Clinical Cardiac Transplantation Research Articles

Donor Heart Preservation with Hydrogen Sulfide: A Systematic Review and Meta-Analysis.

Preclinical studies have shown that postconditioning with hydrogen sulfide (H2S) exerts cardioprotective effects against myocardial ischemia-reperfusion injury (IRI). The aim of this study was to appraise the current evidence of the cardioprotective effects of H2S against IRI in order to explore the future implementation of H2S in clinical cardiac transplantation. The current literature on H2S postconditioning in the setting of global myocardial ischemia was systematically reviewed and analyzed, performing meta-analyses. A literature search of the electronic databases Medline, Embase and Cinahl identified 1835 studies that were subjected to our pre-defined inclusion criteria. Sixteen studies were considered eligible for inclusion. Postconditioning with H2S showed significant robust effects with regard to limiting infarct size (standardized mean difference (SMD) = −4.12, 95% CI [−5.53–−2.71], p < 0.00001). Furthermore, H2S postconditioning consistently resulted in a significantly lower release of cardiac injury markers, lower levels of oxidative stress and improved cardiac function. Postconditioning with slow-releasing H2S donors offers a valuable opportunity for novel therapies within cardiac preservation for transplantation. Before clinical implication, studies evaluating the long-term effects of H2S treatment and effects of H2S treatment in large animal studies are warranted.

Effects of Ex Vivo Perfusion and Il-6 Receptor Blockade on Ischemia Reperfusion Injury in Cardiac Transplantation

Purpose In cardiac transplantation, ischemia reperfusion injury (IRI) remains a major problem associated with poor short and long-term outcomes. Since pro-inflammatory cytokines and endothelial activation have been hallmarks of IRI for decades, they can be addressed by recent therapeutic advancements. Normothermic ex vivo heart perfusion using the Organ Care System, but also pre-emptive treatment strategies for highly immunized recipients using the IL-6 receptor antibody tocilizumab, are in current clinical use. Therefore, we retrospectively studied blood samples from our clinical cardiac transplantation cohort for parameters of IRI. Methods In a novel protocol for heart transplantation (HTx) recipients with preformed donor-specific antibodies (pfDSA), an intraoperative application of tocilizumab (10mg/kg) was given together with 5 sessions of plasmapheresis, followed by one infusion of IgGAM (2g/kg) and a single dose of anti-CD20 antibody (rituximab). This protocol was applied to 6 recipients of OCS-preserved hearts. 10 recipients of OCS-preserved hearts without pfDSA did not receive tocilizumab (group 2). 24 recipients of standard heart transplants preserved on ice represent group 3. Donor or recipient demographics did not differ among the groups. Plasma levels of cytokines and endothelial markers were compared before, immediately after transplant surgery (T0), 24h (T24) and 3 weeks after transplantation. Results IRI could be measured in standard heart recipients at T0 and T24 by significantly increased levels of IL-6, CXCL8/IL-8, IL-18, Ang-2 and IGFBP-1 (all p Conclusion Normothermic ex vivo preservation in the OCS reduces IRI at the cytokine and endothelial level in recipient blood immediately after transplantation. Application of tocilizumab during transplantation blocks binding of IL-6 to its receptor leading to higher plasma levels. These effects may have a clinically relevant impact on ischemia reperfusion injury post cardiac transplantation.

Insights into the Development of a Life-Sustaining Orthotopic Cardiac Xenotransplant Model

