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D I Godfrey

Publications and source records attributed to D I Godfrey.

At least 19 recordsLinked to original sources
PubMed1998

alpha/beta-T cell receptor (TCR)+CD4-CD8- (NKT) thymocytes prevent insulin-dependent diabetes mellitus in nonobese diabetic (NOD)/Lt mice by the influence of interleukin (IL)-4 and/or IL-10.

We have previously shown that nonobese diabetic (NOD) mice are selectively deficient in alpha/beta-T cell receptor (TCR)+CD4-CD8- NKT cells, a defect that may contribute to their susceptibility to the spontaneous development of insulin-dependent diabetes mellitus (IDDM). The role of NKT cells in protection from IDDM in NOD mice was studied by the infusion of thymocyte subsets into young female NOD mice. A single intravenous injection of 10(6) CD4-/lowCD8- or CD4-CD8- thymocytes from female (BALB/c x NOD)F1 donors protected intact NOD mice from the spontaneous onset of clinical IDDM. Insulitis was still present in some recipient mice, although the cell infiltrates were principally periductal and periislet, rather than the intraislet pattern characteristic of insulitis in unmanipulated NOD mice. Protection was not associated with the induction of "allogenic tolerance" or systemic autoimmunity. Accelerated IDDM occurs after injection of splenocytes from NOD donors into irradiated adult NOD recipients. When alpha/beta-TCR+ and alpha/beta-TCR- subsets of CD4-CD8- thymocytes were transferred with diabetogenic splenocytes and compared for their ability to prevent the development of IDDM in irradiated adult recipients, only the alpha/beta-TCR+ population was protective, confirming that NKT cells were responsible for this activity. The protective effect in the induced model of IDDM was neutralized by anti-IL-4 and anti-IL-10 monoclonal antibodies in vivo, indicating a role for at least one of these cytokines in NKT cell-mediated protection. These results have significant implications for the pathogenesis and potential prevention of IDDM in humans.

Animals↗
PubMed1997

Recent thymic emigrants are distinct from most medullary thymocytes.

In the mouse thymus, newly formed single positive (SP) cells spend an average of 14 days in the thymic medulla. During this time, phenotypic and functional maturation occurs with down-regulation of CD69 and heat stable antigen (HSA), and up-regulation of Qa-2. Very little is known about the final steps that allow or direct these T cells to emigrate and join the recirculating peripheral T cell pool. Currently available data suggest that not all recent thymic emigrants (RTE) complete this maturational sequence in the medulla and that emigration may occur at any time during the medullary maturation stage. In this study, we have compared adhesion and activation marker expression on SP thymocytes, RTE and peripheral T cells to determine more precisely which SP medullary thymocytes are exported. Although RTE were heterogeneous for HSA and Qa-2 expression, they were quite uniform with regard to the expression of other molecules. In contrast to medullary SP thymocytes, most RTE were L-selectin(high) and CD69-. In addition, CD4+ CD8- and CD4- CD8+ RTE were phenotypically distinct from each other in that the former were beta7 integrin(-/low), CD45RB(intermediate) and CD45RC-, while the latter were beta7 integrin(high), CD45RB(high) and CD45RC(low). These phenotypes were comparable to only a minor (as little as 6%) subpopulation of medullary SP thymocytes. Overall, the data indicate that export of cells from the medullary pool of SP thymocytes is not random, but that a series of maturational events within the SP stage are necessary before export can occur.

Animals↗
PubMed1997

Thymic T cell export is not influenced by the peripheral T cell pool.

The peripheral T cell pool is maintained both by export of naive T cells from the thymus and by post-thymic expansion of activated/memory T cells. However, it is not known whether the thymus can alter its output following peripheral T cell depletion. Using intrathymic injection of fluorescein isothiocyanate to detect recent thymic emigrants (RTE), we directly tested whether the thymus is able to alter the number of RTE or the CD4:CD8 ratio of RTE emigrating to the periphery in response to in vivo depletion of total peripheral T cells or CD4 T cells, respectively. Depletion of peripheral T cells was achieved with anti-Thy-1 or anti-CD4, at doses that did not affect thymocyte numbers. Depletion of greater than 70% of peripheral T cells by treatment with anti-Thy-1 in vivo did not alter the number or cell cycle status of RTE trafficking to lymph nodes or spleen during the peripheral reconstitution phase (6, 9, 12 days). Similarly, depletion of the majority of CD4 T cells, which significantly reduced the peripheral CD4:CD8 T cell ratio, did not alter the total number or the proportion of CD4+ CD8- RTE in peripheral lymphoid organs. These data clearly indicate that thymic output is not influenced by downstream alterations in peripheral T cell pool size or CD4:CD8 ratio. Rather we contend that thymic T cell export is internally regulated by as yet undefined mechanisms.

