Partial blockade of T-cell differentiation during ontogeny and marked alterations of the thymic microenvironment in transgenic mice with impaired glucocorticoid receptor function (original) (raw)
Related papers
Immunology Letters, 2003
Thymocyte maturation in the thymus is controlled by stromal and humoral components. Among the humoral regulators locally produced glucocorticoids (GCs) seem to have a key role in the positive selection of thymocytes. Our previous studies have shown that the administration of GCs or the stimulation through the CD3 complex can induce apoptosis of double positive (DP) cells, but the combined presence of these stimuli induces positive selection. In this work our aim was to investigate the effects of antigen exposure and synthetic GC hormone (dexamethasone, DX) administration on the selection processes of DP cells in TcR transgenic mice. In our model, AND-pigeon cytochrome c (PCC)-specific I-E k (MHC-II) restricted V3, V␣11 TcR expressing transgenic mice were treated with PCC, with high or low dose DX, or with PCC and DX together, followed by the analysis of total thymocyte numbers, thymocyte composition, with regard to their CD69, V3 and Annexin V expression. The administration of PCC and/or DX for 2 days resulted in a decreased DP cell number and a significantly increased CD4 SP cell ratio. However, in both cases the total thymocyte numbers decreased. CD69 expression increased on both DP and CD4 SP cells after PCC and/or DX treatments. We found that after DX or combined treatment, the percentage of Annexin V positive cells increased. The ratio of V3 TcR bearing DP thymocytes showed no change after DX or PCC administrations alone, but it decreased significantly after combined treatment. MHC-II bound PCC peptides in the presence of GCs enhanced the maturation of V3+ DP cells into CD4 SP stage, therefore, the V3− cells remained mostly in the DP immature stage. These data indicate that both antigen and low dose GC alone are capable of inducing positive selection of DP cells, but together they gave a stronger effect in promoting positive selection. From these we conclude that GCs influence the maturation and selection processes of thymocytes.
Glucocorticoid receptor deficient thymic and peripheral T cells develop normally in adult mice
European Journal of Immunology, 2002
The involvement of glucocorticoid receptor (GR) signaling in T cell development is highly controversial, with several studies for and against. We have previously demonstrated that GR -/mice, which usually die at birth because of impaired lung development, exhibit normal T cell development, at least in embryonic mice and in fetal thymus organ cultures. To directly investigate the role of GR signaling in adult T cell development, we analyzed the few GR -/mice that occasionally survive birth, and irradiated mice reconstituted with GR -/fetal liver precursors. All thymic and peripheral T cells, as well as other leukocyte lineages, developed and were maintained at normal levels. Anti-CD3-induced cell death of thymocytes in vitro, T cell repertoire heterogeneity and T cell proliferation in response to anti-CD3 stimulation were normal in the absence of GR signaling. Finally, we show that metyrapone, an inhibitor of glucocorticoid synthesis (commonly used to demonstrate a role for glucocorticoids in T cell development), impaired thymocyte development regardless of GR genotype indicating that this reagent inhibits thymocyte development in a glucocorticoid-independent fashion. These data demonstrate that GR signaling is not required for either normal T cell development or peripheral maintenance in embryonic or adult mice.
Endocrinology, 2005
We and others have previously reported that thymic epithelial cells produce glucocorticoids (GCs). In vitro studies have also suggested that thymic-derived GCs play a role in the development of thymocytes. However, until now it has not yet been established whether thymic-derived GCs play a role in thymopoiesis in vivo. To investigate this, we conditionally overexpressed the GC receptor (GR) in thymocytes using transgenic mice with a tetracycline-inducible expression system. The influence of systemic GCs was excluded by adrenalectomizing the transgenic mice before the GR induction. Conditional expression of transgenic GR in the thymocytes of adre-nalectomized transgenic mice led to a decrease in the thymocyte number. This was associated with increased thymocyte apoptosis. The effect of thymic-derived GCs on the thymocytes was confirmed after transgenic GR induction in a thymic organ culture system. Finally, the GR antagonist RU486 increased thymocyte number in adrenalectomized mice in vivo and prevented a reduction in thymocyte number in thymic organ culture after transgenic GR induction. These observations further confirmed a role for the thymic-derived GCs in regulating thymocyte homeostasis in vivo. (Endocrinology 146: 2501-2507, 2005)
Glucocorticoid Resistance in Thymocytes from Mice Expressing a T Cell Receptor Transgene
Biochemical and Biophysical Research Communications, 2000
A majority of thymocytes undergo apoptosis during differentiation due to lack of survival signals provided by T cell receptor (TCR) activation. As glucocorticoids (GC) have been suggested to be involved in this process, we have investigated the GC sensitivity in thymocytes from mice expressing a transgenic selecting TCR. We now report that immature CD4(+)CD8(+) double-positive thymocytes from these mice are comparatively more resistant to corticosterone-induced apoptosis. This is associated with reduced glucocorticoid receptor (GR) expression, increased levels of membrane CD28, increased NF-kappaB DNA binding activity, and increased binding to the CD28 response element in the interleukin-2 gene promoter. Analysis of NF-kappaB/Rel proteins from nuclear extracts demonstrated altered levels of some of these proteins. Our results suggest that TCR recognition of self major histocompatibility antigens generates intracellular signals which alter the thymocyte GC sensitivity and thereby protect them against apoptosis induced by endogenous GC.
