[Development of immune tolerance in liver transplantation]. / Desarrollo de inmunotolerancia en el trasplante hepático.

The liver is a privileged organ and has a lower incidence of rejection than other organs. However, immunosuppressive regimens are still required to control the alloreactive T-lymphocyte response after transplantation. These treatments may lead to severe complications, such as infectious diseases, cancers, cardiovascular diseases and chronic renal insufficiency. In clinical transplantation there is increasing evidence that some liver transplant recipients who cease taking immunosuppressive (IS) drugs maintain allograft function, suggesting that tolerance is already present. This strategy is feasible in 25-33% of liver transplant recipients. A series of experimental and clinical observations indicates that liver allografts can even provide "tolerogenic" properties for other organ grafts. In the clinical setting, clinical operational tolerance (COT) is defined as the absence of acute and chronic rejection and graft survival with normal function and histology in an IS-free, fully immunocompetent host, usually as an end result of a successful attempt at IS withdrawal. The exact mechanisms involved in achieving transplant tolerance remain unknown, although animal models suggest a possible role for regulatory T cells (Treg). Recent data have demonstrated an increase in the frequency of CD4+ CD25(high) T cells and FoxP3 transcripts during IS withdrawal in operationally tolerant liver transplant recipients. The data obtained from transcriptional profiling of the peripheral blood of IS-free liver transplant recipients suggest that there is a molecular signature of tolerance that could be employed to identify tolerant liver transplant recipients and that innate immune cells are likely to play a major role in the maintenance of COT after liver transplantation.

Persistent differences in the level of chimerism in B versus T cells of Freemartin cattle.

Parabiosis during pregnancy regularly results in an exchange of hematopoietic stem cells between cattle twins. We have exploited this phenomenon and show differences in the levels of chimerism between the descendant cell types. Female recipients were screened for the levels of male donor contribution in surface IgM-bearing B lymphocytes versus CD3(+) T lymphocytes using immunomagnetic fractionation and Y-chromosome specific in situ hybridization. Two calves of 15 were discovered to have less than 10% of B cells but over 70% of T cells and other blood leukocytes of male origin. The donor cell ratios remained stable for 9 months. Analysis of lymphoid tissues revealed a similar cell type specific pattern of male cell ratios in both female calves and one twin brother. These findings are in agreement with the existence of an essentially self-sufficient population of developing B cells that gives rise to the peripheral pool of B lymphocytes in young cattle.

Oligoclonal Peyer's patch follicles in the terminal small intestine of cattle.

In small ruminants, the development of B cells differs from that in mice or in man. The anti-body repertoire is expanded in the Peyer's patches of the terminal ileum where each B-cell follicle is found by a few cells. To investigate the amount of founder clones in bovine ileal follicles, we have used sex mismatched cattle twins. These animals are chimeric due to placental anastomoses. Y-chromosome targeted in situ hybridization was used to trace donor-derived cells of the male genotype in a female recipient (called a freemartin). A strong clustering of lymphoid cells originating from either twin was seen in the ileal Peyer's patches (IPPs). Furthermore, the follicles displayed a low amount of immunoglobulin heavy chain gene configurations in comparison with the splenic or jejunal follicles. These findings strongly suggest that as in sheep, the B-cell follicles in cattle IPPs develop oligoclonally.

Chimerism and tolerance: from freemartin cattle and neonatal mice to humans.

Bone marrow transplantation (BMT) results in hematopoietic chimeras that demonstrate donor specific tolerance to tissue and cellular grafts. The clinical application of chimerism to induce tolerance is limited by the morbidity associated with human BMT: failure of engraftment, graft-versus-host disease (GVHD), and toxic host conditioning. BMT in an immunologically mature host has until recently been believed to require complete ablation of the host's immune system to allow donor engraftment. Lethal conditioning is associated with significant morbidity and mortality. Stable multilineage mixed allogeneic chimerism has more recently been achieved in mice using partial myeloablation prior to BMT. Chimeras prepared in this fashion exhibit donor specific tolerance in vitro and in vivo similar to lethally-conditioned recipients. A second factor that has limited the widespread application of BMT to nonmalignant disease, including attempts to induce tolerance, is GVHD. Although T-cell depletion of donor marrow reduces the incidence of GVHD, engraftment is often jeopardized. Although highly purified stem cells (SC) engraft at relatively low doses in syngeneic recipients, they do not durably engraft in MHC-disparate recipients. It has recently become clear that a second cell (facilitating cell) that enhances bone marrow engraftment and minimizes the occurrence of GVHD is required for SC to engraft in MHC-disparate recipients. Methods to optimize engraftment yet minimize GVHD may provide an approach to apply BMT clinically. With decreased morbidity through incomplete recipient conditioning and the ability to engineer a bone marrow graft to contain only the desired cells to optimize engraftment, BMT may provide a reasonable strategy to treat nonmalignant diseases including enzyme deficiencies, hemoglobinopathies, autoimmune diseases, and species-specific viral infections such as HIV. BMT-induced donor specific tolerance may benefit recipients of solid organ transplants by eliminating the need for nonspecific immunosuppression and by preventing chronic rejection. This review will focus on approaches to enable BMT yet minimize recipient morbidity and mortality.

