Ischemic preconditioning of cadaver donor livers protects allografts following transplantation

BACKGROUND: Ischemic preconditioning (IP) has been shown in animal models to protect livers against ischemia/reperfusion injury. The aim of this clinical study is to investigate whether IP of cadaver livers prior to retrieval confers protection on the allografts. METHODS: Cadaveric donor livers were subjected to IP prior to retrieval by clamping of the hepatic pedicle for 10 min followed by reperfusion. Biopsies were obtained from the preconditioned (n=9) and control nonpreconditioned (n=14) liver transplants prior to and 2 hr following reperfusion. Cryosections were stained with antibodies against neutrophils and platelets. RESULTS: IP livers were associated with significantly lower serum levels of aspartate aminotransferase (240+/-98 IU/L vs. 382+/-163 IU/L; P>0.016) and lactate (0.81+/-0.07 mmol/L vs. 1.58+/-0.9 mmol/L; P>0.018) 24 hr following transplantation. Furthermore, recipients of IP livers spent a significantly shorter time in the intensive care unit following transplantation compared to those given nonpreconditioned allografts (1 vs. 2.8+/-1.6 days; P=0.0008). Increases in neutrophil infiltration were detected in 6/14 (43%; P=0.022) and in CD41 deposition in 5/14 (36%; P=0.042) of nonpreconditioned livers. However, none of the IP allografts showed any change in the levels of platelets or neutrophil infiltration following transplantation. CONCLUSION: IP is an effective method of protecting cadaver donor allografts from cold ischemia and subsequent reperfusion injury. IP is also associated with a reduction in the nonspecific inflammatory response.     

Activation of mitochondrial apoptotic pathways in human renal allografts after ischemiareperfusion injury

BACKGROUND: Ischemia-reperfusion injury in cadaveric (CAD) kidney allografts is associated with tubular cell injury, delayed graft function, and an increased incidence of acute and chronic rejection. We tested the hypothesis that activation of specific apoptotic pathways represents a mechanism for tubular cell death after CAD kidney transplantation. METHODS: Serial tissue sections from paraffin-embedded needle biopsy specimens obtained at approximately 1 hr of reperfusion after transplantation of 13 CAD and 12 living-related donor (LRD) renal allografts were examined by using the terminal deoxynucleotide transferase-mediated dUTP nick-end labeling assay to detect apoptosis and by immunohistochemistry for expression of key pro-apoptotic molecules (Bax, Bak, tumor necrosis factor receptor [TNFR]-1, Fas, and cytochrome c). RESULTS: Apoptosis was detected primarily in tubular cells, with a mean+/-standard deviation of 6.8+/-2.2 apoptotic cells per 100 cells examined in CAD renal allografts compared with 1.8+/-2.7 cells per 100 in LRD (P<0.001) renal allografts. There was a significant correlation between apoptosis rate and cold ischemia time in CAD (r=0.86, P<0.001) renal allografts. Bax was expressed in 100% of CAD versus 17% of LRD renal allografts (P<0.001), Bak in 92% of CAD versus 17% of LRD renal allografts (P<0.001), and TNFR-1 in 100% of CAD versus 58% of LRD renal allografts (P<0.05). Fas was expressed in only a small number of samples (23% of CAD and 17% of LRD renal allografts, P=not significant). Bax and Bak were expressed predominantly in apoptotic cells. Cytochrome c was detected as a mitochondrial pattern in LRD renal allografts, but in a diffuse cytosolic distribution in CAD renal allografts. CONCLUSIONS: Ischemia-reperfusion injury in CAD kidney transplants is associated with a duration-dependent increase in tubular cell apoptosis, mediated at least in part by activation of mitochondrial pathways.     

Cadaver versus living donor kidneys: impact of donor factors on antigen induction before transplantation.

BACKGROUND: It is widely recognized that living-related donor (LRD) renal allografts have a higher overall graft survival than cadaver donor transplants. We tested the hypothesis that part of this is attributable to LRD kidneys being obtained under optimal conditions from healthy donors, whereas cadaveric kidneys may have experienced injury as a result of inflammatory events around the time of brain death. METHODS: We have performed a comparative immunohistochemical analysis of pretransplant donor biopsies from cadaveric (N = 65) and LRD (N = 29) kidneys to determine any differences that may predispose them to subsequent damage. Cryostat sections were stained with antibodies to leukocytes, adhesion molecules, and human leukocyte antigen (HLA)-DR antigens, and the expression was assessed semiquantitatively. RESULTS: High levels of endothelial E-selectin and proximal tubular expression of HLA-DR antigens, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 were detected in biopsies from cadaveric kidneys, whereas expression of these markers was markedly reduced in LRD kidneys. High levels of tubular antigen expression were significantly associated with traumatic death, prolonged ventilation, and episodes of infection in cadaver donors. Furthermore, the expression of pretransplant tubular antigens in cadaver donor kidneys was significantly associated with early acute rejection following transplantation, suggesting that such kidneys are predisposed to subsequent immune-mediated attack following transplantation. CONCLUSIONS: These results may explain, in part, the superior outcome of LRD allografts compared with cadaver renal allografts.     

