Regulation of cardiac allograft immune responses by microRNA-155.

INTRODUCTION: A better understanding of the immune mechanisms involved in allograft rejection after transplantation is urgently needed to improve patient outcomes. As microRNA-155 (miR155) plays a critical role in inflammation, we postulated that a deficiency of miR155 will improve cardiac allograft survival and enhance tolerance induction after heart transplantation. METHODS: We developed an acute rejection mouse model through heterotopic BALB/c cardiac transplantation to C57BL/6 (wild-type) and C57BL/6 miR155 knock-out (miR155KO) mice. Further, we induced tolerance in both groups through a costimulatory blockade with CTLA4-Ig (200 µg; post-transplant day 2) and MRI antibodies (250 µg; post-transplant day 0), targeting CD28/B7 and CD40/CD154 signals, respectively. Finally, we examined the effects of injecting 100 µg of small extracellular vesicles (sEVs) isolated from wild-type mice undergoing rejection into tolerant miR155KO mice. RESULTS: Mean survival time (MST) of the cardiac allografts in wild-type and miR155KO mice was 7 and 15 days, respectively (p < 0.0001). Costimulatory blockade increased MST to 65 days and > 100 days in the wild-type and miR155KO recipients, respectively (p < 0.001). Injection of sEVs isolated from wild-type mice undergoing rejection into tolerant miR155KO mice decreased the allograft to 9 days, significantly lower than the tolerant miR155KO mice without injection of sEVs (>100 days; p < 0.0001). CONCLUSION: miR155KO mice have improved cardiac allograft survival and enhanced induction of tolerance after heterotopic cardiac transplantation. Injection of sEVs from wild-type mice undergoing rejection into the miR155KO mice reversed these benefits.

Enhanced enrichment of extracellular vesicles for laboratory and clinical research from drop-sized blood samples.

In the realm of biomedical advancement, extracellular vesicles (EVs) are revolutionizing our capacity to diagnose, monitor, and predict disease progression. However, the comprehensive exploration and clinical application of EVs face significant limitations due to the current isolation techniques. The size exclusion chromatography, commercial precipitation reagents, and ultracentrifugation are frequently employed, necessitating skilled operators and entailing challenges related to consistency, reproducibility, quality, and yields. Notably, the formidable challenge of extracellular vesicle isolation persists when dealing with clinical samples of limited availability. This study addresses these challenges by aiming to devise a rapid, user-friendly, and high-recovery EVs isolation technique tailored for blood samples. The NTI-EXO precipitation method demonstrated a 5-fold increase in the recovery of serum EVs compared to current methodologies. Importantly, we illustrate that a mere two drops of blood (∼100 µL) suffice for the recovery of enriched EVs. The integrity and quality of these isolated EVs were rigorously assessed for the size, purity, and contaminants. This method was validated through the successful isolation of EVs from organ transplant recipients to detect disease-specific exosomal markers, including LKB1, SARS-CoV-2 spike protein, and PD-L1. In conclusion, NTI-EXO method can be used for small clinical samples, thereby advancing discoveries in the EV-centric domain and propelling the frontiers of biomedical research and clinical applications.

Immune responses of lung transplant recipients against SARS-CoV-2 and common respiratory coronaviruses: Evidence for pre-existing cross-reactive immunity.

Humoral and cellular immune responses to SARS-CoV-2 and other coronaviruses in lung transplant recipients are unknown. We measured antibodies and T cell responses against the SARS-CoV-2 spike S2 and nucleocapsid antigens and spike antigens from common respiratory coronaviruses (229E, NL63, OC43, and HKU1) after vaccination or infection of LTxRs. 148 LTxRs from single center were included in this study: 98 after vaccination and 50 following SARS-CoV-2 infection. Antibodies were quantified by enzyme-linked immunosorbent assay. The frequency of T cells secreting IL2, IL4, IL10, IL17, TNFα, and IFNγ were enumerated by enzyme-linked immunospot assay. Our results have shown the development of antibodies to SARS-CoV-2 spike protein in infected LTxRs (39/50) and vaccinated LTxRs (52/98). Vaccinated LTxRs had higher number of T cells producing TNFα but less cells producing IFNγ than infected LTxRs in response to the nucleocapsid antigen and other coronavirus spike antigens. We didn't find correlation between the development of antibodies and cellular immune responses against the SARS-CoV-2 spike protein after vaccination. Instead, LTxRs have pre-existing cellular immunity to common respiratory coronaviruses, leading to cross-reactive immunity against SARS-CoV-2 which likely will provide protection against SARS-Cov-2 infection.