Interspecific chimerism--the characterization and immunological responsiveness of Bos taurus-Bos indicus haemopoietic chimeras produced by embryo transfer.

Bos taurus-Bos indicus twin calves were produced by embryo transfer and their degree of haemopoietic chimerism and immune responsiveness examined. All 10 pairs of Friesian-Brahman twins were chimeric compared with only 6 out of 12 pairs of Jersey-Brahman twins. In the Friesian-Brahman twins, Friesian lymphocytes predominated in all calves while in the Jersey-Brahman twins there was no uniform dominance of one cell type over another. All calves that had chimeric lymphocyte populations were also erythrocyte chimeras, although in two pairs of twins the genotype of the erythrocytes was different from the genotype of the lymphocytes. The lymphocytes of all chimeric calves failed to respond in mixed lymphocyte culture (MLC) tests to the lymphocytes of their co-twins but gave vigorous responses to cells from other calves. Primary and secondary immune responses to keyhole limpet haemocyanin (KLH) antigen were studied in the efferent prefemoral lymph in normal Jersey, Friesian and Brahman calves, in 1 set of Jersey-Brahman twins and in 5 sets of Friesian-Brahman twins. The immune responses of chimeric twins with genetically identical lymphocyte populations were quite dissimilar and were characteristic of the somatic genotype of the calf and not of the genotype of the lymphocyte population. The chimeric status of the cell population in the lymph did not change significantly during the response. These results suggest that either the genetic capacity of a population of lymphocytes to recognize and respond to antigens can be modified during ontogeny in an alien genetic environment or that a crucial determinant of the final outcome of an immune response is not the immunocompetent cell itself but the physiological and anatomical environment in which it exists.

[Role of HY antigen in human and mammalian gonadal organogenesis (author's transl)]. / Rôle de l'antigène HY dans l'organogenèse des gonades chez l'homme et les mammifères.

H-Y (histocompatibility Y) antigen plays a role in gonadal organogenesis, which is poorly understood. Indeed, it is not immunologically well-defined. The use of cytotoxicity tests cannot lead to a right quantification of its expression. In vertebrates, it is a marker of the heterogametic sex. As it is also detected in invertebrates, H-Y antigen is therefore ubiquitous and has a high phylogenic conservation. Its role, as an organizer of mammal testes, was carried out by controversial experiments of dissociation-reorganization, in gonadal cells. In gonads. H-Y antigen can be studied under 3 diverse aspects : secretion, fixation and expression, Recently, Ohno proposed a very attractive pattern of testicular and ovarian organogenesis. In males, organization of seminiferous tubules is the result of the interaction between H-Y antigen and its specific gonadal receptor. In females, primordial follicle formation is induced by the interaction between an hypothetic ovary-organizing antigen (similar to H-Y antigen) and the same specific receptor as in testis. These different hypotheses are discussed. In addition, it is underlined how expression of H-Y antigen (anchorage site of H-Y antigen on plasma membrane associated with beta 2-m) can be distinguished from its fixation on gonadal specific receptors. In view of controversial data, the masculinisation of bovine free-martin gonads by H-Y antigen is discussed. In XX males and XY females, H-Y expression which is variable is correlated with its gonadal organizing role. Finally, at present time location of H-Y structural and control genes remains unsolved. In view of all hypotheses postulated, it is not clearly demonstrated how H-Y antigen can act on gonadal organogenesis. Nevertheless, priority of cell-cell recognition, via H-Y antigen, has to be emphasized.

Serum-borne H-Y antigen in the fetal bovine freemartin.

In mammals, ovarian cells that have been exposed to soluble H-Y antigen acquire the H-Y+ cellular phenotype. They absorb H-Y antibody in serological tests. To determine whether masculinization of the bovine freemartin, the synchorial female twin of a bull, could be due to attachment of circulating bull-twin-derived H-Y in the freemartin gonad, we exposed normal fetal ovarian cells to serum from fetal bulls, fetal cows and fetal freemartins. Ovarian target cells became H-Y+ after exposure to serum from fetal bulls or fetal freemartins, but not after exposure to serum from fetal cows. In a new competitive binding radioassay, uptake of Daudi-secreted tritiated H-Y was inhibited in fetal ovarian target cells first exposed to mouse testis supernatant, a demonstrated source of soluble H-Y. It was also inhibited in ovarian target cells exposed to serum from fetal bulls or fetal freemartins; uptake was unaffected by exposure to serum from fetal cows. Since Daudi-secreted H-Y is known to induce precocious testicular organogenesis in XX indifferent gonads in culture, we infer that initial transformation of the freemartin gonad is due to H-Y antigen that is secreted in the fetal bull, transmitted in the common chorionic vasculature and bound by gonadal receptors of the fetal cow.