Mechanism of skin allograft enhancement across an H-2 class I mutant difference. Evidence for involvement of veto cells

Intravenous injection of spleen cells across mutant class I H-2 incompatibility results in a drastic donor-specific prolongation of skin allograft survival and a marked decrease in the donor-specific cytotoxic T lymphocyte precursor (CTLp) frequency. This immunosuppressive effect depends on the presence of radiosensitive T cells in the donor cell inoculum. It was excluded that a graft-vs.-host reaction was responsible for the observed effects. In mixing experiments, spleen cells from animals transfused with allogeneic lymphocytes could not suppress a normal CTL response against the alloantigen, despite an excess of putative recipient-derived spleen suppressor cells. The data are compatible with the idea that donor T cells function as veto cells which inactivate recipient CTLp directed against the alloantigen expressed by the veto cell.     

Comparison of tolerance inducibility to class I or class II antigens between cyclophosphamide (CP)-induced tolerance and transfusion with donor cells: general effectiveness of CP-induced tolerance and difference of skin graft prolongation in each class I antigen-disparate combination.

Transfusion with allogeneic cells alone was reported to prolong skin allograft survival in the MHC class I antigen alone-disparate combination of B6.C-H-2bm1 (bm1; Kbm1, IAb, IE-, Db)-->C57BL/6 CrSlc (B6; H-2b). Using 6 class I antigen-disparate and 2 class II antigen-disparate combinations, we compared the effectiveness for skin graft prolongation between transfusion with donor cells (TDC) and our system of CP-induced tolerance, which comprises intravenous (i.v.) injection of allogeneic cells followed by cyclophosphamide (CP), i.e., TDC followed by CP. TDC was effective only in the combination of bm1-->B6, but not at all in the other combinations. On the other hand, CP-induced tolerance was effective in the 5 class I antigen-disparate and 2 class II antigen-disparate combinations except for bm1-->B6 combination. These results suggest that CP-induced tolerance may be more general for the induction of unresponsiveness than TDC.     

Induction of tolerance across major barriers using a two-step method with genetic analysis of tolerance induction

Using a murine skin allograft tolerance induction system that consists of intravenous injection of 1 x 10(8) allogeneic spleen cells followed by intraperitoneal (i.p.) injection of 200 mg/kg cyclophosphamide (CP) 2 days later, sensitivity to tolerance induction was examined across various histocompatibility (H) barriers. Although each group of class I, class II or multiminor H antigens was not by itself a prohibitively strong barrier, resistance to tolerance induction increased when the three types of barriers were combined in various ways. When the donor-recipient combinations were disparate at the entire spectrum of both H-2 plus non H-2 antigens (fully allogeneic), profound tolerance to skin allografts was not induced by this method in any of the combinations examined. Based on these results, induction of tolerance across fully allogeneic barriers was attempted in C57BL/10SnJ (B10; H-2b) mice against C3H/HeSnJ (C3H; H-2k) strain by addressing the 11 barriers as two separate challenges. B10 mice were first given B10.BR/SgSnJ (B10.BR; H-2k) spleen cells plus CP to make them tolerant to the H-2k component represented among C3H antigens, and then later were given C3H spleen cells plus CP to establish a tolerant state to the remainder of the disparate antigens of the C3H donors. After these two separate manipulations, C3H skin was accepted in the B10 mice, and normal hair growth was observed in the grafted C3H skin. By contrast, B10 mice given C3H spleen cells plus CP and then again another injection of C3H spleen cells plus CP were not rendered tolerant to C3H skin. In B10 mice, tolerance to C3H induced with B10.BR spleen cells plus CP and then C3H spleen cells plus CP was specific to C3H, and the tolerant B10 mice rejected third-party skin from DBA/2J (DBA; H-2d) strain in a normal fashion. In transfer experiments, the mechanism of tolerance was found to be based largely on reduction of the effector cells rather than on a mechanism involving active suppression. Assays for chimerism revealed that maintaining the tolerant state required persistence of cells of donor origin. These data indicate that in a primary immune response to a certain dose of allogeneic cells (tolerogen), the existence of a relatively large proportion of potentially reactive clones in the host may trigger proliferation of only a part of the population and some of the potentially reactive cells may differentiate rapidly without a prolonged period of proliferation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Immune response to H-2 class I antigens on platelets. II. Specific decrease of H-2 class I-specific antibody response induced by treatment with allogeneic platelets

Mice pretreated with injections of allogeneic platelets were found to mount a decreased antibody response upon challenge by lymphocytes of the same donor strain. This decrease was mediated by platelets themselves, and not by leucocytes and red cells contaminating the platelet suspension. It affected specifically antibodies reactive with H-2 class I antigens present on donor platelets. This phenomenon may be related to the lack of class II or some non-H-2 antigens on platelets, and/or to properties of their class I antigens (soluble molecules adsorbed from the plasma). These findings emphasize the potential usefulness of purified platelet transfusions preceding organ transplantation in man.     