Downregulation of a tumor suppressor gene LKB1 in lung transplantation as a biomarker for chronic murine lung allograft rejection.

BACKGROUND: We recently demonstrated decreased tumor suppressor gene liver kinase B1 (LKB1) level in lung transplant recipients diagnosed with bronchiolitis obliterans syndrome. STE20-related adaptor alpha (STRADα) functions as a pseudokinase that binds and regulates LKB1 activity. METHODS: A murine model of chronic lung allograft rejection in which a single lung from a B6D2F1 mouse was orthotopically transplanted into a DBA/2J mouse was employed. We examined the effect of LKB1 knockdown using CRISPR-CAS9 in vitro culture system. RESULTS: Significant downregulation of LKB1 and STRADα expression was found in donor lung compared to recipient lung. STRADα knockdown significantly inhibited LKB1, pAMPK expression but induced phosphorylated mammalian target of rapamycin (mTOR), fibronectin, and Collagen-I, expression in BEAS-2B cells. LKB1 overexpression decreased fibronectin, Collagen-I, and phosphorylated mTOR expression in A549 cells. CONCLUSIONS: We demonstrated that downregulation of LKB1-STRADα pathway accompanied with increased fibrosis, results in development of chronic rejection following murine lung transplantation.

Extracellular vesicles: a potential new player in antibody-mediated rejection in lung allograft recipients.

Identification of recipients with pre-existing antibodies and cross-matching of recipient sera with donor lymphocytes have reduced the incidence of antibody-mediated rejection (AMR) after human lung transplantation. However, AMR is still common and requires not only immediate intervention but also has long-term consequences including an increased risk of chronic lung allograft dysfunction (CLAD). The mechanisms resulting in AMR remain largely unknown due to the variation in clinical and histopathological features among lung transplant recipients; however, several reports have demonstrated a strong association between the development of antibodies against mismatched donor human leucocyte antigens [donor-specific antibodies (DSAs)] and AMR. In addition, the development of antibodies against lung self-antigens (K alpha1 tubulin and collagen V) also plays a vital role in AMR pathogenesis, either alone or in combination with DSAs. In the current article, we will review the existing literature regarding the association of DSAs with AMR, along with clinical diagnostic features and current treatment options for AMR. We will also discuss the role of extracellular vesicles (EVs) in the immune-related pathogenesis of AMR, which can lead to CLAD.

Direct Correlation of Soluble HLA and HLA-Containing Exosomes and Inverse Correlation of Tolerance Marker-Containing Exosomes With Antibody-Mediated Rejection After Simultaneous Liver-Kidney Transplantation: A Case Study.

