The effects of brain death and ischemia on tolerance induction are organ-specific.

We have previously shown that 12 days of high-dose calcineurin inhibition induced tolerance in MHC inbred miniature swine receiving MHC-mismatched lung, kidney, or co-transplanted heart/kidney allografts. However, if lung grafts were procured from donation after brain death (DBD), and transplanted alone, they were rejected within 19-45 days. Here, we investigated whether donor brain death with or without allograft ischemia would also prevent tolerance induction in kidney or heart/kidney recipients. Four kidney recipients treated with 12 days of calcineurin inhibition received organs from donors rendered brain dead for 4 hours. Six heart/kidney recipients also treated with calcineurin inhibition received organs from donors rendered brain dead for 4 hours, 8 hours, or 4 hours with 4 additional hours of cold storage. In contrast to lung allograft recipients, all isolated kidney or heart/kidney recipients that received organs from DBD donors achieved long-term survival (>100 days) without histologic evidence of rejection. Proinflammatory cytokine gene expression was upregulated in lungs and hearts, but not kidney allografts, after brain death. These data suggest that the deleterious effects of brain death and ischemia on tolerance induction are organ-specific, which has implications for the application of tolerance to clinical transplantation.

Donor brain death inhibits tolerance induction in miniature swine recipients of fully MHC-disparate pulmonary allografts.

We have previously shown that a short course of high-dose tacrolimus induces long-term tolerance to fully mismatched lung allografts procured from healthy MHC-inbred miniature swine. Here, we investigate whether donor brain death affects tolerance induction. Four recipient swine were transplanted with fully mismatched lung grafts from donors that were rendered brain dead and mechanically ventilated for 4 h before procurement (Group 1). These recipients were compared to two control groups (Group 2: 4 h of donor ventilation without brain death [n = 5]; and Group 3: no donor brain death with <1 h of ventilation [n = 6]). All recipients were treated with a 12-day course of tacrolimus. In contrast to both groups of control animals, the swine transplanted with lung allografts from brain dead donors all rejected their grafts by postoperative day 45 and showed persistent responsiveness to donor antigen by MLR. Several additional swine underwent brain death induction and/or mechanical ventilation alone to determine the effects of these procedures on the expression of proinflammatory molecules. Significant increases in serum concentrations of IL-1, TNF-α and IL-10 were seen after brain death. Upregulation of IL-1 and IL-6 gene expression was also observed.

The indirect alloresponse impairs the induction but not maintenance of tolerance to MHC class I-disparate allografts.

We studied the effects of indirect allorecognition on the induction and maintenance phases of tolerance in miniature swine cotransplanted with heart and kidney allografts. MHC class I-mismatched heart and kidney grafts were cotransplanted in recipients receiving CyA for 12 days. Recipients were unimmunized or immunized with a set of donor-derived or control third-party MHC class I peptides either 21 days prior to transplantation or over 100 days after transplantation. T-cell proliferation, delayed type hypersensitivity reaction (DTH) and antibody production were assessed. All animals injected with donor MHC class I peptides developed potent indirect alloresponses specific to the immunizing peptides. While untreated recipients developed stable tolerance, all animals preimmunized with donor allopeptides rejected kidney-heart transplants acutely. In contrast, when peptide immunization was delayed until over 100 days after kidney-heart transplantation, no effects were observed on graft function or in vitro measures of alloimmunity. Donor peptide immunization prevented tolerance when administered to recipients pre transplantation but did not abrogate tolerance when administered to long-term survivors post transplantation. This suggests that the presence of T cells activated via indirect allorecognition represent a barrier to the induction but not the maintenance of tolerance.

Myocardial tissue engineering and regeneration as a therapeutic alternative to transplantation.

Ischemic cardiomyopathy leading to congestive heart failure remains the leading source of morbidity and mortality in Western society and medical management of this condition offers only palliative treatment. While allogeneic heart transplantation can both extend and improve the quality of life for patients with end-stage heart failure, this therapeutic option is limited by donor organ shortage. Even after successful transplantation, chronic cardiac rejection in the form of cardiac allograft vasculopathy can severely limit the lifespan of the transplanted organ. Current experimental efforts focus on cellular cardiomyoplasty, myocardial tissue engineering, and myocardial regeneration as alternative approaches to whole organ transplantation. Such strategies may offer novel forms of therapy to patients with end-stage heart failure within the near future.

Multiparameter flow cytometric approach for simultaneous evaluation of T lymphocyte-endothelial cell interactions.

