Redefining the molecular rejection states in 3230 heart transplant biopsies: Relationships to parenchymal injury and graft survival.

The first-generation Molecular Microscope (MMDx) system for heart transplant endomyocardial biopsies used expression of rejection-associated transcripts (RATs) to diagnose not only T cell-mediated rejection (TCMR) and antibody-mediated rejection (ABMR) but also acute injury. However, the ideal system should detect rejection without being influenced by injury, to permit analysis of the relationship between rejection and parenchymal injury. To achieve this, we developed a new rejection classification in an expanded cohort of 3230 biopsies: 1641 from INTERHEART (ClinicalTrials.gov NCT02670408), plus 1589 service biopsies added to improve the power of the machine learning algorithms. The new system used 6 rejection classifiers instead of RATs and generated 7 rejection archetypes: No rejection, 48%; Minor, 24%; TCMR1, 2.3%; TCMR2, 2.7%; TCMR/mixed, 2.7%; early-stage ABMR, 3.9%; and fully developed ABMR, 16%. Using rejection classifiers eliminated cross-reactions with acute injury, permitting separate assessment of rejection and injury. TCMR was associated with severe-recent injury and late atrophy-fibrosis and rarely had normal parenchyma. ABMR was better tolerated, seldom producing severe injury, but in later biopsies was often associated with atrophy-fibrosis, indicating long-term risk. Graft survival and left ventricular ejection fraction were reduced not only in hearts with TCMR but also in hearts with severe-recent injury and atrophy-fibrosis, even without rejection.

The adapted Heart Donor Score.

The Heart Donor Score (HDS) predicts donor organ discard for medical reasons and survival after heart transplantation (HTX) in the Eurotransplant allocation system. Our aim was to adapt the HDS for application in the United Network for Organ Sharing (UNOS) registry. To adjust for differences between the Eurotransplant and UNOS registries, the "adapted HDS" was created (aHDS) by exclusion of the covariates "valve function," "left-ventricular hypertrophy," and exclusion of "drug abuse" from the variable "compromised history." Two datasets were analyzed to evaluate associations of the aHDS with donor organ discard (n = 70 948) and survival (n = 19 279). The aHDS was significantly associated with donor organ discard [odds ratio 2.72, 95% confidence interval (CI) 2.68-2.76, P < 0.001; c-statistic: 0.937). The score performed comparably in donors <60 and ≥60 years of age. The aHDS was a significant predictor of survival as evaluated by univariate Cox proportional hazards analysis (hazard ratio 1.04, 95% CI 1.01-1.07, P = 0.023), although the association lost significance in a multivariable model. The aHDS predicts donor organ discard. Negative effects of most aHDS components on survival are likely eliminated by highly accurate donor selection processes.

Molecular-level HLA mismatch is associated with rejection and worsened graft survival in heart transplant recipients - a retrospective study.

The aim was to evaluate the association of molecular-level human leukocyte antigen (HLA) mismatching with post-transplant graft survival, rejection, and cardiac allograft vasculopathy (CAV). We retrospectively analyzed all primary cardiac transplant recipients between 01/1984-06/2016. 1167 patients fulfilled inclusion criteria and had HLA typing information available. In 312 donor-recipient pairs, typing at serological split antigen level was available. We used the Epitope MisMatch Algorithm to calculate the number of amino acid differences in antibody-verified HLA eplets (amino acid mismatch load (AAMM)) between donor and recipient. Patients with a higher HLA-DR AAMM load had inferior 1-year graft survival (hazard ratio [HR], 1.14; 95% confidence interval [CI], 1.01-1.28). The HLA-AB AAMM load showed no impact on graft survival. In the subgroup with available split-level information, we observed an inferior graft survival for a higher HLA-DR AAMM load 3 months after transplantation (HR, 1.22; 95% CI, 1.04-1.44) and a higher risk for rejection for an increasing HLA-AB (HR, 1.70; 95% CI, 1.29-2.24) and HLA-DR (HR, 1.32; 95% CI, 1.09-1.61) AAMM load. No impact on the development of CAV was found. Molecular-level HLA mismatch analysis could serve as a tool for risk stratification after heart transplantation and might take us one step further into precision medicine.