Introduction Advances in clinical immunosuppression and characterization of optimal donor genetics continue to move the field of xenotransplantation toward a clinical reality. Cardiac xenograft survival in a non-life supporting heterotopic model has been extended to greater than 900 days as previously described by our lab. Orthotopic life supporting models for cardiac xenografts are being developed with the major barrier being significant peri-operative mortality. We detail the development of an orthotopic pig-to-baboon cardiac xenotransplant model, offering insights gained over time for replication of the model. Materials and Methods Specific pathogen free baboons were transplanted with GTKO.CD46 donor pig hearts in the orthotopic position (n = 3) by cardiac transplant and congenital cardiac surgeons. The clinical team also included a perfusionist and cardiac anesthesiologist. Detailed reports were generated regarding equipment and medications required for the cases to approximate clinical orthotopic cardiac transplantation. Monitoring capabilities were optimized as well as resuscitative access and strategy. These baboons were recovered from prior experiments and per institutional protocol were approved for non-survival studies; cases were performed with intended short term survival under anesthesia followed by elective euthanasia. Results and Discussion All recipient baboons were transplanted and weaned off of cardiopulmonary bypass with life sustaining cardiac function. Low dose inotropic support was continued in all cases; vasopressor support was continued in one case. Two cases were electively terminated after wean from bypass; one recipient suffered arrest three hours post wean from bypass. Recipients were monitored using femoral arterial lines, transesophageal echocardiogram, external EKG, continuous pulse oximetry, and serial bloodwork including ABG, CBC, BMP, and ACT. Central venous access with multiple lumens was critical to post-operative care. Invasive hemodynamic monitoring, pretreatment with anti-arrhythmics, and availability of cardioversion for the donor pig allowed rapid reversal of fibrillation episodes during procurement. Ideal cold ischemia time was found to be close to one hour; shorter ischemia time resulted in poorer xenograft function (n = 1) as evidenced by difficult wean from bypass and persistent need for vasopressor support. Atrial and ventricular pacing wires proved integral to the cardiopulmonary bypass weaning process. Conclusion Cardiac xenotransplantation is a potential solution to the paucity of human organs. Technical refinement of the life-supporting orthotopic cardiac xenotransplant with expert clinical help and applying human standards of post-operative care are the critical next step toward a clinical reality. Here we detail our experience developing an operative and critical care approach to the orthotopic cardiac xenotransplantation model.

Donor Heart Procurement From the Donation after Circulatory Death Pathway

Until recently, only isolated cases of cardiac transplantation utilizing hearts procured from the donation after circulatory death pathway, between co-located donor and recipient, have been reported. The strategy of utilizing supplemented cardioplegia in conjunction with a rapid surgical protocol to reduce warm ischemia, combined with subsequent donor heart evaluation on a portable ex-situ perfusion system, has permitted the use of these extended-criteria organs for successful clinical orthotopic cardiac transplantation in a reliable and reproducible manner. We describe the technique developed and used at St Vincent's Hospital in Sydney, Australia, for distantly procured donation after circulatory death hearts.

Scoring System Validity for Short-term Outcomes of Coronary Artery Bypass Surgery in Indigenous Australians

tion. A better understanding of the withdrawal period, DCDheart physiology andEVP science is required to improve outcomes. We aimed to define the derangements following withdrawal, noting their impact on donor blood use in EVP. Methodology: In a clinically relevant porcine asphyxia model (n=24) and initial human trials, haemodynamic (MAP, CVP, intra-ventricular pressures(IVP)), electrical/circulatory arrest times, metabolic (pH, O2, lactate, trop-T), biochemical (K+) and endocrine changes (noradrenaline & adrenaline measured in coronary sinus and systemic sites) duringwithdrawal periods (20-40 min) were analysed. Blood collection methods for EVP, and perfusion strategies (pressure vs. flow) were tested. Results: From pre-withdrawal to post-arrest, IVP increased in the RV, while LV IVP fell (PV loop studies). Early rapid O2 desaturations were noted (14 mmHg +/-1 at 5 min post withdrawal). By 30 min post-withdrawal, pHwas 10mmol/L, troponin-T >200 mg/L and K+ >10mmol/L. Adrenaline and noradrenaline (NA) surges were noted at 4 min post withdrawal (assoc. HR increase) with ongoing amine release (NA) in the coronary sinus, but no further increase systemically (p<0.05). Conclusion: Dramatic changes occurred during the withdrawal period, notably RV distension, profound acidosis, myocardial ischaemia & hyperkalaemia all of which have important implications for clinical cardiac transplantation/ EVP. A greater understanding of these processes provides insight for post-mortem interventions to limit impact and optimise EVP perfusate.