There is a lower incidence of antibody-mediated rejection (AMR) after simultaneous liver-kidney transplantation (SLKT) than after kidney-only transplantation. It has been suggested that soluble human leukocyte antigen (sHLA) produced by the liver protects the kidney from AMR. However, this hypothesis has not been tested after SLKT. We present a case of SLKT with 2 donor-specific antibodies (DSAs) (DR53, 12,364 mean fluorescence intensity [MFI]; DQ7, 1253 MFI) that displayed a decrease by day 7 (DR53, 2747 MFI; DQ7, 107 MFI). On day 351, the patient was diagnosed with kidney AMR associated with high levels of DSA (DR53, 18,542 MFI; DQ7, 22,007 MFI) that persisted until day 531. High levels of sHLA-DR/DQ and HLA-DR/DQ-containing exosomes were also detected on day 398. Consequently, the patient underwent treatment with plasmapheresis, intravenous immunoglobulin, prednisone, and rituximab. On day 752, biopsy results were negative for AMR. Moderate levels of DSA (DR53, 9798 MFI; DQ7, 1271 MFI), and baseline levels of sHLA-DR/DQ and HLA-DR/DQ-containing exosomes were observed. Increases in CD4+CD25+FOXP3+ regulatory T cell marker-containing exosomes (CD73, programmed death-ligand 1) were observed on day 752 compared to day 398. These data show a direct correlation between sHLA and HLA-containing exosomes and an inverse correlation between tolerance marker-containing exosomes and kidney AMR after SLKT.

Role for exosomes with self-antigens and immune regulatory molecules in allo- and auto-immunity leading to chronic immune injury following murine kidney transplantation.

OBJECTIVE: Antibodies against donor human leukocyte antigen are a risk factor for chronic immune injury (CII) following renal transplantation; however, it is often not detectable. The main goal of this study is to gain new insights into the kinetics of exosome release and content in sensitized vs non-sensitized recipients. Towards this, we investigated the role for circulating exosomes with allo and self-antigens as well as immunoregulatory molecules in the development of CII and acute rejection. METHODS: Using murine kidney allograft rejection models, we investigated the role of exosomes on immune responses leading to allo- and auto-immunity to self-antigens resulting in rejection. Exosomes were analyzed for kidney self-antigens (Collagen-IV, fibronectin, angiotensin II receptor type 1), and immune-regulatory molecules (PD-L1, CD73) using western blot. Antibodies to donor MHC in serum samples were detected by immunofluorescence, self-antigens by enzyme-linked immunosorbent assay and kidney tissue infiltrating cells were determined by immunohistochemistry. RESULTS: BALB/c; H2d to C57BL/6; H2b renal transplantation (BALB/c), resulted in tubulitis and cellular infiltration by day 14, suggestive of acute inflammation, that was self-limiting with functioning graft. This contributed to CII on post-transplant day >100, which was preceded by induction of exosomes with donor and self-antigens leading to antibodies and immune-regulatory molecules. The absence of acute rejection in this allogenic transplant model is likely due to the induction of splenic and, graft-infiltrating CD4 + FoxP3+ T regulatory cells. In contrast, prior sensitization by skin graft followed by kidney transplantation induced antibodies to MHC and self-antigens leading to acute rejection. CONCLUSION: We demonstrate a pivotal role for induction of exosomes with immune-regulatory molecules, allo- and auto-immunity to self-antigens leading to chronic immune injury following murine kidney transplantation.

Circulating exosomes induced by respiratory viral infections in lung transplant recipients activate cellular stress, innate immune pathways and epithelial to mesenchymal transition.

BACKGROUND: Chronic lung transplant rejection occurs in over 50% of lung transplant recipients and mechanism of chronic rejection is unknown. Evaluation of potential mechanism of exosomes from lung transplant recipients diagnosed with respiratory viral infection (RVI) in inducing chronic lung allograft dysfunction (CLAD). METHOD: Exosomes were isolated from lung transplant recipients followed by DNA and RNA isolation from exosomes. Cell signaling mechanisms were studied by co-culturing exosomes with human epithelial cells. Mice were immunized with exosomes and lung homogenates were studied for immune signaling proteins. RESULTS: Exosomes from lung transplant recipients with RVI carry nucleic acids which are capable of inducing innate immune signaling, endoplasmic reticulum stress, and epithelial mesenchymal transition. CONCLUSION: Therefore, we propose that RVI can lead to induction of exosomes that initiate the process leading to CLAD in mice models. These novel findings identified the molecular mechanisms by which RVI increases the risk of CLAD.

