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A novel clinically relevant strategy to abrogate autoimmunity and regulate alloimmunity in NOD mice.

Vergani A, D'Addio F, Jurewicz M, Petrelli A, Watanabe T, Liu K, Law K, Schuetz C, Carvello M, Orsenigo E, Deng S, Rodig SJ, Ansari JM, Staudacher C, Abdi R, Williams J, Markmann J, Atkinson M, Sayegh MH, Fiorina P - Diabetes (2010)

Bottom Line: Using allogeneic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, we addressed the therapeutic efficacy and immunomodulatory mechanisms associated with a new immunoregulatory protocol based on prolonged low-dose mATG plus CTLA4-Ig.Immunologic analysis of mice receiving transplants revealed a complete abrogation of autoimmune responses and severe downregulation of alloimmunity in response to treatment.The capacity to regulate alloimmunity and to abrogate the autoimmune response in NOD mice in different settings confirmed that prolonged mATG+CTLA4-Ig treatment is a clinically relevant strategy to translate to humans with type 1 diabetes.

View Article: PubMed Central - PubMed

Affiliation: Transplantation Research Center, Children's Hospital and Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA. paolo.fiorina@childrens.harvard.edu

ABSTRACT

Objective: To investigate a new clinically relevant immunoregulatory strategy based on treatment with murine Thymoglobulin mATG Genzyme and CTLA4-Ig in NOD mice to prevent allo- and autoimmune activation using a stringent model of islet transplantation and diabetes reversal.

Research design and methods: Using allogeneic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, we addressed the therapeutic efficacy and immunomodulatory mechanisms associated with a new immunoregulatory protocol based on prolonged low-dose mATG plus CTLA4-Ig.

Results: BALB/c islets transplanted into hyperglycemic NOD mice under prolonged mATG+CTLA4-Ig treatment showed a pronounced delay in allograft rejection compared with untreated mice (mean survival time: 54 vs. 8 days, P < 0.0001). Immunologic analysis of mice receiving transplants revealed a complete abrogation of autoimmune responses and severe downregulation of alloimmunity in response to treatment. The striking effect on autoimmunity was confirmed by 100% diabetes reversal in newly hyperglycemic NOD mice and 100% indefinite survival of syngeneic islet transplantation (NOD.SCID into NOD mice).

Conclusions: The capacity to regulate alloimmunity and to abrogate the autoimmune response in NOD mice in different settings confirmed that prolonged mATG+CTLA4-Ig treatment is a clinically relevant strategy to translate to humans with type 1 diabetes.

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Diabetic mice were treated with prolonged mATG+CTLA4-Ig the second day of glycemia >250 mg/dl. In 10 of 10 mice, diabetes was reversed, and 7 of 10 reverted in the first week. Although some fluctuation was observed, a stable reversal of diabetes was observed in treated mice (A). Untreated mice or CTLA4-Ig-treated mice never displayed reversal of hyperglycemia (B and C); 3 of 6 mice stably reverted in the prolonged mATG group (D), whereas 8 of 12 reverted in the induction mATG+CTLA4-Ig group. Histologic analysis showed a clear protection of islets in treated mice, with an almost complete clearance of the lymphocyte infiltrate, which was confined to the islet periphery. Notably, insulin staining was well represented (F1, H&E; F2, insulin; F3, glucagon; F4, B220; F5, CD3; F6, FoxP3). Insulitis score confirmed the reversal of the infiltrate by the treatment (G). IFN-γ production by splenocytes was tested in response to BDC2.5 peptide and to IGRP (206–214) peptide in reverted mice at day 60 after treatment; prolonged mATG+CTLA4-Ig completely suppressed the autoimmune response (NS vs. normo; P < 0.05 vs. hyper, n = 3) (H). The proportion of Tregs was increased in treated mice (P < 0.05 vs. normo or hyper, n = 3) (spleen: I; PLN: L), whereas the Teff proportion was reduced (P < 0.05 vs. normo or hyper, n = 3) (spleen: M; PLN: N); quantification of autoreactive T cells by tetramer staining confirmed the reduction of autoimmunity in prolonged mATG+CTLA4-Ig treated mice (P < 0.05 vs. hyper, n = 3) (O). *P < 0.05. (A high-quality digital representation of this figure is available in the online issue.)
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Figure 7: Diabetic mice were treated with prolonged mATG+CTLA4-Ig the second day of glycemia >250 mg/dl. In 10 of 10 mice, diabetes was reversed, and 7 of 10 reverted in the first week. Although some fluctuation was observed, a stable reversal of diabetes was observed in treated mice (A). Untreated mice or CTLA4-Ig-treated mice never displayed reversal of hyperglycemia (B and C); 3 of 6 mice stably reverted in the prolonged mATG group (D), whereas 8 of 12 reverted in the induction mATG+CTLA4-Ig group. Histologic analysis showed a clear protection of islets in treated mice, with an almost complete clearance of the lymphocyte infiltrate, which was confined to the islet periphery. Notably, insulin staining was well represented (F1, H&E; F2, insulin; F3, glucagon; F4, B220; F5, CD3; F6, FoxP3). Insulitis score confirmed the reversal of the infiltrate by the treatment (G). IFN-γ production by splenocytes was tested in response to BDC2.5 peptide and to IGRP (206–214) peptide in reverted mice at day 60 after treatment; prolonged mATG+CTLA4-Ig completely suppressed the autoimmune response (NS vs. normo; P < 0.05 vs. hyper, n = 3) (H). The proportion of Tregs was increased in treated mice (P < 0.05 vs. normo or hyper, n = 3) (spleen: I; PLN: L), whereas the Teff proportion was reduced (P < 0.05 vs. normo or hyper, n = 3) (spleen: M; PLN: N); quantification of autoreactive T cells by tetramer staining confirmed the reduction of autoimmunity in prolonged mATG+CTLA4-Ig treated mice (P < 0.05 vs. hyper, n = 3) (O). *P < 0.05. (A high-quality digital representation of this figure is available in the online issue.)