The Downtime Donor Heart: Analysis of Pre-Transplant Function and Current Utilization for Transplantation

Purpose Previous investigation reveals that donors resuscitated following cardiorespiratory arrest can be utilized for cardiac transplantation with satisfactory post-transplant survival within recipients of these organs. We analyzed this donor group to determine its contribution to transplantation and evaluate donor heart function after resuscitation. Methods and Materials We reviewed 225 consecutive donor organs offered to our institution for cardiac transplantation. Demographic and clinical data was obtained from UNOS. Functional data was obtained by analyzing donor echocardiograms. “Downtime” donor hearts (DD) were compared with organs from conventional donors without a period of arrest (CD). Results A history of resuscitation following circulatory arrest was present in 99 donors (44%). The mean downtime was 30 +/- 21 minutes. Organ utilization was 48% in the DD group vs. 45% for CD; p=NS. Donor age was significantly lower in the DD group (41+/-15 vs. 36+/-14;p=0.01), there was no difference in height or weight. Peak troponin I concentration (DD 2.3+/-4.3 vs. CD 1.1 +/-3.6 ng/ml; p=0.04) was elevated in the DD group. Left ventricular ejection fraction was significantly greater in the CD group. (DD 58.6+/-9% vs. CD 61.3+/-8.7%; p=0.02). The percentage of donor hearts with LV dysfunction (LVEF Conclusions Organs procured from donors resuscitated following cardiocirculatory arrest currently contribute a substantial proportion of organs for cardiac transplantation. Cardiac function in these donors is acceptable, albeit marginally impaired in comparison to those without a history of arrest. The potential for recovery and utilization of donor hearts in this setting may further support the prospect of clinical DCD cardiac transplantation.

Sildenafil Citrate Augments Myocardial Protection in Heart Transplantation

Sildenafil citrate has been shown to induce myocardial protective effects in a variety of experimental settings. Whether these effects could be used to enhance myocardial protection afforded by crystalloid cardioplegia, volatile anesthesia and hypothermia during cardiac transplantation remains to be established. We investigated the use of sildenafil-mediated cardioprotection in a rat model of heterotopic cardiac transplantation. Sildenafil citrate (0.7 mg/kg) was administered intravenously to the donor 30 min before onset of ischemia or 5 min before reperfusion in the recipient. In situ cardioplegic arrest was followed by an ischemic time of 3 or 6 hr, transplantation, and blood reperfusion. Myocardial functional recovery was studied in vivo by using a left ventricular balloon and cellular injury quantified by assay of troponin I release and apoptosis. Sildenafil preconditioning but not postconditioning significantly improved initial myocardial systolic and diastolic function after 3 hr of hypothermic cardioplegic arrest (114+/-4 mm Hg vs. 83+/-4 mm Hg generated pressure, [P<0.01]). The protective effect of sildenafil declined over a 3-hr period of reperfusion along with overall myocardial function, no longer reaching statistical significance at 3 hr. The protective effects of sildenafil were abolished by the putative blocker of the mitochondrial ATP sensitive potassium channel, 5-hydroxydecanoate, before sildenafil administration. Protein kinase C delta showed significant translocation after sildenafil administration in the donor. We conclude that sildenafil citrate pretreatment augments myocardial functional recovery after an ischemic time relevant to clinical cardiac transplantation. This effect is associated with protein kinase C activation/translocation and inhibited by 5-hydroxydecanoate.

Anti–LFA-1 Monotherapy Prevents Neointimal Formation in a Murine Model of Transplant Intimal Hyperplasia

