Regulatory T Cell Infusion Can Enhance Memory T Cell and Alloantibody Responses in Lymphodepleted Nonhuman Primate Heart Allograft Recipients.

The ability of regulatory T cells (Treg) to prolong allograft survival and promote transplant tolerance in lymphodepleted rodents is well established. Few studies, however, have addressed the therapeutic potential of adoptively transferred, CD4(+) CD25(+) CD127(-) Foxp3(+) (Treg) in clinically relevant large animal models. We infused ex vivo-expanded, functionally stable, nonselected Treg (up to a maximum cumulative dose of 1.87 billion cells) into antithymocyte globulin-lymphodepleted, MHC-mismatched cynomolgus monkey heart graft recipients before homeostatic recovery of effector T cells. The monkeys also received tacrolimus, anti-interleukin-6 receptor monoclonal antibodies and tapered rapamycin maintenance therapy. Treg administration in single or multiple doses during the early postsurgical period (up to 1 month posttransplantation), when host T cells were profoundly depleted, resulted in inferior graft function compared with controls. This was accompanied by increased incidences of effector memory T cells, enhanced interferon-γ production by host CD8(+) T cells, elevated levels of proinflammatory cytokines, and antidonor alloantibodies. The findings caution against infusion of Treg during the early posttransplantation period after lymphodepletion. Despite marked but transient increases in Treg relative to endogenous effector T cells and use of reputed "Treg-friendly" agents, the host environment/immune effector mechanisms instigated under these conditions can perturb rather than favor the potential therapeutic efficacy of adoptively transferred Treg.

Communication gaps associated with donor-derived infections.

The detection and management of potential donor-derived infections is challenging, in part due to the complexity of communications between diverse labs, organ procurement organizations (OPOs), and recipient transplant centers. We sought to determine if communication delays or errors occur in the reporting and management of donor-derived infections and if these are associated with preventable adverse events in recipients. All reported potential donor-derived transmission events reviewed by the Organ Procurement and Transplantation Network Ad Hoc Disease Transmission Advisory Committee from January 2008 to June 2010 were evaluated for communication gaps between the donor center, OPO and transplant centers. The impact on recipient outcomes was then determined. Fifty-six infection events (IEs; involving 168 recipients) were evaluated. Eighteen IEs (48 recipients) were associated with communication gaps, of which 12 resulted in adverse effects in 69% of recipients (20/29), including six deaths. When IEs and test results were reported without delay, appropriate interventions were taken, subsequently minimizing or averting recipient infection (23 IEs, 72 recipients). Communication gaps in reported IEs are frequent, occur at multiple levels in the communication process, and contribute to adverse outcomes among affected transplant recipients. Conversely, effective communication minimized or averted infection in transplant recipients.

Cancers among US organ donors: a comparison of transplant and cancer registry diagnoses.

Transmission of cancer is a life-threatening complication of transplantation. Monitoring transplantation practice requires complete recording of donor cancers. The US Scientific Registry of Transplant Recipients (SRTR) captures cancers in deceased donors (beginning in 1994) and living donors (2004). We linked the SRTR (52,599 donors, 110,762 transplants) with state cancer registries. Cancer registries identified cancers in 519 donors: 373 deceased donors (0.9%) and 146 living donors (1.2%). Among deceased donors, 50.7% of cancers were brain tumors. Among living donors, 54.0% were diagnosed after donation; most were cancers common in the general population (e.g. breast, prostate). There were 1063 deceased donors with cancer diagnosed in the SRTR or cancer registry, and the SRTR lacked a cancer diagnosis for 107 (10.1%) of these. There were 103 living donors with cancer before or at donation, diagnosed in the SRTR or cancer registry, and the SRTR did not have a cancer diagnosis for 43 (41.7%) of these. The SRTR does not record cancers after donation in living donors and so missed 81 cancers documented in cancer registries. In conclusion, donor cancers are uncommon, but lack of documentation of some cases highlights a need for improved ascertainment and reporting by organ procurement organizations and transplant programs.