Diminished impact of cytomegalovirus infection on graft vasculopathy development in the antiviral prophylaxis era - a retrospective study.

Evidence concerning an association between cytomegalovirus (CMV) infection and accelerated cardiac allograft vasculopathy (CAV) is inconclusive. Data were analyzed retrospectively from 297 consecutive heart transplants between 1.1.2002 and 31.12.2012. Patients ≤18 years of age, survival, and follow-up ≤1-year post-transplant and patients with early CAV were excluded. CMV-infection was diagnosed and monitored closely in the first year. CAV was diagnosed by coronary angiography via left heart catheterization, and results were categorized according to the International Society of Heart and Lung Transplantation (ISHLT) scoring system. Risk factors for CAV were tested in a multivariable model. Median follow-up was 7.5 years (IQR: 5.6-10.3). CMV infection in the first year after transplantation occurred in 26% of patients (n = 78), CMV disease in 5% (n = 15). CAV ≥1 ISHLT was detected in 36% (n = 108). Incidence of CAV >1 ISHLT and severity of CAV increased over time. No statistically significant association between CMV infection and disease within the first year and risk of CAV after 1-year post-HTx was detected in the univariate (P = 0.16) and multivariable [hazard ratio (HR), 1.36; confidence interval (CI), 0.89-2.07; P = 0.16] Cox regression. In the multivariable Cox regression, donor age (HR, 1.04; 95% CI, 1.02-1.06; P < 0.01) and acute cellular rejection (ACR) ≥2R in the first year after HTx (HR, 1.77; 95% CI, 1.06-2.95; P = 0.03) were independent risk factors for CAV development. In our cohort, CMV infection and disease in the first year after transplantation did not significantly influence the risk of CAV in the long-term follow-up.

Molecular states associated with dysfunction and graft loss in heart transplants.

BACKGROUND: We explored the changes in gene expression correlating with dysfunction and graft failure in endomyocardial biopsies. METHODS: Genome-wide microarrays (19,462 genes) were used to define mRNA changes correlating with dysfunction (left ventricular ejection fraction [LVEF] ≤ 55) and risk of graft loss within 3 years postbiopsy. LVEF data was available for 1,013 biopsies and survival data for 779 patients (74 losses). Molecular classifiers were built for predicting dysfunction (LVEF ≤ 55) and postbiopsy 3-year survival. RESULTS: Dysfunction is correlated with dedifferentiation-decreased expression of normal heart transcripts, for example, solute carriers, along with increased expression of inflammation genes. Many genes with reduced expression in dysfunction were matrix genes such as fibulin 1 and decorin. Gene ontology (GO) categories suggested matrix remodeling and inflammation, not rejection. Genes associated with the risk of failure postbiopsy overlapped dysfunction genes but also included genes affecting microcirculation, for example, arginase 2, which reduces NO production, and endothelin 1. GO terms also reflected increased glycolysis and response to hypoxia, but decreased VEGF and angiogenesis pathways. T cell-mediated rejection was associated with reduced survival and antibody-mediated rejection with relatively good survival, but the main determinants of survival were features of parenchymal injury. Both dysfunction and graft loss were correlated with increased biopsy expression of BNP (gene NPPB). Survival probability classifiers divided hearts into risk quintiles, with actuarial 3-year postbiopsy survival >95% for the highest versus 50% for the lowest. CONCLUSIONS: Dysfunction in transplanted hearts reflects dedifferentiation, decreased matrix genes, injury, and inflammation. The risk of short-term loss includes these changes but is also associated with microcirculation abnormalities, glycolysis, and response to hypoxia.

Prioritizing direct heart procurement in organ donors after circulatory death does not jeopardize lung transplant outcomes.