Cardiac allograft vasculopathy (CAV) is the pre-eminent cause of late cardiac allograft failure. It is characterized by a concentric intimal hyperplasia, which we designate transplant intimal hyperplasia (TIH). To date, blockade of the adhesion molecule lymphocyte function-associated antigen-1 (LFA-1) has been shown to be effective in preventing TIH in experimental models of transplantation, but only when combined with other immunosuppressants. In this study we explored the impact of monotherapy against LFA-1 in a carotid artery allograft model of TIH. B10A(2R) (H-2(h2)) mice were used as donors and C57BL/6 (H-2(b)) mice used as recipients. The recipients were treated with a monoclonal antibody against LFA-1alpha (M17/4) or isotype-matched control immunoglobulin. Grafts were harvested after 35 days and analyzed by histomorphometry and immunohistochemistry. Blood samples were taken and analyzed by differential cell count and alloantibody levels. We found that treatment with M17/4 resulted in a significant reduction in TIH compared with controls. Immunostaining revealed that LFA-1alpha blockade inhibited CD45+ leukocyte infiltration, prevented intimal smooth muscle cell (SMC) proliferation, and preserved the medial SMC population. Finally, we demonstrated a reduction in the serum alloantibody titer in the group treated with anti-LFA-1alpha when compared with controls. We have demonstrated for the first time that LFA-1alpha blockade on its own can prevent development of TIH in an experimental model. The concept of modulating LFA-1alpha-mediated leukocyte migration and T-cell activation may therefore be of relevance to clinical cardiac transplantation and, as such, represents a potential target for therapeutic intervention against clinical CAV.

Th1 Adaptive Immune Responses in Cardiac Graft Arteriosclerosis

Heart transplants are not rejected in the absence of adaptive immune responses, eg, in transplantations between genetically identical donors and hosts or to recipients with severe combined immunodeficiencies. Activation of adaptive immunity to an allograft causes destructive responses against donor parenchymal or vascular cells, called acute rejection. In the presence of adequate immunosuppression, acute rejection is suppressed; however, chronic rejection may occur, manifesting as conduit arterial lumen loss resulting from expansion of the tunica intima and remodeling of vessel size, called graft arteriosclerosis.1 The differences in pathological findings and in sensitivity to immunosuppressive agents suggest that chronic graft failure arises from different mechanisms operative in separate allograft compartments than in acute graft rejection. The precise pathogenesis of graft arteriosclerosis is unknown, although there is broad consensus that it is, at least in part, an alloimmune process in that the arteriosclerotic changes are limited to the vascular bed of the allograft and spare the host vasculature. Adaptive immunity may activate a variety of effector mechanisms that cause chronic graft rejection in murine transplantation models.1 Although it is not the only factor that can cause graft arteriosclerosis, the evidence for the signature Th1 cytokine, interferon (IFN)-γ, is particularly compelling. In mouse transplantation models, serological neutralization or the genetic absence of IFN-γ reduces intimal expansion of graft arteries.2,3 Moreover, in a chimeric human-mouse model, antibody neutralization of IFN-γ prevents allogeneic T cell–mediated endothelial dysfunction, intimal thickening, and outward vascular remodeling; administration of IFN-γ accelerates these effects; and exogenous IFN-γ in the absence of leukocytes is sufficient to cause the arteriosclerotic changes.4–6 However, definitive evidence for a pathogenetic role of IFN-γ in clinical cardiac transplantation is lacking. Articles pp 1599 and 1608 In this issue of Circulation , van Loosdregt et al7 performed a comprehensive inventory of cytokines, chemokines, …

Aortic allograft vasculopathy is mediated by CD8 + T cells in Cyclosporin A immunosuppressed mice

We investigated the role of CD4 + T cells and CD8 + T cells in mediating allograft vasculopathy in Cyclosporin A (CyA) immunosuppressed mice. We first established that a dose of 50 mg/kg/d CyA was required to prevent acute rejection in C57BL/6 mice. CyA given at 50 mg/kg/d did not prevent allograft vasculopathy in either cardiac or aortic transplants in these mice. Using CD4 −/− and CD8 −/− mice, we established that CyA immunosuppression at this dose was only effective at preventing allograft vasculopathy in mice lacking CD8 + T cells. This implicates CD8 + T cells in the development of AV in situations of clinical cardiac transplantation where CyA is still the mainstay of immunosuppressive therapy. We confirmed the important role for CD8 + T cells in AV in the face of CyA immunosuppression by allopriming mice in the presence of CyA and transferring alloprimed T cells into RAG1 −/− immunodeficient mice. The RAG1 −/− mice were also treated with CyA. In this situation (CyA present during the allopriming and in the recipient), only primed CD8 + T cells could mediate AV, primed CD4 + T cells could not. Alloprimed CD8 + T cells raised in the presence of CyA exhibited markedly reduced direct recognition responses (as measured by MLR) and effector responses (as measured by cytotoxic activity). In contrast indirect activation was retained. We interpret these data to suggest that in the face of CyA immunosuppression CD4 + T cell effector function is ablated while CD8 + T cell function remains partially intact. The in vitro data suggest that the indirect pathway remains intact in this population of CyA resistant CD8 + T cells.