Re-examination of the lymphocytotoxic crossmatch in liver transplantation: can C4d stains help in monitoring?

C4d-assisted recognition of antibody-mediated rejection (AMR) in formalin-fixed paraffin-embedded tissues (FFPE) from donor-specific antibody-positive (DSA+) renal allograft recipients prompted study of DSA+ liver allograft recipients as measured by lymphocytotoxic crossmatch (XM) and/or Luminex. XM results did not influence patient or allograft survival, or cellular rejection rates, but XM+ recipients received significantly more prophylactic steroids. Endothelial C4d staining strongly correlates with XM+ (<3 weeks posttransplantation) and DSA+ status and cellular rejection, but not with worse Banff grading or treatment response. Diffuse C4d staining, XM+, DSA+ and ABO- incompatibility status, histopathology and clinical-serologic profile helped establish an isolated AMR diagnosis in 5 of 100 (5%) XM+ and one ABO-incompatible, recipients. C4d staining later after transplantation was associated with rejection and nonrejection-related causes of allograft dysfunction in DSA- and DSA+ recipients, some of whom had good outcomes without additional therapy. Liver allograft FFPE C4d staining: (a) can help classify liver allograft dysfunction; (b) substantiates antibody contribution to rejection; (c) probably represents nonalloantibody insults and/or complete absorption in DSA- recipients and (d) alone, is an imperfect AMR marker needing correlation with routine histopathology, clinical and serologic profiles. Further study in late biopsies and other tissue markers of liver AMR with simultaneous DSA measurements are needed.

An update on donor-derived disease transmission in organ transplantation.

Several recent donor-to-recipient disease transmissions have highlighted the importance of this rare complication of solid organ transplantation. The epidemiology of donor-derived disease transmissions in the United States has been described through reports to the Organ Procurement and Transplant Network (OPTN); these reports are reviewed and categorized by the ad hoc Disease Transmission Advisory Committee (DTAC); additional data comes through the published literature. From these reports, it is possible to estimate that donor-derived disease transmission complicates less than 1% of all transplant procedures but when a transmission occurs, significant morbidity and mortality can result. Only through continued presentation of the available data can continuous quality improvements be made. As the epidemiology of donor-derived disease transmission has become better understood, several groups have been working on methods to further mitigate this risk.

Donor-transmitted malignancies in organ transplantation: assessment of clinical risk.

The continuing organ shortage requires evaluation of all potential donors, including those with malignant disease. In the United States, no organized approach to assessment of risk of donor tumor transmission exists, and organs from such donors are often discarded. The ad hoc Disease Transmission Advisory Committee (DTAC) of the Organ Procurement and Transplantation Network/United Network for Organ Sharing (OPTN/UNOS) formed an ad hoc Malignancy Subcommittee to advise on this subject. The Subcommittee reviewed the largely anecdotal literature and held discussions to generate a framework to approach risk evaluation in this circumstance. Six levels of risk developed by consensus. Suggested approach to donor utilization is given for each category, recognizing the primacy of individual clinical judgment and often emergent clinical circumstances. Categories are populated with specific tumors based on available data, including active or historical cancer. Benign tumors are considered in relation to risk of malignant transformation. Specific attention is paid to potential use of kidneys harboring small solitary renal cell carcinomas, and to patients with central nervous system tumors. This resource document is tailored to clinical practice in the United States and should aid clinical decision making in the difficult circumstance of an organ donor with potential or proven neoplasia.

Impact of HIV on liver fibrosis in men with hepatitis C infection and haemophilia.