Cyclophosphamide-induced chimera-type tolerance to allografts: an overview of drug-induced immunological tolerance

A CP-induced tolerance induction method in mice that consists of donor spleen cell injection followed 2 days later by CP treatment was found useful for inducing permanent skin allograft tolerance. In order to regularly overcome fully allogeneic (major H-2 plus multiminor H) barriers, both a reduction of recipient T cells with anti-T cell antibody before tolerance induction and a supplement of the spleen cells with bone marrow cells were required. The reduction of recipient T cells was necessary because the antigen-stimulated-and-proliferating cell destruction with CP, which is considered to be the main mechanism of this system, is not completed due to the less proliferative quick maturation of T cells when a relatively large number of reactive T cells exist. The bone marrow cells were required probably because the establishment of stable mixed chimerism is necessary to ensure skin graft survival. This system is considered to be potentially applicable to human transplantation.     

Drug-induced tolerance to allografts in mice. X. Augmentation of split tolerance in murine combinations disparate at both H-2 and non-H-2 antigens by the use of spleen cells from donors preimmunized with recipient antigens

In a fully allogeneic murine combination of C3H/HeSlc (C3H) (H-2k) and C57BL/6CrSlc (B6) (H-2b), C3H mice were primed i.v. with 1 X 10(8) spleen cells from B6 mice preimmunized i.v. with 5 X 10(7) C3H spleen cells and then were given i.p. 200 mg/kg cyclophosphamide (CP) 2 days later (Im-B6-Sc plus CP group). The tolerant state in those recipient mice was compared with that in mice made tolerant conventionally with 1 X 10(8) naive B6 spleen cells plus 200 mg/kg CP (naive-B6-Sc plus CP group). B6 skin was rejected in an almost normal fashion in both the naive-B6-Sc plus CP group and the Im-B6-Sc plus CP group. However, EL4 tumor allografts (B6 origin) inoculated after complete rejection of B6 skin grafts were specifically accepted in both groups. Moreover, the tumor growth in the Im-B6-Sc plus CP group was faster than that in the naive-B6-Sc plus CP group. Mixed lymphocyte reaction, cytotoxic T lymphocyte activity, and antibody production against the tolerogen were depressed more profoundly in the Im-B6-Sc plus CP group than in the naive-B6-Sc plus CP group. These observations were consistent with the results from tumor allografting. The other immunological parameters examined in the present study, including helper T cell activity and delayed foot-pad reaction, were retained in the Im-B6-Sc plus CP group at the same levels as in the naive-B6-Sc plus CP group. These observations were consistent with the results from skin allografting. In conclusion, tumor allograft tolerance was made more profound by the use of spleen cells from donors preimmunized with recipient antigens as the tolerogen than by the use of naive spleen cells. However, skin allograft tolerance was not achieved at all by these same treatments. The contribution of graft-versus-host disease to this phenomenon was excluded by the chimeric analysis in AKR/JSea (H-2k) mice given the preimmunized (with AKR antigens) B6 spleen cells plus CP. These results strongly support the existence of a less proliferative lymphocyte population which does not evoke cell divisions to mature even after the strong stimulation with the preimmunized spleen cells and is resistant to tolerance induction.     

ADULT TRANSPLANTATION TOLERANCE INDUCED BY LENTIL LECTIN: III. INDUCTION OF TRANSPLANTATION TOLERANCE BY LENTIL LECTIN IN MOUSE STRAIN COMBINATIONS WITH DIFFERENT H-2 DISPARITIES: TOLEROGENIC EFFECT OF H-2D REGION ANTIGENS

The survival time of skin allografts was investigated in 28 combinations of mouse strains differing at various loci, in adult recipients treated with lentil seed lectin (LCA). The recipients were given 1 mg of LCA daily (i.v. or i.p.) after transplantation of the skin graft. The immunosuppressive effect of LCA was generally inversely related to the conventional strength of the allotransplantation reaction, i.e. the rate of rejection of allografts on untreated recipients. In the case of H-2D antigenic disparities, the grafts usually survived for more than 100 days. The H-2A or H-2K disparity appeared to be relatively resistant to induction of tolerance by LCA treatment. The relative resistance of various H-2 antigens toward the induction of tolerance by LCA differs to some extent from that observed in experiments on the induction of neonatal tolerance by semiallogeneic lymphocytes as described in literature.     