Is biopsy-proven cellular rejection an important clinical consideration in heart transplantation?

Immunosuppression strategies to prevent allograft rejection represent the cornerstone of long-term survival after heart transplantation. Endomyocardial biopsy has defined rejection in clinical cardiac transplantation and established a threshold for therapy. With the development of more effective immunosuppression modalities and the asymptomatic nature of most histologic rejection episodes, controversy exists regarding the need to augment immunosuppression based purely on histologic findings. The frequency of histologic rejection has declined with current immunosuppression. Resolution of lower grades of histologic rejection without treatment is the norm in both pediatric and adult heart transplant studies. Recurrent rejection episodes have been linked to the subsequent development of allograft coronary artery disease, and late rejection (even if asymptomatic) is associated with decreased survival in pediatric heart transplant recipients. Black race is a risk factor for recurrent rejection and reduced survival after late cellular rejection. Apoptosis of inflammatory cells is more evident during and after histologic rejection treated with corticosteroids. Despite numerous noninvasive modalities evaluated for the detection of rejection, to date noninvasive methods cannot reliably predict histologic rejection. Histologic rejection appears less common with current immunosuppressive strategies, and controversy exists about the need to treat asymptomatic rejection. It remains unproven whether non-treatment of moderate or greater rejection (>/=3A) increases the likelihood of recurrent rejection, which if present, may increase the risk of allograft coronary disease and/or reduced long-term survival.

Heat shock protein, inducible nitric oxide synthase and apoptotic markers in the acute phase of human cardiac transplantation.

Solid organ transplantation is associated with activation of apoptotic pathways and other stress markers. We aimed to describe the expression of Bax, Bcl-2, iNOs and Hsp-70 in the endothelium and myocytes of both ventricles and to see if there is any relationship with clinical donor organ failure. Twelve patients undergoing heart or heart-lung transplantation (including three domino cases) were studied with transmural biopsies from the right (RV) and the left ventricles (LV) at the following points: after donor optimisation; at the end of ischaemic time; and after 10 min of reperfusion. The 1-week endomyocardial RV biopsy was also examined. Five donor hearts turned down purely on functional grounds were analysed also. There was no difference between the RV and the LV for any of the markers at intraoperative assessment. The pattern of expression was not predictive of allograft failure. Donor hearts, however, have a strong pro-apoptotic phenotype, which is largely unopposed by the protective factors Bcl-2 and Hsp-70. Furthermore, the intensity of myocyte staining increases over time for Bax (P<0.001) and iNOs (P=0.02). Domino hearts showed a similar pattern. Compared to usable organs, poorly functioning donor hearts have stronger myocardial staining for Bax (P=0.002) and iNOs (P=0.01). Clinical cardiac transplantation is associated with activation of the Bax and iNOs pathways in both ventricles. The myocardium is affected in time-dependent fashion but this is compatible, to a certain extent, with satisfactory allograft function. Donor hearts turned down on the basis of poor haemodynamic performance have significantly higher expression of Bax and iNOs.

Induction of immunologic tolerance to cardiac allograft by simultaneous blockade of inducible co-stimulator and cytotoxic T-lymphocyte antigen 4 pathway.