Hepatitis C virus (HCV) is the major cause of liver disease in haemophilia. Few data exist on the proportion with liver fibrosis in this group after long-term HCV and HIV co-infection. We conducted a cross-sectional multi-centre study to determine the impact of HIV on the prevalence and risk factors for fibrosis in haemophilic men with chronic hepatitis C. Biopsies were independently scored by Ishak, Metavir and Knodell systems. Variables were tested for associations with fibrosis using logistic regression and receiver operating curves (ROC). Of 220 biopsied HCV(+) men, 23.6% had Metavir ≥ F3 fibrosis, with higher mean Metavir fibrosis scores among HIV/HCV co-infected than HCV mono-infected, 1.6 vs. 1.3 (P = 0.044). Variables significantly associated with fibrosis included AST, ALT, APRI score (AST/ULN × 100/platelet × 10(9) /L), alpha-fetoprotein (all P < 0.0001), platelets (P = 0.0003) and ferritin (P = 0.0008). In multiple logistic regression of serum markers, alpha-fetoprotein, APRI and ALT were significantly associated with ≥ F3 fibrosis [AUROC = 0.77 (95% CI 0.69, 0.86)]. Alpha-fetoprotein, APRI and ferritin were significant in HIV(-) [AUROC = 0.82 (95% CI 0.72, 0.92)], and alpha-fetoprotein and platelets in HIV(+) [AUROC = 0.77 (95% CI 0.65, 0.88]. In a multivariable model of demographic and clinical variables, transformed (natural logarithm) of alpha-fetoprotein (P = 0.0003), age (P = 0.006) and HCV treatment (P = 0.027) were significantly associated with fibrosis. Nearly one-fourth of haemophilic men have Metavir ≥ 3 fibrosis. The odds for developing fibrosis are increased in those with elevated alpha-fetoprotein, increasing age and past HCV treatment.

Ex vivo application of carbon monoxide in UW solution prevents transplant-induced renal ischemia/reperfusion injury in pigs.

I/R injury is a major deleterious factor of successful kidney transplantation (KTx). Carbon monoxide (CO) is an endogenous gaseous regulatory molecule, and exogenously delivered CO in low concentrations provides potent cytoprotection. This study evaluated efficacies of CO exposure to excised kidney grafts to inhibit I/R injury in the pig KTx model. Porcine kidneys were stored for 48 h in control UW or UW supplemented with CO (CO-UW) and autotransplanted in a 14-day follow-up study. In the control UW group, animal survival was 80% (4/5) with peak serum creatinine levels of 12.0 +/- 5.1 mg/dL. CO-UW showed potent protection, and peak creatinine levels were reduced to 6.9 +/- 1.4 mg/dL with 100% (5/5) survival without any noticeable adverse event or abnormal COHb value. Control grafts at 14 days showed significant tubular damages, focal fibrotic changes and numerous infiltrates. The CO-UW group showed significantly less severe histopathological changes with less TGF-beta and p-Smad3 expression. Grafts in CO-UW also showed significantly lower early mRNA levels for proinflammatory cytokines and less lipid peroxidation. CO in UW provides significant protection against renal I/R injury in the porcine KTx model. Ex vivo exposure of kidney grafts to CO during cold storage may therefore be a safe strategy to reduce I/R injury.

Highly active antiretroviral therapy improves ESLD-free survival in HIV-HCV co-infection.

The impact of highly active antiretroviral therapy (HAART) on progression to end-stage liver disease (ESLD) in human immunodeficiency virus (HIV)/hepatitis C virus (HCV) co-infection remains controversial. We studied 157 HCV+ haemophilic men (85 HIV+ and 72 HIV-), on whom dates of HIV and HCV seroconversion and clinical outcomes were known. Time to ESLD was determined by Kaplan-Meier product-limit methods and risk factors for ESLD progression were analysed by a Cox proportional hazards model. Among HIV+ men, ESLD was more common, 17 of 85 (20.0%) than in HIV-, eight of 72 (11.1%) and median ESLD-free survival significantly shorter, P = 0.009, hazard ratio 3.00 [95% confidence interval (CI): 1.27-7.08]. HAART treated HIV+ had longer ESLD-free survival than HIV+ untreated, 30.3 vs. 20.0 years, P = 0.043, hazard ratio, 3.14 (95% CI: 1.27-7.08), comparable with survival in HIV- men, P = 0.13, hazard ratio 2.20 (95% CI: 0.76-2.35). Progression was unrelated to HAART toxicity (n = 0) or HCV antiviral therapy (n = 7). HIV+ HAART Rx and HIV- did not differ in HCV duration, age at ESLD, age at death or present, overall or AIDS mortality, all P > 0.05. These data suggest that HAART improves ESLD-free survival, approaching that in HIV- men.

