Esther Tamayo

CD4+CD25+ T Cell-Dependent Inhibition of Autoimmunity in Transgenic Mice Overexpressing Human Bcl-2 in T Lymphocytes

Regulation of lymphocyte survival is essential for the maintenance of lymphoid homeostasis preventing the development of autoimmune diseases. Recently, we described a systemic lupus erythematosus associated with an IgA nephropathy in autoimmune-prone (NZW × C57BL/6)F1 overexpressing human Bcl-2 (hBcl-2) in B cells (transgenic (Tg) 1). In the present study, we analyze in detail a second line of hBcl-2 Tg mice overexpressing the transgene in all B cells and in a fraction of CD4+ and CD8+ T cells (Tg2). We demonstrate here that the overexpression of hBcl-2 in T cells observed in Tg2 mice is associated with a resistance to the development of lupus disease and collagen type II-induced arthritis in both (NZW × C57BL/6)F1 and (DBA/1 × C57BL/6)F1 Tg2 mice, respectively. The disease-protective effect observed in autoimmune-prone Tg2 mice is accompanied by an increase of peripheral CD4+CD25+ hBcl-2+ regulatory T cells (Tregs), expressing glucocorticoid-induced TNFR, CTLA-4, and FoxP3. Furthermore, the in vivo depletion of CD4+CD25+ Tregs in (DBA/1 × C57BL/6)F1 Tg2 mice promotes the development of a severe collagen type II-induced arthritis. Taken together, our results indicate that the overexpression of hBcl-2 in CD4+ T cells alters the homeostatic mechanisms controlling the number of CD4+CD25+ Tregs resulting in the inhibition of autoimmune diseases.

Involvement of the intrinsic and extrinsic cell-death pathways in the induction of apoptosis of mature lymphocytes by the Escherichia coli heat-labile enterotoxin

Escherichia coli heat‐labile enterotoxin (LT) exhibits a broad range of immunomodulatory activities, including the induction of lymphocyte‐programmed cell death. In previous studies, we have demonstrated that in vivo LT promotes apoptosis of immature T and B cells through the stimulation of endogenous glucocorticoids. In the present study, we show that the extrinsic cell‐death pathway as well as the apoptosis‐inducing factor do not participate in the LT‐induced elimination of thymocytes. In contrast to developing lymphocytes, LT promotes the death of mature lymphocytes by both glucocorticoid‐ and Fas death receptor/Fas ligand‐dependent mechanisms. However, the dependency of these mechanisms in the LT‐induced cell‐death activity seems to be different among CD4+ and CD8+ T cells. Altogether, our study shows that the same bacterial toxin can induce apoptosis of lymphoid cells through several mechanisms depending on the status of differentiation of these cells.

The Escherichia coli heat-labile enterotoxin induces apoptosis of immature lymphocytes in vivo via a glucocorticoid-dependent pathway

Escherichia coli heat‐labile enterotoxin (LT) exhibits a broad range of immunomodulatory activities, including the induction of lymphocyte‐programmed cell death. However, the nature of the lymphoid populations sensitive to LT‐induced apoptosis and the mechanisms used by this toxin to promote such activity are still unclear. In this study, we demonstrate that LT induces in mice a rapid increase in the levels of circulating corticosterone, resulting in a dramatic induction of cell death of immature CD4+CD8+, B220+IgM– and IgM+IgD– T and B cell progenitors, respectively. Apoptosis of these cell populations is similar to that reported after experimental treatment with corticosteroids, it is inhibited by mifepristone, a glucocorticoid receptor antagonist, and does not occur in adrenalectomized animals. These results clearly indicate that endogenous glucocorticoids are the mediators of the LT‐induced cell death, which involves Bcl‐2‐dependent apoptotic pathways. The LT‐mediated programmed cell death requires systemic exposure and the enzymatic activity of LT, since a mutant devoid of any enzymatic activity have no pro‐apoptotic effect at any dose tested.

p27Kip1 inhibits systemic autoimmunity through the control of Treg cell activity and differentiation