Role of Glucocorticoids in Early T-Cell Differentiation
Annals of The New York Academy of Sciences, 2000
Abstract: The results of the t-cell differentiation in the progeny of adrenalectomized pregnant rats (adx fetuses), an experimental model that ensures the absence of glucocorticoids (GCs) during the first stages of development, are summarized. in Adx thymuses there is an accelerated maturation of thymocytes that is reversed by in vivo GC replacement. in addition, Adx thymuses show decreased cell content, which correlates with both the increased numbers of apoptotic cells and an early migration of DP (CD4+CD8+) and SP (both CD4+CD8− and CD4−CD8+) thymocytes to the spleen. As shown by in vitro recolonization assays, accelerated T-cell differentiation is a consequence of changes in the biology of lymphoid precursors occurring in the fetal liver of Adx fetuses. They arrive at the thymic primordium earlier and mature faster than the fetal liver lymphoid progenitors from Sham control fetuses. After the establishment of a fetal hypothalamus-pituitary-gland-adrenal-gland (HPA) axis, there is a gradual normalization of the T-cell development Adx fetuses.
Glucocorticoid (GC) sensitivity and GC receptor expression differ in thymocyte subpopulations
International Immunology, 2002
Positive and negative selection steps in the thymus prevent non-functional or harmful T cells from reaching the periphery. To examine the role of glucocorticoid (GC) hormone and its intracellular receptor (GCR) in thymocyte development we measured the GCR expression in different thymocyte subpopulations of BALB/c mice with or without previous dexamethasone (DX), anti-CD3 mAb, RU-486 and RU-43044 treatment. Four-color labeling of thymocytes allowed detection of surface CD4/ CD8/CD69 expression in parallel with intracellular GCR molecules by¯ow cytometry. Doublepositive (DP) CD4 + CD8 + thymocytes showed the lowest GCR expression compared to doublenegative (DN) CD4 ± CD8 ± thymocytes and mature single-positive (SP) cells. DX treatment caused a concentration-dependent depletion of the DP cell population and increased appearance of mature SP cells with reduced GCR levels. GCR antagonists (RU-486 or RU-43044) did not in¯uence the effect of DX on thymocyte composition; however, RU-43044 inhibited the high-dose GC-induced GCR down-regulation in SP and DN cells. GCR antagonists alone did not in¯uence the maturation of thymocytes and receptor numbers. Combined low-dose anti-CD3 mAb and DX treatment caused an enhanced maturation (positive selection) of thymocytes followed by the elevation of CD69 + DP cells. The sensitivity of DP thymocytes with a GCR low phenotype to GC action and the ineffectiveness of the GCR antagonist treatment may re¯ect a non-genomic GC action in the thymic selection steps.
Immunity, 2000
Commercial Road to either activate or repress transcription of target Prahran, 3181, Victoria genes. The GR can also modify gene transcription via Australia direct protein-protein interactions with other transcrip- † Baker Medical Research Institute tion factors, such as AP1 and NF-B (Cato and Wade, Commercial Road 1996). Prahran, Victoria, 3181 The exact role of glucocorticoids in intrathymic T cell Australia development has recently been investigated by two different groups using transgenic mice bearing an antisense construct of the GR gene (King et al., 1995; Morale Summary et al., 1995; Sacedon et al., 1999) Although both groups reported significant changes in T cell development, the Glucocorticoids are believed to play a role in T cell results from one group contrasted with those from the development and selection, although their precise other. The reasons for these discrepancies are unclear. function is controversial. Glucocorticoid receptor (GR)-Glucocorticoids have also been proposed as a key factor in determining positive versus negative selection. deficient mice were used to directly investigate this Two recent studies have provided data showing that problem. GR-deficient thymocytes were resistant to inhibitors of steroidogenic enzymes increase thymocyte dexamethasone-mediated apoptosis, confirming the sensitivity to deletion (Vacchio and Ashwell, 1997; Vacabsence of glucocorticoid responsiveness. An absence chio et al., 1999). However, these findings have also of GR signaling had no impact on thymocyte developbeen contrasted by other reports showing decreased ment either in vivo or in vitro. T cell differentiation, thymocyte sensitivity to deletion after inhibition of GR including positive selection, was normal as assessed signaling (Jondal et al., 1993; Xue et al., 1996; Paziranby normal development of CD4 ؉ CD8 ؉ , ␣TCR ؉ CD4 ؉ , deh et al., 1999). Collectively, while these studies sugand ␣TCR ؉ CD8 ؉ thymocytes. Negative selection, gest a possible function for glucocorticoids in T cell mediated by the superantigen staphylococcal enterodevelopment, their precise role remains unclear.