Identification of Phospho-Tyrosine Targets as a Strategy for the Treatment of Esophageal Adenocarcinoma Cells.

INTRODUCTION: Esophageal cancer (EC) is an aggressive cancer type that is increasing at a high rate in the US and worldwide. Extensive sequencing of EC specimens has shown that there are no consistent driver mutations that can impact treatment strategies. The goal of this study was to identify activated tyrosine kinase receptors (TKRs) in EC samples as potential targets in the treatment of EC. METHODS: Activated tyrosine kinase receptors were detected using a dot-blot array for human TK receptors. Human esophageal cancer cell lines were transplanted into immunocompromised mice, and tumor xenografts were subjected to tyrosine kinase inhibitors based on the dot-blot array data. RESULTS: Using the OE33 esophageal cancer cell line, we identified activated EGF receptor (EGFR), as well as ErbB2 and ErbB3. Treatment of this cell line with erlotinib, a specific inhibitor of EGFR, did not impact the growth of this tumor cell line. Treating the OE33 cell line with afatinib, a pan-EGFR family inhibitor resulted in the growth inhibition of OE33, indicating that the ErbB2 and ErbB3 receptors were contributing to tumor cell proliferation. Afatinib treatment of mice growing OE33 tumors inhibited growth of the OE33 tumor cells. DISCUSSION: Activated tyrosine kinase receptors were readily detected in both cancer cell lines and human esophageal cancer samples. By identifying the activated receptors and then using the appropriate tyrosine kinase inhibitors, we can block tumor growth in vitro and in animal xenografts. We propose that identifying and targeting activated TKRs can be used as a personalized EC tumor treatment strategy.

Decline in Club Cell Secretory Proteins, Exosomes Induction and Immune Responses to Lung Self-antigens, Kα1 Tubulin and Collagen V, Leading to Chronic Rejection After Human Lung Transplantation.

BACKGROUND: Chronic lung allograft dysfunction (CLAD), is a major hurdle for long-term lung allograft survival after lung transplant and roughly 50% of lung transplant recipients (LTxRs) develop CLAD within 5 years. The mechanisms of CLAD development remain unknown. Donor-specific immune responses to HLA and lung self-antigens (SAgs) are vital to the pathogenesis of CLAD. Reduction in Club cell secretory protein (CCSP) has been reported in bronchoalveolar lavage (BAL) fluid samples from LTxRs with bronchiolitis obliterans syndrome (BOS). CCSP levels in BAL fluid and development of antibodies to lung SAgs in plasma were determined by ELISA. Cytokines in BAL fluid were analyzed by 30-plex Luminex panel. Exosomes from BAL fluid or plasma were analyzed for SAgs, natural killer (NK) cells markers, and cytotoxic molecules. RESULTS: We demonstrate that LTxRs with BOS have lower CCSP levels up to 9 months before BOS diagnosis. LTxRs with antibodies to SAgs 1-year posttransplant also developed DSA (43%) and had lower CCSP. BOS with lower CCSP also induced Interleukin-8 and reduced vascular endothelial growth factor. Exosomes from BOS contained increased SAgs, NK cells markers, and cytotoxic molecules. CONCLUSIONS: We conclude lower CCSP leads to inflammation, pro-inflammatory cytokine production, immune responses to HLA and SAgs, and induction of exosomes. For the first time, we demonstrate that CCSP loss results in exosome release from NK cells capable of stimulating innate and adaptive immunity posttransplant. This increases the risk of BOS, suggesting a role of NK cell exosomes in CLAD development.

Self-reactive antibodies associated with bronchiolitis obliterans syndrome subtype of chronic lung allograft dysfunction.

