Intestinal Allograft Rejection Research Articles

Calprotectin

To identify a noninvasive screening test for intestinal allograft monitoring. Intestinal allograft rejection is difficult to distinguish from other causes of diarrhea and can rapidly lead to severe exfoliation or death. Protocol biopsies are standard for allograft monitoring but may cause serious complications. No noninvasive test has shown clinical utility for monitoring of the intestinal allograft. Calprotectin levels (n = 68) were measured in this pilot study from ileostomy effluent in patients with histologic evidence of acute rejection (n = 12), viral enteritis (n = 5), and nonspecific inflammation (n = 16) and compared with those with normal allograft histology (n = 35). Median stool calprotectin levels from patients with rejection were significantly higher than those from patients with viral enteritis or normal biopsies [198 mg/kg compared with 7 and 19 mg/kg, respectively (P = 0.0002)]. Receiver operator characteristics suggest the optimal cut-off level to distinguish rejection from other diagnoses is 92 mg/kg with specificity of 77% and sensitivity of 83%. Although false-positive results occurred in 26% of patients with normal biopsies and 30% with nonspecific changes, no treated episode of acute rejection was below the cutoff. In addition, in 2 patients with serial levels, elevations in the calprotectin levels preceded histologic changes by 6 to 18 days. Low stool calprotectin levels correlate well with a low risk for intestinal allograft rejection. If confirmed, biopsies may be reserved in the future for confirmation of rejection, eliminating protocol biopsies, and immunosuppressive changes could potentially be made before allograft injury.

Innovative Approaches to Improving Organ Availability for Small Bowel Transplant Candidates

Small bowel transplant candidates have unique features that create special challenges for providing them with organs in a timely manner. More than half of intestinal transplant candidates also need livers. Because there is a greater overall demand for livers than for small bowels, organ allocation in this candidate subset is dictated by the liver allocation policies. However, the extremely high mortality rate associated with patients on the waiting list for combined liver-intestinal transplantation (LIT) attests to the inadequacies of the liver allocation system in recognizing the unique risk factors in patients with intestinal failure who develop end-stage liver disease (ESLD). This report addresses some of the organ allocation issues underlying the discrepancies in waiting list outcomes seen in LIT candidates and discusses possible contributory factors and potential solutions to this problem. The use of living donor and segmental grafts from deceased donors are discussed specifically as potential surgical solutions to the problem, and the potential utility, limitations, and ethical concerns associated with these innovative strategies are discussed.

Utilization of Dried Blood Spot Citrulline Level as a Noninvasive Method for Monitoring Graft Function Following Intestinal Transplantation

Citrulline concentrations have been proposed as a marker for intestinal allograft rejection. We instituted dried blood spot (DBS) specimen monitoring of citrulline to simplify sample collection posttransplant. This study demonstrates the correlation between plasma and dried blood spot specimen citrulline concentrations after intestinal transplantation. Plasma and DBS samples were analyzed by hydrophilic interaction chromatography tandem mass spectrometry. Comparison of the strength of linear correlation was made according to the type of surgery, sonication time, DBS citrulline levels, and the time interval between the blood sample collection and the assay date. A very strong linear correlation exists between the plasma and DBS citrulline concentrations (r=0.87; P<0.001). The correlation between plasma and DBS citrulline concentrations was maintained when evaluating only the intestinal transplant recipients. There was no significant difference in the strength of linear correlation according to sonication time, cirtrulline concentrations, or length of time to assay date. DBS citrulline monitoring will ease sample collection following intestinal transplantation and improve the ability to detect intestinal dysfunction and rejection by a noninvasive means.