Immunobiology, 2019
Functional disturbances in regulatory T cells (Treg) have been described in autoimmune diseases, and their potential therapeutic use is intensively studied. Our goal was to investigate the influence of glucocorticoid hormone on the in vitro differentiation of Treg cells from thymic and splenic CD4 + T cells under different conditions to establish methods for generating stable and functionally suppressive iTregs for future use in adoptive transfer experiments. Methods: Thymic and splenic CD4 + T lymphocytes were isolated from 3 to 4 week-old control and in vivo dexamethasone (DX) pretreated BALB/c mice using magnetic bead negative selection, followed by CD25 positive selection. The cells were cultured with anti-CD3/CD28 beads and IL-2 in the presence or absence of TGFβ and/or DX for 3-6 days. Multiparametric flow cytometry was performed using CD4, CD25, CD8, TGFβ (LAP) cell surface and Foxp3, IL-4, IL-10, IL-17 and IFNγ intracellular staining. Quantitative RT-PCR was performed to measure IL-10, TGFβ cytokine and Foxp3 mRNA levels. Results: Differentiation of thymus-derived CD4 + cells in vitro into iTreg cells was most effective (24-25%) when anti-CD3/CD28 beads, IL-2, and TGFβ were present. Splenic CD4 + T cell expansion under same conditions resulted in a higher (44-45%) iTreg cell ratio that further increased (up to 50% Treg) in the presence of DX. Elevated immunosuppressive cytokine (IL-10 and TGFβ) production by iTregs could be measured both at protein and mRNA levels without elevation of Th1/Th2 or Th17 cytokine production. We got the highest iTreg ratio (74%) and TGFβ production when CD4 + CD25 + splenic T cells were stimulated in the presence of TGFβ. In vivo 4 days DX pretreatment resulted in enhanced in vitro expansion and Foxp3 expression of thymus-derived iTregs and decreased differentiation of spleen-derived iTreg cells. In these Tregs the relative expression of IL-10 mRNA significantly decreased under all in vitro stimulation conditions, while TGFβ mRNA level did not change. Conclusion: DX promotes the expansion of thymic and splenic Treg cells, and enhances Foxp3 + expression and the production of immunosuppressive cytokines IL-10 and TGFβ in vitro. In vivo pretreatment of mice with DX inhibited the immunosuppressive cytokine production of in vitro differentiated Treg cells. We hypothesize that patients receiving GC therapy may need special attention prior to in vitro expansion and transplantation of Treg cells.
Histochemical and molecular overview of the thymus as site for T-cells development
Progress in Histochemistry and Cytochemistry, 2008
The thymus represents the primary site for T cell lymphopoiesis, providing a coordinated set for critical factors to induce and support lineage commitment, differentiation and survival of thymus-seeding cells. One irrefutable fact is that the presence of non-lymphoid cells through the thymic parenchyma serves to provide coordinated migration and differentiation of T lymphocytes. Moreover, the link between foetal development and normal anatomy has been stressed in this review. Regarding thymic embryology, its epithelium is derived from the embryonic endodermal layer, with possible contributions from the ectoderm. A series of differentiating steps is essential, each of which must be completed in order to provide the optimum environment for thymic development and function. The second part of this article is focused on thymic T-cell development and differentiation, which is a stepwise process, mediated by a variety of stromal cells in different regions of the organ. It depends strongly on the thymic microenvironment, a cellular network formed by epithelial cells, macrophages, dendritic cells and fibroblasts, that provide the combination of cellular interactions, cytokines and chemokines to induce thymocyte precursors for the generation of functional T cells. The mediators of this process are not well defined but it has been demonstrated that some interactions are under neuroendocrine control. Moreover, some studies pointed out that reciprocal signals from developing T cells also are essential for establishment and maintenance of the thymic microenvironment. Finally, we have also highlighted the heterogeneity of the lymphoid, non-lymphoid components and the multi-phasic steps of thymic differentiation. In conclusion, this review contributes to an understanding of the complex mechanisms in which the foetal and postnatal thymus is involved. This could be a prerequisite for developing new therapies specifically aimed to overcome immunological defects, linked or not-linked to aging.
Cell Immunol, 1992
This study examined the involvement of c-for protooncogene in thymocyte development from lymphohemopoietic T cell progenitors, within the thymic microenvironment. We first analyzed the thymocytes developing in vitro in the fetal thymus from the c-j& transgenic mice and found a high proportion of CD4+ single positive (SP) cells. We then seeded either fetal liver or hone marrow (BM) cells from normal donors onto lymphocyte-depleted fetal thymus explants of c-fos transgenic mice. The results showed an increased proportion of mature CD4+ SP and decreased CD4+CD8+ double positive (DP) cells. A similar pattern of CD4/CD8 thymocyte subsets was observed when either thymus or BM cells from c-j& transgenic mice developed within a normal thymic stroma. The kinetics of thymocyte development in organ culture (from Days 3 to 11) suggested that the SP cells obtained under these conditions may have bypassed the CD4+CD8+ DP phase. It appears that the altered pattern of thymocyte development manifested in adult cfis transgenic mice can be induced by the early embryonic thymic stroma, and may also involve cells in the lymphohemopoietic tissues. 0 1992 Academic PKSS, IX.