Noriko Murase

Cell migration, chimerism, and graft acceptance

The chimeric nature of the transplanted liver was first shown in our long-surviving human recipients of orthotopic hepatic allografts in 1969.1 When liver grafts were obtained from cadaveric donors of the opposite sex, karyotyping studies showed that hepatocytes and endothelium of major blood vessels retained their donor specificity, whereas the entire macrophage system, including Kuppfer cells, was replaced with recipient cells.2 Where donor cells that had left the liver had gone was unknown, but their continued presence was confirmed by the acquisition and maintenance in recipient blood of new donor-specific immunoglobulin (Gm) types1,3 and red-blood-cell alloantibodies, if donors with ABO non-identity were used.4 Davies et al5 attributed the secretion of new soluble HLA class I antigens of donor type to transplanted hepatocytes. However, these HLA molecules come from bone-marrow-derived macrophages and/or dendritic cells,6 and probably have the same origin from migrated donor cells as the additional Gm types and red-cell antibodies.

Tolerogenic immunosuppression for organ transplantation

BACKGROUND Insight into the mechanisms of organ engraftment and acquired tolerance has made it possible to facilitate these mechanisms, by tailoring the timing and dosage of immunosuppression in accordance with two therapeutic principles: recipient pretreatment, and minimum use of post-transplant immunosuppression. We aimed to apply these principles in recipients of renal and extrarenal organ transplants. METHODS 82 patients awaiting kidney, liver, pancreas, or intestinal transplantation were pretreated with about 5 mg/kg of a broadly reacting rabbit antithymocyte globulin during several hours. Post-transplant immunosuppression was restricted to tacrolimus unless additional drugs were needed to treat breakthrough rejection. After 4 months, patients on tacrolimus monotherapy were considered for dose-spacing to every other day or longer intervals. FINDINGS We frequently saw evidence of immune activation in graft biopsy samples, but unless this was associated with graft dysfunction or serious immune destruction, treatment usually was not intensified. Immunosuppression-related morbidity was virtually eliminated. 78 (95%) of 82 patients survived at 1 year and at 13-18 months. Graft survival was 73 (89%) of 82 at 1 year and 72 (88%) of 82 at 13-18 months. Of the 72 recipients with surviving grafts, 43 are on spaced doses of tacrolimus monotherapy: every other day (n=6), three times per week (11), twice per week (15), or once per week (11). INTERPRETATION The striking ability to wean immunosuppression in these recipients indicates variable induction of tolerance. The simple therapeutic principles are neither drug-specific nor organ-specific. Systematic application of these principles should allow improvements in quality of life and long-term survival after organ transplantation.

Clinical Intestinal Transplantation: A Decade of Experience at a Single Center

ObjectiveTo assess the long-term efficacy of intestinal transplantation under tacrolimus-based immunosuppression and the therapeutic benefit of newly developed adjunct immunosuppressants and management strategies. Summary Background DataWith the advent of tacrolimus in 1990, transplantation of the intestine began to emerge as therapy for intestinal failure. However, a high risk of rejection, with the consequent need for acute and chronic high-dose immunosuppression, has inhibited its widespread application. MethodsDuring an 11-year period, divided into two segments by a 1-year moratorium in 1994, 155 patients received 165 intestinal allografts under immunosuppression based on tacrolimus and prednisone: 65 intestine alone, 75 liver and intestine, and 25 multivisceral. For the transplantations since the moratorium (n = 99), an adjunct immunosuppressant (cyclophos-phamide or daclizumab) was used for 74 transplantations, adjunct donor bone marrow was given in 39, and the intestine of 11 allografts was irradiated with a single dose of 750 cGy. ResultsThe actuarial survival rate for the total population was 75% at 1 year, 54% at 5 years, and 42% at 10 years. Recipients of liver plus intestine had the best long-term prognosis and the lowest risk of graft loss from rejection (P = .001). Since 1994, survival rates have improved. Techniques for early detection of Epstein-Barr and cytomegaloviral infections, bone marrow augmentation, the adjunct use of the interleukin-2 antagonist daclizumab, and most recently allograft irradiation may have contributed to the better results. ConclusionThe survival rates after intestinal transplantation have cumulatively improved during the past decade. With the management strategies currently under evaluation, intestinal transplant procedures have the potential to become the standard of care for patients with end-stage intestinal failure.