Protection of Mouse Small Bowel Allografts by FTY720 and Costimulation Blockade

The clinical application of small bowel transplantation (SBTx) is hampered by its pronounced immunogenicity. We aimed to test the hypothesis that prolonged sequestration of lymphocytes in secondary lymphoid organs may enhance the alloprotective effect of costimulation blockade. For this purpose, recipients of intestinal allografts were treated with MR1, FTY720, combined FTY720 plus MR1, or were left untreated. Grafts were examined 6 and 14 days after transplantation by applying a histologic rejection score, multiparameter-immunofluorescent staining, and flow cytometry. FTY720 or MR1 monotherapy did not prevent the rejection of mouse intestinal allografts, whereas combined therapy with FTY720 plus MR1 profoundly inhibited rejection at day 6 and day 14 after transplantation. In FTY720-treated mice infiltration of host lymphocytes in graft mesenteric lymph nodes, Peyer's patches, intraepithelial lymphocytes, and lamina propria lymphocytes (LPLs) was reduced on day 6. Anti-CD40L antibody improved the rejection score at day 14 but had no effect at day 6. Importantly, host CD8 T-cell infiltration in graft LPLs was significantly reduced compared with all other groups. FTY720 plus MR1 effectively inhibited intestinal allograft rejection in mice, possibly by enhancing the alloprotective effects of costimulation blockade by prolonged sequestration of lymphocytes in secondary lymphoid organs.

Carcinoma of donor origin after liver‐intestine transplantation in a child

Tumor-related complications after intestinal transplantation in children have been principally EBV driven post-transplant disorders. We describe the clinical course of a child, with a diagnosis of microvillus inclusion disease who received a liver and intestine allograft at the age of 9 months. His postoperative course was significant for multiple episodes of acute intestinal allograft rejection and eventually the development of post-transplant lymphoproliferative disorder (PTLD), which resolved. At 8 yr post-transplant he presented with masses in the intestine allograft mesentery and in the right lobe of the allograft liver, biopsy of which revealed a relatively undifferentiated tumor, suggestive of a carcinoma. In situ hybridization for X and Y chromosomes, revealed his tumor to be of donor origin. Treatment included debulking of the mesenteric mass with segmental enterectomy of the intestinal allograft, and stopping his immunosuppression for a period of 4 months; this resulted in complete resolution of his malignancy. Immunosuppression with tacrolimus and steroids was restarted because of intestinal allograft rejection; he died suddenly of unknown causes at 17 months post-diagnosis of carcinoma. The severely immunosuppressed state produced in this patient allowed for the development of an unusual donor derived carcinoma, which resolved spontaneously with withdrawal of immunosuppression. The mechanism of such regression of tumor may be related to restitution of immunologic competence, but is yet to be determined.

Control of intestinal allograft rejection by FTY720 and costimulation blockade

Introduction The clinical application of small bowel transplantation (SBTx) is hampered byits pronounced immunogenicity. In this study we examined the effects of the novelimmunosuppressant FTY720 and costimulation blockade by an anti-CD40L mAb (MR-1) ina stringent mouse model of SBTx. Methods SBTx was performed in mice with a full MHC mismatch (donors: C3H=H-2 k;recipients: C57BL/6=H-2 b). Recipients were divided into four groups: 1, untreated group; 2,MR1 monotherapy (500 μg IV on days 0, 2, 4, and 7); 3, FTY720 monotherapy (1 mg/kg bodyweight PO for 14 consecutive days after transplantation); 4, FTY720 plus MR1–treated group.Graft rejection grades were assessed by H&E staining. Graft mesenteric lymph nodes (MLNs),Peyer's patches (PPs), as well as intraepithelial lymphocytes (IELs) and lamina proprialymphocytes (LPLs) were analyzed by flow cytometry and three-color immunofluorescencestaining. Results Neither FTY720 nor MR1 monotherapy was efficient in preventing the rejection ofmouse intestinal allografts, whereas FTY720 plus MR1 profoundly inhibited the rejectionresponse at the 14th posttransplant day. The infiltration of host lymphocytes was reduced ingraft MLNs, PPs, IELs, and LPLs by FTY720 therapy. FTY720 plus MR1 inhibited hostCD8 + T-cell infiltration in graft LPLs when compared with grafts treated with FTY720 only.Additionally, two subpopulations, CD11b +high Gr1 − and CD11b +intermediate Gr1 + cells, weredecreased in FTY720-treated grafts. Conclusions FTY720 plus MR1 efficiently delayed intestinal allograft rejection in a mousemodel by preventing the infiltration of host lymphocytes, particularly of CD8 + cells.

