Specific and nonspecific resistance to local graft-versus-host reaction in F1 hybrids pretreated intravenously with parent-strain spleen cells. I. Two distinct mechanisms

B6D2F1 hybrid mice pretreated i.v. with 5 X 10(7) spleen cells from B6 donors seven days earlier (B6-pretreated B6D2F1 hybrids) develop resistance to local GVHR induced by the subcutaneous injection of spleen cells of either B6 (GVHR-B6) or D2 (GVHR-D2) origin. This resistance has specific and a nonspecific components that concern the GVHR-B6 and the GVHR-D2, respectively. The two types of resistance to GVHR are neither induced under the same conditions nor mediated by the same mechanism. Specific resistance to GVHR is observed in B6D2F1 hybrids pretreated with unseparated, anti-Lyt-1.2+C' treated or 1000 rads-irradiated B6 cells, but not in B6D2F1 hybrids pretreated with anti-Thy-1.2+C' or anti Lyt-2.2+C'-treated B6 cells. In contrast, nonspecific resistance to GVHR is induced only by pretreatment with unseparated B6 cells. Treatment of B6 cells with anti-Thy-1.2, anti-Lyt-1.2, or anti-Lyt-2.2 moAb plus C', or their irradiation at 1000 rads completely abolishes their capacity to induce the nonspecific resistance to GVHR. Moreover, specific resistance to GVHR can be transferred to normal B6D2F1 mice by injection of nylon-adherent, anti-Thy-1.2+C'-treated or 2000-rads-irradiated, but not unseparated or nylon-nonadherent, B6-pretreated B6D2F1 spleen cells. Treatment of nylon-adherent B6-pretreated B6D2F1 cells with anti H-2d antiserum plus C' does not affect their capacity to transfer specific resistance to GVHR. Nonspecific resistance to GVHR can be transferred by unseparated, anti-Lyt-1.1+C' or anti Lyt-2.1+C'-treated, but not by anti-Thy-1.2+C' anti-Lyt-1.2+C', anti-Lyt-2.2+C'-treated or 2000-rads-irradiated B6-pretreated B6D2F1 spleen cells. Both types of resistance are observed in B6D2F1 hybrids pretreated with more than 2.5 X 10(7) B6 spleen cells.     

Suppressor cells in rhesus monkeys treated with antithymocyte globulin

Treatment of rhesus monkey skin allograft recipients with a brief course of rabbit antithymocyte globulin (ATG) produces an enduring immunosuppressive effect on the cellular immune system. Despite early recovery of circulating T cells, in vitro mitogen-induced lymphoproliferative responses of peripheral blood mononuclear cells (PBMCs) remain abnormally depressed for months. In this study we show that depressed mitogen-induced lymphoproliferative responses in these animals are attributable to regulatory effects of adherent PBMCs. Removal of the adherent fraction of PBMCs on Sephadex G-10 produced a significant restoration of the mitogen-induced lymphoproliferative responses in ATG-treated monkeys. Addition of the prostaglandin synthetase inhibitor indomethacin to cultures of unfractionated PBMCs from these animals also caused a significant recovery of the lymphoproliferative response. Indomethacin did not enhance the response of control animals or the response of the nonadherent PBMC fraction of ATG-treated animals. These data suggest that a prostaglandin-dependent mechanism is involved in the suppressive action of the adherent cells. PBMCs from ATG-treated monkeys cocultured with normal cryopreserved autologous cells induced a dose-dependent suppression in the response to both concanavalin A (Con A) and phytohemagglutinin (PHA). The suppressive activity depended upon the adherent cell fraction and was found to be resistant to low-dose gamma irradiation. These data indicate that administration of rabbit ATG induces nonspecific suppressor cells in the rhesus monkey. Preliminary characterization studies suggest the involvement of suppressor monocytes. The possible role of this suppressor cell system in the immunosuppressive action of rabbit ATG is discussed.     

Treatment of human acute graft-versus-host disease with antithymocyte globulin and cyclosporine with or without methylprednisolone

Forty-eight patients with hematologic malignancies treated by allogeneic marrow transplantation developed acute graft-versus-host disease (GVHD), grades II-IV, despite prophylaxis with methotrexate. They were treated with a combination of antithymocyte globulin (ATG) and cyclosporine (CsA), with or without the addition of methylprednisolone (MP). Thirty patients had received HLA-identical and 18 HLA-nonidentical transplants. Median onset of GVHD was day 13 (range 8-60) for patients with HLA-nonidentical grafts and day 18 (range 7-48) for patients given HLA-identical grafts (P = 0.01). Forty-five patients could be evaluated for response on day 7 of therapy. Among these, 13 of 27 given ATG/CSP and 6 of 18 given ATG/CSP/MP improved. Among 33 patients evaluable on day 14 of therapy 13 of 19 given ATG/CSP and 5 of 14 given ATG/CSP/MP showed improvement of GVHD. Patients given HLA-nonidentical grafts responded somewhat (although not significantly) less frequently than patients given HLA-identical grafts. Chronic GVHD developed in 16 of 18 evaluable patients given ATG/CSP and in 5 of 6 given ATG/CSP/MP. Viral, bacterial, and fungal infections were the major cause of death in both groups. Interstitial pneumonitis was more frequent among patients given ATG/CSP/MP. Survival beyond 6 months was 67% among patients treated with ATG/CSP and 25% with ATG/CSP/MP. These data indicate that a regimen of ATG/CSP is of value in the treatment of acute GVHD. The addition of MP was not beneficial and resulted in decreased survival--presumably because of excessive immunosuppression and associated complications.     

