1. ¹Department of Laboratory Medicine and Pathobiology, University of Toronto and St. Michael's Hospital, Toronto, ON, Canada M5B 1W8
2. ²Division of Immunogenetics, Department of Pediatrics, University of Pittsburgh School of Medicine, Rangos Research Center, Children's Hospital of Pittsburgh, Pittsburgh, PA 15213, USA

Correspondence: Yelena Glinka, Department of Laboratory Medicine, St. Michael's Hospital, 30 Bond Street, Room 2-013CC, Toronto, ON, Canada M5B 1W8. Fax: +1 416 864 5648. E-mail: glinkay@smh.toronto.on.ca.

Received 14 October 2005; Revised 16 March 2006; Accepted 24 March 2006.

Abstract

DNA vaccination of autoimmune diabetes-prone NOD mice with unmodified target islet antigens, i.e., preproinsulin (PPIns) or glutamic acid decarboxylase 65 (GAD65), is poorly protective. However, in this study, we demonstrate protection against disease by covaccination with a mutant B7-1 molecule (B7-1wa) that binds the negative T cell regulator CTLA-4 (CD152), but not CD28. Codelivery of plasmids encoding a PPIns–GAD65 fusion construct and B7-1wa protected against both insulitis and diabetes. In vitro, the T cells of covaccinated mice had negative responses to both insulin and GAD65, and this was restored by adding blocking antibodies to transforming growth factor 1 (TGF-1), suggesting a role for this cytokine. Adoptive transfer experiments revealed that DNA vaccination generated protective CD4⁺ regulatory T cells (Tr) of either CD25⁺ or CD25^- phenotype. Furthermore, vaccinated mice had increased numbers of T cells with Tr-associated markers, such as CTLA-4, Foxp3, and membrane-bound TGF-1. Tr cells inhibited the responses of diabetogenic T cells to islet antigens, and depletion of T cells expressing membrane-bound TGF-1 abolished the suppressive effect. Thus, selective engagement of CTLA-4 during islet-antigen DNA vaccination induces Tr cells that protect against this autoimmune disease.