Effect of specific neutrophil elastase inhibitor on ischemia/reperfusion injury in rat liver transplantation.

Activated neutrophils have been implicated as playing an important role in ischemia/reperfusion injury of the liver by releasing toxic mediators such as oxygen free radicals and elastases. In the present study, we evaluated the effect of a novel, specific neutrophil elastase inhibitor (ONO-5046) on cold-ischemia/reperfusion injury of the liver allograft in rodents. Livers from male Lewis rats were procured and stored cold (4 degrees C) in lactated Ringer's solution and transplanted orthotopically. Recipients were divided into three groups: Vehicle group, 5-h preservation and vehicle (n = 8); ONO-5046 group, 5-h preservation and administration of ONO-5046 (n = 8); and Control group, minimum preservation only (n = 8). Bile output after reperfusion was significantly larger in the ONO-5046 group compared to the Vehicle group (P < 0.05 or less). Sinusoidal endothelial cell function represented by the serum hyaluronic acid concentration at 120 min after reperfusion of the ONO-5046 group was significantly lower than that in the Vehicle group (17.0 +/- 7.9 vs 36.2 +/- 14.9 ng/ml, P < 0.05), whereas serum transaminase levels 120 min after reperfusion were comparable between the two groups. Liver tissue energy charge 120 min after reperfusion was significantly better in the ONO-5046 group compared to the Vehicle group (P < 0.05). Furthermore, the number of neutrophils infiltrating the allograft after reperfusion was significantly depressed in the ONO-5046 group compared to the Vehicle group (P < 0. 02). These data suggest that the neutrophil elastase might cause liver damage early after reperfusion in cold-stored liver, which can be ameliorated by the administration of a specific neutrophil elastase inhibitor, ONO-5046.     

Expression of IL-8 during reperfusion of renal allografts is dependent on ischemic time

BACKGROUND: Ischemia/reperfusion injury is an inherent consequence of solid organ transplantation that increases tissue inflammation and negatively impacts organ transplant function and survival. This study investigated the expression levels of chemokine and chemokine receptor genes in living versus cadaver donor renal allografts before and after reperfusion. METHODS: This study involved 39 renal transplant patients (19 cadaveric and 20 living donor). The ischemia biopsy was taken just before graft declamping and the reperfusion biopsy 30 min after declamping. Whole-cell RNA was isolated and chemokine (IL-8, CCL2/MCP-1, CXCL10/IP-10 and CCL5/RANTES) and chemokine receptor (CCR2 and CCR5) expression was tested by quantitative PCR. RESULTS: Just prior to declamping, ischemic cadaveric donor grafts had higher expression of CXCL10/IP-10 but not IL-8 or CCL2/MCP-1 than living donor grafts. IL-8 expression increased 50% from ischemia to reperfusion in living donor grafts but increased more than 13-fold during reperfusion of cadaver donor grafts. Increased total ischemia time induced greater IL-8 expression during reperfusion. MCP-1 expression also increased during reperfusion of living and cadaver donor grafts but differences were not observed between the two groups of grafts. RANTES, CCR2, and CCR5 expression did not change in ischemic vs. reperfusion biopsies. CONCLUSIONS: The expression of chemokines directing neutrophil and macrophage recruitment increases during reperfusion of living and cadaveric donor renal allografts. Expression levels of IL-8 correlate with the ischemic time imposed on the renal graft. Early tissue injury may be attenuated by strategies antagonizing chemokines directing the recruitment of neutrophils and macrophages into kidney grafts.     