Animals↗
PubMed1997

Flow cytometric study of T cell development in NOD mice reveals a deficiency in alphabetaTCR+CDR-CD8- thymocytes.

As a result of failed induction of T cell tolerance to pancreatic B cells, non-obese diabetic (NOD) mice develop spontaneous autoimmune insulin-dependent diabetes mellitus (IDDM). The thymic stroma, which plays a crucial role in thymic T cell maturation, undergoes extensive premature disorganization in NOD mice, so it is of interest to examine NOD T cell development. In this study, both major and minor developmental populations of thymocytes of NOD/Lt mice were studied and compared to those of BALB/c, C57BL/6 and CBA mice by multiparameter flow cytometry (FACS). These results are described in detail and reveal that most thymocyte subsets were normally represented, including alphaTcR-CD4-CD8- (triple negative; TN), alphabetaTcR-CD4+CD8- and alphabetaTcR-CD4-CD8+ (immature single positive; ISP), alphabetaTcR-/lowCD4+CD8+ (double positive; DP) and alphabetaTcR+CD4+CD8- and alphabetaTcR+CD4-CD8+ (mature single positive; SP) as well as gammadelta T cells. However, NOD mice exhibited a marked deficiency of thymic alphabetaTcR+CD4-CD8- (alphabeta+DN) T cells. alphabeta+DN T cells, which are included among NK1+ T cells in C57BL/6 mice, produce large amounts of IL-4 on primary stimulation. Given the potential significance of NKT cells in immunoregulation, it is possible that the scarcity of these cells in NOD mice plays a role in the pathogenesis of IDDM.

Animals↗
PubMed1997

Association between alphabetaTCR+CD4-CD8- T-cell deficiency and IDDM in NOD/Lt mice.

NOD mice develop spontaneous IDDM as a result of T-cell-mediated autoimmune destruction of pancreatic beta-cells. It is not known why these T-cells become autoreactive, nor is it clear whether the breakdown in self-tolerance reflects a general problem in T-cell development or a selective defect in an as yet undefined regulatory cell population. In this study, we showed that NOD mice, although relatively normal with regard to most thymocyte subsets, exhibit a marked deficiency in alphabetaTCR+CD4-CD8- (alphabeta+DN) T-cells in the thymus and, to a lesser extent, in the periphery. These T-cells have been termed NKT cells (NK1.1+-like T-cells) because they share some cell surface markers with conventional natural killer (NK) cells. To examine the role of these cells in the pathogenesis of IDDM, semiallogeneic or syngeneic double-negative (DN) thymocytes, enriched for NKT cells, were transferred into intact 4-week-old NOD recipients; the onset of diabetes was then monitored over the ensuing 30 weeks. Mice receiving NKT-enriched thymocytes did not develop diabetes, whereas mice receiving unfractionated thymocytes or phosphate-buffered saline developed diabetes at the normal rate. NKT cells represent a distinct T-cell lineage that has been shown to play a role in immunoregulation in vivo. The deficiency of these cells observed in NOD mice may therefore contribute to destruction of pancreatic islet cells by conventional T-cells.

Adoptive Transfer↗
PubMed1995

Thymic emigration: conveyor belts or lucky dips?

The thymic medulla has always seemed a rather uncomplicated compartment, simply storing mature thymocytes until they are exported to the peripheral lymphoid organs. However, as discussed here by Roland Scollay and Dale Godfrey, a careful look at recent data suggests that events in the medulla may be more complex and protracted than previously thought.