Since vascular endothelium is now recognized as an immunologically active tissue, a better understanding of the relationship between endothelial cells and T lymphocytes is critical to the field of solid organ transplantation. Investigations of endothelial cell-T cell interactions have been limited by methodology. We developed a flow cytometric method allowing for concurrent investigation of multiple cell populations within the same culture that can be applied to these complex interactions. Allogeneic CD8+ or CD4+ T cells labeled with 5,6-carboxyfluorescein diacetate succinimidyl ester (CFSE) were added to a murine endothelial cell monolayer, in which endothelial proliferation was not inhibited by irradiation or addition of a cell cycle-blocking agent. At specific time points, the coculture was analyzed by flow cytometry. T-cell proliferation could be detected by gating on the T-cell subset and evaluating the CFSE fluorescence peaks. By directly analyzing cellular division, we minimized erroneous interpretation of the data encountered by previous studies, which utilized (3)H-thymidine incorporation as sole measure of proliferation. Further subgating on cells that divided facilitated the study of CD8+ lymphocyte activation, differentiation, and acquisition of effector function. By gating on the endothelial cell population, phenotypic changes such as upregulation of surface MHC molecules or immune-mediated apoptosis could be detected. In conclusion, we present a flow cytometric approach that could have important applications for clinical immunological monitoring in allogeneic or xenogeneic transplantation, and might provide the requisite information to better tailor immunotherapy to prevent chronic rejection.

Transmission of donor cancer into cardiothoracic transplant recipients.

BACKGROUND: The demand for transplantable organs exceeds donor supply. Patients with central nervous system (CNS) or other tumors are controversial donors, and the donor cancer transmission rates in cardiothoracic transplant recipients have not been determined. The Israel Penn International Transplant Tumor Registry (IPITTR) was queried to define the risk of donor cancer transmission in cardiothoracic transplant recipients. METHODS: All heart, lung, or heart-lung recipients of organs from donors with a history of malignancy were reviewed. Donor and recipient demographics, histologic findings, and recurrence were reviewed. RESULTS: Twenty-two patients received 17 hearts, 3 lungs, and 2 heart-lung transplants from donors with known CNS or other malignancies. No malignancy transmissions were noted with astrocytomas (n = 3) or glioblastomas (n = 1), except a medulloblastoma that recurred at 6 months. The transmission rate for CNS tumors was 17% (1 of 6), and 1- and 3-year survivals were 67% and 50%, respectively. The most common non-CNS donor cancer was renal cell carcinoma (n = 5). Two renal cell cancer transmissions occurred, both when vascular extension was present. The most aggressive tumor transmission was choriocarcinoma (n = 2) and melanoma (n = 2). Two of 3 choriocarcinomas metastasized with 67% mortality, and both melanomas were transmitted and resulted in death. Other donor cancers included angiosarcoma (n = 2), cervical (n = 1), lung (n = 1), prostate (n = 1), and a liver adenocarcinoma. The transmission rate for all non-CNS groups was 56% (9 of 16) with a 2-year survival of 40%. CONCLUSIONS: The IPITTR experience indicates that tumor transmission is high (10 of 22, 45%) in cardiothoracic transplant recipients. Similar to intra-abdominal organ recipients in the IPITTR, (1) renal cell carcinomas without capsular invasion appear safe with no transmission, (2) vascular invasion in renal cell carcinoma appears to result in early tumor transmission, and (3) melanoma and choriocarcinoma have high rates of transmission with early and almost universal death.

Bronchogenic carcinoma complicating lung transplantation.

BACKGROUND: Malignancy is a well-recognized complication of solid-organ transplantation. Although a variety of malignancies have been reported in lung transplant recipients, a paucity of information exists regarding the incidence and clinical course of bronchogenic carcinoma in this patient population. METHODS: We conducted a retrospective cohort study of our lung transplant experience at the University of Pennsylvania. RESULTS: We identified 6 patients with bronchogenic carcinoma detected at the time of, or developing after, transplantation. The incidence of bronchogenic carcinoma was 2.4%. All patients with lung cancer had a history of smoking, with an average of 79 +/- 39 pack-years. A total of 5 patients had chronic obstructive pulmonary disease, and 1 had idiopathic pulmonary fibrosis. Lung cancers were all of non-small-cell histology and first developed in native lungs. Three patients had bronchogenic carcinoma at the time of surgery. The remaining 3 patients were diagnosed between 280 and 1,982 days post-transplantation. Of the 6 patients, 4 presented with a rapid course suggestive of an infectious process. The 1- and 2-year survival rates after diagnosis were 33% and 17%, respectively. CONCLUSION: Lung transplant recipients are at risk for harboring or developing bronchogenic carcinoma in their native lungs. Rapid progression to locally advanced or metastatic disease commonly occurs, at times mimicking an infection. Bronchogenic carcinoma should be considered in the differential diagnosis of pleuroparenchymal processes involving the native lung.

Characteristics and outcomes of patients with sarcoidosis listed for lung transplantation.