Intraoperative fluid management of living donor versus cadaveric liver transplant recipients

Living donor liver transplantation has increasingly become an alternative to cadaveric donor liver transplants for select adult patients. Because these cases can be performed electively, living donor recipients may have better compensated liver disease at the time of surgery than cadaver donor recipients. However, it is unknown if this difference would have a significant effect on their intraoperative course. Therefore, we compared the intraoperative fluid management of patients receiving liver grafts from either living or cadaveric donors (n = 25, each group). Patient groups did not differ in demographics or baseline laboratory values. The duration of anesthesia and anhepatic phases were significantly longer in living donor cases (651 +/- 80 minutes vs 409 +/- 20 and 55 +/- 14 vs 45 +/- 6, P < .05). Adjusted for anesthesia time and patient weight, fluid administration (crystalloid and albumin) was not different between the two groups. Intraoperative transfusion requirements were also not significantly different in recipients from living donors versus cadaveric donors with regard to red blood cells, fresh frozen plasma, platelets, and cryoprecipitate. However, arterial oxygenation was better preserved in recipients from living donors. The PaO2/FiO2 (P/F) ratio at the end of the procedure was significantly better in patients receiving livers from living rather than from cadaveric donors (P/F ratio 335 +/- 114 mm Hg vs 271 +/- 174, P < .05). Our results indicate that while intraoperative fluid and transfusion requirements are similar, the impact of transplantation on pulmonary gas exchange is more pronounced in patients receiving organs from cadaveric donors. This difference may arise from longer cold ischemia times present in the cadaveric donor group.     

Leukocyte infiltration and inflammatory antigen expression in cadaveric and living-donor livers before transplant

BACKGROUND: There is evidence to indicate that organs obtained from cadaveric donors may be injured as a result of inflammatory events occurring at around the time of brain death. The aim of this study was to investigate whether there are differences in the expression of proinflammatory molecules between cadaveric and living-donor livers before transplant and to determine whether there is any association with donor factors and posttransplant graft function. METHODS: A comparison of biopsies obtained before implantation from cadaveric (n=22) and living-related donor (LRD) (n=10) livers was performed. Cryostat tissue sections were stained with antibodies to leukocyte subpopulations, adhesion molecules, and human leukocyte antigen class II antigens. RESULTS: Significantly higher levels of CD3+ lymphocytes (1.5%+/-0.8% vs. 0.5%+/-0.3%; P=0.00004), CD68+ monocytes and macrophages (4.0%+/-1.2% vs. 2.7%+/-0.6%; P=0.0003), and Fas-ligand staining (4.2%+/-2.6% vs. 1.5%+/-1.1%; P=0.0003) were detected in cadaveric livers compared with LRD livers before transplantation. Furthermore, higher levels of intercellular adhesion molecule-1 expression were detected in cadaveric donor livers and found to be associated with longer periods of ventilation (P=0.01), infection in the donor (P=0.013), and administration of dopamine (P=0.03). Although there were no differences in neutrophil infiltration between cadaveric and LRD livers, significantly higher levels were found in cadaveric donors with infection (P=0.01). CONCLUSION: This study demonstrates that inflammatory changes occur in cadaveric donor livers and are associated with events occurring during the period of intensive care. These proinflammatory changes did not seem to affect the short-term clinical outcome of cadaveric liver allografts but may contribute to alloimmune responses and impairment of graft function in the long term.     

Non-heart-beating versus cadaveric and living-donor livers: differences in inflammatory markers before transplantation

BACKGROUND: Liver transplantation from non-heart-beating donors (NHBD) has been reintroduced into clinical practice to increase the donor pool; however, little is known about the immune status of NHBD livers. The aim of this study was to assess intragraft cell populations and inflammatory markers in NHBD and to compare the findings with cadaveric and living-related donor (LRD) livers. METHODS: Biopsy specimens were obtained from controlled NHBD (n=9), conventional cadaveric (n=22), and living-donor (n=10) livers at the end of cold storage. Cryostat sections were stained for monocytes-macrophages, T lymphocytes, and intercellular adhesion molecule (ICAM)-1. RESULTS: The levels of leukocyte infiltration in NHBD reflected those found in conventional cadaver donors and were significantly higher than in LRD livers. Similar levels of CD68+ monocytes-macrophages were detected in cadaver (4.0+/-1.2%) and NHBD livers (4.6+/-1.2%) and were significantly greater than in the LRD livers (2.6+/-0.5%, P<0.01). Furthermore, the levels of T lymphocytes in NHBD (1.1+/-0.6%) and cadaver donors (1.5+/-0.8%) were similar, and were higher than in LRD (vs. 0.47+/-0.3%, P<0.05). Twelve of 22 (60%) cadaver livers had high levels of ICAM-1 expression (grade 3), compared with only 1 of 10 (10%) LRD livers (P=0.02). Four of nine (44%) controlled NHBD livers expressed high levels of ICAM-1. CONCLUSIONS: The results demonstrate that livers obtained from controlled NHBD before transplantation are similar to conventional cadaver donors regarding the level of leukocyte infiltration. Nevertheless, lower levels of ICAM-1 were detected in NHBD, suggesting less exposure to inflammatory mediators than conventional cadaver donor livers.     