Mentions: Considering the effectiveness of prolonged mATG+CTLA4-Ig in abrogating the autoimmune response in a stringent model of allogeneic and syngeneic islet transplantation, we tested prolonged mATG+CTLA4-Ig for its effect in reversing established diabetes. Hyperglycemic NOD mice were treated after 2 consecutive days of glucose measurements >240 mg/dl with prolonged mATG+CTLA4-Ig. Notably, normoglycemia was restored in 100% of treated mice (10 of 10 mice) (Fig. 7A). Reversal was, for the most part, achieved rapidly: 7 mice reverted within the first week after the initiation of treatment; the other 3 mice reverted in 2, 3, and 6 weeks, respectively. Mice that reverted at later time points generally exhibited higher glucose levels at the initiation of treatment. Normoglycemia was maintained for 60 days of follow-up (Fig. 7A). Mice in the control group and the group treated with CTLA4-Ig alone did not show any sign of restoration of normoglycemia (Fig. 7B and C). On the contrary, 3 of 6 in the group treated with prolonged mATG alone (Fig. 7D) and 8 of 12 in the induction mATG+CTLA4-Ig group stably reverted from hyperglycemia (Fig. 7E).


A novel clinically relevant strategy to abrogate autoimmunity and regulate alloimmunity in NOD mice.

Vergani A, D'Addio F, Jurewicz M, Petrelli A, Watanabe T, Liu K, Law K, Schuetz C, Carvello M, Orsenigo E, Deng S, Rodig SJ, Ansari JM, Staudacher C, Abdi R, Williams J, Markmann J, Atkinson M, Sayegh MH, Fiorina P - Diabetes (2010)