Evolutionary experience with immunosuppression in pediatric intestinal transplantation

Background/Purpose Intestinal transplantation has developed to become the standard of care for patients with irreversible intestinal failure who are not responding to total parenteral nutrition. Once considered experimental, it has taken time and much effort for the procedure to become a clinical reality, with final acceptance primarily because of the vastly improved outcomes. Advances and novel modifications in immunosuppression have been at the forefront of these improvements. The authors review their evolutionary experience with intestinal transplantation, particularly relating changes in immunosuppression protocols to improved outcomes. Methods From July 1990 to December 2003, 122 children received 129 intestinal containing allografts (70 liver/intestine, 42 isolated intestine, 17 multivisceral). Mean age was 5.3 ± 5.2 years, and 55% were boys. Indications for transplantation were mostly short gut syndrome. The allografts were cadaveric, ABO identical (except one), with no immunomodulation. Bone marrow augmentation was used in 29% of the recipients since 1995. T-cell lymphoctytotoxic crossmatch was positive in 24% cases. Immunosuppression protocols can be divided into 3 categories: (i) maintenance tacrolimus and steroids (n = 52, 1990-1995, 1997-1998); (ii) addition of induction therapy with cyclophosphamide (n = 16, 1995-1997) then daclizumab (n = 24, 1998-2001). A third immunosuppressive agent was added in either group where increased immunosuppression was indicated; (iii) pretreatment/induction with antilymphocyte conditioning and steroid-free posttransplantation tacrolimus monotherapy (n = 37, 2002-2003). In this later group, if clinically stable at 60 to 90 days posttransplantation, and no recent rejection, the tacrolimus was weaned by decreasing frequency of dosing. Results The overall Kaplan-Meier patient/graft survival was 81%/76% at 1 year, 62%/60% at 3 years, and 61%/51% at 5 years. Survival continues to improve, with 1-year patient/graft survival being 71%/62%, 77%/75%, and 100%/100% for groups (i), (ii), and (iii), respectively. Acute intestinal allograft rejection has decreased markedly in group (iii). The rate of infectious diseases, such as cytomegalovirus and Epstein-Barr virus, is lowest in group (iii). Graft-versus-host disease has not significantly increased with the latest protocol. Most importantly, the overall level of immunosuppression requirements has decreased markedly, with most patients in group (iii) being on monotherapy. Of these, most had their monotherapy weaned down to spaced doses, something never systematically attempted or achieved in pediatric intestinal transplantation. Conclusions Intestinal transplantation has progressed markedly over the last 13 years. Although there have been modifications in all aspects of the procedure, the story of intestinal transplantation has been the evolution of successful immunosuppression regimens. Our latest pretreatment/induction conditioning and posttransplantation monotherapy strategy improves graft acceptance and lowers subsequent immunosuppression dosing requirements. It is expected this will overcome many of the complications related to the previously high immunosuppression requirements. Minimization of immunosuppression with avoidance of steroid therapy offers profound long-term benefits, especially in the pediatric population. The patients still remain challenging and complex in every aspect; however, these advances offer significant hope to both patients and caregivers alike.

