Long-term abrogation of autoimmune diabetes in nonobese diabetic mice by immunotherapy with anti-lymphocyte serum.

Abstract

We investigated the therapeutic effect of antilnphocyte serum (ALS) on clinically overt diabetes by using a nonobese diabetic (NOD) Immunotherapy with ALS. ALS was obtained by immunizing rabbits with a pool of lymph node cells prepared from NOD, C3H/He, DBA/2, and (C57BL/6 x A)F1 mice (11). A 0.4-ml injection of ALS was given intraperitoneally to diabetic NOD mice on days 0 (the first day of treatment regardless of the duration of diabetes) and +2. Control mice were given 0.4 ml ofnormal rabbit serum (NRS) on days 0 and +2. Blood glucose levels were monitored every 2-3 days initially and once a week thereafter. Achievement of euglycemia (<11 mM) and independence from exogenous insulin treatment after immunotherapy was defined as clinical remission.Immunotherapy with Monoclonal Antibodis (mAbs). Hybridoma cells producing anti-CD4 (anti-L3T4, clone GK 1.5) mAb were kindly provided by Kevin J. Lafferty (Barbara Davis Center for Juvenile Diabetes, Denver). Hybridomas producing anti-CD8 (anti-Lyt 2.1, clone 116-13.1, and antiLyt 2.2, clone 41-3.48) mAb were obtained from American Type Culture Collection. Ascites fluid that contained mAbs at -5 mg/ml was used. Diabetic NOD mice were treated with anti-CD4 mAb at 0.2 ml on days 0, 7, 14, and 21 and/or 0.1 ml of anti-CD8 (anti-Lyt 2.1) mAb on days 0 and 14. This treatment protocol has been shown (12) to achieve long-term removal of CD4' and/or CD8+ T lymphocytes from peripheral lymphoid organs in other inbred mouse strains. Control diabetic NOD mice were treated with 0.2 ml of anti-Lyt 2.2 mAb on days 0, 7, 14, and 21.Flow Cytometric Analysis. Peripheral lymph node cells were stained with fluorescein isothiocyanate conjugates of mAb 53-6.7 (anti-CD8) and phycoerythrin-conjugates of mAb GK1.5 (anti-CD4), both of which were obtained from Becton Dickinson. Two-color analysis was performed with a singlebeam flow cytometer, FACScan (Becton Dickinson) and the FACSCAN Research Software (Becton Dickinson).Islet Isolation and Transplantation. Because insulitis generally starts at 4 weeks of age (1), NOD islet donors were Abbreviations: NOD, nonobese diabetic; ALS, anti-lymphocyte serum; NRS, normal rabbit serum; mAb, monoclonal antibody.tTo whom reprint requests should be addressed at: CRI, New

Comparison of a T Cell Clone and of T Cells from a TCR Transgenic Mouse: TCR Transgenic T Cells Specific for Self-Antigen Are Atypical

“…We previously reported that the BDC-2.5 T cell clone, which was isolated from the spleen and lymph nodes of a diabetic NOD mouse, requires CD8 ϩ T cells from a diabetic NOD mouse to transfer diabetes to adult NOD-scid mice (9). Thus, the activity of this clone is in keeping with studies demonstrating that both CD4 ϩ and CD8 ϩ T cells are required for the development of spontaneous diabetes in the NOD mouse (51) and for the transfer of diabetes to immunodeficient adult NOD recipients by spleen cells from diabetic NOD mice (52)(53)(54). Specifically, both CD4 ϩ and CD8 ϩ T cells have been shown to be absolutely required for the transfer of diabetes to ␥-irradiated NOD (52) and NOD nude (53) FIGURE 9.…”

“…The BDC-2.5 T cell clone and spleen cells from diabetic 2.5 TCR Tg/ NOD-scid mice were activated for 48 h at 37°C in 10% CO 2 in CM containing EL-4 supernatant (equivalent to 7 U/ml IL-2), 10 ng/ml PMA, and 1 g/ml ionomycin before use as effector cells in the 51 Cr release assay. K562-FasL Ϫ effector cells, G10-K562-FasL ϩ effector cells, and L1210-Fas ϩ target cells (28 -30) were a gift from R.C.…”

“…Duke (Ceres Pharmaceuticals, Denver, CO). 51 Cr release assays were performed essentially as previously described (28 -30). Briefly, L1210-Fas ϩ target cells were labeled with 100 Ci 51 Cr for 2 h at 37°C in 10% CO 2 .…”

A novel apoptosis-inducing anti-PSGL-1 antibody for T cell-mediated diseases

“…While in most of these approaches mice were treated prior to or upon disease induction, only few were reported to be able to reverse established diabetes by acting on diabetogenic effector cells [31,32] or by harboring a dominant downregulatory mechanism [33]. In our studies, TAB4 functions through killing the disease-causing effector cells rather than through inducing a global immunosuppression.…”

Adoptive cell therapy using antigen-specific CD4−CD8− T regulatory cells to prevent autoimmune diabetes and promote islet allograft survival in NOD mice

“…However, transplantation of islet allografts into spontaneously diabetic NOD hosts provides an even more daunting challenge because of the generalised defect in tolerance mechanisms and the presence of pre-activated cytopathic islet-specific autoimmune T cells [24,25]. Rapamycin is a macrolide antibiotic with potent immunosuppressive and immunoregulatory properties.…”