A retrospective analysis of the safety and efficacy of apixaban use after lung transplant.

Direct acting oral anticoagulants (DOACs) have become the mainstay of treatment for patients requiring anticoagulation for atrial arrhythmias and venous thromboembolism (VTE) but safety and efficacy has not been established in lung transplantation. This is a retrospective review of 28 adult lung transplant patients who were prescribed apixaban for stroke prevention in atrial arrhythmias or treatment of VTE between October 15, 2015 and December 31, 2018. The primary outcome was a composite of efficacy and safety measured by recurrence or breakthrough of stroke or thromboembolism and bleeding events. Seven patients were treated for atrial arrhythmias and 21 treated for VTE. Fifteen patients received CYP3A4 or P-gp inhibitors at initiation of anticoagulation, and 4 of these patients received strong CYP3A4 inhibitors. During the follow-up period, one breakthrough DVT and one clinically relevant non-major bleed were observed. These data suggest that apixaban may be safe to use for lung transplant patients, and larger studies are warranted to assess long-term outcomes as well as safety and efficacy of alternative DOACs.

Perioperative anidulafungin combined with triazole prophylaxis for the prevention of early invasive candidiasis in lung transplant recipients.

BACKGROUND: Invasive candidiasis (IC) is a substantial cause of morbidity and mortality among lung transplant recipients (LTRs). Postoperative factors include prolonged hospital stay, central lines, delayed chest closure, and dehiscence increase IC risk. Correspondingly, current guidelines propose targeted IC coverage early posttransplant with fluconazole or an echinocandin. METHODS: This retrospective analysis was performed on LTRs from January 2016 to January 2020 and evaluated effectiveness of a recent protocol utilizing perioperative anidulafungin for early IC prevention in addition to long-term triazole antifungal prophylaxis. Prior to this protocol, patients were primarily established on itraconazole prophylaxis alone. The primary endpoint was proven or probable IC within 90 days after transplant. Multivariable logistic regression modeling was used to assess risk factors for invasive fungal infection (IFI). RESULTS: Among 144 LTRs, there was a numerically lower incidence of IC in the protocol group, although not statistically significant (6% vs. 13%, p = 0.16). Incidence of proven or probable IFI was 7.5% in the protocol cohort and 19.5% in the pre-protocol cohort (p = 0.038). In multivariable analysis, when controlling for lung allocation score (OR 1.04, 95% CI 1.01-1.08), donor perioperative culture with fungal growth (OR 2.92, 95% CI 1.02-8.92), and dehiscence (OR 3.54, 95% CI 1.14-10.85), protocol cohort was not significantly associated with IFI (OR 0.41, 95% CI 0.12-1.23). CONCLUSIONS: To our knowledge, this is the first study investigating combination triazole/echinocandin use in the early post-lung transplant period. These findings demonstrate that in-hospital anidulafungin offers unclear benefit for early IC prevention when used in combination with triazole prophylaxis.

Anti-LFA-1 induces CD8 T-cell dependent allograft tolerance and augments suppressor phenotype CD8 cells.

The induction of tolerance to transplanted organs is a major objective in transplantation immunology research. Lymphocyte function-associated antigen-1 (LFA-1) interactions have been identified as a key component of the T-cell activation process that may be interrupted to lead to allograft tolerance. In mice, αLFA-1 mAb is a potent monotherapy that leads to the induction of donor-specific transferable tolerance. By interrogating important adaptive and innate immunity pathways, we demonstrate that the induction of tolerance relies on CD8+T-cells. We further demonstrate that αLFA-1 induced tolerance is associated with CD8+CD28-T-cells with a suppressor phenotype, and that while CD8 cells are present, the effector T-cell response is abrogated. A recent publication has shown that CD8+CD28- cells are not diminished by cyclosporine or rapamycin, therefore CD8+CD28- cells represent a clinically relevant population. To our knowledge, this is the first time that a mechanism for αLFA-1 induced tolerance has been described.

Cutting Edge: Roles for Batf3-Dependent APCs in the Rejection of Minor Histocompatibility Antigen-Mismatched Grafts.