CONTROL OF INTESTINAL ALLOGRAFT REJECTION BY FTY720 AND COSTIMULATION BLOCKADE

P1038 Aims: The clinical application of small bowel transplantation (SBTx) is hampered by its pronounced immunogenicity. To date no immunosuppressive protocol has proven to prevent intestinal allograft rejection in mouse models. Therefore, in this study we examined the effects of the novel immunosuppressant FTY-720 and costimulation blockade by an anti-CD40L mAb (MR-1) in a heterotopic model of small bowel transplantation. Methods: SBTx was performed in mice with a full MHC haplotype mismatch (donors: C3H=H-2k; recipients: C57BL/6=H-2b). Recipients were divided into 4 groups: 1, non-treated group; 2, MR1 monotherapy (500 μg iv on day 0, and 500 μg i.p. on days 2, 4 and 7); 3, FTY720 monotherapy (1 mg/kg body weight po for 14 consecutive days after transplantation); 4, FTY720 plus MR1 treated group. Each group contained 6 mice for histological assessment at day 14, and another 6 mice for FACS analysis and histology at day 6 after transplantation. Analyses were performed on donor and recipient mesenteric lymph nodes, Peyer’s Patches, as well as Intraepithelial Lymphocytes (IEL) and Lamina Propria Lymphocytes (LPL) that were purified by differential collagenase digestion. Tissue sections were analyzed by three colour immunofluorescence staining on an Axiovert M200 microscope. Lymphocytes were stained with monoclonal antibodies, and were studied by three-colour flow cytometry on a FACSCalibur (Becton Dickinson). Results: Both FTY720 and MR1 monotherapy were not efficient in alleviating the rejection of mouse intestinal allografts, whereas FTY720 plus MR1 profoundly improved the rejection response both at early (day 6) and later (day 14) time points. In the latter, the infiltration of host lymphocytes was profoundly reduced in graft mesenteric lymph nodes (MLNs), Peyer’s patches (PPs), intraepithelial lymphocytes (IELs), and lamina propria lymphocytes (LPLs) by FTY720 therapy. FTY720 plus MR1 inhibited host CD8+ T cell infiltration in graft LPLs when compared with grafts treated with FTY720 only. Infiltration by CD44+ T cells was inhibited in most of the FTY720 plus MR1 treated grafts (4/6). Additionally, two sub-populations, CD11b+high Gr1+ and CD11b+high Gr1+ cells were decreased in FTY720 treated grafts. Conclusions: FTY720 plus MR1 efficiently delayed intestinal allograft rejection in a mouse model by preventing the infiltration of host lymphocytes, particularly of CD8+ cells.

Richard Wood and intestinal transplantation

Richard Wood (1943–2003) developed an interest in small bowel transplantation on his appointment to the Chair of Surgery at the Medical College of St Bartholomew's Hospital, London, UK, in 1984 and he organized the first international symposium on the subject in October 1989. His research which initially focused on intestinal function evolved to assess the role of migration of leukocytes between graft and host tissues in the development of rejection and graft-versus-host disease (GvHD). His group explored avenues to prevent rejection and GvHD, but none of these initiatives were totally effective. They did show that ischemia, rejection, and GvHD in heteropic small bowel grafts were all associated with shifts in intestinal microflora toward potentially pathogenic organisms. When the research program relocated to the University of Sheffield in 1994, studies were undertaken to assess adhesion molecule and heat shock protein expression during intestinal allograft rejection. An interest in ischemia–reperfusion (I/R) injury also developed and in vivo microscopy was used to directly visualize the intestinal villus microcirculation after intestinal I/R injury. Hypothermia and the nitric oxide donor FK 409 were shown to prevent mortality, and to attenuate localized and remote tissue damage and leukocyte adherence in the villus microcirculation. In contrast, tacrolimus had no such effect. These studies were ongoing when ill health forced his retirement in 2002. He maintained a keen interest in, and enthusiasm for, the experimental program despite his progressive illness. This support continued to inspire all investigators around him.

Histamine-degrading enzymes as cellular markers of acute small bowel allograft rejection

Abstract Intestinal histaminedegrading enzymes diamine oxidase (DAO) and histamine N-methyltransferase (HNMT) activities are relatively constant per individual and bowel segment, and they reflect the functional integrity of the intestinal mucosa. It was, therefore, hypothesised that a decrease in these enzymes could be indicative of acute rejection of an intestinal allograft. Enzymatic activities of DAO and HNMT were determined in mucosal biopsies of isogeneic (Lewis-to-Lewis, n=48) and allogeneic (Brown Norway-to-Lewis, n=48) heterotopic small bowel transplants in a rat model at various time periods. Allograft recipients were not given any immunosuppression. While no changes in enzyme activities were observed in isografts up to day 8 following transplantation, significantly reduced activities of both enzymes were found in all allografts 6–8 days after transplantation. Activities of both DAO and HNMT exhibited a strong negative correlation with the histological rejection score (P<0.01). We can conclude that DAO and HNMT activities in gut mucosa are reliable quantitative markers of acute intestinal allograft rejection in the rat that support histopathological analysis.

Impaired rejection and mucosal injury of small intestinal allografts lacking the interferon-gamma receptor.