Suppression of the in vitro parent antihybrid reaction by spleen cells from F1 hybrids pretreated with spleen cells from the opposite parent strain

Spleen cells from B6C3F1 hybrid mice pretreated i.v. with 5 X 10(7) C3H spleen cells seven days earlier (C3H-pretreated B6C3F1) suppress the in vitro B6 anti-B6C3F1 proliferative and cytotoxic responses, when they are added to cultures of B6 responding and B6C3F1 stimulating spleen cells. This suppression is mediated by a Thy-1+Lyt-1+2+ cell of C3H origin that is radiosensitive at 2000 rads. This suppressor cell is not induced by the injection to B6C3F1 hybrids of spleen cells from the other parent strain (B6) or an allogeneic strain (D2). It does not suppress either the response of the other parent (C3H) or an allogeneic strain (D2) to B6C3F1 antigens, or the response of B6 cells to an allogeneic strain (D2). Its induction depends upon the number and the subpopulation of C3H spleen cells injected since suppression is observed after the injection of more than 2.5 X 10(7) C3H cells, and the suppression inducing cells have the phenotype Thy-1+Lyt-1+2+. This phenomenon is not limited to the C3H-B6C3F1 genetic combination, since it has been observed in all parent hybrid combinations tested to date.     

Specific resistance to local graft-versus-host reaction in F1 hybrids pretreated intravenously with parent-strain spleen cells. Role of cytotoxic T lymphocytes directed against receptors for the major histocompatibility complex

Pretreatment of B6D2F1 hybrids by injection of B6 spleen cells by subcutaneous or intravenous routes induces specific resistance to the local graft versus host reaction provoked by s.c. engraftment of B6 spleen cells. This resistance has been attributed to the presence in the pretreated F1 hybrids of cytotoxic T lymphocytes directed against receptors that recognize the D2 alloantigens (anti-D2-receptor CTL). However, this hypothesis would seem to be challenged, at least partially, by our previously published results showing that a) when tested before induction of GVHR, the anti-D2-receptor CTL are detectable in F1 hybrids pretreated only by the s.c. route but not by the i.v. route; and b) specific resistance to GVHR observed in i.v.-pretreated F1 hybrids is mediated by a nylon-adherent, Thy-1-, radioresistant (2000 rads) suppressor cell of B6 origin that does not manifest any anti-D2-receptor CTL activity. However, these results did not allow us to exclude the possibility of the presence, in the i.v.-pretreated F1 hybrids, of anti-D2-receptor precursor CTL that could be reactivated during the GVHR by the D2-receptors expressed on the proliferating clone of grafted B6 cells, then differentiate to the receptor-specific CTL effectors that control the development of the GVHR. That is why we have studied in the present work the CTL activity developed against D2-receptors after induction of GVHR in either normal or resistant F1 hybrids. Our results show that F1 hybrids protected against GVHR by i.v. pretreatment with B6 cells or by a transfer of nylon-adherent spleen cells from i.v.-pretreated syngeneic F1 mice do not manifest enhanced anti-D2-receptor CTL activity. When considered along with our previous observations, these results favor our hypothesis that anti-D2-receptor CTL are not involved in the specific resistance to GVHR observed in the i.v.-pretreated F1 hybrids.     

Induction immunosuppression with rabbit antithymocyte globulin in pediatric liver transplantation.

Routine use of rabbit antithymocyte globulin (RATG) induction therapy remains controversial in pediatric liver transplantation. We reviewed our experience of 18 cadaveric liver transplants in 18 children over a span of 2 years. All patients received the same immunosuppression: perioperative steroid therapy with taper, 3 doses of RATG, and maintenance therapy of steroids and tacrolimus started on postoperative day 3. Mean follow-up was 2.2 +/- 0.2 years. End-stage liver disease was secondary to biliary atresia in 10 patients (56%) and metabolic disorders in 4 patients (22%). Graft and patient survival were 89%. Serum bilirubin was 1.2 mg/dL, 1.1 mg/dL, 0.5 mg/dL, and 0.5 mg/dL at 1, 3, 6, and 12 months, respectively. The 2 mortalities were secondary to multiple organ system failure. Overall rejection rate was 17% (3/18). Rejection episodes occurred at 4, 6, and 7 months. Two patients were treated with steroids; the third was treated with OKT3. No patient has developed posttransplant lymphoproliferative disease. Serum creatinine was 0.7 mg/dL, 0.6 mg/dL, 0.6 mg/dL, and 0.6 mg/dL at 1, 3, 6, and 12 months, respectively, among surviving patients. In conclusion, our data suggest that RATG induction with steroid and tacrolimus maintenance therapy is safe, easy to use, and effective in the prevention of rejection.     

Extended treatment with antithymocyte globulin (ATGAM) in renal allograft recipients.

Twenty renal allograft recipients were treated with antithymocyte globulin (ATGAM; ATG) for up to 16 weeks in addition to azathioprine and prednisone, while 20 controls received no ATG. The ATG group showed a lower incidence of first rejection episodes during the first month after transplantation, and also a better functional graft survival rate up to 2 years after transplantation. The results in this early ATG trial were better than those in subsequent trials which used 14-day treatment regimens. Longer treatment deserves another look.