Intracoronary E-/L-selectin blockade reduces neutrophil infiltration in heart transplantation

BACKGROUND: This study examined the effect of local intracoronary delivery of a unique monoclonal antibody (mAb) to both E- and L-selectin (EL-246) on neutrophil infiltration after global ischemia during cardiac transplantation. METHODS: In 12 ovine heart transplants, allograft coronary arteries were locally perfused with EL-246 (n = 6), or isotype-matched control antibodies (n = 2) or saline (n = 4). At 24 hours posttransplant, myocardium was analyzed for neutrophil infiltration and myocardial water content. RESULTS: The mean number of intramyocardial neutrophils per area (PMN/hpf) was greatly reduced in the allografts perfused with EL-246 (3.45 +/- 0.4 PMN/hpf), compared with an average 6.5 +/- 0.97 PMN/hpf in control hearts (p = 0.004). Peripheral leukocyte counts were unaffected; myocardial water content was not significantly reduced. CONCLUSIONS: Local perfusion of cardiac allografts with blocking antibody EL-246 before reperfusion significantly reduced the neutrophilic infiltration that occurs early after transplantation. Prohibiting neutrophil-endothelial adhesion and transmigration may be useful in decreasing neutrophil-dependent post-reperfusion injury in transplantation and routine cardiac surgery.     

Cold ischemia augments allogeneic-mediated injury in rat kidney allografts

BACKGROUND: Some clinical studies demonstrate that kidney grafts with prolonged cold ischemia experience early acute rejection more often than those with minimal ischemia. The mechanism, however, is putative. Therefore, the aim of this study was to unravel the impact of ischemia on the immune response in rat kidney allografts compared with that in isografts. METHODS: To induce ischemic injury, donor kidneys were preserved for 24 hours in 4 degrees C University of Wisconsin solution before transplantation. No immunosuppression was administered. The histomorphology according to the BANFF criteria for acute rejection and infiltrating cells were assessed at days 1, 2, 3, 4, 6, and 8 post-transplantation. RESULTS: In allografts, exposure of the kidney to ischemia led to a significantly earlier onset of interstitial cell infiltration and tubulitis compared with nonischemic allografts. The BANFF score of interstitial cell infiltration was 1 +/- 0 vs. 0.25 +/- 0.29 at day 3 and 2 +/- 0 vs. 1.25 +/- 0.25 at day 4. In contrast, in isografts, the effect of ischemia on the histology was not significant. From day 6, the histologic differences between ischemic and nonischemic grafts disappeared. Ischemia led to a more intense expression of P-selectin (day 1), intercellular adhesion molecule-1 (ICAM-1; day 2), and major histocompatibility complex (MHC) class II on endothelium and proximal tubular cells (day 2) in both allografts and isografts. Concurrently with the up-regulated ICAM-1 and MHC expression, significantly more CD4(+) cells and macrophages infiltrated the ischemic allografts at days 2 and 3 and the ischemic isografts at day 4. Importantly, the influx of these cells after ischemia was significantly greater in allografts than in isografts. CONCLUSIONS: Cold ischemia augments allogeneic-mediated cell infiltration in rat kidney allografts. The earlier onset of acute rejection in 24-hour cold preserved allografts may be prevented by better preservation or treatment using tailored immunosuppression.     

Activation of intracellular neutrophil elastase in the transplantation of ischemic liver.

Ischemia-reperfusion injury is an important cause of primary nonfunction of transplanted organs, and neutrophil elastase has been implicated in the pathophysiology of ischemia-reperfusion injury. We assessed the kinetics of intracellular neutrophil elastase (INE) activity in canine liver transplantation. Mongrel dogs underwent orthotopic whole-liver transplantation. The animals in group I (n = 6) received fresh liver grafts, and all of the dogs survived longer than 24 h. The animals in group II (n = 5) received liver grafts injured by 30 min of warm ischemia. Only 1 animal survived longer than 24 h after reperfusion. A significant increase in the serum ALT and LDH levels was observed in group II after reperfusion of the graft. Isolated peripheral neutrophils were homogenized, and the neutrophil elastase activity in the supernatant was determined by using a spectrophotometric assay. The INE activity was expressed as the neutrophil elastase value per 1 x 10(10) peripheral neutrophils. In group I, the INE activity 10 min and 2 h after reperfusion was 7.6 +/- 2.6 and 6.1 +/- 2.4 U, respectively. In group II, this activity was 25.9 +/- 7.4 and 44.3 +/- 23.7 U, respectively. There was a significant correlation between serum LDH levels and INE activity 10 min after reperfusion (gamma = 0.70, p < 0.02). In conclusion, the INE activity increased more sharply after the reperfusion of ischemically injured liver grafts. The INE activity correlates with serum LDH levels immediately after reperfusion, suggesting that the increase in the INE activity depends on the severity of ischemic damage.     