Effect of FK 506 on spontaneous diabetes in BB rats.

From days 30-120 after birth, 59 BB rats were treated with water (n = 20) or FK 506 in intragastric doses of 1 mg.kg-1.day-1 (n = 19) or 2 mg.kg-1.day-1 (n = 20). Diabetes developed in 75, 15, and 0% of the 3 groups, respectively. Animals protected from diabetes by FK 506 had normal intraperitoneal glucose tolerance tests, virtual absence histopathologically of autoimmune insulitis, and normal pancreatic insulin content. Forty-five to 75 days after stopping FK 506, approximately 75% of the rats that were diabetes free at 120 days remained so.

Suppressed T-cell receptor zeta chain expression and cytokine production in pancreatic cancer patients.

Suppression of various functions of T cells derived from cancer patients has been linked previously to changes in the T-cell receptor (TCR)-associated signal transduction molecules, in particular the zeta chain of the TCR complex. In this study, we have examined the TCRzeta chain expression and cytokine production in vivo and in vitro in T cells of patients with metastatic adenocarcinomas of the pancreas that participated in a Phase I clinical trial of the MUC1 peptide plus bacillus Calmette-Guerin cancer vaccine. A majority of the patients had reduced TCRzeta chain expression and interleukin 4 production by T cells, and all of the patients showed decreased production of IFN-gamma of their peripheral T cells when compared with healthy individuals. Peripheral blood T cells were activated with the phorbol ester phorbol myrisate acetate and ionomycin to show that although aberrant TCRzeta chain expression and decreased cytokine production were often correlated, the reduced cytokine production was not simply a consequence of an impaired TCRzeta chain expression. Rather, these are two separate but parallel defects in signal transduction in T cells, which are potentially modulated by the same mechanisms. Half of the patients showed an improvement for TCRzeta chain or IFN-gamma expression after vaccination.

Involvement of the gastrointestinal tract by Epstein-Barr virus--associated posttransplant lymphoproliferative disorders.

Posttransplant lymphoproliferative disorders (PTLD) are abnormal growths of lymphoid cells that occur in immunosuppressed organ transplant recipients. Most cases are of B-lymphocyte origin and are associated with Epstein-Barr virus infection. Twelve of 72 allograft recipients with PTLD in our series have had disease predominantly involving the gastrointestinal tract. The lesions are most often multiple and preferentially involve the distal small bowel. The appearance of the lymphoid cells ranges from nonuniform (polymorphic) to uniform (monomorphic). Most tumors contain a clonal component of B-lymphocytes, with or without a nonclonal background. Appropriate primary treatment includes surgical resection and reduction of immunosuppression. Occasional PTLD of the gastrointestinal tract do not respond to this regimen, these represent a more advanced state of tumor progression. Currently, 11 of 12 patients are alive 10-13 months after diagnosis. The surgical pathologist must be aware of the appropriate setting in which to consider a diagnosis of PTLD and be able to distinguish this condition from other lymphoproliferative disorders of the gastrointestinal tract.

Herpesvirus 6 variant A infection after heart transplantation with giant cell transformation in bile ductular and gastroduodenal epithelium.

Herpesvirus 6 (HHV-6) is a ubiquitous virus known to cause febrile syndromes and exanthema subitum in children. Less commonly, and particularly in organ transplant recipients, it may result in hepatitis, bone marrow suppression, interstitial pneunonitis, and meningoencephalitis. This report expands the spectrum of clinical disease associated with HHV-6 by documenting viral infection in a 44-year-old heart transplant recipient presenting with gastroduodenitis, pancreatitis, and hepatitis. On histopathologic examination, the gastric, duodenal, and bile ductular epithelium showed a multinucleate giant cell transformation similar to the cytopathic effect caused by the virus in human T-lymphocytes infected in vitro. Electron microscopy showed herpes particles with a thick tegument layer in the duodenum. Polymerase chain reaction amplified HHV-6 variant A sequences from multiple sites. Serology confirmed the presence of an acute HHV-6 infection. Thus, HHV-6 variant A can cause gastroduodenitis and pancreatitis in immunosuppressed individuals. Multinucleate giant cells and enveloped virions with a prominent tegument can be used as morphologic criteria to raise the possibility of HHV-6 infection in human biopsy tissue.

