigen 1 prevent crescent formation in rat autoimmune glomerulonephritis.

In patients with glomerulonephritis widespread crescents are associated with a poor prognosis. Crescent formation appears to depend on the migration of mononuclear cells into Bowman's space, and therefore the interaction between leukocytes and glomerular endothelium may be a critical event in the genesis of crescents. We performed the present study to determine the effects of mouse monoclonal antibodies to the adhesion molecules intercellular adhesion molecule 1 (ICAM-1) and lymphocyte function-associated antigen 1 (LFA-1) in a model of crescentic glomerulonephritis in Wistar-Kyoto rats, induced by immunization with bovine glomerular basement membrane (GBM). By 10-14 d after immunization, the rats had developed circulating anti-GBM antibodies, reactive with the alpha 3 chain of type IV collagen (the Goodpasture antigen), accompanied by proteinuria, accumulation of rat immunoglobulin (Ig)G in the GBM, increased expression of ICAM-1 by glomerular endothelial cells, infiltration of glomerular tufts with LFA-1+ T cells and monocyte/macrophages, and early crescents. At 5 wk all rats had diffuse fibrocellular crescents, glomerular sclerosis, and tubulointerstitial damage. All rats developed severe renal insufficiency and died by 5 or 6 wk. The administration of monoclonal antibodies to rat ICAM-1 and LFA-1 markedly decreased the severity of the renal disease. In a group of rats injected three times a week with the monoclonal antibodies, from 2 d before immunization with GBM to day 14, glomerular abnormalities and proteinuria were virtually absent at day 14; even at 5 wk glomerular disease was quite mild, with only slight crescent formation and with only a mild decrease in renal function. When treatment was continued until 5 wk, the beneficial effects were even more marked, with virtual absence of crescents and with preservation of normal renal function. In a group of rats in which treatment was initiated on day 14, shortly after the appearance of glomerular abnormalities, progression of the disease was appreciably retarded, and the decrease in renal function was inhibited. The kidneys of rats treated from days -2 to 14 with antibodies to ICAM-1 and LFA-1 showed bright linear staining for rat IgG along the GBM, which did not differ in intensity from that seen in untreated rats. Furthermore, the titers of anti-GBM antibodies at 2 wk in treated rats were not lower than that seen in most of the untreated rats. There was, however, moderate reduction of anti-GBM antibodies at 5 wk in the treated rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Hyaluronate is a component of crescents in rat autoimmune glomerulonephritis.

BACKGROUND: Crescent formation is a major feature of rapidly progressive glomerulonephritis and is generally associated with a poor prognosis. Crescents are formed by accumulation of monocyte/macrophages and plasma proteins in Bowman's space, by proliferation of parietal epithelial cells and fibroblasts, and by deposition of the extracellular matrix. Interactions of components of the extracellular matrix with surface receptors of inflammatory cells may be important in crescent formation. One such receptor is the glycoprotein, CD44, whose main ligand is hyaluronate. We performed the present study to determine if hyaluronate is a component of crescents in a model of autoimmune anti-glomerular basement membrane nephritis in rats. EXPERIMENTAL DESIGN: Wistar-Kyoto rats were immunized with bovine glomerular basement membrane, that resulted in severe crescentic glomerulonephritis. Sections of renal tissue were studied with two probes to detect hyaluronate: (a) a soluble CD44-human immunoglobulin fusion protein; and (b) a hyaluronic acid-binding protein. Both probes were revealed by immunofluorescence techniques. The specificity of the reactions was established by selective enzymatic digestions. RESULTS: Marked accumulation of hyaluronate was demonstrated in developing and sclerosing crescents, in association with local infiltration of T lymphocytes and monocyte/macrophages, cells known to express CD44. Lesser amounts of hyaluronate were found in periglomerular infiltrates. CONCLUSIONS: Hyaluronate is an abundant extracellular component of crescents, and may play a critical role in their formation, by its effects on migration and activation of CD44+ lymphocytes, monocyte/macrophages, fibroblasts and epithelial cells.

Antineutrophil cytoplasmic autoantibody-associated vasculitis presenting as Sjögren's syndrome.

A 63-year-old woman, in whom a diagnosis of Sjögren's syndrome was initially made, proved to have systemic vasculitis with salivary gland involvement and necrotizing and crescentic glomerulonephritis. Antineutrophil cytoplasmic autoantibodies (ANCA) against myeloperoxidase were positive. ANCA-associated vasculitis should be considered in the differential diagnosis of Sjögren's syndrome. A positive finding on immunoassay for ANCA against myeloperoxidase or proteinase 3 may help establish the diagnosis.

Coexpression of CD15 and CD20 by Reed-Sternberg cells in Hodgkin's disease.

The immunophenotype of the Reed-Sternberg cells in Hodgkin's disease is heterogeneous among different cases; this heterogeneity has contributed to the continuing uncertainty regarding the normal counterpart of the Reed-Sternberg cell. In this study, the authors demonstrate coexpression of the B-cell marker, CD20, and the granulocyte associated antigen, CD15, by Reed-Sternberg cells in three of 20 cases of nodular sclerosis and mixed cellularity Hodgkin's disease using a double-labelling technique in one case and staining of serial sections in three cases. Additionally, the authors found that expression of CD20 occurred more often in tumors with a monomorphous proliferation of mononuclear and binucleate Hodgkin's and Reed-Sternberg cells, without numerous eosinophils or polymorphonuclear neutrophils. In contrast, expression of CD15 by Reed-Sternberg cells was associated with a greater granulocyte infiltrate. The presence or absence of fibrosis, plasma cells, and histiocytes did not correlate with antigen expression. These results suggest that there may be a continuum of antigen expression by Reed-Sternberg cells, with some cells expressing CD20, some CD15, and others expressing both antigens; cells coexpressing both CD15 and CD20 may represent an unstable intermediate in the process of antigen switching. The possibility that antigen expression by the neoplastic cells in a given case may modulate depending on the background infiltrate could explain the heterogeneity of immunophenotype among cases of Hodgkin's disease.