Inducible co-stimulator (ICOS) is one of the most recently described members of the CD28 family, and it plays an important role in immune responses. To investigate the role of ICOS in allograft rejection, the authors studied graft survival after cardiac transplantation in mice. Hearts from BALB/c mice were transplanted into C3H/He mice. Immunohistochemical staining and flow cytometry were performed. Monoclonal antibody to ICOS or ICOS-immunoglobulin (Ig) was injected intraperitoneally. The authors performed mixed lymphocyte reaction (MLR). ICOS was expressed strongly by graft-infiltrating cells during rejection of the allograft. Blockade of the ICOS pathway with anti-ICOS antibody and ICOSIg significantly prolonged graft survival time relative to that in untreated mice; however, all cardiac allografts were eventually rejected by a single treatment. Treatment with both ICOSIg and cytotoxic T-lymphocyte antigen 4 (CTLA4) Ig induced not only long-term acceptance of the cardiac allograft but also donor-specific tolerance, which was shown by acceptance of donor but not third-party skin. Graft arterial intimal hyperplasia in these cardiac allografts was remarkably less than that in cardiac allografts treated with tacrolimus. Addition of anti-ICOS antibody or ICOSIg to MLR resulted in inhibition of T-cell proliferation. Inhibition of T-cell proliferation with ICOSIg and CTLA4Ig was more effective than that with ICOSIg alone. Thus, ICOS appears to be an important regulator of T-cell activation, and may be an effective therapy in clinical cardiac transplantation.

Forty-hour heart preservation by intermittent perfusion

The shortage of donor hearts due to the limited duration of organ storage remains a major restricting factor for clinical cardiac transplantation. This study evaluated the efficacy of intermittent perfusion (IP) in preserving functonal viability of the cardiac explant. Mongrel dogs (n=10, 20-25 kg) of either sex were randomly paired as donors and recipients of orthotopic heart transplantation. An additional 4 hearts were used for magnetic resonance spectroscopic measurement of myocardial high energy phosphates during storage. The anesthetized donor dog was mechanically ventilated, the chest was opened, and the heart was cardioplegically arrested. After excision, the heart was stored at 0°C for 40 hr by immersion interrupted by IP at predetermined intervals. Myocardial ATP decreased during storage but was repleted by IP. At 40 hr, ATP remained at 73±8% of the prestorage level; heart wet weight (175±20 g) increased insignificantly. After storage, the donor heart was orthotopically transplanted into the recipient maintained on cardiopulmonary bypass. After reestablishment of coronary perfusion, the grafted heart was defibrillated and paced until a sinuous rhythm was obtained. All five recipients were then successfully weaned off the bypass. Heart rate (HR, bpm), systolic and diastolic arterial pressure (SAP and DAP, mm Hg), and the rate of change of aortic pressure during systole and diastole (mm Hg/sec) were recorded for 6 hr, then the animals were euthanized. At the end of 6 hr of observation, HR was 96±4; SAP, 111±6; DAP, 53±8; dp/dt, 1031±152; -dp/dt, -350±52. In contrast, dogs receiving hearts preserved by cold storage alone for the equivalent period would not be expected to regain cardiac function. In conclusion, IP significantly enhanced the functional and metabolic preservation of the canine cardiac explant, and offers a promising intervention for long-term preservation of the human donor heart. (Supported by American Heart Association, New York Affiliated, Inc. and 21st Century Medicine, Inc.).

Morphometric analysis of miniature swine hearts as potential human xenografts.

Miniature swine are considered to be potential donors for clinical cardiac transplantation. However, it is unclear how an appropriately sized porcine donor will be selected for a particular human recipient. To address this issue, we performed a morphometric study of the swine heart using transthoracic echocardiography (n = 26) to determine the diameters of the aortic annulus and root, pulmonary artery annulus, and mitral valve annulus. We also obtained direct ex vivo measurements of swine heart weight and linear dimensions (n = 71). Relationships between a swine's height, weight, length, chest circumference and these internal and external cardiac dimensions are described. The strongest correlations were found between a pig's body length and its aortic annulus and root diameters (r-values = 0.97). These relationships are accurately described by univariate linear regression models. By cross-relating our morphometric measurements of aortic annulus diameter in the miniature swine with normative human data, we were able to develop a nomogram, relating swine length and human height, which predicts which miniature swine would donate the best size-matched heart for a particular human recipient.