Induction of tolerance to non-H-2 alloantigens is not restricted by the MHC molecules expressed on the donor cells in cyclophosphamide-induced tolerance

A long-lasting skin tolerance to non-H-2 alloantigens has been reported to be readily induced, when the recipient mice were primed intravenously (i.v.) with 5 x 10(7) spleen cells (SC) plus 1.5 x 10(7) bone marrow cells (BMC) from the H-2 identical strains of mice, and treated intraperitoneally (i.p.) with 200 mg/kg CP 2 days later. The present study was conducted in order to clarify whether or not tolerance induction to non-H-2 alloantigens in cyclophosphamide (CP)-induced tolerance is restricted by H-2 alloantigens on donor cells. When BALB/c (BALB; H-2d) female mice were primed i.v. with fully allogeneic [H-2 plus minor histocompatibility (H) antigen-disparate C57BL/10 (B10; H-2b) female SC plus BMC and treated i.p. with CP 2 days later, the survival of H-2 identical B10.D2 nSnSlc (B10.D2; H-2d) female skin was moderately prolonged, but the survival of fully allogeneic B10 female skin was not. When semiallogeneic (B10 x B10.D2) (H-2bxd) female cells or H-2 identical B10.D2 female cells were used as the tolerogen, the survival of B10.D2 female skin was prolonged moderately or permanently, respectively. There was no significant difference between the prolongations of B10.D2 female skin graft survival in the BALB mice treated with the B10 cells followed by CP, and the (B10 x B10.D2) cells followed by CP. Similar results were observed in H-Y antigen-disparate combinations. These results strongly suggest that tolerance induction to non-H-2 alloantigens is not restricted by the products of donor MHC in CP-induced tolerance.     

Instillation of allogeneic lung antigen-presenting cells deficient in expression of major histocompatibility complex class I or II antigens have differential effects on local cellular and humoral immunity and on pathology in recipient murine lungs

Recognition of allogeneic major histocompatibility complex (MHC) molecules expressed on donor lung antigen-presenting cells (APCs) by host T lymphocytes is believed to stimulate lung allograft rejection. However, the specific roles of donor MHC molecules in the rejection response is unknown. We report a murine model in which instilling allogeneic lung APCs into recipient lungs induces pathology analogous to acute rejection, and the production of interferon (IFN)-gamma, immunoglobulin (Ig) G2a, and alloantibodies in recipient lungs. Using allogeneic lung APCs (C57BL/6, I-a(b), H-2(b)) deficient in MHC class I, II, or both for instillation into lungs of BALB/c mice (I-a(d), H-2(d)), the purpose of the current study was to determine the specific roles of donor MHC molecules in stimulating local alloimmune responses. The data show that MHC class I or II on donor APCs induced IFN-gamma and IgG2a synthesis locally, though less than that induced by wild-type cells. Both MHC class I and II were required to induce alloantibody production. Instillation of wild-type or class I- or class II-deficient APCs induced comparable pathologic lesions in recipient lungs, and more severe than that induced by MHC-deficient cells. These data show that donor MHC class I and II molecules have differential effects in the stimulation of local alloimmune responses.     

Donor and recipient specific tolerance in cells from semi-allogeneic, H-2 subregion compatible or fully allogeneic bone marrow chimeras attributable to clonal deletion