BACKGROUND: Chronic Lung Allograft Dysfunction (CLAD) remains the major limitation in long term survival after lung transplantation. Our objective is to evaluate for the presence of autoantibodies to self-antigens, which is a pathway along with complex interplay with immune as well as non-immune mechanisms that leads to a fibroproliferative process resulting in CLAD. METHODS: Serum profiles of IgG autoantibodies were evaluated using customized proteomic microarray with 124 antigens. Output from microarray analyzed as antibody scores is correlated with bronchiolitis obliterans (BOS) subtype of CLAD using Mann-Whitney U test or Fisher exact test. Autoantibodies were evaluated for their predictive value for progressive BOS using a Cox proportional hazard model. BOS free survival and overall survival was analyzed using Kaplan-Meier survival analysis. RESULTS: Forty- two patients included in the study are grouped into "stable BOS" and "progressive BOS" for comparisons. Pulmonary fibrosis is the major indication for lung transplantation in our cohort. Progressive BOS group had significantly worse survival (p < 0.005). Sixteen IgG autoantibodies are significantly elevated at baseline in progressive BOS group. Six among them correlated with worse BOS free survival (p < 0.05). In addition, these six IgG autoantibodies remain elevated at three months and one year after lung transplantation. CONCLUSION: Pre-existing IgG autoantibodies correlate with progressive BOS and survival in a single center, small cohort of lung transplant recipients. Further validation with larger sample size, external cohort and confirmation with additional tissue, bronchoalveolar lavage samples are necessary to confirm the preliminary findings in our study.

The role of miRNA-155 in the immunopathogenesis of obliterative airway disease in mice induced by circulating exosomes from human lung transplant recipients with chronic lung allograft dysfunction.

Human lung transplant recipients undergoing rejection induce circulatory exosomes with lung self-antigens (SAgs), K-alpha 1 Tubulin and Collagen V, and immunization of C57BL/6 mice with exosomes induced obliterative airway disease (HEI-OAD). We analyzed whether exosomes with SAgs induced immunity in microRNA-155 knockout mice (miR-155KO), as microRNA-155 is an immune regulator. C57BL/6 and miR-155KO were immunized with exosomes from stable or chronic rejection (bronchiolitis obliterans syndrome (BOS) and on day 30, induction of exosomes, antibodies (Abs) to SAgs and cellular immunity were determined. C57BL/6 immunized with exosomes from BOS developed OAD. These immunized animals also developed Abs to SAgs and increased frequency of SAg-specific IFNγ and IL17- producing cells. In contrast, Abs to SAgs did not develop in miR-155KO and there was reduction in frequency of cells producing IL10. Upregulation of suppressor of cytokine signaling for lung inflammation was also noted resulting in abrogation of induction of exosomes with SAgs OAD.

Pre-existing self-reactive IgA antibodies associated with primary graft dysfunction after lung transplantation.

BACKGROUND: Primary graft Dysfunction (PGD) results in significant mortality and morbidity after lung transplantation (LT). The objective of this study was to evaluate if pre-existing antibodies to self-antigens in sera of LT recipients are associated with PGD. METHODS: The serum profiles of IgG and IgA autoantibodies were analyzed using a customized proteomic microarray bearing 124 autoantigens. Autoantibodies were analyzed using Mann-Whitney U test or Fisher exact test. The association of the autoantibodies with clinical phenotypes and survival was analyzed by Kaplan-Meier Survival Analysis. Receiver operating curve characteristics (ROC) were calculated to evaluate the predictive value of the autoantibodies for PGD. RESULTS: 51 patients were included in this study. Autoantigen microarray analysis on the pre-transplantation samples identified 17 IgA and 3 IgG autoantibodies which were significantly higher in recipients who developed PGD compared to those who did not (adjusted p < .05 and fold change>1.5). 6 IgA Abs were significantly associated with survival. Taken as a panel, an elevation of 6 IgA Abs had significant predictive value for PGD. Area under the curve value for the panel was 0.9413 for PGD with ROC analysis. Notably, 6 of the 17 IgA autoantigen targets are belong to proteoglycan family of extracellular matrix proteins. CONCLUSION: Pre-existing IgG and IgA autoantibodies in LT patients correlate with PGD and with survival in a single center, small cohort of lung transplant recipients. Further validation is needed to confirm the findings in the study.