Editorial: Cardiac transplantation for auxiliary circulatory support

In this issue, Khaghani and associates report their experience with heterotopic cardiac transplantation in infants and children. This study is a welcome addition to the literature from the group at Harefield, which has assembled an extensive experience with this procedure. The concept of using the transplanted heart as an auxiliary pump was introduced in 1964.1Reemtsma K The heart as a test organ in transplantation studies.Ann NY Acad Sci. 1964; 120: 778-785PubMed Google Scholar, 2McGough EC Brewer PL Reemtsma K The parallel heart: studies of intrathoracic auxiliary cardiac transplants.Surgery. 1966; 60: 153-158Google Scholar These studies demonstrated that the auxiliary heart, placed in parallel with the animal's own heart, could sustain the circulation when the native heart was nonfunctional. This concept of assisted circulation now has been widely applied in the use of mechanical support, such as ventricular assist devices. It has been applied sporadically in clinical cardiac transplantation, and this report summarizes the experience in infants and children. The authors report that in a total of 1278 cardiac transplants, 104 (8.1% of the total experience) were placed in the intrathoracic, heterotopic position. This report focuses on 12 infants and children, nine of whom survive. The indications that the authors cite are (1) presence of fixed high pulmonary vascular resistance, (2) availability of undersized donors, and (3) the expectation of a degree of recipient heart recovery. It is encouraging that all patients showed a decrease in pulmonary artery pressure and all showed a normal growth pattern. It seems probable that the application of auxiliary cardiac support, both with transplants and with mechanical devices, will continue. The use of this approach for transplantation into patients with high, fixed pulmonary vascular resistance seems established. The further development of ventricular assist devices similarly will usually use this approach in preference to in situ replacement of the heart. Its usefulness in treating patients with reversible myocardial disease is intriguing but not yet established. The increasingly severe shortage of donor hearts has resulted in the use of marginally acceptable organs, and the use of such transplants as auxiliary pumps has certain advantages. Similarly, as cardiac xenografts approach clinical use, the auxiliary position rather than in situ replacement has appeal in this experimental area with uncertain outcome. Finally, a comment about nomenclature: The term heterotopic is nonspecific and refers to placement of the heart in any abnormal position. In the literature of experimental cardiac transplantation, heterotopic often refers to cervical or abdominal placement of the heart. The term piggyback is flippant and should be discarded. I recommend the terms auxiliary and parallel, as used in the original publications of this technique. These terms appropriately describe the function of the transplant, which supplements cardiac function by working in parallel with the native heart.

Immunomodulation by an Adenylate Cyclase Activator, NKH477,in Vivoandin Vitro

Cyclic adenosine monophosphate (cAMP) is an intracellular second messenger which modulates T cell function. NKH477 is a direct adenylate cyclase activator derived from forskolin and now under clinical investigation as a positive inotropic agent. While the immunosuppressive effects of forskolin on lymphocytes have been reported, little is known about its effectsin vivo.In this study, we investigated whether NKH477 has immunosuppressive effects in mice, namely on cardiac allograft survival, and on the generation of cytotoxic T lymphocytes (CTL), T cell proliferation in mixed lymphocyte reaction (MLR), and production of interleukin-2 (IL-2) in MLR and in mitogen response. We assessed the effects of standard immunosuppressant cyclosporin A (CsA) on IL-2 production and on allograft survival to estimate the intensity of rejection in this acute rejection model. Saline-treated C57BL/6 (H-2b) mice rejected DBA/2 (H-2d) cardiac allografts with a median graft survival time of 10 days. In contrast, median graft survival was prolonged to 12 and 15 days in mice treated with NKH477 at 1 and 3 mg/kg/day, respectively (P< 0.01 vs control). The equivalent dose of CsA (40 mg/kg/day) to the maintenance dose after clinical cardiac transplantation prolonged median graft survival time to 15.5 days, indicating that high dose of NKH477 was as efficacious as lower dose of CsA. Addition of NKH477 to the culture medium suppressed the generation of CTL, T cell proliferation in MLR, and production of IL-2 in MLR and in mitogen response. These results suggest that NKH477 exerts a beneficial effect on murine cardiac allograft survival by modulating T cell function.