Small intestinal mucosal T cell activation results in villous atrophy and crypt hyperplasia. There is conflicting evidence as to whether a Th1 IFN-gamma response may be involved. Using a murine intestinal transplant model of T cell-mediated enteropathy we aimed to study the role of IFN-gamma on the development of villous atrophy and crypt hyperplasia. Isografts or allografts of foetal small intestine from 129SV-/- IFN-gamma receptor knockout mice or wild type mice were implanted under the kidney capsule of Balb/c recipient mice. Grafts were examined histologically at intervals from 2 to 9 days post implantation for signs of rejection. Quantitative rtPCR for IFN-gamma, TNFalpha and IL-4 was conducted on grafts at 5 and 9 days post implantation. In allografts, rejection accompanied by the development of villous atrophy and crypt hyperplasia, occurred in a time-dependent manner. However this process was markedly slower in the IFN-gamma receptor knockout grafts compared to the wild type grafts at 5 days (chi2 = 10.08, P = 0.007) and 9 days post implantation (chi2 = 13.25, P = 0.004). There were also significantly fewer TNFalpha transcripts in allografts of IFN-gamma-/- intestine than in wild type allografts (P = 0.02). IFN-gamma has a partial, but not obligatory, role in the development of villous atrophy and crypt hyperplasia during T cell mediated rejection of intestinal allografts.

Role of 4-1BB in Allograft Rejection Mediated by CD8+ T Cells

Blockade of traditional costimulatory molecules fails to inhibit rejection in many models where CD8+ T cells are sufficient to mediate rejection. This observation demonstrates that in many settings CD8+ T cells are not dependent upon CD28 or CD154 signals to mediate rejection. 4-1BB (CD137) has been shown to be an important regulatory molecule for CD8+ T cells in a variety of nontransplant models. Here we show that blocking the 4-1BB pathway significantly inhibited rejection of intestinal allografts by CD8+ but not CD4+ T cells. This effect was associated with significantly decreased expression of the genes encoding TNFalpha and secondary lymphoid chemokine (SLC) within the spleens of recipient mice. Disruption of the 4-1BB pathway also impaired the priming of alloantigen-specific CD8+ T cells and the accumulation of recipient dendritic cells within the spleen. These data directly demonstrate an important role for 4-1BB in allograft rejection; particularly rejection mediated by CD8+ T cells. Our data suggest that in addition to providing a direct costimulatory signal to T cells, the 4-1BB pathway may regulate other important steps in the immune response such as the migration of T cells and dendritic cells.

Histamine-degrading enzymes as cellular markers of acute small bowel allograft rejection.

Intestinal histamine-degrading enzymes diamine oxidase (DAO) and histamine N-methyltransferase (HNMT) activities are relatively constant per individual and bowel segment, and they reflect the functional integrity of the intestinal mucosa. It was, therefore, hypothesised that a decrease in these enzymes could be indicative of acute rejection of an intestinal allograft. Enzymatic activities of DAO and HNMT were determined in mucosal biopsies of isogeneic (Lewis-to-Lewis, n=48) and allogeneic (Brown Norway-to-Lewis, n=48) heterotopic small bowel transplants in a rat model at various time periods. Allograft recipients were not given any immunosuppression. While no changes in enzyme activities were observed in isografts up to day 8 following transplantation, significantly reduced activities of both enzymes were found in all allografts 6-8 days after transplantation. Activities of both DAO and HNMT exhibited a strong negative correlation with the histological rejection score ( P<0.01). We can conclude that DAO and HNMT activities in gut mucosa are reliable quantitative markers of acute intestinal allograft rejection in the rat that support histopathological analysis.

Monoclonal antibody against beta7 integrins, but not beta7 deficiency, attenuates intestinal allograft rejection in mice.