Diabetic mice were treated with prolonged mATG+CTLA4-Ig the second day of glycemia >250 mg/dl. In 10 of 10 mice, diabetes was reversed, and 7 of 10 reverted in the first week. Although some fluctuation was observed, a stable reversal of diabetes was observed in treated mice (A). Untreated mice or CTLA4-Ig-treated mice never displayed reversal of hyperglycemia (B and C); 3 of 6 mice stably reverted in the prolonged mATG group (D), whereas 8 of 12 reverted in the induction mATG+CTLA4-Ig group. Histologic analysis showed a clear protection of islets in treated mice, with an almost complete clearance of the lymphocyte infiltrate, which was confined to the islet periphery. Notably, insulin staining was well represented (F1, H&E; F2, insulin; F3, glucagon; F4, B220; F5, CD3; F6, FoxP3). Insulitis score confirmed the reversal of the infiltrate by the treatment (G). IFN-γ production by splenocytes was tested in response to BDC2.5 peptide and to IGRP (206–214) peptide in reverted mice at day 60 after treatment; prolonged mATG+CTLA4-Ig completely suppressed the autoimmune response (NS vs. normo; P < 0.05 vs. hyper, n = 3) (H). The proportion of Tregs was increased in treated mice (P < 0.05 vs. normo or hyper, n = 3) (spleen: I; PLN: L), whereas the Teff proportion was reduced (P < 0.05 vs. normo or hyper, n = 3) (spleen: M; PLN: N); quantification of autoreactive T cells by tetramer staining confirmed the reduction of autoimmunity in prolonged mATG+CTLA4-Ig treated mice (P < 0.05 vs. hyper, n = 3) (O). *P < 0.05. (A high-quality digital representation of this figure is available in the online issue.)
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Figure 7: Diabetic mice were treated with prolonged mATG+CTLA4-Ig the second day of glycemia >250 mg/dl. In 10 of 10 mice, diabetes was reversed, and 7 of 10 reverted in the first week. Although some fluctuation was observed, a stable reversal of diabetes was observed in treated mice (A). Untreated mice or CTLA4-Ig-treated mice never displayed reversal of hyperglycemia (B and C); 3 of 6 mice stably reverted in the prolonged mATG group (D), whereas 8 of 12 reverted in the induction mATG+CTLA4-Ig group. Histologic analysis showed a clear protection of islets in treated mice, with an almost complete clearance of the lymphocyte infiltrate, which was confined to the islet periphery. Notably, insulin staining was well represented (F1, H&E; F2, insulin; F3, glucagon; F4, B220; F5, CD3; F6, FoxP3). Insulitis score confirmed the reversal of the infiltrate by the treatment (G). IFN-γ production by splenocytes was tested in response to BDC2.5 peptide and to IGRP (206–214) peptide in reverted mice at day 60 after treatment; prolonged mATG+CTLA4-Ig completely suppressed the autoimmune response (NS vs. normo; P < 0.05 vs. hyper, n = 3) (H). The proportion of Tregs was increased in treated mice (P < 0.05 vs. normo or hyper, n = 3) (spleen: I; PLN: L), whereas the Teff proportion was reduced (P < 0.05 vs. normo or hyper, n = 3) (spleen: M; PLN: N); quantification of autoreactive T cells by tetramer staining confirmed the reduction of autoimmunity in prolonged mATG+CTLA4-Ig treated mice (P < 0.05 vs. hyper, n = 3) (O). *P < 0.05. (A high-quality digital representation of this figure is available in the online issue.)
Mentions: Considering the effectiveness of prolonged mATG+CTLA4-Ig in abrogating the autoimmune response in a stringent model of allogeneic and syngeneic islet transplantation, we tested prolonged mATG+CTLA4-Ig for its effect in reversing established diabetes. Hyperglycemic NOD mice were treated after 2 consecutive days of glucose measurements >240 mg/dl with prolonged mATG+CTLA4-Ig. Notably, normoglycemia was restored in 100% of treated mice (10 of 10 mice) (Fig. 7A). Reversal was, for the most part, achieved rapidly: 7 mice reverted within the first week after the initiation of treatment; the other 3 mice reverted in 2, 3, and 6 weeks, respectively. Mice that reverted at later time points generally exhibited higher glucose levels at the initiation of treatment. Normoglycemia was maintained for 60 days of follow-up (Fig. 7A). Mice in the control group and the group treated with CTLA4-Ig alone did not show any sign of restoration of normoglycemia (Fig. 7B and C). On the contrary, 3 of 6 in the group treated with prolonged mATG alone (Fig. 7D) and 8 of 12 in the induction mATG+CTLA4-Ig group stably reverted from hyperglycemia (Fig. 7E).

Bottom Line: Using allogeneic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, we addressed the therapeutic efficacy and immunomodulatory mechanisms associated with a new immunoregulatory protocol based on prolonged low-dose mATG plus CTLA4-Ig.Immunologic analysis of mice receiving transplants revealed a complete abrogation of autoimmune responses and severe downregulation of alloimmunity in response to treatment.The capacity to regulate alloimmunity and to abrogate the autoimmune response in NOD mice in different settings confirmed that prolonged mATG+CTLA4-Ig treatment is a clinically relevant strategy to translate to humans with type 1 diabetes.

View Article: PubMed Central - PubMed

Affiliation: Transplantation Research Center, Children's Hospital and Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA. paolo.fiorina@childrens.harvard.edu

ABSTRACT

Objective: To investigate a new clinically relevant immunoregulatory strategy based on treatment with murine Thymoglobulin mATG Genzyme and CTLA4-Ig in NOD mice to prevent allo- and autoimmune activation using a stringent model of islet transplantation and diabetes reversal.

Research design and methods: Using allogeneic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, we addressed the therapeutic efficacy and immunomodulatory mechanisms associated with a new immunoregulatory protocol based on prolonged low-dose mATG plus CTLA4-Ig.

Results: BALB/c islets transplanted into hyperglycemic NOD mice under prolonged mATG+CTLA4-Ig treatment showed a pronounced delay in allograft rejection compared with untreated mice (mean survival time: 54 vs. 8 days, P < 0.0001). Immunologic analysis of mice receiving transplants revealed a complete abrogation of autoimmune responses and severe downregulation of alloimmunity in response to treatment. The striking effect on autoimmunity was confirmed by 100% diabetes reversal in newly hyperglycemic NOD mice and 100% indefinite survival of syngeneic islet transplantation (NOD.SCID into NOD mice).

Conclusions: The capacity to regulate alloimmunity and to abrogate the autoimmune response in NOD mice in different settings confirmed that prolonged mATG+CTLA4-Ig treatment is a clinically relevant strategy to translate to humans with type 1 diabetes.

Show MeSH
Related in: MedlinePlus