In transplantation, a major obstacle for graft acceptance in MHC-matched individuals is the mismatch of minor histocompatibility Ags. Minor histocompatibility Ags are peptides derived from polymorphic proteins that can be presented by APCs on MHC molecules. The APC subtype uniquely responsible for the rejection of minor Ag-mismatched grafts has not yet been identified. In this study, we examined graft rejection in three mouse models: 1) mismatch of male-specific minor Ags, 2) mismatch of minor Ags distinct from male-specific minor Ags, and 3) skin transplant. This study demonstrates that in the absence of pathogen-associated molecular patterns, Batf3-dependent dendritic cells elicit the rejection of cells and grafts expressing mismatched minor Ags. The implication of our findings in clinical transplantation may be significant, as minor Ag reactivity has been implicated in the pathogenesis of multiple allograft tissues.

Ectopic expression of Fas Ligand on cardiomyocytes renders cardiac allografts resistant to CD4(+) T-cell mediated rejection.

Fas Ligand limits inflammatory injury and permits allograft survival by inducing apoptosis of Fas-bearing lymphocytes. Previous studies have shown that the CD4(+) T-cell is both sufficient and required for murine cardiac allograft rejection. Here, utilizing a transgenic mouse that over-expresses Fas Ligand specifically on cardiomyocytes as heart donors, we sought to determine if Fas Ligand on graft parenchymal cells could resist CD4(+) T-cell mediated rejection. When transplanted into fully immunocompetent BALB/c recipients Fas Ligand transgenic hearts were acutely rejected. However, when transplanted into CD4(+) T-cell reconstituted BALB/c-rag(-/-) recipients, Fas Ligand hearts demonstrated long-term survival. These results indicate that Fas Ligand over-expression on cardiomyocytes can indeed resist CD4(+) T-cell mediated cardiac rejection and suggests contact dependence between Fas Ligand expressing graft parenchymal cells and the effector CD4(+) T-cells.

Acute myocardial infarction secondary to mucormycosis after lung transplantation.

We present a case of a 57-year-old man who underwent bilateral lung transplantation for idiopathic pulmonary fibrosis. His immediately post-operative course was complicated by fever and cardiac arrest. Despite supportive care and broad-spectrum antibiotics, he experienced continued clinical decline. Autopsy results indicated angioinvasive mucormycosis and coronary arteritis resulting in acute myocardial infarction as the cause of death.

Impact of rituximab-associated B-cell defects on West Nile virus meningoencephalitis in solid organ transplant recipients.

Evidence suggests that West Nile virus (WNV) neuroinvasive disease occurs more frequently in both solid organ and human stem cell transplant recipients. The effect of concomitant anti-B-cell therapy with rituximab, a CD20(+) monoclonal antibody, on WNV infection in this population, however, has not been reported. We describe a case of a patient with alpha-1-antitrypsin deficiency who underwent single lung transplantation in 2005 and was maintained on tacrolimus, cytoxan and prednisone. More recently, she had received two courses of rituximab for recurrent A2-A3 grade rejection with concomitant capillaritis and presented six months later with rapid, fulminant WNV meningoencephalitis. Her diagnosis was made by cerebrospinal fluid (CSF) PCR but serum and CSF WNV IgM and IgG remained negative. She received WNV-specific hyperimmune globulin (Omr-Ig-Am) through a compassionate protocol. She experienced a rapidly progressive and devastating neurological course despite treatment and died three wk after onset of her symptoms. Autopsy revealed extensive meningoencephalomyelitis.

Spontaneous allograft tolerance in B7-deficient mice independent of preexisting endogenous CD4+CD25+ regulatory T-cells.

BACKGROUND: Acute cardiac allograft rejection requires host, but not donor, expression of B7-1/B7-2 costimulatory molecules. However, acute cardiac rejection requires direct antigen presentation by donor-derived antigen presenting cells to CD4 T-cells and does not require indirect antigen presentation to CD4 T-cells. Given this discrepancy in the literature and that the consequence of allograft exposure in B7-deficient mice is unknown; the goal of the study was to examine the antidonor status of allografted B7-1/B7-2-deficient hosts. METHODS: C57Bl/6 B7-1/B7-2-/- mice were grafted with heterotopic BALB/c hearts. Recipients bearing long-term surviving allografts were used to examine the status of antidonor reactivity in vitro and in vivo. Tolerance was examined in vivo through adoptive transfer of splenocytes from graft-bearing animals to secondary immune-deficient Rag-1-/- hosts bearing donor-type or third-party cardiac allografts and by regulatory T-cell depletion with anti-CD25 antibody. RESULTS: When transferred to B7-replete Rag-1-/- recipients, cells from naïve B7-1/B7-2-/- mice readily initiated cardiac allograft rejection. However, splenocytes transferred from long-term allograft acceptor B7-1/B7-2-/- hosts failed to reject donor-type hearts but acutely rejected third-party allografts. In addition, such cells did not reject (donorxthird-party) F1 allografts. Finally, in vivo depletion of regulatory T-cells did not prevent long-term acceptance. CONCLUSIONS: Results demonstrate that B7-deficient T-cells are capable of acute cardiac allograft rejection in a B7-replete environment. Importantly, results also show that B7-deficient hosts do not simply ignore cardiac allografts, but rather spontaneously develop transferable, donor-specific tolerance and linked suppression in vivo. Interestingly, this tolerant state does not require endogenous CD4+CD25+ regulatory T-cells.