beta7 integrins mediate homing and retention of lymphocytes to the normal and inflamed small bowel in a tissue-selective fashion. In the present study, we investigated the expression of beta7 integrins after small bowel transplantation (SBT) and tested the effects of blocking beta7-mediated pathways by using monoclonal antibody (mAb) or knockout mice. Heterotopic SBT from BALB/c to C57BL/6 (B6) was used as a surgical model. Expression of beta7 integrins was measured on recipient lymphocytes (CD4 and CD8 ) in spleen, blood, and mesenteric lymph nodes (MLN) using flow cytometry. To analyze the effects of blocking beta7 integrins on graft survival, either beta7-deficient B6 or wild-type B6 mice that were treated with mAb against beta7 were studied. After allogeneic SBT, there were markedly increased levels of alpha4beta7 recipient CD8 lymphocytes in MLN, blood, and spleen as early as 3 days postoperatively. In contrast, alpha4beta7 integrin levels in isograft recipients were similar to those of normal mice. Mean survival time of intestinal allografts was not affected by beta7 deficiency (7.3+/-1 days) compared with wild-type mice (7.5+/-0.8 days). However, mAb against beta7 integrins significantly prolonged graft survival (12.8+/-1 days) compared with treatment with control mAb (7.3+/-1 days, <0.001). Histologic changes of SBT rejection were significantly attenuated when mice were given mAb against beta7. As indicated by the increased levels of alpha4beta7 CD8 lymphocytes, activation of this integrin contributes to the immune response in SBT rejection. Furthermore, blocking beta7 integrins with mAb provides a suitable target for immunosuppressive therapy. The discrepancy in survival data obtained by mAb and beta7 deficiency may be a result of the more rapid activation of compensatory mechanisms in the knockout mice.

Diffuse mesenterial sclerosis: a characteristic feature of chronic small-bowel allograft rejection.

Chronic rejection is the major cause of late intestinal allograft dysfunction. The aim of this study was to analyze in detail the histopathological features of chronic rejection in the ACI-to-Lewis rat model of intestinal transplantation. Chronic rejection was achieved in orthotopic small-bowel allografts (ACI-Lewis) by limited immunosuppression with cyclosporin A (CyA). Isogeneic transplants (ACI-ACI) as well as native bowels (ACI) with and without immunosuppression served as controls. Bowels were removed together with the mesenteries 90 days postoperatively and analyzed using sections stained with hematoxylin and eosin as well as Masson's trichrome. The slides were coded, randomized and analyzed by grading of histological abnormalities. The most striking alterations of the allografts were noticed in the mesenteries exhibiting an extensive infiltration by mononuclear cells accompanied by a progressive diffuse fibrosis with shrinking of the mesenteries. These changes were most pronounced in the perivascular areas of the mesenteric arteriae and venae rectae. Three of five allografts showed vasculitis with myointimal proliferation of the arteriae rectae. Focally, there was spill-over of the inflammatory cells onto the intestinal muscularis propria. The mucosa of the allografts showed mild blunting, lymphocytic infiltration of the crypt epithelium and increased crypt cell apoptoses. The submucosa was unaffected, and there were no detectable abnormalities of the enteric ganglion cells. The present data support the view that chronic rejection of intestinal allografts is characterized by a diffuse sclerosing mesenteritis which may significantly contribute to late graft dysfunction. The present model may be useful to study the pathomechanisms of this inflammatory fibrosing process.

Successful tolerance induction under CD40 ligation in a rodent small bowel transplant model: first report of a study with the novel antibody AH.F5.

Intestinal transplantation has been hampered by high rates of intestinal allograft rejection. One mechanism of altering rejection in other organ transplant models has been blockade of second set T-cell costimulatory signals. AH.F5, a novel hamster anti-rat monoclonal antibody to CD154, blocks CD40-dependent T-cell costimulation. We hypothesized that blockade of this pathway might abrogate rejection in a rodent orthotopic survival model of intestinal transplantation. Eight groups were studied with different dosing schema, including syngeneic transplants (group 1), untreated allogeneic transplants (group 2), allogeneic transplants plus multiple doses of AH.F5 alone given IV or s.c. (groups 3 and 4), allogeneic transplants plus donor splenocyte preconditioning with and without single dose AH.F5 (groups 5 and 6), and donor splenocyte preconditioning followed by multiple doses of AH.F5 with and without thymectomy (groups 7 and 8). Control animals all died within 12 days of transplantation, whereas antibody-alone and splenocytes-alone resulted in modest prolongation of survival to 16 days. Only animals treated with splenocytes before transplantation and AH.F5 survived long-term (>60 days, group 8). These animals tolerated donor-specific skin grafts, rejected third-party grafts, and fed normally. However, their weight gain was subnormal and they demonstrated intestinal muscular thickening, which might represent chronic rejection. Thymectomy prevented the induction of tolerance. AH.F5 prevents acute intestinal allograft rejection in combination with donor-specific splenocyte preconditioning. We achieved long-term survival and the animals appeared tolerant. Central conditioning is essential for success with this antibody when used alone. Further studies with different dosing regimens or second agents seem warranted.