Donor heart endothelial cells as targets for graft infiltrating lymphocytes after clinical cardiac transplantation

The in vitro cytotoxic reactivity of allograft infiltrating cells cultured from endomyocardial biopsies was tested against endothelial cells (EC) isolated from the own donor heart. EC derived from pieces of atrium were found to be proper targets for graft infiltrating cytotoxic T cells from four patients. The specificity of this cytotoxicity was further analysed by cold target inhibition studies and blocking with anti-CD3, anti-CD4 or anti-CD8 monoclonal antibodies. The experiments revealed that, besides a clearly HLA directed recognition, a more heterogeneous, multispecific response can be found, which might be partially explained by the activity of EC specific T cell clones. We conclude that this system provides a valuable approach to investigate the reactivity of graft infiltrating cells against EC in relation to the clinical course of the transplantation.

Superior qualities of University of Wisconsin solution for ex vivo preservation of the pig heart

The components of the University of Wisconsin solution have the potential to enhance and extend heart preservation. We have evaluated University of Wisconsin solution by comparing it with St. Thomas' Hospital cardioplegic solution in the isolated pig heart subjected to 8 hours of ischemia at 4 degrees C (n = 6 in each). The hearts were perfused ex vivo with enriched autologous blood for the control and the postpreservation assessments. Morphologic, metabolic, and functional evaluations were performed. Left and right ventricular function as assessed by the slope values of systolic and diastolic pressure-volume relationships of isovolumically contracting isolated heart was better preserved by University of Wisconsin solution (percent reduction: left ventricular systolic, 52.4% +/- 5.5% versus 17.7% +/- 6.7% [p less than 0.001]; right ventricular systolic, 125.6% +/- 46.4% versus 65.5% +/- 31.4% [p less than 0.05]; right ventricular diastolic, 112.3% +/- 48.7% versus 40.2% +/- 31.3% [p less than 0.02] after St. Thomas' Hospital and University of Wisconsin preservation, respectively). Postischemic recovery of left ventricular rate of rise of pressure and myocardial oxygen consumption were significantly improved after University of Wisconsin preservation (percent reduction, rate of rise of pressure: St. Thomas' Hospital 39.3% +/- 8.1%; University of Wisconsin 18.1% +/- 4.6%; percent reduction, myocardial oxygen consumption St. Thomas' Hospital 55.1% +/- 6.9%, University of Wisconsin 24.8% +/- 6.7%; p less than 0.001). Microvascular functional integrity as assessed by coronary vascular resistance was well maintained throughout the postischemic period and was similar to the preischemic control value in the University of Wisconsin group. By contrast, a significant increase was found at the beginning of postpreservation reperfusion, with a progressive rise thereafter in the St. Thomas' Hospital group (p less than 0.001). Preservation of myocardial adenosine triphosphate was improved and energy charge was unchanged after 8 hours of ischemia and reperfusion in the University of Wisconsin-preserved hearts compared with the St. Thomas' Hospital-preserved hearts (p less than 0.01). Electron microscopic examination revealed substantially better preservation of the contractile apparatus after preservation with University of Wisconsin solution. Myocytes from hearts receiving University of Wisconsin solution, unlike those given St. Thomas' Hospital solution, showed relaxed myofibrils with prominent I-bands. We conclude that University of Wisconsin solution has the potential to improve the preservation of the heart and possibly prolong the ischemic period in clinical cardiac transplantation.

Current Issues in Clinical and Experimental Cardiac Transplantation

Cardiac transplantation is now an accepted therapy for end-stage heart failure not amenable to usual medical or surgical therapy. Despite excellent survival, important unresolved problems remain. This review focuses on current issues in cardiac transplantation. These controversial areas of active interest include the relationship between histocompatibility matching and the results of transplantation, the causes and treatment of graft coronary arteriopathy, and the mechanism of reversible heart failure that is caused by reversible transplant rejection. These fascinating unresolved issues remain significant challenges to those involved in the care of heart transplant recipients. Their resolution will improve significantly the results of this form of therapy.