Epstein-Barr virus-negative post-transplant lymphoproliferative disorders: a distinct entity?

Post-transplant lymphoproliferative disorders (PTLDs) are usually but not invariably associated with Epstein-Barr virus (EBV). The reported incidence, however, of EBV-negative PTLDs varies widely, and it is uncertain whether they should be considered analogous to EBV-positive PTLDs and whether they have any distinctive features. Therefore, the EBV status of 133 PTLDs from 80 patients was determined using EBV-encoded small ribonucleic acid (EBER) in situ hybridization stains with or without Southern blot EBV terminal repeat analysis. The morphologic, immunophenotypic, genotypic, and clinical features of the EBV-negative PTLDs were reviewed, and selected features were compared with EBV-positive cases. Twenty-one percent of patients had at least one EBV-negative PTLD (14% of biopsies). The initial EBV-negative PTLDs occurred a median of 50 months post-transplantation compared with 10 months for EBV-positive cases. Although only 2% of PTLDs from before 1991 were EBV negative, 23% of subsequent PTLDs were EBV negative (p <0.001). Of the EBV-negative PTLDs, 67% were of monomorphic type (M-PTLD) compared with 42% of EBV-positive cases (p <0.05). The other EBV-negative PTLDs were of infectious mononucleosis-like, plasma cell-rich (n = 2), small B-cell lymphoid neoplasm, large granular lymphocyte disorder (n = 4) and polymorphic (P) types. B-cell clonality was established in 14 specimens and T-cell clonality was established in three (two patients). None of the remaining specimens were studied with Southern blot analysis and some had no ancillary studies. Rearrangement of c-MYC was identified in two M-PTLDs with small noncleaved-like features, and rearrangement of BCL-2 was found in one large noncleaved-like M-PTLD. Ten patients were alive at 3 to 63 months (only three patients received chemotherapy). Seven patients, all with M-PTLDs, are dead at 0.3 to 6 months. Therefore, EBV-negative PTLDs have distinct features, but some do respond to decreased immunosuppression, similar to EBV-positive cases, suggesting that EBV positivity should not be an absolute criterion for the diagnosis of a PTLD.

Recurrent hepatitis C in liver allografts: prospective assessment of diagnostic accuracy, identification of pitfalls, and observations about pathogenesis.

RATIONALE AND DESIGN: The accuracy of a prospective histopathologic diagnosis of rejection and recurrent hepatitis C (HCV) was determined in 48 HCV RNA-positive liver allograft recipients enrolled in an "immunosuppression minimization protocol" between July 29, 2001 and January 24, 2003. Prospective entry of all pertinent treatment, laboratory, and histopathology results into an electronic database enabled a retrospective analysis of the accuracy of histopathologic diagnoses and the pathophysiologic relationship between recurrent HCV and rejection. RESULTS: Time to first onset of acute rejection (AR) (mean, 107 days; median, 83 days; range, 7-329 days) overlapped with the time to first onset of recurrent HCV (mean, 115 days; median, 123 days; range, 22-315 days), making distinction between the two difficult. AR and chronic rejection (CR) with and without co-existent HCV showed overlapping but significantly different liver injury test profiles. One major and two minor errors occurred (positive predictive values for AR = 91%; recurrent HCV = 100%); all involved an overdiagnosis of AR in the context of recurrent HCV. Retrospective analysis of the mistakes showed that major errors can be avoided altogether and the impact of unavoidable minor errors can be minimized by strict adherence to specific histopathologic criteria, close clinicopathologic correlation including examination of HCV RNA levels, and a conservative approach to the use of additional immunosuppression. In addition, histopathologic diagnoses of moderate and severe AR and CR were associated with relatively low HCV RNA levels, whereas relatively high HCV RNA levels were associated with a histopathologic diagnosis of hepatitis alone, particularly the cholestatic variant of HCV. CONCLUSIONS: Liver allograft biopsy interpretation can rapidly and accurately distinguish between recurrent HCV and AR/CR. In addition, the histopathologic observations suggest that the immune mechanism responsible for HCV clearance overlap with those leading to significant rejection.