Systemic chimerism in human female recipients of male livers

We have previously reported data from clinical and laboratory animal observations which suggest that organ tolerance after transplantation depends on a state of balanced lymphodendritic cell chimerism between the host and donor graft. We have sought further evidence to support this hypothesis by investigating HLA-mismatched liver allograft recipients. 9 of 9 female recipients of livers from male donors had chimerism in their allografts and extrahepatic tissues, according to in-situ hybridisation and molecular techniques 10 to 19 years posttransplantation. In 8 women with good graft function, evidence of the Y chromosome was found in the blood (6/8), skin (8/8), and lymph nodes (7/8). A ninth patient whose transplant failed after 12 years from recurrent chronic viral hepatitis had chimerism in her lymph nodes, skin, jejunum, and aorta at the time of retransplantation. Although cell migration is thought to take place after all types of transplantation, the large population of migratory cells in, and the extent of their seeding from, hepatic grafts may explain the privileged tolerogenicity of the liver compared with other organs.

Chimerism and donor-specific nonreactivity 27 to 29 years after kidney allotransplantation

Chimerism was demonstrated with immunocytochemical and/or polymerase chain reaction techniques in kidney allografts and in the native skin, lymph nodes, or blood of 5 of 5 patients who received continuously functioning renal transplants from 1 or 2 haplotype HLA mismatched consanguineous donors (4 parents, 1 aunt) 27–29 years ago. In the 4 cases where the kidney donor still was alive to provide stimulator lymphocytes for testing, these provoked no (n=2) or modest (n=2) MLR in contrast to vigorous MLR to third party lymphocytes. In all 4 cases, the donor cells failed to generate in vitro cytotoxic effector cells (cell-mediated lymphocytotoxicity). These findings are in accord with the hypothesis that cell migration, repopulation, and chimerism are seminal events that define graft acceptance and ultimately can lead to acquired donor-specific nonresponsiveness (tolerance).

Acute rejection is associated with antibodies to non-Gal antigens in baboons using Gal-knockout pig kidneys.

We transplanted kidneys from α1,3-galactosyltransferase knockout (GalT-KO) pigs into six baboons using two different immunosuppressive regimens, but most of the baboons died from severe acute humoral xenograft rejection. Circulating induced antibodies to non-Gal antigens were markedly elevated at rejection, which mediated strong complement-dependent cytotoxicity against GalT-KO porcine target cells. These data suggest that antibodies to non-Gal antigens will present an additional barrier to transplantation of organs from GalT-KO pigs to humans.

The lost chord: microchimerism and allograft survival

Recent evidence suggests that passenger leukocytes migrate after organ transplantation and produce persistent chimerism, which is essential for sustained survival of the allografts. Here, Thomas Starzl and colleagues argure that this hematolymphopoietic chimerism provides an important framework for the interpretation of basic and therapeutically oriented transplantataion research.

Five hundred intestinal and multivisceral transplantations at a single center: major advances with new challenges.

Objective:To assess the evolution of visceral transplantation in the milieu of surgical technical modifications, new immunosuppressive protocols, and other management strategies. Summary Background Data:With the clinical feasibility of intestinal and multivisceral transplantation in 1990, multifaceted innovative tactics were required to improve outcome and increase procedural practicality. Methods:Divided into 3 eras, 453 patients received 500 visceral transplants. The primary used immunosuppression was tacrolimus-steroid-only during Era I (5/90–5/94), adjunct induction with multiple drug therapy during Era II (1/95–6/01), and recipient pretreatment with tacrolimus monotherapy during Era III (7/01–11/08). During Era II/III, donor bone marrow was given (n = 79), intestine was ex vivo irradiated (n = 44), and Epstein-Barr-Virus (EBV)/cytomegalovirus (CMV) loads were monitored. Results:Actuarial patient survival was 85% at 1-year, 61% at 5-years, 42% at 10-years, and 35% at 15-years with respective graft survival of 80%, 50%, 33%, and 29%. With a 10% retransplantation rate, second/third graft survival was 69% at 1-year and 47% at 5-years. The best outcome was with intestine-liver allografts. Era III rabbit antithymocyte globulin or alemtuzumab pretreatment-based strategy was associated with significant (P < 0.0001) improvement in outcome with 1- and 5-year patient survival of 92% and 70%. Conclusion:Survival has greatly improved over time as management strategies evolved. The current results clearly justify elevating the procedure level to that of other abdominal organs with the privilege to permanently reside in a respected place in the surgical armamentarium. Meanwhile, innovative tactics are still required to conquer long-term hazards of chronic rejection of liver-free allografts and infection of multivisceral recipients.