Lobar torsion complicating bilateral lung transplantation.

We report a case of left lower lobe torsion in a patient who had undergone bilateral lung transplantation for alpha(1)-antitrypsin deficiency. The patient experienced acute pulmonary hypertension and hypoxemia on post-operative Day 3 and the chest X-ray showed bilateral alveolar infiltrates and a new focal consolidation of the left lower lobe. Fiberoptic bronchoscopy showed complete obstruction of the left lower lobe bronchus and abnormal rotation of the left upper lobe bronchus suggesting torsion, which was confirmed by pulmonary angiography and ultimately at thoracotomy. The possibility of acute lobar torsion should be considered in lung transplant recipients who demonstrate evidence of acute respiratory insufficiency in the early post-operative period.

Murine heterotopic heart transplant technique.

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique was developed and reported in 2001 by Niimi. Described here are the techniques that have evolved over more than a decade in the hands of three surgeons (Plenter, Grazia, Pietra) in our center. These techniques are now being passed on to a younger generation of surgeons and researchers. Based largely on the Niimi experience, the procedures used have evolved in the fine details - details which we will endeavor to relate here in such a way that others may be able to use this very useful model. Like Niimi, we have found that a video aid to learning is a priceless resource for the beginner.

Four decades of vascularized heterotopic cardiac transplantation in the mouse.

Since the first clinical heart transplant in 1967, there has been a heightened need to understand immune and inflammatory responses to "foreign" tissues. Research efforts in those early days were based on species that would now be considered "large" and were typically out-bred individuals. While this closely mirrors the clinical scenario, where genetic mismatches of donors and recipients can only be minimized in the selection process, these were not ideal models for studying the complexities and nuances of the immune system. Even when the rat was considered the standard model those early endeavors were limited by a small number of rat strains. The mouse model has provided us with an overwhelming array of strains, knockouts, knockins and transgenics that allow us to investigate the many layers of the innate and adaptive immune systems leading to a much greater understanding of immune responses. Fully vascularized heterotopic cardiac transplantation in the mouse has now been with us for four decades; the original papers describing this technique being published by Corry in 1973. In the subsequent 40 years, this technique has been used by many laboratories, including our own, and has become a powerful tool for the investigation of transplant immunity and ischemia reperfusion injury. Given the modern availability of mouse strains and mouse-related reagents, our current understanding of transplant immunity undoubtedly would not exist without such a technique.

Pre-transplant presence of antibodies to MICA and HLA class I or II are associated with an earlier onset of bronchiolitis obliterans syndrome in lung transplant recipients.

Previous reports using cell-based methods (CDC-AHG) suggest that the presence of pre-transplant HLA Class I and II antibodies are associated with worse survival following lung transplantation. Similarly, antibodies to major histocompatibility complex Class I chain-related gene A (MICA) have been associated with increased graft failure following kidney transplantation. Using highly sensitive solid phase assays, we sought to determine whether the pre-transplant presence of antibodies to MICA or HLA Class I or II predicted short or long-term lung allograft function. Pre-transplant sera screened for antibodies to MICA by Labscreen Single Antigen format and HLA by Luminex (n = 192) revealed antibody presence in 31 (16.1%) and 70 (36.4%) patients, respectively. HLA antibody presence correlated with increased bronchiolitis Obliterans syndrome (BOS)-1 development at 3 years [32.9% (23/70) vs. 18.9% (23/122), p = 0.03] while MICA antibodies correlated with BOS-2 development [32.3% (10/31) vs. 14.9% (24/161), p = 0.02]. The presence of HLA or MICA antibodies correlated with BOS-1 development [32.5% (26/81) vs.18.0% (20/111), p = 0.02] and BOS-2 [24.7% (20/81) vs. 12.6% (14/111), p = 0.02] at 3 years. We found no correlation between antibody presence and episodes of acute cellular rejection or overall survival. We conclude that the presence of pre-transplant HLA or MICA antibodies is associated with earlier BOS onset following lung transplantation.

