Cytolytically Defective Tregs Can Prevent Spontaneous Autoimmune Disease and Gvhd, but Fail to Suppress Autochthonous Lymphoproliferation

Abstract

The role CD4+CD25+FoxP3+ (Treg) cells play in the establishment and maintenance of immune tolerance makes them attractive candidates for therapeutic prevention and treatment of autoimmune and transplant rejection responses. Autoimmune diseases have diverse pathologies/underlying effector mechanisms and how Treg cells control each disorder remains unclear. Granzyme B in murine and perforin in human Treg cells have been reported to be involved with suppression of proliferation and lysis of activated T cells, respectively. Perforin/fas-ligand double cytotoxic deficient (B6-cdd) mice develop a spontaneous lymphoproliferative disorder with selective tissue infiltration, principally in the pancreas and ovaries, together with weight loss and death by ~15 weeks of age (i.e. cdd syndrome). Uncontrolled expansion of the effector T cell pool is proposed to result as a consequence of cytotoxically impaired, activated T cells unable to lyse Ag-presenting monocytes and macrophages.Since CD4+CD25+FoxP3+ cells have the capacity to regulate lymphocyte proliferation, this compartment was examined in B6-cdd (H2b) mice. At 6–7 weeks of age-prior to complete onset of the syndrome-splenic CD4+CD25+ T cells are present and functionally suppressive in vitro. We were therefore interested in investigating their capacity to regulate auto and allo-reactive T cell responses in situ in the context of defined spontaneous autoimmune disease (IL2Rβ−/−)and GVHD (MHC Class I/II disparate) models. Purified CD4+CD25+ T cells isolated from B6-cdd mice were injected into B6-IL2Rβ−/− (CD122−/−) newborns and effectively prevented autoimmune disease assessed by hematocrit, absence of lymphoproliferation and clinical signs. B6-cdd Tregs were identified and collected several (~3) months post-injection from the rescued animals and found to exhibit functional suppression in vitro. Interestingly, identical results were obtained in allogeneic BALB/c (H2d) IL2Rβ−/− recipients. Purified CD4+CD25+ T cells isolated from B6-cdd or B6-wt mice were transplanted together with T cell replete B6 BMC into lethally irradiated, MHC-mismatched BALB/c recipients. B6-cdd Tregs were found to be comparable to B6-wt Tregs in their ability to prevent development of ac ute GVHD as assessed by weight loss, clinical signs and survival post-BMT.In total, these observations demonstrate that CD4+CD25+FoxP3+ Treg cells do not require the cytolytic effector molecules perforin and fas-ligand for in vitro suppression, in vivo regulation of a spontaneous (CD122−/−) autoimmune disease as well as for acute GVHD. Their inability to control autochthonous B6-cdd lymphoproliferative syndrome could ultimately be a result of death or dysregulation of Treg cells in these mice. Alternatively, distinct Treg effector molecules may be required to control different T cell responses associated with various autoimmune/non-self reactivity and ongoing experiments are investigating these possibilities.