Specificities of tolerance induced in allogeneic bone marrow (BM) chimeras which had been established by injecting allogeneic BM cells pretreated with anti-Thy-1 mAb alone (without complement (C)) were analyzed using Simonsen's splenomegaly assay. Lymphocytes from fully allogeneic, semi-allogeneic and H-2 subregion compatible BM chimeras were specifically unresponsive to donor and recipient antigens (Ag). However, cells from H-2 subregion compatible chimeras initiated as vigorously a GVHR in F1 recipient mice, which were disparate at H-2K and I-A regions, as did spleen cells of donor mice, which were incompatible at the entire H-2 and minor histocompatibility regions of the recipients. The donor cells from such chimeras that initiated these considerable GVHR were either CD4+ or CD8+ T cells. Furthermore, synergistic effects by the CD4+ and CD8+ T lymphocytes were also observed. We found no evidence for a suppressive mechanism(s) in maintenance of the specific tolerance in allogeneic chimeras. Further, when lymphoid cells from these chimeras were adoptively transferred to irradiated mice of the donor strain and maintained for 5 days in the absence of recipient Ag (tolerogen), the adoptively transferred cells were shown to retain their unresponsiveness to the recipient Ag. These results reveal that T lymphocytes from allogeneic BM chimeras prepared by our method had been specifically induced to a tolerant state to both donor and recipient Ag and that the major mechanism of induction and maintenance of long-lasting tolerance is attributable to clonal deletion of both CD4+ and CD8+ T cell subsets rather than to the development of a population of suppressor cells of any sort.     

供体脾细胞输注诱导小鼠移植耐受及其机理的研究

目的以持续供体脾细胞输注的方法建立异基因嵌合体动物模型,并探讨移植耐受形成的机制。方法BALB/c（H-2     

Genetic and stimulatory cell type requirements for inducing class I major histocompatibility complex alloantigen-specific in vivo cytotoxic T cell immunity

Current interpretation based on analytical in vitro works that actions of Ia antigens and accessory cells such as macrophages and dendritic cells are crucial for inducing cytotoxic T cell responses to class I major histocompatibility complex (MHC) alloantigens has been challenged by experiments performed in a newly developed system handling in vivo cytotoxic T cell immunity. We first characterized the transplantation immunity for second-set rejection of ascitic tumor allografts as principally induced by allogeneic stimulator cells via direct pathway, and as exclusively mediated by class I MHC alloantigen-specific in vivo cytotoxic T cell activity. By comparison of activities of limiting effective doses (10(4)-10(5) cells per mouse) of various stimulator cells in this defined system, we could demonstrate that genetic disparity at the D region of H-2 to the recipient is just enough for inducing the immunity, and presence of allogeneic or syngeneic Ia antigens in addition to H-2D alloantigens on stimulator cells does not give any premium effect. Further study revealed that allogeneic peritoneal cells rich in macrophages or glass-adherent spleen cells enriched for dendritic cells are not stronger stimulators than allogeneic adherent cell-depleted spleen cells and semi-allogeneic thymocytes. These results fit with the alternative concept that the physiological pathway inducing in vivo cytotoxic T cell immunity for graft rejection entirely depends on class I MHC antigens on live lymphocytes as self-supported stimulators, and does not crucially involve additional stimulator activities of Ia antigens and special accessory cell types, which must be in vivo concerned with induction of other types of transplantation immunity.     

Islet Transplantation in Mice Differing in the I and S Subregions of the H-2 Complex

Pancreatic islets from A.TH mice were transplanted into the spleen or streptozotocin (SZ)-diabetic A.TL mice. The two strains of mice are congenic inbred strains, differing only in the I and S subregions of the H-2 complex. The allogeneic islet grafts decreased blood glucose temporarily, but the islets were rejected after 21 ± 7 days (mean ± SD). The effect of skin presensitization was tested by giving both allogeneic and syngeneic skin grafts to each of a second set of A.TL mice before streptozotocin treatment and islet transplantation. The time course of rejection of the allogeneic islets in animals that received initial skin grafts was decreased to 8 ± 3 days. In both skin-presensitized and non-presensitized mice syngeneic islet grafts were able to restore normoglycaemia, even in animals that had previously rejected an islet allograft. These observations demonstrate that transplantation of pancreatic islet allografts across the I and S subregions of the H-2 complex is sufficient to induce rejection of the islets. The islet rejection was markedly accelerated by prior sensitization with allogeneic skin grafting. It is suggested that elements in allogeneic skin grafts serve as inducers of cytotoxic T-cell responses directed against gene products of the I and/or S subregions present on cells in the allogeneic islets.     

H-2 restriction for lymphocyte homing into lymph nodes.