CD4 T cells mediate cardiac xenograft rejection via host MHC Class II.

BACKGROUND: Previous studies have shown that acute CD4 T-cell-mediated cardiac allograft rejection requires donor major histocompatibility complex (MHC) Class II expression and can be independent of "indirect" antigen presentation. However, other studies suggested that indirect antigen presentation to CD4 T cells may play a primary role in cellular xenograft immunity. Thus, the relative roles of direct/indirect CD4 T cell reactivity against cardiac xenografts are unclear. In this study we set out to determine the role for indirect CD4 T cell reactivity in cardiac xenograft rejection. METHODS: Rat hearts were transplanted heterotopically into wild-type and immunodeficient mice. Recipients were untreated, treated with depleting antibodies, or reconstituted with wild-type cells. RESULTS: Antibody depletion confirmed that rat heart xenograft rejection in C57Bl/6 mice was CD4 T-cell-dependent. Also, heart xenografts survived long term in B6 MHC Class II (C2D)-deficient mice. Graft acceptance in C2D mice was not secondary to CD4 T cell deficiency alone, because transferred B6 CD4 T cells failed to trigger rejection in C2D hosts. Furthermore, purified CD4 T cells were sufficient for acute rejection of rat heart xenografts in immune-deficient B6rag1(-/-) recipients. Importantly, CD4 T cells did not reject rat hearts in C2Drag1(-/-) hosts, in contrast to results using cardiac allografts. "Direct" xenoreactive CD4 T cells were not sufficient to mediate rejection despite vigorous reactivity to rat stimulator cells in vitro. CONCLUSIONS: Taken together, our results show that CD4 T cells are both necessary and sufficient for acute cardiac xenograft rejection and that host MHC Class II is critical in this process.

Acute cardiac allograft rejection by directly cytotoxic CD4 T cells: parallel requirements for Fas and perforin.

BACKGROUND: CD4 T cells can suffice as effector cells to mediate primary acute cardiac allograft rejection. Although CD4 T cells can readily kill appropriate target cells in vitro, the corresponding role of such cytolytic activity for mediating allograft rejection in vivo is unknown. Therefore, we determined whether the cytolytic effector molecules perforin (PFP) and/or FasL (CD95L) were necessary for CD4 T cell-mediated rejection in vivo. METHODS: Wild-type C3H(H-2) or Fas (CD95)-deficient C3Hlpr (H-2) hearts were transplanted into immune-deficient C57B6rag (H-2) mice. Then, recipients were reconstituted with naïve purified CD4 T cells from wild-type, PFP-deficient, or FasL (gld)-deficient T-cell donors. RESULTS: In vitro, alloreactive CD4 T cells were competent to lyse donor major histocompatibility complex class II+ target cells, largely by a Fas-dependent mechanism. In vivo, the individual disruption of donor Fas expression (lpr) or CD4 T-cell-derived PFP had no significant impact on acute rejection. However, FasL-deficient (gld) CD4 T cells demonstrated delayed allograft rejection. Importantly, the simultaneous removal of both donor Fas expression and CD4 T-cell PFP completely abrogated acute rejection, despite the persistence of CD4 T cells within the graft. CONCLUSIONS: Results demonstrate that the direct rejection of cardiac allografts by CD4 effector T cells requires the alternative contribution of graft Fas expression and T cell PFP expression. To our knowledge, this is the first demonstration that cytolytic activity by CD4 T cells can play an obligate role for primary acute allograft rejection in vivo.

Perturbation of leukocyte function-associated antigen-1/intercellular adhesion molecule-1 results in differential outcomes in cardiac vs islet allograft survival.