Clinical intestinal transplantation: new perspectives and immunologic considerations.

Background: Although tacrolimus-based immunosuppression has made intestinal transplantation feasible, the risk of the requisite chronic high-dose treatment has inhibited the widespread use of these procedures. We have examined our 1990–1997 experience to determine whether immunomodulatory strategies to improve outlook could be added to drug treatment. Study Design: Ninety-eight consecutive patients (59 children, 39 adults) with a panoply of indications received 104 allografts under tacrolimus-based immunosuppression: intestine only (n = 37); liver and intestine (n = 50); or multivisceral (n = 17). Of the last 42 patients, 20 received unmodified adjunct donor bone marrow cells; the other 22 were contemporaneous control patients. Results: With a mean followup of 32 ± 26 months (range, 1–86 months), 12 recipients (3 intestine only, 9 composite grafts) are alive with good nutrition beyond the 5-year milestone. Forty-seven (48%) of the total group survive bearing grafts that provide full (91%) or partial (9%) nutrition. Actuarial patient survival at 1 and 5 years (72% and 48%, respectively) was similar with isolated intestinal and composite graft recipients, but the loss rate of grafts from rejection was highest with intestine alone. The best results were in patients between 2 and 18 years of age (68% at 5 years). Adjunct bone marrow did not significantly affect the incidence of graft rejection, B-cell lymphoma, or the rate or severity of graft-versus-host disease. Conclusions: These results demonstrate that longterm rehabilitation similar to that with the other kinds of organ allografts is achievable with all three kinds of intestinal transplant procedures, that the morbidity and mortality is still too high for their widespread application, and that the liver is significantly but marginally protective of concomitantly engrafted intestine. Although none of the endpoints were markedly altered by donor leukocyte augmentation (and chimerism) with bone marrow, establishment of the safety of this adjunct procedure opens the way to further immune modulation strategies that can be added to the augmentation protocol.The advent of tacrolimus allowed clinical intestinal transplantation to become a feasible procedure for patients with irreversible intestinal failure. Over last 5 years, 71 patients underwent intestinal transplantation. Forty-one recipients were children, and 30 recipients were adults. Twenty-five patients received an isolated intestinal graft, 34 patients received a combined liver-intestinal graft, and 12 received a multivisceral graft. The colon was included the intestinal graft in 29 patients. One-year, 2-year, and 4-year actuarial patient survival is 72%, 57%, and 45%, respectively. Our experience has shown that infectious, and immunological problems have caused significant morbidity and mortality. In this paper, we present our clinical experience and overview with intestinal transplantation.

Chimerism after liver transplantation for type IV glycogen storage disease and type 1 Gaucher's disease.

BACKGROUND Liver transplantation for type IV glycogen storage disease (branching-enzyme deficiency) results in the resorption of extrahepatic deposits of amylopectin, but the mechanism of resorption is not known. METHODS We studied two patients with type IV glycogen storage disease 37 and 91 months after liver transplantation and a third patient with lysosomal glucocerebrosidase deficiency (type 1 Gauchers disease), in whom tissue glucocerebroside deposition had decreased 26 months after liver replacement, to determine whether the migration of cells from the allograft (microchimerism) could explain the improved metabolism of enzyme-deficient tissues in the recipient. Samples of blood and biopsy specimens of the skin, lymph nodes, heart, bone marrow, or intestine were examined immunocytochemically with the use of donor-specific monoclonal anti-HLA antibodies and the polymerase chain reaction, with preliminary amplification specific to donor alleles of the gene for the beta chain of HLA-DR molecules, followed by hybridization with allele-specific oligonucleotide probes. RESULTS Histopathological examination revealed that the cardiac deposits of amylopectin in the patients with glycogen storage disease and the lymph-node deposits of glucocerebroside in the patient with Gauchers disease were dramatically reduced after transplantation. Immunocytochemical analysis showed cells containing the HLA phenotypes of the donor in the heart and skin of the patients with glycogen storage disease and in the lymph nodes, but not the skin, of the patient with Gauchers disease. Polymerase-chain-reaction analysis demonstrated donor HLA-DR DNA in the heart of both patients with glycogen storage disease, in the skin of one of them, and in the skin, intestine, blood, and bone marrow of the patient with Gauchers disease. CONCLUSIONS Systemic microchimerism occurs after liver allotransplantation and can ameliorate pancellular enzyme deficiencies.