Migratory patterns into lymph nodes of labeled spleen lymphocytes, injected intravenously, showed considerable differences according to the genetic identity or disparity of donor and recipient mice. The use of mice carrying different H-2 haplotypes in the same B 10 background, or, conversely, the same H-2b haplotype in different backgrounds, showed that no homing was observed in the case of complete disparity at the H-2 complex, contrasting with optimal homing in the case of H-2 identity. Homing was not influenced by disparity at the genetic background. Irradiation of recipient mice did not change the results and suggested that no immunological host-vs.-graft reaction was involved in the H-2 restriction observed. The use of donor-recipient pairs carrying recombinant H-2 haplotypes showed that a single H-2 K and/or H-2 D identity allowed almost optimal homing. The results indirectly suggested that an immunological graft-vs.-host reaction was not involved in the phenomenon but rather a positive recognition of H-2 K or H-2 D identity between donor and recipient cells.     

Mls allo-determinants are recognized in an MHC class II antigen-dependent but unrestricted fashion by a discrete set of T cells

Blocking studies carried out with anti-H-2 class II antigen antibodies show that H-2 class II molecules are intimately involved in the recognition of Mlsa determinants by unprimed, specifically responsive T cells. The blocking of the anti-Mlsa response by these antibodies were not due to inhibition of IL-1 production by H-2 class II antigen positive macrophages. A strain analysis indicates that the response to Mlsa is regulated by H-2-linked genes and that this effect is exerted at the level of the stimulator cells; however, the response to Mls epitopes does not appear to be H-2 restricted. Finally, the ability of spleen cells from Mls-incompatible mice to induce a state of Mls-specific, clonal deletion type tolerance by neonatal injection, shows that Mls determinants exist in qualitatively allelic forms and suggests that these determinants are recognized by specific clones of T cells.     

Tolerance to class I major histocompatibility complex antigens in chicken B cell chimeras. Effect of B cell depletion on transferability of tolerance

B cells from bursa of Fabricius of newly hatched chickens are able to reconstitute the B cell compartment of chemically bursectomized chickens. The resulting B cell chimerism can be detected with monoclonal antibodies against donor B cell alloantigen. Chimeric chickens accept donor-type skin grafts and are unresponsive to donor major histocompatibility complex (MHC) antigens in graft-vs.-host splenomegaly assay and mixed lymphocyte reaction. To study the capability of B cells to induce tolerance to selected MHC antigens, we transplanted class I or total MHC-incompatible bursa cells into cyclophosphamide-treated recipients. The recipients of class I or total MHC-incompatible bursa cells were equally tolerant of donor-MHC antigens. To further analyze the mechanisms of tolerance to class I antigens vs. total MHC, spleen cells from tolerant chickens were transferred to irradiated, histocompatible secondary hosts. The secondary recipients were also unresponsive to bursa cell donor-strain MHC antigens. However, if the chimeric B cells were depleted before the spleen cell transfer, the transfer of tolerance to total MHC was severely inhibited. Instead, most recipients of B cell-depleted spleen cells tolerant of class I antigens were still tolerant of bursa cell donor MHC. Our results indicate differences in the transferability of tolerance to class I antigens vs. entire MHC, although in primary recipients of bursa cells the tolerance is similar. These data suggest that a mechanism that is not dependent on the presence of donor cell chimerism contributes to the maintenance of tolerance to donor class I antigens. The transfer of tolerance to total MHC disparity requires the presence of chimeric cells indicating that donor alloantigen expression is needed for induction of tolerance in the secondary hosts.     

Attenuation of the Fas-L independent B16BL6 melanoma lymphocidic capacity by H-2K class I molecules

We have previously reported that the capacity of highly malignant B16BL6 murine melanoma cells to induce cell death in naive syngeneic lymphocytes stems from the absence of major histocompatibility complex (MHC) class I glycoproteins in these melanoma cells. Our present study provides evidence that the above-mentioned lymphocidic activities of B16BL6 cells are selectively attenuated when the expression of H-2K (but not H-2D or H-2L) MHC class I glycoproteins is reconstituted in these cells. The induction of apoptosis in naive lymphocytes by H-2K-deficient melanoma cells does not involve the Fas ligand (Fas-L)/FAS signaling module, as demonstrated by employing lymphocytes derived from Fas-L(gld)- or Fas(lpr)-deficient mice in co-culture experiments. Furthermore, these tumor cells fail to induce Fas-L-mediated fratricide in co-cultured lymphocytes and do not express Fas-L either when grown alone or co-cultured with lymphocytes. These findings explain the previously widely reported selective down-regulation of certain MHC class I-encoded glycoproteins (H-2K, bur not H-2D or H-2L) during tumor progression. Namely, the initiation of an effective immune response against H-2K-deficient cells could be abrogated at very early steps, as the result of the induction of Fas-L/Fas-independent cell death among naive lymphoid cells.     