BACKGROUND: Various studies indicate the requirements for tolerance induction may vary between different transplanted tissues and organs. Consequently, we compared the efficacy of anti-leukocyte function-associated antigen-1 (LFA-1)/anti-intercellular adhesion molecule-1 (ICAM-1) monoclonal antibody therapy for facilitating cardiac vs islet long-term allograft acceptance in mice. METHODS: BALB/c (H-2d) mouse cardiac or islet allografts were transplanted into recipient CBA/J (H-2k) mice. Monoclonal anti-body therapy with anti-LFA-1, anti-ICAM-1, the combination, or control rat immunoglobulin (Ig) was administered intraperitoneally on Days 0 to 5. Cardiac allograft function was assessed by palpation and islet graft function by blood glucose monitoring. Mixed lymphocyte assays were performed to assess proliferation of CD4 and CD8 T-cells under conditions of stimulator-cell ICAM-1 and/or LFA-1 deficiency. RESULTS: Anti-ICAM-1 therapy resulted in a modest prolongation of cardiac allografts but in pronounced survival of islet allografts. Anti-LFA-1 therapy promoted significant long-term survival of both cardiac and islet allografts. Surprisingly, combined anti-LFA-1/anti-ICAM-1 therapy abrogated long-term islet, but not cardiac, allograft acceptance relative to either monotherapy. Mixed lymphocyte reactions demonstrated complete blockade of CD4 and CD8 T-cell proliferation under conditions of ICAM-1 deficiency alone or in combination with anti-LFA-1 therapy. CONCLUSION: These results indicate that optimal therapies for some allografts (vascularized-heart) may not translate to other types of allografts (cellular-islet). Thus, the type of transplant represents an independent variable for optimizing strategies to promote indefinite allograft acceptance. Complete inhibition of CD4 and CD8 T-cell proliferation during ICAM-1/LFA-1 blockade suggests a threshold signal may be dependent upon ICAM-1/LFA-1 for regulatory tolerance to occur and that this signal may be lost under conditions of minimal graft cellular mass.

A two-step model of acute CD4 T-cell mediated cardiac allograft rejection.

CD4 T cells are both necessary and sufficient to mediate acute cardiac allograft rejection in mice. This process requires "direct" engagement of donor MHC class II molecules. That is, acute rejection by CD4+ T cells requires target MHC class II expression by the donor and not by the host. However, it is unclear whether CD4+ T cell rejection requires MHC class II expression on donor hemopoietic cells, nonhemopoietic cells, or both. To address this issue, bone marrow transplantation in mice was used to generate chimeric heart donors in which MHC class II was expressed either on somatic or on hemopoietic cells. We report that direct recognition of hemopoietic and nonhemopoietic cells are individually rate limiting for CD4+ T cell-mediated rejection in vivo. Importantly, active immunization with MHC class II(+) APCs triggered acute rejection of hearts expressing MHC class II only on the somatic compartment. Thus, donor somatic cells, including endothelial cells, are not sufficient to initiate acute rejection; but they are necessary as targets of direct alloreactive CD4 T cells. Taken together, results support a two-stage model in which donor passenger leukocytes are required to activate the CD4 response while direct interaction with the somatic compartment is necessary for the efferent phase of acute graft rejection.

Increasing plasmalogen levels protects human endothelial cells during hypoxia.

Supplementation of cultured human pulmonary arterial endothelial cells (PAEC) with sn-1-O-hexadecylglycerol (HG) resulted in an approximately twofold increase in cellular levels of plasmalogens, a subclass of phospholipids known to have antioxidant properties; this was due, primarily, to a fourfold increase in the choline plasmalogens. Exposure of unsupplemented human PAEC to hypoxia (PO(2) = 20-25 mmHg) caused an increase in cellular reactive oxygen species (ROS) over a period of 5 days with a coincident decrease in viability. In contrast, HG-supplemented cells survived for at least 2 wk under these conditions with no evidence of increased ROS. Hypoxia resulted in a selective increase in the turnover of the plasmalogen plasmenylethanolamine. Human PAEC with elevated plasmalogen levels were also more resistant to H(2)O(2), hyperoxia, and the superoxide generator plumbagin. This protection was seemingly specific to cellular stresses in which significant ROS were generated because the sensitivity to lethal heat shock or glucose deprivation was not altered in HG-treated human PAEC. HG, by itself, was not sufficient for protection; HG supplementation of bovine PAEC had no effect upon plasmalogen levels and did not rescue these cells from the cytotoxic effects of hypoxia. This is the initial demonstration that plasmalogen content can be substantially enhanced in a normal cell. These data also demonstrate that HG can protect cells during hypoxia and other ROS-mediated stress, likely due to the resulting increase in these antioxidant phospholipids.