Living unrelated donors in kidney transplants: better long-term results than with non-HLA-identical living related donors?

BACKGROUND: Given the severe organ shortage and the documented superior results obtained with living (vs. cadaver) donor kidney transplants, we have adopted a very aggressive policy for the use of living donors. Currently, we make thorough attempts to locate a living related donor (LRD) or a living unrelated donor (LURD) before proceeding with a cadaver transplant. METHODS: We compared the results of our LURD versus LRD transplants to determine any significant difference in outcome. RESULTS: Between 1/1/84 and 6/30/98, we performed 711 adult kidney transplants with non-HLA-identical living donors. Of these, 595 procedures used LRDs and 116 used LURDs. Immunosuppression for both groups was cyclosporine-based, although LURD recipients received 5-7 days of induction therapy (antilymphocyte globulin or antithymocyte globulin), whereas LRD recipients did not. LURD recipients tended to be older, to have inferior HLA matching, and to have older donors than did the LRD recipients (all factors potentially associated with decreased graft survival). Short-term results, including initial graft function and incidence of acute rejection, were similar in the two groups. LURD recipients had a slightly higher incidence of cytomegalovirus disease (P=NS). We found no difference in patient and graft survival rates. However, the incidence of biopsy-proven chronic rejection was significantly lower among LURD recipients (16.7% for LRD recipients and 10.0% for LURD recipients at 5 years posttransplant; P=0.05). LRD recipients also had a greater incidence of late (>6 months posttransplant) acute rejection episodes than did the LURD recipients (8.6% vs. 2.6%, P=0.04). The exact reason for these findings is unknown. CONCLUSION: Although LURD recipients have poorer HLA matching and older donors, their patient and graft survival rates are equivalent to those of non-HLA-identical LRD recipients. The incidence of biopsy-proven chronic rejection is lower in LURD transplants. Given this finding and the superior results of living donor (vs. cadaver) transplants, a thorough search should be made for a living donor-LRD or LURD-before proceeding with a cadaver transplant.     

Reduction of hepatic ischemia/reperfusion injury by a soluble P-selectin glycoprotein ligand-1.

OBJECTIVE: The authors' goal was to determine the effects of specific binding and blockade of P- and E-selectins by a soluble P-selectin glycoprotein ligand-1 (PSGL-1) in rat models of hepatic in vivo warm ischemia and ex vivo cold ischemia. The authors also sought to determine the effect of selectin blockade on isograft survival in a syngeneic rat orthotopic liver transplant model. SUMMARY BACKGROUND DATA: Ischemia/reperfusion (I/R) injury is a major factor in poor graft function after liver transplantation, which may profoundly influence early graft function and late changes. It is hypothesized that I/R injury leads to the upregulation of P-selectin, which is then rapidly translocated to endothelial cell surfaces within 5 minutes of reperfusion of the liver, initiating steps leading to tethering of polymorphonuclear neutrophil leukocytes to the vascular intima. Local production by leukocytes of interleukin-1, tumor necrosis factor-alpha, or both induces P-selectin expression on the endothelium and continues the cascade of events, which increases cell adherence and infiltration of the organ. METHODS: To examine directly the effects of selectins in a warm hepatic I/R injury model, 100 microg of PSGL-1 or saline was given through the portal vein at the time of total hepatic inflow occlusion. The effects of PSGL-1 in cold ischemia were assessed using an isolated perfused rat liver after 6 hours of 4 degrees C storage in University of Wisconsin (UW) solution, with or without the instillation of PSGL-1 before the storage. To evaluate the effect of selectin blockade on liver transplant survival, syngeneic orthotopic liver transplants were performed between inbred male Sprague-Dawley rats after 24 hours of cold ischemic storage in UW solution. A separate group of animals received two doses of 100 microg of PSGL-1 through the portal vein before storage and before reperfusion of the transplanted liver. Recipient survival was assessed at 7 days, and the Kaplan-Meier product limit estimate method was used for univariate calculations of time-dependent recipient survival events. RESULTS: In an in vivo warm rat liver ischemia model, perfusion with PSGL-1 afforded considerable protection from I/R injury, as demonstrated by decreased transaminase release, reduced histologic hepatocyte damage, and suppressed neutrophil infiltration, versus controls (p < 0.05). When cold stored livers were reperfused, PSGL-1 reduced the degree of hepatocyte transaminase release, reduced neutrophil infiltration, and decreased histologic hepatocyte damage (p < 0.05 vs. UW-only controls). On reperfusion, livers treated with PSGL-1 demonstrated increased portal vein blood flow and bile production (p < 0.05 vs. UW-only controls). In addition, 90% of the rats receiving liver isografts stored in UW solution supplemented with PSGL-1 survived 7 days versus 50% of those whose transplanted syngeneic livers had been stored in UW alone (p < 0.05). CONCLUSIONS: Selectins play an important role in I/R injury of the liver. Early modulation of the interaction between P-selectin and its ligand decreases hepatocyte injury, neutrophil adhesion, and subsequent migration in both warm and cold rat liver ischemia models. In addition, the use of PSGL-1 before ischemic storage and before transplantation prevents hepatic injury, as documented by a significant increase in liver isograft survival. These findings have important clinical ramifications: early inhibition of alloantigen-independent mechanisms during the I/R damage may influence both short- and long-term survival of liver allografts.     