Renal cortical neoplasms in long term survivors of solid organ transplantation.

BACKGROUND: Renal cortical neoplasms have been reported after organ transplantation, but the level of risk as well as the histological features are poorly defined. METHODS: A retrospective autopsy-based study was performed to evaluate renal neoplasms occurring in patients who underwent solid organ transplantation, died, and received an autopsy from 1981 to 1997 (383 liver, 125 heart, 52 lung, 39 heart/lung, 98 kidney, 4 bowel). Patients were divided into those with short (less than 101 days), medium (101 days to 5 years), and long-term survival (more than 5 years). The control group consisted of hospital autopsies on nontransplanted patients from the odd-numbered years, 1983 through 1997. RESULTS: Renal cortical neoplasms were identified in 32/1325 of nontransplanted patients and 15/701 transplanted patients. In transplanted patients, neoplasms were identified in 14 native and 1 allograft kidney: 2/391 in short-term survivors, 3/234 in medium, and 10/76 in long term survivors. While transplant patients with short and medium length survival had no increased risk for neoplasms, patients with long-term survival showed a 9-fold increase in cortical neoplasms. Transplant patients with neoplasms averaged 47 years of age at death, significantly younger than the average age of 70 for nontransplanted control patients with renal neoplasms. The neoplasms in transplanted patients were all tubulopapillary, except for one clear cell neoplasm and ranged in size from 0.1 to 2 cm. CONCLUSIONS: Long-term survivors of solid organ transplants have an 9-fold increased risk of developing tubulopapillary renal cortical neoplasms.

Sequential analyses of the development of lymphoproliferative disorders in rats receiving cyclosporine.

Cyclosporine (Cys)-associated, atypical lymphoproliferation in gut-associated lymphoid tissue (GALT) was reproducibly observed in male Sprague Dawley rats by feeding a diet containing 0.015% Cys. Sequential morphological changes occurring in the lymphoid organs of the rats on a Cys diet for 8-10 weeks were examined, and these changes were correlated with those observed in electrophoretic patterns of serum proteins. Proliferative lymphoid alterations were detected in the GALT after 2 weeks of feeding, and these progressed through 8 weeks. Atypical lymphoid proliferation was also seen in lymph nodes after 8 weeks of feeding; thymuses showed marked medullary atrophy, and spleens had a diminution of their periarterial lymphoid sheaths. Elevated levels of gamma-globulin were detected after 4 weeks of Cys administration, and became more pronounced by 8 weeks. The lymphoid lesions induced in rats bear striking similarities to the post transplant lymphoproliferative lesions in patients receiving Cys and in patients with the acquired immune deficiency syndrome.

Occult nonhematopoietic malignancies present at autopsy in solid organ transplant patients who died within 100 days.

BACKGROUND: Patients are at an increased risk for developing malignancies after transplantation. Lymphomas, skin malignancies, Kaposi's sarcomas, and cervical/vulvar neoplasms are the most common, but visceral malignancies are also well documented, with a reported frequency ranging from 1% to 6%. These visceral tumors represent a mix of neoplasms that were clinically occult at the time of transplantation and those that arise de novo after transplantation. Little information, however, is available on the frequency of clinically occult malignancies at the time of transplantation and their contribution to the number of posttransplant malignancies. METHODS: A retrospective study was performed of all patients who received an organ transplant from January 1981 to June 1997 and died within 100 days, a time interval in which epithelial malignancies found at autopsy were presumed to have been present, but clinically occult, at the time of transplantation. RESULTS: A total of 375 patients were studied who received the following organ transplants: 231 liver, 52 heart, 26 heart and lung, 32 lung, and 34 kidney. Eleven malignancies were identified for an overall frequency of 2.9% and included three thyroid carcinomas, three carcinoids of the small bowel, two lung carcinomas, one laryngeal carcinoma, one renal cell carcinoma, and one seminoma. CONCLUSION: The 2.9% frequency of malignancies seen in this study suggests that a small, but significant, number of patients have occult malignancies at the time of transplantation and that these occult tumors contribute substantially to the number of malignancies that present clinically after transplantation.