CD3+ T cells in severe combined immunodeficiency (scid) mice. IV. Graft-vs.-host resistance of H-2d scid mice to intravenous injection of allogeneic H-2b (C57BL/6) spleen cells.

Intraperitoneal injection of 2 x 10(7) nonfractionated spleen cells (SC) from C57BL/6 (B6, H-2b) mice into completely allogeneic immunodeficient H-2d scid mice induced clinical and histological signs of acute graft-vs.-host disease (GVHD), with all transplanted severe combined immunodeficiency (scid) mice dying in the 3rd week post-transfer. In contrast four out of five scid mice survived for greater than 7 weeks after intravenous (i.v.) injections of equal numbers of B6 SC. Intravenously allotransplanted scid mice analyzed in the 8th week post-transfer had engrafted donor-type CD4+ and CD8+ T cells in the spleens but showed no clinical or histological evidence of GVHD. i.v. injection of 10(7) or 10(6)O B6 SC engrafted allogeneic T cells in spleens of scid recipients; in contrast, i.v. injection of 10(5) nonfractionated B6 SC or 3 x 10(5) cell sorter-purified, naive or anti-H-2d-primed splenic CD4+ or CD8+ B6 T cells led to rejection by young scid recipient mice. B6 T cells engrafted into spleens of scid mice after i.v. injection showed proliferative anti-host alloreactivity in vitro. No cytotoxic reactivity against host-type alloantigens was found in standard 4-h 51Cr-release assays. These data demonstrate that allogeneic T cells injected i.v. into immunodeficient scid mice are partially tolerized against host-type alloantigens.     

In vivo induction of H-2K/D antigens by recombinant interferon-gamma

B10.BR mice received i.v. increasing doses of recombinant interferon-gamma (rIFN-gamma) on three consecutive days. Using an immunoperoxidase technique the distribution of H-2K/D antigens was studied in frozen tissue sections of thirteen organs (kidney, liver, pancreas, esophagus, stomach, small intestine, colon, lungs, heart, brain, thymus, lymph node and spleen). Class I antigens were shown to be induced or enhanced in almost every organ after exposure to IFN-gamma. This effect was particularly conspicuous for renal tubular cells, hepatocytes, bronchiolar epithelial cells, gastric mucous cells, thymic cortical lymphocytes and capillary endothelial cells in heart and kidney. Neurons, glial cells, gastric chief and parietal cells, and pancreas cells were not inducible. The findings show that i.v. application of IFN-gamma leads to strong induction or enhancement of major histocompatibility complex class I antigens in a wide variety of tissues.     

CD3+ T Cells in Severe Combined Immunodeficiency (SCID) Mice. V. Allogeneic T Cells Engrafted into SCID Mice Do Not Induce Graft-Versus-Host Disease in spite of the Absence of Host Veto and Natural Suppressor Cell Activity

Intravenous injection of 10(6) to 10(7) non-fractionated spleen cells (SC) from C57BL/6 (B6, H-2b) mice into completely allogeneic, immunodeficient H-2d severe combined immuno deficiency (scid) mice leads to engraftment of allogeneic donor T cells. Mice analysed in the tenth week posttransfer had engrafted donor-type CD4+ and CD8+ T cells in the spleens but showed no clinical evidence of graft-versus-host disease (GVHD). Transfer of allogeneic T cells engrafted in scid recipients did not induce GVHD upon i.v. injection into secondary scid recipients and lead in most recipients to engraftment of a pure CD4+ T-cell population. Experiments were carried out to investigate the reason(s) for the lack of GVHD in recipient scid mice, i.e. the presence of allotolerance in the engrafted donor T cells. Scid spleen cells (SC) efficiently stimulated alloreactive responses of B6 T cells: scid SC stimulated H-2d-specific cytotoxic responses in a B6 anti-scid mixed lymphocyte culture in vitro, and scid SC injected i.v. into B6 mice efficiently primed splenic cytotoxic lymphocyte precursors against H-2d alloantigens. Moreover when assayed in vitro, no veto activity or natural suppressor activity was detectable in scid SC. These data demonstrate that tolerizing mechanisms currently believed to operate in vivo can not explain the fact that allogeneic T cells injected i.v. into immunodeficient scid mice become tolerized against host-type alloantigens. Our results are discussed in the light of clinical experience of allogeneic T-cell transfer in scid infants.