Functional recovery of preserved livers following warm ischemia: improvement by machine perfusion preservation.

BACKGROUND: Hypothermic machine perfusion preservation has the potential to relieve the current donor shortage problem by reclaiming and preserving marginal donor organs including those from viable non-heart-beating donors. A number of problems exist with the current machine perfusion technology for preserving livers, and much research is needed to determine the clinical impact of this technology in preserving non-heart-beating donor livers. METHODS: This study was conducted to compare the poststorage function and microcirculation of simple cold stored and machine perfusion preserved livers that had experienced 30 min of warm ischemia followed by a 10 hr preservation period. In an isolated rat liver perfusion model, lactate dehydrogenase activity, indocyanine green secretion, and portal pressure values were determined at major time points. An intravital microscopy was conducted to assess microcirculation. RESULTS: The results showed an increase in flow homogeneity of machine perfused livers, which correlated with the reduction in portal pressure when compared with simple cold storage (5.4+/-0.4 vs. 8.7+/-0.6 mm Hg). A reduction in lactate dehydrogenase levels in the perfusate (333+/-22 vs.103+/-8 U/L) and an increase in bile production of the machine perfused livers (4.9+/-0.5 vs. 33.2+/-1.7 microg/min/g liver) and indocyanine green secretion (11.7+/-1.7 vs. 21.2+/-2.1 Abs/g bile) were observed at all time points (mean+/-SE of final point given). Intravital microscopic examination indicated that large regions of non flow, as indicated by the absence of fluorescein isothiocyanate-labeled albumin, were observed in the simple cold stored tissue, whereas machine perfused liver showed increase flow homogeneity. Values of bile production, indocyanine secretion, and cellular damages were comparable with controls. Histologic examination confirmed that simple cold stored tissue displayed increased vacuolization, and machine perfused tissue showed regions of normal hepatic structure. CONCLUSION: These results suggest that machine perfusion for 10 hr improves both poststorage function and microcirculation while reducing cellular damage of liver tissue that has experienced 30 min of warm ischemia, when compared with simple cold storage. Further studies need to be conducted, but this study suggests that machine perfusion preservation has the potential to reclaim and preserve liver tissues after warm ischemic insult.

Lack of donor hyporesponsiveness and donor chimerism after clinical transplantation of the hand.

BACKGROUND: Composite tissue allografts offer great potential in reconstructive surgery. However, the risks of immunosuppression and graft-versus-host disease (GVHD) after transplantation of vascularized bone in these grafts are significant. Transplantation of vascularized bone also may confer donor hematopoietic chimerism and, potentially, tolerance. We have followed two hand transplant recipients for more than 1 year to determine the level of chimerism and possible donor-specific tolerance, in addition to possible GVHD. METHODS: We performed kinetic studies on peripheral blood of two subjects after hand transplantation that included portions of the radius and ulna. We evaluated donor-specific reactivity, chimerism, and antibody production. RESULTS: Donor-specific tolerance did not develop clinically or in mixed lymphocyte reaction. The first subject recovered an excellent in vitro response to phytohemagglutinin, donor and third-party alloantigen, and by month 4 and at month 12 also recovered the ability to respond to Epstein-Barr virus. The second subject also demonstrated good in vitro proliferative responses, which were attenuated by immunosuppression. No phenotypic changes in mature hematopoietic lineages were detected by four-color flow cytometry other than those expected in response to immunosuppression. Donor chimerism was not detectable using four-color flow cytometry. Microchimerism (approximately 1:75,000 cells) was observed at the level of detection in some of the early posttransplantation specimens and was undetectable thereafter. CONCLUSIONS: In this particular transplantation and immunosuppressive regimen, the composite tissue allograft with vascularized bone marrow did not provide the immunologic benefit of tolerance induction nor cause GVHD.

Survival transplantation of preserved non-heart-beating donor rat livers: preservation by hypothermic machine perfusion.

BACKGROUND: Non-heart-beating donor (NHBD) livers are an untapped source with the potential to provide relief to the current donor shortage problem. Hypothermic machine perfusion (MP) has the potential to reclaim and preserve these marginal donor organs. METHODS: This study compared 5-day survival in a rat NHBD liver transplantation model with simple cold storage (SCS) and MP-preserved tissues that had experienced 30 min of warm ischemia followed by a 5-hr preservation period with the University of Wisconsin solution. Total release of lactate dehydrogenase (LDH) and alanine aminotransferase (ALT) were determined at major time points. Bilirubin levels and histology were examined after 5-day survival. RESULTS: Six of seven control livers and five of six MP livers survived, whereas SCS tissues had survival in zero of seven. The results showed that MP livers had reduced release of LDH and ALT after 5 hr of storage, 5.07+/-1.42 and 2.02+/-0.69 U (mean+/-SE), respectively, compared with SCS, 15.54+/-0.81 and 3.41.3+/-0.73 U, respectively. Bilirubin values after 5-day survival of MP livers (1.17+/-0.49 mg/dL) were comparable to controls (0.91+/-0.36 mg/dL). Histology confirms that SCS displayed increased necrosis and MP tissue showed regions of near normal hepatic structure. CONCLUSIONS: These results suggest that MP for 5 hr improves survival and reduces cellular damage of liver tissue that has experienced 30 min of warm ischemia when compared with SCS tissues. Further studies need to be conducted, but this study suggests that MP preservation has the potential to reclaim and preserve NHBD liver tissues.

Ex-vivo study of flow dynamics and endothelial cell structure during extended hypothermic machine perfusion preservation of livers.

Liver transplantation is often the only effective treatment for end stage liver diseases resulting from cirrhosis, hepatitis, progressive jaundice, and biliary atresia. Hypothermic machine perfusion (HMP) preservation may enhance donor pool by extending preservation time and reclaiming marginal donor livers including those from non-heart beating donors (NHBD), as demonstrated in the kidney. However, current HMP protocols have not been successful in improving extended preservation of livers and the major cause of preservation injury remains unknown. An intravital microscopy study was conducted to understand the flow dynamics of sinusoidal perfusion during 24h HMP with cold modified University of Wisconsin (UW) solution. Fluorescein isothiocynate (FITC) labeled albumin was utilized to visualize microvascular space and FITC labeled red blood cells (RBCs) were used to visualize flow dynamics during HMP. A heterogeneous flow pattern with regions of red cell stasis was observed after 24-h HMP. To examine the cause of red cell stasis, intravital and confocal microscopy studies of endothelial cells (ECs) structure labeled with DiI acetylated low-density lipoprotein (DiI acLDL) were conducted. These studies suggest that morphological changes in EC structures occurred during 24h HMP, which may cause obstruction to the sinusoidal flow. Histological findings confirm these results. As a result, heterogeneous flow pattern, red cell stasis, and edema occur, which may lead to the failure of these tissues following extended HMP.