Oxygen free radical content and neutrophil infiltration are important determinants in mucosal injury after rat small bowel transplantation.

Mucosal injury is an immediate event following revascularization of small intestinal grafts in the context of transplantation (SBTx). The generation of oxygen free radicals (OFR) and tissue infiltration by activated neutrophils are consequences of ischemia and reperfusion and known causative factors of tissue injury; to delineate their role in the reperfusion injury occurring after cold preservation of the intestine and subsequent transplantation was the aim of this study. Prior to orthotopic SBTx in Sprague-Dawley rats, grafts were stored in cold (4 degrees C) Ringer's lactate solution for 1 (n=6), 2 (n=7), and 4 hr (n=7). Small bowel biopsy specimens were obtained before harvesting, at the end of the (cold) ischemic period and immediately before unclamping (i.e., before revascularization) and 30, 60, 120 min, and 24 hr after transplantation to evaluate tissue injury by histology, OFR production, (measured by luminol-enhanced chemiluminescence [LCL]), and the degree of neutrophil infiltration by myeloperoxidase staining. Reperfusion of the graft significantly worsened the histologically graded mucosal injury compared with that seen before unclamping. However, 24 hr after engraftment, mucosal morphology was restored almost completely. OFR production increased significantly during the early phases of reperfusion (30, 60, and 120 min) and returned to control values after 24 hr. Reperfusion of the graft was associated with a marked increase in the number of mucosal neutrophils. The present study indicates that OFR production and neutrophilic infiltration commence and progressively increase with graft reperfusion. These changes parallel the mucosal injury. Ischemic intervals of 4 hr were not associated with a statistically significant greater ischemic-injury patterns compared with 1- and 2-hr intervals. The profound changes associated with reperfusion probably overshadow the minor, yet likely, progressive injury patterns associated with longer ischemia times.

Cross-species graft-versus-host-disease is accompanied by a donor-derived cellular immune response.

Acute graft-versus-host-disease is classically described as reactivity of donor lymphocytes to recipient alloantigens. To date there is debate as to whether GVHD can be induced across a species barrier. We recently reported the induction of stable xenogeneic chimerism (rat-->mouse) and donor-specific transplantation tolerance using the transplantation of untreated rat bone marrow cells into B10 mouse recipients. Survival of chimeras was excellent, and there was no evidence of GVHD. We now describe the induction of xenogeneic GVHD by transplanting large numbers of donor rat spleen cells with the bone marrow inoculum. All chimeras that received bone marrow and untreated spleen cells developed an external appearance compatible with GVHD and had a median survival time of 14 days. Mice that received equivalent numbers of untreated rat bone marrow alone appeared healthy, had no evidence for GVHD, and survived > 90 days. The usual epithelial target tissues for allogeneic GVHD in those mice that received xenogeneic bone marrow and spleen cells showed the presence of tissue injury and histologic features compatible with GVHD. Donor rat MHC class I and class II positive cells were prominent cell types present in the tongues of mice that developed features of GVHD, and this was accompanied by a significant inflammatory tissue response with loss of the dermal-epidermal architecture. In contrast, fully xenogeneic chimeras without GVHD had no evidence for tissue injury or pathologic cellular infiltrates when examined by immunohistochemical analysis. These data suggest that although fully xenogeneic chimeras resist GVHD, GVHD can be induced across a species barrier if sufficient numbers of donor rat spleen cells are added to the bone marrow inoculum. Further comparisons of these models may provide an approach to study the mechanisms responsible for xenoreactivity in vivo.