Background: Controlled donation after circulatory death (cDCD) has become a standard in liver, kidney, and lung transplantation (LTx). Based on recent innovations in ex vivo heart preservation, heart transplant centers have started to accept cDCD heart allografts. Because the heart has very limited tolerance to warm ischemia, changes to the cDCD organ procurement procedures are needed. These changes entail delayed ventilation and prolonged warm ischemia for the lungs. Whether this negatively impacts lung allograft function is unclear. Methods: A retrospective analysis of cDCD lungs transplanted between 2012 and February 2022 at the Medical University of Vienna was performed. The heart + lung group consisted of cases in which the heart was procured by a cardiac team for subsequent normothermic ex vivo perfusion. A control group (lung group) was formed by cases where only the lungs were explanted. In heart + lung group cases, the heart procurement team placed cannulas after circulatory death and a hands-off time, collected donor blood for ex vivo perfusion, and performed rapid organ perfusion with Custodiol solution, after which the heart was explanted. Up to this point, the lung procurement team did not interfere. No concurrent lung ventilation or pulmonary artery perfusion was performed. After the cardiac procurement team left the table, ventilation was initiated, and lung perfusion was performed directly through both stumps of the pulmonary arteries using 2 large-bore Foley catheters. This study analyzed procedural explant times, postoperative outcomes, primary graft dysfunction (PGD), duration of mechanical ventilation, length of intensive care unit (ICU) stay, and early survival after LTx. Results: A total of 56 cDCD lungs were transplanted during the study period. In 7 cases (12.5%), the heart was also procured (heart + lung group); in 49 cases (87.5%), only the lungs were explanted (lung group). Basic donor parameters were comparable in the 2 groups. The median times from circulatory arrest to lung perfusion (24 minutes vs 13.5 minutes; P = .002) and from skin incision to lung perfusion (14 minutes vs 5 minutes; P = .005) were significantly longer for the heart + lung procedures. However, this did not affect post-transplantation PGD grade at 0 hours (P = .851), 24 hours (P = .856), 48 hours (P = .929), and 72 hours (P = .874). At 72 hours after transplantation, none of the lungs in the heart + lung group but 1 lung (2.2%) in lung group was in PGD 3. The median duration of mechanical ventilation (50 hours vs 41 hours; P = .801), length of ICU stay (8 days vs 6 days; P = .951), and total length of hospital stay (27 days vs 25 days; P = .814) were also comparable in the 2 groups. In-hospital mortality occurred in only 1 patient of the lung group (2.2%). Conclusions: Although prioritized cDCD heart explantation is associated with delayed ventilation and significantly longer warm ischemic time to the lungs, post-LTx outcomes within the first year are unchanged. Prioritizing heart perfusion and explantation in the setting of cDCD procurement can be considered acceptable.

Many heart transplant biopsies currently diagnosed as no rejection have mild molecular antibody-mediated rejection-related changes.

BACKGROUND: The Molecular Microscope (MMDx) system classifies heart transplant endomyocardial biopsies as No-rejection (NR), Early-injury, T cell-mediated (TCMR), antibody-mediated (ABMR), mixed, and possible rejection (possible TCMR, possible ABMR). Rejection-like gene expression patterns in NR biopsies have not been described. We extended the MMDx methodology, using a larger data set, to define a new "Minor" category characterized by low-level inflammation in non-rejecting biopsies. METHODS: Using MMDx criteria from a previous study, molecular rejection was assessed in 1,320 biopsies (645 patients) using microarray expression of rejection-associated transcripts (RATs). Of these biopsies, 819 were NR. A new archetypal analysis model in the 1,320 data set split the NRs into NR-Normal (N = 462) and NR-Minor (N = 359). RESULTS: Compared to NR-Normal, NR-Minor were more often histologic TCMR1R, with a higher prevalence of donor-specific antibody (DSA). DSA positivity increased in a gradient: NR-Normal 24%; NR-Minor 34%; possible ABMR 42%; ABMR 66%. The top 20 transcripts distinguishing NR-Minor from NR-Normal were all ABMR-related and/or IFNG-inducible, and also exhibited a gradient of increasing expression from NR-Normal through ABMR. In random forest analysis, TCMR and Early-injury were associated with reduced LVEF and increased graft loss, but NR-Minor and ABMR scores were not. Surprisingly, hearts with MMDx ABMR showed comparatively little graft loss. CONCLUSIONS: Many heart transplants currently diagnosed as NR by histologic or molecular assessment have minor increases in ABMR-related and IFNG-inducible transcripts, associated with DSA positivity and mild histologic inflammation. These results suggest that low-level ABMR-related molecular stress may be operating in many more hearts than previously estimated. (ClinicalTrials.gov #NCT02670408).