Kidney NGAL is a novel early marker of acute injury following transplantation

Acute kidney injury secondary to ischemia–reperfusion in renal allografts often results in delayed graft function. We tested the hypothesis that expression of neutrophil gelatinase-associated lipocalin (NGAL) is an early marker of acute kidney injury following transplantation. Sections from paraffin-embedded protocol biopsy specimens obtained at approximately one hour of reperfusion after transplantation of 13 cadaveric (CAD) and 12 living-related (LRD) renal allografts were examined by immunohistochemistry for expression of NGAL. The staining intensity was correlated with cold ischemia time, peak post-operative serum creatinine, and dialysis requirement. There were no differences between the LRD and CAD groups in age, gender or preoperative serum creatinine. Using a scoring system of 0 (no staining) to 3 (most intense staining), NGAL expression was significantly increased in CAD specimens (2.3±0.8 versus 0.8±0.7 in LRD, p<0.001). There was a strong correlation between NGAL staining intensity and cold ischemia time (R=0.87, p<0.001). Importantly, NGAL staining in these early CAD biopsies was strongly correlated with peak postoperative serum creatinine, which occurred days later (R=0.86, p<0.001). Four patients developed delayed graft function requiring dialysis during the first week posttransplantation; all of these patients displayed the most intense NGAL staining in their first protocol biopsies. We conclude that NGAL staining intensity in early protocol biopsies represents a novel predictive biomarker of acute kidney injury following transplantation.     

Perflubron emulsion prevents PMN activation and improves myocardial functional recovery after cold ischemia and reperfusion

BACKGROUND: In cardiopulmonary bypass, extracorporeal circulation activates neutrophils, which contribute to ischemia reperfusion injury and postoperative myocardial dysfunction. Perfluorocarbons (PFCs) are compounds that dissolve oxygen and have anti-inflammatory and neutrophil-stabilizing properties. We hypothesized that perflubron emulsion (PFE), a PFC, would attenuate neutrophil activation during simulated extracorporeal circulation (SECC) and would preserve myocardial functional recovery during reperfusion after cold ischemia. METHODS: In a SECC, diluted blood was circulated for 120 min and subsequently used to reperfuse isolated rat hearts after 2 h of cold (12 degrees C) ischemia. Three groups were studied: noncirculated control; SECC/no additive; and SECC/PFE added. In control and SECC/no additive groups, whole blood was diluted 1:1 with plasmalyte. SECC/PFE blood was diluted 1:1 with plasmalyte and PFE (0.075 mL/mL diluted whole blood). Blood counts and neutrophil activation experiments were performed before and after 120 min of SECC. Reperfusion was accomplished using a modified Langendorff preparation. Left ventricular developed pressure, dP/dt, and coronary flow were measured at 10, 15, and 20 min of reperfusion. RESULTS: After 120 min SECC, neutrophil activation was significantly reduced in the SECC/PFE group compared to the SECC/no additive group (38.1 +/- 2.3% versus 51.7 +/- 1.0%, P < 0.05). Compared to cold ischemic hearts reperfused with fresh, non-recirculated blood, left ventricular developed pressure and dP/dt were significantly impaired in the cold ischemic hearts reperfused with SECC/no additive blood (P < 0.05). In contrast, myocardial functional recovery was not impaired in the hearts reperfused with SECC/PFE blood. CONCLUSIONS: SECC-induced neutrophil activation was attenuated with Perflubron treatment. In addition, the progressive impairment in myocardial functional recovery after cold ischemia was significantly improved with treatment. PFE has clinical potential to limit neutrophil-mediated reperfusion injury following cold ischemia.     

