Generation of Antigen Microarrays to Screen for Autoantibodies in Heart Failure and Heart Transplantation (original) (raw)
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Profiling non-HLA antibody responses in antibody-mediated rejection following heart transplantation
American Journal of Transplantation, 2020
Antibody-mediated rejection (AMR) driven by the development of donor-specific antibodies (DSA) directed against mismatched donor HLA is a major risk factor for graft loss in cardiac transplantation. Recently, the relevance of non-HLA antibodies has become more prominent as AMR can be diagnosed in the absence of circulating DSA. Here, we assessed a single-center cohort of 64 orthotopic heart transplant recipients transplanted between 1994 and 2014. Serum collected from patients with ≥pAMR1 (n=43) and non-AMR (n=21) were tested for reactivity against a panel of 44 non-HLA autoantigens. The AMR group had a significantly greater percentage of patients with elevated reactivity to autoantigens compared to non-AMR (p=0.002) and healthy controls (n=94, p<0.0001). DSA-positive AMR patients exhibited greater reactivity to autoantigens compared to DSA-negative (p<0.0001) and AMR patients with DSA and PRA>10% were identified as the subgroup with significantly elevated responses. Four antigens, vimentin, tubulin alpha 1B, lamin A/C and apolipoprotein L2, were significantly different between AMR and non-AMR. Moreover, increased reactivity to these antigens was associated with graft failure. These results suggest that antibodies to non-HLA are associated with DSA-positive AMR although their specific role in mediating allograft injury is not yet understood.
American Journal of Transplantation, 2020
We analyzed humoral immune responses to non-HLA antigens after cardiac transplantation to identify antibodies associated with allograft rejection. Protein microarray identified 366 non-HLA antibodies (>1.5 fold, p<0.5) from a discovery cohort of HLA antibody negative, endothelial cell crossmatch positive sera obtained from twelve cardiac allograft recipients at the time of biopsy proven rejection. From these, 19 plasma membrane proteins and 10 autoantigens identified from gene ontology analysis were combined with 48 proteins identified through literature search to generate a multiplex bead array. Longitudinal sera from a multicenter cohort of adult cardiac allograft recipients (samples: n=477 no rejection; n=69 rejection) identified 18 non-HLA antibodies associated with rejection (p<0.1) including 4 newly-identified non-HLA antigenic targets (DEXI, EMCN, LPHN1 and SSB). CART analysis showed 5/18 non-HLA antibodies distinguished rejection vs. non-rejection. Antibodies to 4/18 non-HLA antigens synergize with HLA DSA and significantly increase the odds of rejection (p<0.1). The non-HLA panel was validated using an independent adult cardiac transplant cohort (n=21 no rejection; n=42 rejection, >1R) with an area under the curve (AUC) of .87 (p<0.05) with 92.86% sensitivity and 66.67% specificity. We conclude that multiplex bead array assessment of non-HLA antibodies identifies cardiac transplant recipients at risk of rejection.
The Journal of Heart and Lung Transplantation, 2011
BACKGROUND: Recent efforts are being undertaken to update and refine current diagnostic criteria for antibody-mediated rejection (AMR) in heart transplantation. We believe that the appropriate reactants are those that best predict the adverse consequences of AMR and therefore tested various models using different reactants to find the best predictors of cardiovascular mortality in pathologically defined AMR.
Antibodies aggravate the development of ischemic heart failure
American journal of physiology. Heart and circulatory physiology, 2018
Heart-specific antibodies have been widely associated with myocardial infarction (MI). However, it remains unclear whether autoantibodies mediate disease progression or are a byproduct of cardiac injury. To disambiguate the role of immunoglobulins in MI, we characterized the development of ischemic heart failure (HF) in agammaglobulinemic mice (AIDμS). While these animals can produce functional B-cells, they cannot synthesize secretory IgM (μS) or perform immunoglobulin class-switching (AID), leading to complete antibody deficiency. Agammaglobulinemia did not affect overall post-MI survival but resulted in a significant reduction in infarct size. Echocardiographic analyses showed that compared to the WT infarcted controls, the AIDμS mice exhibited improved cardiac function and reduced remodeling at day 56 post-MI. These differences remained significant even after animals with matched infarct sizes were compared. Infarcted AIDμS mice also showed reduced myocardial expression levels o...
Transplant International, 2004
Autoimmune mechanisms play an important role in the pathogenesis of allograft vasculopathy following heart transplantation, but the autoantigens involved have been only sparsely studied. Citrate synthase (CS) enzyme is a conserved molecule, and, as an important mitochondria1 autoantigen, it is protected by the "immunological homunculus". Tissue destruction and alteration of the immune regulatory mechanisms can induce pathological immune response against CS in other autoimmune diseases. In our present study we aimed to detect CS-specific autoantibodies in heart transplant patients, therefore, prospective, randomised clinical tests were conducted on 33 heart transplant patients and compared with -1 30 healthy blood donors. The level and isotype of CS antibodies were detected by simple binding indirect enzyme-linked immunosorbent assay (ELISA). The epitope specifici-ties of the autoantibodies were measured on synthetic overlapping peptide sequences of CS enzyme by an indirect multi-pin ELISA method. Mainly IgM isotype CS autoantibodies were found in healthy controls, while IgG was found at higher levels and frequency (fourtimes higher) in heart transplant patients. Autoantibodies of IgG isotype recognise different epitopes than do autoantibodies of IgM isotype, even within the same group and individual. New epitope-specific IgG and IgM isotype autoantibodies appeared in heart transplant patients when compared with the controls. Our findings suggest a possible role of CS-specific autoantibodies in the pathomechanism of allograft vasculopathy .
Circulation, 2002
Background-Immunoadsorption capable of removing circulating autoantibodies represents an additional therapeutic approach in dilated cardiomyopathy (DCM). The role played by autoantibodies belonging to the immunoglobulin (Ig) subclass G-3 in cardiac dysfunction remains to be elucidated. Methods and Results-Patients with DCM (left ventricular ejection fraction Ͻ30%) participated in this case-control study. Nine patients underwent immunoadsorption with protein A (low affinity to IgG-3), and 9 patients were treated with anti-IgG, which removes all IgG subclasses. Immunoadsorption was performed in 4 courses at 1-month intervals until month 3. In the 2 groups, immunoadsorption induced comparable reduction of total IgG (Ͼ80%). IgG-3 was effectively eliminated only by anti-IgG adsorption (eg, during the first immunoadsorption course; protein A, Ϫ37Ϯ4%; anti-IgG, Ϫ89Ϯ3%; PϽ0.001 versus protein A). The  1-receptor autoantibody was effectively reduced only by anti-IgG (PϽ0.01 versus protein A). Hemodynamics did not change in the protein A group. In the anti-IgG group during the first immunoadsorption course, cardiac index increased from 2.3Ϯ0.1 to 3.0Ϯ0.1 L • min Ϫ1 • m Ϫ2 (PϽ0.01 versus protein A). After 3 months, before the last immunoadsorption course, cardiac index was 2.2Ϯ0.1 L • min Ϫ1 • m Ϫ2 in the protein A group and 3.0Ϯ0.2 L • min Ϫ1 • m Ϫ2 in the anti-IgG group (PϽ0.01 versus protein A). Left ventricular ejection fraction increased only in the anti-IgG group (PϽ0.05 versus protein A). Conclusions-Autoantibodies belonging to IgG-3 may play an important role in cardiac dysfunction of DCM. The removal of antibodies of the IgG-3 subclass may represent an essential mechanism of immunoadsorption in DCM. (Circulation.