The role of donor-derived exosomes in lung allograft rejection.

Lung transplant recipients (LTxRs) with acute or chronic rejection release circulating exosomes that mostly originate from donor lung tissue and express mismatched human leucocyte antigens (HLA) and lung-associated self-antigens (SAgs), Collagen-V and K alpha 1 Tubulin. During lung transplant (LTx), donor lungs often undergo injuries that increase the antigenicity of the transplanted organ. 30% of LTxRs also have pre-transplant antibodies (Abs) to HLA and lung SAgs, which may induce conditions that increase the risk of chronic lung allograft dysfunction (CLAD). Post-transplant, some recipients experience de novo development of Abs to mismatched donor HLA (donor-specific antibody [DSA]) and Abs to lung SAgs, which have been implicated in CLAD pathogenesis. Because most LTxRs who develop DSA also develop Abs to SAgs, some have suggested a synergistic relationship between alloimmunity and autoimmunity in CLAD immunopathogenesis. These processes likely occur from stress-induced exosome release. Exosomes carry allo-antigens, lung SAgs, several micro RNAs, proteasome, co-stimulatory molecules, and pro-inflammatory transcription factors-resulting in efficient antigen presentation by direct, semidirect, and indirect pathways, leading to immune responses to both allo-antigens and lung-associated SAgs. This review summarizes recent findings on the role of exosomes, and processes triggering immune responses to allo-antigens and lung SAgs that ultimately culminate in CLAD.

Molecular events contributing to successful pediatric cardiac transplantation in HLA sensitized recipients.

Antibodies to HLA resulting in positive cytotoxicity crossmatch are generally considered a contraindication for cardiac transplantation. However, cardiac transplantations have been performed in children by reducing the Abs and modifying immunosuppression. To identify mechanisms leading to allograft acceptance in the presence of Abs to donor HLA, we analyzed priming events in endothelial cells (EC) by incubating with sera containing low levels of anti-HLA followed by saturating concentration of anti-HLA. Pre-transplant sera were obtained from children with low levels of Abs to HLA who underwent transplantation. EC were selected for donor HLA and exposed to sera for 72 h (priming), followed by saturating concentrations of anti-HLA (challenge). Priming of EC with sera induced the phosphatidylinositol 3-kinase/Akt mediated by the BMP4/WNT pathway and subsequent challenge with panel reactive antibody sera increased survival genes Bcl2 and Heme oxygenase-1, decreased adhesion molecules, induced complement inhibitory proteins and reduced pro-inflammatory cytokines. In contrast, EC which did not express donor HLA showed decreased anti-apoptotic genes. Primed EC, upon challenge with anti-HLA, results in increased survival genes, decreased adhesion molecules, induction of complement inhibitory proteins, and downregulation of pro-inflammatory cytokines which may result in accommodation of pediatric cardiac allografts despite HLA sensitization.

The role of exosomes in allograft immunity.

Extracellular vesicles are emerging as potent vehicles of intercellular communication. In this review, we focus on a subclass of extracellular vesicles called exosomes. Previously considered an unimportant catch-all, exosomes have recently been recognized for their role in various diseases and their potential for therapeutic use. We have examined the role of exosomes after human lung transplantation and have delineated the composition of circulating exosomes isolated from lung transplant recipients diagnosed with acute and chronic rejection, primary graft dysfunction, and respiratory viral infection. The presence of lung-associated self-antigens (K-alpha 1 Tubulin and collagen V) and mismatched donor HLA in exosomes isolated from lung transplant recipients signifies that these exosomes originated in the transplanted lungs, and therefore dramatically affect transplant biology and immune pathways. Exosomes released from transplanted organs also carry other proteins, costimulatory molecules, and nucleic acids. Therefore, these molecules may be used as biomarkers for allograft rejection and immunity.

Tissue-associated self-antigens containing exosomes: Role in allograft rejection.