Frequency and severity of acute rejection in live- versus cadaveric-donor renal transplants

BACKGROUND: Live donors are an increasingly important source of kidneys for transplantation in Australia. The aim of this study was to compare the rate and severity of rejection between patients receiving kidney transplants from live versus cadaveric donors. METHODS: A retrospective analysis was undertaken of all patients receiving live-donor (n=109) and cadaveric-donor (n=389) renal transplants at our institution between April 1, 1994, and March 31, 2000. Follow-up was completed on all patients until graft loss, death, or May 31, 2001. RESULTS: The baseline characteristics of the live-donor and cadaveric groups were similar, except for recipient age (mean+/-SD, 36.3+/-15.6 vs. 44.5+/-14.4 years, respectively; P<0.001); donor age (46.1+/-11.3 vs. 36.1+/-16.4 years, P<0.001); pretransplant dialysis duration (1.36+/-2.1 vs. 3.4+/-4.4 years, P<0.001); and the proportions of patients receiving first allografts (95% vs. 88%, respectively; P<0.05), antibody induction (8% vs. 20%, P<0.01), and mycophenolate mofetil (MMF) (60% vs. 37%, P<0.001). Acute rejection was observed in 48 (44%) live-donor and 108 (28%) cadaveric transplants (P=0.001). Cadaveric donor type was independently predictive of less acute rejection both on logistic regression (adjusted odds ratio [AOR], 0.47; 95% confidence interval [CI], 0.30-0.73; P=0.001) and multivariate Cox proportional hazards model analysis (hazard ratio, 0.49; 95% CI, 0.34-0.69; P<0.001). Patients receiving cadaveric-donor transplants were also significantly less likely to receive antibody therapy for rejection (univariate, 18% vs. 9%; P=0.006; multivariate AOR, 0.45; 95% CI, -0.25-0.82; P<0.01), independent of recipient age, gender, race, transplant number, human leukocyte antigen mismatch, sensitization, induction therapy, delayed graft function, MMF use, tacrolimus or cyclosporine A use, sirolimus-everolimus use, year of transplant, donor age, or dialysis duration. However, donor type did not independently influence graft survival, immunologic graft survival, or patient survival. CONCLUSIONS: Live-donor kidney transplant recipients had a higher rate and severity of rejection and a shorter rejection-free period than cadaveric renal transplant recipients. Further consideration of the reasons for this difference and the use of alternative immunosuppressive strategies for live-donor transplants are recommended.     

Protective effects of ulinastatin against ischemia-reperfusion injury

We investigated the protective effect of urinary trypsin inhibitor (ulinastatin: UTI) in vitro, in relation to the neutrophil activity in hepatic ischemia/reperfusion (I/R) injury. The rat liver was removed and preserved in cold Ringer's lactate solution for 60 min, followed by 120 min of reperfusion with oxygenated perfusate. The rats were divided into four groups (n = 8 in each group). The livers were perfused with Krebs-Henseleit (K-H) solution containing no additives in group 1, 50,000 U/kg of UTI in group 2, 3.5 x 10(6) of neutrophils in group 3, and both neutrophils and UTI in group 4. In group 3, the AST and ALT levels were always higher than those in other three groups at any point evaluated (P < 0.01) and the LDH levels were observed to be significantly higher than those in other three groups at 0, 5, 10, 60, and 90 min after reperfusion (P < 0. 01). These increase were suppressed by additional pretreatment with UTI in group 4. The bile flow during reperfusion was significantly suppressed in group 3 compared to that of group 4, at both 30 (P < 0. 01) and 60 (P < 0.05) min after reperfusion. The MPO activity after reperfusion in group 3 also significantly increased compared to other three groups (P < 0.01). These data thus suggest that UTI ameliorated the ischemia/reperfusion injury in vitro by inhibiting of neutrophil accumulation in the postischemic liver.     

