Jonathan A. Cardella

Cell Death in Human Lung Transplantation: Apoptosis Induction in Human Lungs During Ischemia and After Transplantation

OBJECTIVE To examine the presence and extent of apoptosis as well as the affected cell types in human lung tissue before, during, and after transplantation. SUMMARY BACKGROUND DATA Apoptosis has been described in various human and animal models of ischemia-reperfusion injury, including heart, liver, and kidney, but not in lungs. Therefore, the presence of apoptosis and its role in human lungs after transplantation is not clear. METHODS Lung tissue biopsies were obtained from 20 consecutive human lungs for transplantation after cold ischemic preservation (1-5 hours), after warm ischemia time (during implantation), and 30, 60, and 120 minutes after graft reperfusion. To detect and quantify apoptosis, fluorescent in situ end labeling of DNA fragments (TUNEL assay) was used. Electron microscopy was performed to verify the morphologic changes consistent with apoptosis and to identify the cell types, which were lost by apoptosis. RESULTS Almost no evidence of apoptosis was found in specimens after immediate cold and warm ischemic periods. Significant increases in the numbers of cells undergoing apoptosis were observed after graft reperfusion in a time-dependent manner. The mean fraction of apoptotic cells at 30, 60, and 120 minutes after graft reperfusion were 16.6%, 22.1%, and 34.9% of total cells, respectively. Most of the apoptotic cells appeared to be alveolar type II pneumocytes, as confirmed by electron microscopy. CONCLUSIONS Programmed cell death (apoptosis) appears to be a significant type of cell loss in human lungs after transplantation, and this may contribute to ischemia-reperfusion injury during the early phase of graft reperfusion. This cell loss might be responsible for severe organ dysfunction, which is seen in 20% of patients after lung transplantation. Therefore, this work is of importance to surgeons for the future development of interventions to prevent cell death in transplantation.

Transtracheal gene transfection of donor lungs prior to organ procurement increases transgene levels at reperfusion and following transplantation

BACKGROUND Gene therapys potential to modify donor organs to better withstand the process of transplantation has yet to be realized. To determine whether gene transfection is feasible to treat the early post-transplant injury of ischemia-reperfusion, we compared transfection of lungs in the donor prior to organ procurement with transfection of harvested ex vivo lungs in a rat single lung transplant model. METHODS Lewis rats (donor transfection [DT]; n = 4) underwent transtracheal adenoviral-mediated transfection with 10(9) plaque forming unit of the beta-galactosidase reporter gene. Donor lungs were harvested following 6 hours of in vivo post-transfection ventilation, and then preserved for 6 hours at 4 degrees C prior to left single-lung transplantation. Ex vivo transfection was performed following organ retrieval; lungs were then preserved at 4 degrees C for 6 hours (EVT6h; n = 6) and 12 hours (EVT12h; n = 6) prior to transplantation. Lung transgene expression was measured by chemiluminescence at reperfusion, and at 2 hours following lung transplantation. RESULTS Donor transfection lungs showed significantly higher levels of transgene expression as compared with EVT lungs at the time of reperfusion (DT = 3,408+/-1,301 relative light units/mg protein; EVT6h = 218+/-7; EVT12h = 213+/-26; p < 0.02) and at 2 hours after lung transplantation (DT = 2900+/-870; EVT6h = 62+/-27; EVT12h = 123+/-21; p < 0.005). Transgene expression measured in the heart, liver, kidney, and serum from DT rats demonstrated virtually no evidence of collateral transfection at 12 hours post-transfection (all <5.0). CONCLUSIONS Gene transfection of donor lungs produces significantly higher levels of transgene expression in lungs at the critical time of reperfusion and in the early period following lung transplantation as compared to ex vivo transfection of cold preserved lungs. Transtracheal donor-lung transfection does not appear to result in collateral transfection of other transplantable organs. Local adenoviral-mediated transfection of the lungs is possible in the multiorgan donor prior to organ procurement and may provide the optimal strategy for gene therapeutic manipulations to address post-transplant ischemia-reperfusion injury.

Compliance, attitudes and barriers to post‐operative colorectal cancer follow‐up

RATIONALE Meta-analyses demonstrate that surveillance following curative-intent colorectal cancer (CRC) surgery can improve survival. Our multidisciplinary team adopted a stringent CRC follow-up (FU) guideline in 2000. The purpose of this study was to assess adherence and barriers to FU for CRC. METHODS Patients with primary CRC aged 19-75 years, treated with curative intent surgery from July 2000 to December 2002 were identified from a prospective database. Compliance with FU was assessed primarily by chart review. We also surveyed patients and providers to explore attitudes and barriers to surveillance adherence using tenets of the Health Belief Model. RESULTS 96 patients met inclusion criteria and were appropriate for FU. Median FU was 34 months. Guideline targets were met for 70% of clinic visits; 49% of carcinoembryonic antigen (CEA) determinations; and 62% of abdominal imaging studies. Post-operative colonoscopy did not occur in 6/93 patients. Seventy per cent of health care providers and 55% of patients completed a survey. Access to testing and confusion about which provider orders investigations were identified as important barriers to FU. CONCLUSION Patterns of CRC FU were widely variable despite implementation of a guideline. Despite patient and provider agreement with the principles of CRC FU, adoption was inhibited by confusion among multiple providers regarding investigation coordination.