Serum citrulline is a potential marker for rejection of intestinal allografts

Serum citrulline is a potential marker for rejection of intestinal allografts

Cutting edge: membrane lymphotoxin regulates CD8(+) T cell-mediated intestinal allograft rejection.

Blocking the CD28/B7 and/or CD154/CD40 costimulatory pathways promotes long-term allograft survival in many transplant models where CD4(+) T cells are necessary for rejection. When CD8(+) T cells are sufficient to mediate rejection, these approaches fail, resulting in costimulation blockade-resistant rejection. To address this problem we examined the role of lymphotoxin-related molecules in CD8(+) T cell-mediated rejection of murine intestinal allografts. Targeting membrane lymphotoxin by means of a fusion protein, mAb, or genetic mutation inhibited rejection of intestinal allografts by CD8(+) T cells. This effect was associated with decreased monokine induced by IFN-gamma (Mig) and secondary lymphoid chemokine (SLC) gene expression within allografts and spleens respectively. Blocking membrane lymphotoxin did not inhibit rejection mediated by CD4(+) T cells. Combining disruption of membrane lymphotoxin and treatment with CTLA4-Ig inhibited rejection in wild-type mice. These data demonstrate that membrane lymphotoxin is an important regulatory molecule for CD8(+) T cells mediating rejection and suggest a strategy to avoid costimulation blockade-resistant rejection.

Serum citrulline and rejection in small bowel transplantation: a preliminary report.

There is no known serum marker for intestinal rejection. Serum concentrations of the amino acid citrulline arise almost exclusively from the intestinal mucosa. We examined the impact of acute cellular rejection (ACR) of intestinal allografts on serum citrulline levels. Citrulline concentrations were assayed in serum samples of healthy volunteers (n=6) and seven patients who underwent small bowel transplants (SBTx). Trends in mean citrulline concentrations versus degree of ACR were assessed by matching posttransplantation citrulline concentrations with patients' grade of ACR at time of serum collection. Rejection was confirmed by biopsy and graded by following standardized criteria. An additional patient had citrulline concentrations determined for 31 sequential specimens 3-60 days posttransplant. Mean citrulline concentrations in controls were significantly higher than posttransplantation samples at any rejection grade. Mean concentrations declined significantly as rejection severity increased. The overall downward trend was statistically significant (P<0.05). In sequential measurements, citrulline levels increased significantly over time with declining severity of rejection. The increase in mean citrulline concentration between posttransplant days 3-16 and 52-60 was significant (P<0.01). Serum citrulline levels decline with increasing grade of ACR and may be a useful serum marker for intestinal rejection.

Small bowel transplantation: selection criteria, operative techniques, advances in specific immunosuppresion, prognosis

Intestinal transplantation is now an accepted therapy for intestinal failure when parenteral nutrition therapy cannot be tolerated. During the past year, evidence has been provided indicating that neither stomach nor colon need to be included in the transplant, even if a primary motility disorder is the indication for surgery. The liver should be included in the composite allograft when there are clinical indications of portal hypertension resulting from parenteral nutrition associated cholestasis. When liver disease develops, operations intended to improve gut function should be avoided in preference of early listing for transplantation. During the past year, initial attempts at adult to child intestinal transplantation were carried out with some success; reduction in the diameter of the adult donor bowel may not be uniformly necessary. New immunosuppressive therapies have been employed recently, but few have been subjected to peer review. Experimental models have clarified the pathology, if not the immunobiology, of chronic intestinal allograft rejection and the ability of the liver to promote tolerance of a cotransplanted intestinal allograft. Treatment of posttransplant lymphoproliferative disease has been augmented by the use of anti-CD 20 antibody that targets Epstein-Barr virus infected B-cells for destruction with high specificity.