Renal transplantation after prolonged dwell peritoneal dialysis in children

Thirty-three children received a total of 38 renal transplants (18 living related donor, and 20 cadaveric) after being on CAPD and/or CCPD (PDPD). Ten patients (12 transplants) were converted to hemodialysis pre-transplant in order to be free of the risk of peritonitis and off antibiotics, whereas 23 patients (26 transplants) were on PDPD at the time of transplant. The latter group of patients are described in greater detail. Within this group there was one episode of catheter colonization with Flavobacterium, and only three patients developed ascites post-transplant. Of the 26 transplants, catheters were removed at the time of transplant in the 13 LRD allografts but left in situ for a mean of 3.8 weeks in the 13 cadaveric transplant recipients. Peritoneal dialysis was required post-transplant in seven patients (two LRD recipients requiring a new catheter placement) without complications. Our policy of removing PD catheters at the time of transplant in LRD recipients and prior to hospital discharge in cadaveric transplant recipients has resulted in the avoidance of additional hospitalizations in 19 of the 26 transplants and avoided extra surgery in 11 of the 13 LRD transplants. We conclude that children who have been on PDPD are suitable candidates for renal transplantation and that the early removal of PD catheters, including removal at the time of transplantation in LRD recipients, is associated with a significant reduction in operative procedures for the patients.     

Cold ischemia time: an independent predictor of increased HLA class I antibody production after rejection of a primary cadaveric renal allograft

BACKGROUND: Cadaveric kidneys experiencing longer cold ischemia time (CIT) are associated with higher levels of delayed graft function, acute rejection, and early graft loss. One mechanism to explain these results is that ischemia/reperfusion (I/R) injury makes the allograft more immunogenic by upregulating molecules involved in the immune response (e.g., HLA Class I/II). METHODS: We evaluated the influence of CIT on the production of HLA Class I antibody level, measured by an antihuman globulin panel reactive antibody (AHG PRA) level, in 90 unsensitized recipients of primary cadaveric renal transplants (from a total of 1442 between 1985 and 1997) who rejected their kidneys. RESULTS: By multivariate analysis, a CIT of 15 hr or more (vs. < 15 hr) independently increased the risk of the AHG Class I PRA level being > or = 20% after unsensitized patients rejected their first kidneys (relative risk=3.57; 95% confidence interval=1.26 to 10.14; P=0.01), despite the same degree of Class I/II mismatch between the two CIT groups. The overall mean peak PRA level after primary kidney rejection was significantly lower for the CIT < 15 hr group (25.9%+/-33.9; n=24) compared with the CIT > or = 15 hr group (46.3%+/-36.5; n=66) (P<0.001). CONCLUSION: Longer CIT induces a humorally more immunogenic kidney.     

Chronic rejection of rat renal allografts. II. The impact of prolonged ischemia time on transplant histology.

The effect of prolonged ischemia time on the generation of chronic rejection was investigated in long-surviving rat renal allografts. Three groups of rats were compared: allografts with a 30-min ischemia time, allografts with a 60-min ischemia time, and syngeneic grafts with a 60-min ischemia time. All transplants were initially immunosuppressed with 5 mg/kg/day of cyclosporine i.m. for 3 weeks postoperatively. The CsA trough levels were similar in the three rat groups. All groups demonstrated an initial rise and fall in serum creatinine; a simultaneous biopsy confirmed acute CsA toxicity. Thereafter the serum creatinine level remained on the normal control level both in the 30-min ischemia allografts and in the 60-min ischemia syngeneic grafts. After the initial decline, the serum creatinine level steadily increased in the 60-min ischemia allograft group and was 181 +/- 64 mumol/L at the end of the observation period, 12 weeks postoperatively, compared with 85 +/- 21 mumol/L in the 30-min ischemia allografts and 89 +/- 25 mumol/L in the 60-min ischemia syngeneic grafts. Quantitative histology that was performed upon autopsy demonstrated that the 60-min ischemia allografts showed persistent inflammation, inflammatory cell pyroninophilia, vascular arteriosclerosis and obliteration, and glomerular sclerosis, which were significantly stronger than in similarly immunosuppressed syngeneic transplants. Reduction of the ischemia time from 60 to 30 min significantly ameliorated the vascular and glomerular changes in the allografts but not the inflammatory alterations. This experimental study confirms previous clinical observations and demonstrates that prolonged ischemia contributes to chronic rejection in renal allografts. The results suggest that the effect of prolonged ischemia on chronic rejection is directed primarily to the parenchymal components of the graft, possibly to the graft vascular endothelium. As prolonged ischemia did not significantly affect the inflammation in the transplants, we conclude that the effect is not directed to the intensity of the host antigraft immune response.