Indications, Complications, and Outcomes of Cardiac Surgery After Heart Transplantation: Results From the Cash Study.

Background: Allograft pathologies, such as valvular, coronary artery, or aortic disease, may occur early and late after cardiac transplantation. Cardiac surgery after heart transplantation (CASH) may be an option to improve quality of life and allograft function and prolong survival. Experience with CASH, however, has been limited to single-center reports. Methods: We performed a retrospective, multicenter study of heart transplant recipients with CASH between January 1984 and December 2020. In this study, 60 high-volume cardiac transplant centers were invited to participate. Results: Data were available from 19 centers in North America (n = 7), South America (n = 1), and Europe (n = 11), with a total of 110 patients. A median of 3 (IQR 2-8.5) operations was reported by each center; five centers included ≥ 10 patients. Indications for CASH were valvular disease (n = 62), coronary artery disease (CAD) (n = 16), constrictive pericarditis (n = 17), aortic pathology (n = 13), and myxoma (n = 2). The median age at CASH was 57.7 (47.8-63.1) years, with a median time from transplant to CASH of 4.4 (1-9.6) years. Reoperation within the first year after transplantation was performed in 24.5%. In-hospital mortality was 9.1% (n = 10). 1-year survival was 86.2% and median follow-up was 8.2 (3.8-14.6) years. The most frequent perioperative complications were acute kidney injury and bleeding revision in 18 and 9.1%, respectively. Conclusion: Cardiac surgery after heart transplantation has low in-hospital mortality and postoperative complications in carefully selected patients. The incidence and type of CASH vary between international centers. Risk factors for the worse outcome are higher European System for Cardiac Operative Risk Evaluation (EuroSCORE II) and postoperative renal failure.

Severe gastroparesis after orthotopic heart transplantation.

We present a case of severe gastroparesis in a recipient of orthotopic heart transplantation. Although a rare condition after heart transplantation, it is a potential cause of significant morbidity, including vomiting, aspiration and pneumonia. Moreover, impaired gastric emptying alters the pharmacokinetics of immunosuppressive medication with increased risk of severe side effects. Herein, we describe a diagnostic and therapeutic strategy that was successfully applied in a patient with gastroparesis.

Donor heart selection and outcomes: An analysis of over 2,000 cases.

BACKGROUND: Decision-making when offered a donor heart for transplantation is complex, and supportive data describing outcomes according to acceptance or non-acceptance choices are sparse. Our aim was to analyze donor heart acceptance decisions and associated outcomes at a single center, and after subsequent acceptance elsewhere. METHODS: This investigation was a retrospective analysis of data obtained from the University of Vienna Medical Center and Eurotransplant centers for the period 2001 to 2015. RESULTS: Our center accepted 31.8% (699 of 2,199) of donor hearts offered. Unlike other centers, the acceptance rate, with or without transplantation, did not increase over time. Of the donor hearts rejected by our center, 38.1% (572 of 1,500) were later accepted elsewhere. Acceptance rates were twice as high for donor hearts initially rejected for non-quality reasons (339 of 601, 56.4%) compared with initial rejection for quality reasons (233 of 899, 25.9%). Three-year patient survival rate was 79% at Vienna; for donor hearts initially rejected by Vienna for non-quality reasons or quality reasons, it was 73% and 63%, respectively (p < 0.001). Outcomes at other centers after transplantation of grafts rejected by Vienna varied according to the reason for rejection, with good 3-year survival rates for rejection due to positive virology (77%), high catecholamines (68%), long ischemic time (71%), or low ejection fraction (68%), but poor survival was observed for hearts rejected for hypernatremia (46%), cardiac arrest (21%), or valve pathology (50%). CONCLUSIONS: A less restrictive policy for accepting donor hearts at our center, particularly regarding rejection for non-quality reasons or for positive virology, high catecholamine levels, longer ischemic time, or low ejection fraction, could expand our donor pool while maintaining good outcomes.