Exosomes are extracellular vesicles that express self-antigens (SAgs) and donor human leukocyte antigens. Tissue-specific exosomes can be detected in the circulation following lung, heart, kidney and islet cell transplantations. We collected serum samples from patients who had undergone lung (n = 30), heart (n = 8), or kidney (n = 15) transplantations to isolate circulating exosomes. Exosome purity was analyzed by Western blot, using CD9 exosome-specific markers. Tissue-associated lung SAgs, collagen V (Col-V) and K-alpha 1 tubulin (Kα1T), heart SAgs, myosin and vimentin, and kidney SAgs, fibronectin and collagen IV (Col-IV), were identified using western blot. Lung transplant recipients diagnosed with bronchiolitis obliterans syndrome had exosomes with higher expression of Col-V (4.2-fold) and Kα1T (37.1-fold) than stable. Exosomes isolated from heart transplant recipients diagnosed with coronary artery vasculopathy had a 3.9-fold increase in myosin and a 4.7-fold increase in vimentin compared with stable. Further, Kidney transplant recipients diagnosed with transplant glomerulopathy had circulating exosomes with a 2-fold increased expression of fibronectin and 2.5-fold increase in Col-IV compared with stable. We conclude that circulating exosomes with tissue associated SAgs have the potential to be a noninvasive biomarker for allograft rejection.

Exosomes expressing the self-antigens myosin and vimentin play an important role in syngeneic cardiac transplant rejection induced by antibodies to cardiac myosin.

Long-term success of heart transplantation is hindered by humoral and cell-mediated immune responses. We studied preexisting antibodies to cardiac self-antigens, myosin and vimentin, and exosomes induced by antibodies to self-antigens in eliciting immune responses to cardiac grafts. After syngeneic heterotopic murine heart transplantation, rabbit anti-myosin or normal rabbit immunoglobulin was administered at day 0 or 7. Sera were collected after heartbeat cessation, cellular infiltration was analyzed, and exosomes were isolated from sera. Histopathologic examination of the controls' transplanted hearts demonstrated normal architecture, and their sera demonstrated neither antibodies to self-antigens nor exosomes expressing self-antigens. Administration of antibodies to cardiac myosin immediately posttransplantation (day 0) but not on day 7 triggered graft failure on day 7, and histopathologic examination revealed marked cellular infiltration with neutrophils and lymphocytes. Histopathologic examination of rejected hearts also demonstrated myocyte damage as sera had increased antibodies to myosin and vimentin and development of exosomes expressing self-antigens. Administration of exosomes isolated from failed grafts containing self-antigens induced graft dysfunction; exosomes isolated from stable mice did not induce graft failure. Antibodies to self-antigens can induce exosomes containing self-antigens, initiating an immune response and causing graft failure after cardiac transplantation.

The role of C4d deposition in the diagnosis of antibody-mediated rejection after lung transplantation.

Antibody-mediated rejection (AMR) is an increasingly recognized form of lung rejection. C4d deposition has been an inconsistent finding in previous reports and its role in the diagnosis has been controversial. We conducted a retrospective single-center study to characterize cases of C4d-negative probable AMR and to compare these to cases of definite (C4d-positive) AMR. We identified 73 cases of AMR: 28 (38%) were C4d-positive and 45 (62%) were C4d-negative. The two groups had a similar clinical presentation, and although more patients in the C4d-positive group had neutrophilic capillaritis (54% vs. 29%, P = .035), there was no significant difference in the presence of other histologic findings. Despite aggressive antibody-depleting therapy, 19 of 73 (26%) patients in the overall cohort died within 30 days, but there was no significant difference in freedom from chronic lung allograft dysfunction (CLAD) or survival between the two groups. We conclude that AMR may cause allograft failure, but that the diagnosis requires a multidisciplinary approach and a high index of suspicion. C4d deposition does not appear to be a necessary criterion for the diagnosis, and although some cases may respond initially to therapy, there is a high incidence of CLAD and poor survival after AMR.