STUDY OBJECTIVES: To characterize the course of patients with advanced sarcoidosis who have been listed for lung transplantation and to identify prognostic factors for death while they are on the waiting list. DESIGN: Retrospective cohort study. SETTING: Tertiary-care university hospital. PATIENTS: Forty-three patients with sarcoidosis who have been listed for lung transplantation at the University of Pennsylvania Medical Center. METHODS: A multivariable explanatory analysis using a Cox proportional hazards model was performed to determine risk factors that are independently associated with mortality while patients await transplantation. RESULTS: Twenty-three of the 43 patients (53%) died while awaiting transplantation. The survival rate of listed patients (as determined by the Kaplan-Meier method) was 66% at 1 year, 40% at 2 years, and 31% at 3 years. In a univariate analysis, the following factors were significantly associated with death on the waiting list: PaO(2) < or = 60 mm Hg (relative risk [RR], 3.4; 95% confidence interval [CI], 1.2 to 9.3); mean pulmonary artery pressure > or = 35 mm Hg (RR, 3.2; 95% CI, 1.1 to 9.5); cardiac index < or = 2 L/min/m(2) (RR, 2.8; 95% CI, 1.2 to 6.6), and right atrial pressure (RAP) > or = 15 mm Hg (RR, 7.6; 95% CI, 3.0 to 19.3). Multivariable analysis revealed that RAP > or = 15 mm Hg was the only independent prognostic variable (RR, 5.2; 95% CI, 1.6 to 16.7; p = 0.006). Twelve patients underwent lung transplantation. Survival after transplantation determined by the Kaplan-Meier method was 62% at both 1 and 2 years, and 50% at 3 years. CONCLUSIONS: Patients with advanced sarcoidosis awaiting lung transplantation have a high mortality rate with a median survival of < 2 years. Mortality is most closely linked to elevated RAP. While earlier referral may diminish the mortality rate of patients on the waiting list for transplantation, further improvements in posttransplantation outcomes will be necessary to ensure that this procedure truly bestows a survival benefit.

Emergent lung retransplantation after discovery of two primary malignancies in the donor.

A 50-year-old woman underwent single lung transplantation for advanced chronic obstructive pulmonary disease. Shortly after the procedure, it was discovered that the donor suffered from both a renal cell carcinoma and a spindle-cell sarcoma of the ascending aorta, which had metastasized to the spleen. The patient was emergently listed for a retransplantation and underwent bilateral lung transplantation after a new donor became available 4 days after the initial transplantation procedure. After 24 months, the patient is without evidence of malignancy. This case illustrates the role of immediate retransplantation for patients who have inadvertently received thoracic organs from donors harboring occult malignancies.

Application of "double bridge mechanical" resuscitation for profound cardiogenic shock leading to cardiac transplantation.

BACKGROUND: In patients with acute profound cardiogenic circulatory failure unresponsive to conventional resuscitation, we instituted immediate aggressive application of extracorporeal membrane oxygenation (ECMO) to restore circulatory stability. Long-term hemodynamic support was accomplished with an early "bridge" to ventricular assist device (VAD) before definitive treatment with cardiac transplantation. METHODS: A respective review of ECMO and VAD data registries was instituted. RESULTS: From May 1996 to July 2000, 23 patients were placed on ECMO support for profound cardiogenic circulatory failure. Eleven patients (47%) were withdrawn from support due to severe neurologic injury or multisystem organ failure. Three patients (13%) were weaned off ECMO with good outcome. Nine patients (39%) were transferred to a VAD. Two patients expired while on VAD support, and 7 of the VAD-supported patients (78%) survived to transplantation. Overall survival was 43%. CONCLUSIONS: Emergent ECMO support is a salvage approach for cardiac resuscitation once conventional measures have failed. In neurologically intact patients, the early transfer to a VAD quickly stabilizes hemodynamics, avoids complications, and is essential for long-term circulatory support before definitive treatment with cardiac transplantation.

A simple method for culturing mouse vascular endothelium.

Vascular endothelium is an important site for a wide array of immunological processes such as inflammation, atherosclerosis and allograft rejection. Culture methods of mouse vascular endothelium would provide an important in vitro correlate to immunological murine in vivo models. We describe a simple method to culture mouse vascular endothelium from thoracic aorta. Our cultured cells express typical phenotypic (CD105, CD31, CD106), morphological and ultrastructural (intercellular junctions, Weibel-Palade bodies) markers of vascular endothelium. They also possess functional receptors for uptake and processing of acetylated low-density lipoproteins. The mouse vascular endothelium within our system expresses high levels of MHC class I and MHC class II after activation with IFN-gamma. In addition, these cells express the accessory molecules CD80 and CD54, while they lack constitutive expression of CD86 and CD40, providing them the means to function as antigen presenting cells. Alloreactive CD4(+) and CD8(+) T lymphocytes demonstrate evidence of DNA synthesis after co-culture with activated vascular endothelium indicating their commitment to proliferation. In conclusion, we describe a simple culture system to isolate and grow mouse vascular endothelium, which provides a powerful tool to study biological interactions in vitro.