OBJECTIVE Despite the importance of Treg cells in the maintenance of immunologic tolerance, the mechanisms that control their generation and activity are unknown. Since the cell cycle inhibitor p27(Kip1) (p27) was involved in T cell anergy, we undertook this study to explore its role in both Treg cell processes. METHODS The development of type II collagen-induced arthritis (CIA) and lupus-like abnormalities was compared between transgenic mice overexpressing human Bcl-2 in T cells (BCL2-TgT mice) and nontransgenic mice that were deficient or not deficient in p27. The contribution of Treg cells to disease evolution was also explored. Finally, the in vitro activity of Treg cells and their differentiation from naive CD4+ cells was compared between these strains of mice. RESULTS BCL2-TgT mice were protected against CIA by a Treg cell-dependent mechanism. In association with this protection, the overexpression of Bcl-2 in T cells enhanced the differentiation and activity of Treg cells. Both Bcl-2 effects were independent of its antiapoptotic activity but dependent on its capacity to induce the expression of p27 that augmented the strength of transforming growth factor β (TGFβ) signaling in T cells. Accordingly, down-modulation of p27 expression in BCL2-TgT mice promoted CIA. In addition, p27 deficiency in aged C57BL/6 mice reduced the number and activity of Treg cells and induced the development of mild lupus-like abnormalities. CONCLUSION Our results point to p27 as a critical regulator of Treg cell differentiation and function through the positive modulation of TGFβ signaling strength in T cells.

GITR contributes to the systemic adjuvanticity of the Escherichia coli heat-labile enterotoxin

The Escherichia coli heat‐labile enterotoxin (LT) possesses a powerful mucosal and systemic adjuvant effect. However, little is known about the cellular and molecular basis of the immunostimulatory activity of LT at the mucosal level, and even less information is available on the mechanisms underlying its systemic adjuvant activity. In this study, we show that distinct mechanisms are responsible for the parenteral and mucosal adjuvanticity of LT. Indeed, the systemic administration of LT upregulates the expression of glucocorticoid‐induced TNFR‐related protein (GITR), but not other activation markers, in naive T cells. Using WT and GITR‐deficient mice and LT and its enzymatically inactive mutant LTK63 as adjuvants, we show that the induction of GITR expression in T cells accounts for the systemic immunostimulatory capacity of LT, which requires an intact enzymatic activity. In contrast, the mucosal administration of LT does not induce GITR expression on Peyers patche T cells and accordingly no differences are observed in the mucosal adjuvanticity of LT between WT and GITR‐deficient mice. Altogether, our results demonstrate the distinct effect of LT after parenteral administration when compared with the mucosal delivery, and describe a new mechanism of LT adjuvanticity related to its ability to induce the expression of GITR in CD4+ T cells.

Modulation of autoimmune arthritis severity in mice by apolipoprotein E (ApoE) and cholesterol.

Apolipoprotein E (ApoE) deficiency promoted an exacerbation of autoimmune arthritis in mice by inducing proinflammatory immune responses. In this study we analysed the contribution of hypercholesterolaemia and/or the absence of ApoE anti‐inflammatory properties, unrelated to its function in the control of cholesterol metabolism, towards the acceleration of arthritis in these mutant animals. The induction and severity of collagen type II‐induced arthritis (CIA) were compared for B10.RIII wild‐type (WT), B10.RIII.ApoE+/–, B10.RIII.ApoE–/– and B10.RIII.low‐density lipoprotein receptor (LDLR–/–) mice with different concentrations of circulating ApoE and cholesterol. A 50–70% reduction in serum levels of ApoE was observed in heterozygous B10.RIII.ApoE+/– mice in comparison to B10.RIII.WT, although both strains of mice exhibited similar circulating lipid profiles. This ApoE reduction was associated with an increased CIA severity that remained lower than in homozygous B10.RIII.ApoE–/– mice. An important rise in circulating ApoE concentration was observed in hypercholesterolaemic B10.RIII.LDLR–/– mice fed with a normal chow diet, and both parameters increased further with an atherogenic hypercholesterolaemic diet. However, the severity of CIA in B10.RIII.LDLR–/– mice was similar to that of B10.RIII.WT controls. In conclusion, by comparing the evolution of CIA between several strains of mutant mice with different levels of serum ApoE and cholesterol, our results demonstrate that both hypercholesterolaemia and ApoE regulate the intensity of in‐vivo systemic autoimmune responses.