Animals↗
PubMed1995

CD38 expression on mouse T cells: CD38 defines functionally distinct subsets of alpha beta TCR+CD4-CD8- thymocytes.

We have examined CD38 expression on mouse lymphocytes using the rat mAb NIM-R5 and demonstrate that CD38 expression is restricted to approximately 8% of thymocytes. Although CD38 is absent from the majority of CD4+CD8- and CD4-CD8+ T cells, we detected a strong correlation between CD38 expression and alpha beta+CD4-CD8- T cells in the thymus, with nearly 80% of alpha beta TCR+CD4-CD8- thymocytes being CD38+. Using heat stable antigen (HSA) and CD38, we divided alpha beta+CD4-CD8- thymocytes into four subsets: HSA+CD38-, HSA-CD38hi, HSA-CD38low and HSA-CD38-. Two established characteristics of alpha beta TCR+CD4-CD8- cells, bias towards V beta 8.2 TCR expression and high levels of IL-4 production, were used to establish a possible relationship between the above thymocyte subsets. Our present data show that the HSA+CD38- subset is not biased towards V beta 8.2 TCR expression whereas the HSA-CD38- subset does show this bias (approximately 47%). Neither of these subsets make IL-4 upon CD3 mediated stimulation. In contrast, the CD38+ subsets are heavily biased toward V beta 8.2 expression and produce large amounts of IL-4 upon stimulation, particularly the CD38low cells. Taken together, these data suggest that these four subsets represent various stages of a possible differentiation pathway for alpha beta TCR+CD4-CD8- cells, with the HSA+CD38- subset being the most immature while the HSA-CD38low subset is the most functionally mature. These characteristics support the view that alpha beta TCR+CD4-CD8- T cells represent an independent lineage with a distinct, but as yet obscure, role in immunity.

ADP-ribosyl Cyclase↗
PubMed1994

Onset of TCR-beta gene rearrangement and role of TCR-beta expression during CD3-CD4-CD8- thymocyte differentiation.

TCR-beta gene rearrangement or expression is necessary and sufficient for the progression of early alpha beta thymocyte differentiation from the CD3-CD4-CD8- triple negative (TN)3 to the CD4+CD8+ double positive stage. The onset of TCR-beta rearrangement is currently thought to occur gradually. Some thymocytes were reported to be rearranged at the earliest (CD44+CD25-) TN stage, whereas other thymocytes did not initiate TCR-beta rearrangement until the latest (CD44-CD25-) TN stage. Here, we have isolated subsets of TN thymocytes on the basis of surface expression of CD44 and CD25, with c-kit as an additional marker. We present a revised model of early T cell development in which TCR-beta and TCR-gamma rearrangements occur abruptly, at the CD44lowCD25+ c-kitlowTN stage. A high level of c-kit expression defines pro-T cells which have not yet rearranged their TCR genes. Germ-line TCR-beta transcripts, and transcripts of recombination activating genes (RAG)-1 and 2, are detected before TCR-beta gene rearrangement. Analyses of TN thymocytes of RAG-1 mutant mice, and of various TCR mutant and TCR transgenic RAG-1 mutant mice, indicate the existence of a control point at the CD44-CD25+TN stage at which cells expressing a productively rearranged TCR-beta chain are selected for further differentiation.

Animals↗
PubMed1994

IL-12 influences intrathymic T cell development.

IL-12 has been implicated in the maturation and activation of peripheral T lymphocytes and NK cells. In the present study we have investigated the potential role of IL-12 in intrathymic T cell development. Treatment of mouse fetal thymic organ culture with IL-12 caused a significant reduction in size and cell number compared with the untreated controls. Flow cytometric analysis of the thymocytes recovered from these lobes showed differential effects on individual thymocyte subsets, most but not all of which were significantly decreased. In contrast, however, we observed an increase in both the percentage and cell number of alpha beta TCR+CD4-CD8+ thymocytes. This effect could be neutralized with an anti-IL-12 antibody, demonstrating the specificity of the influence of IL-12 in the fetal thymic organ culture system. IL-12 caused proliferation of isolated thymocyte subsets, particularly CD3+CD4-CD8+ cells in the presence of IL-2 and IL-4. Additionally, IL-12 induced significant proliferation of early CD3-CD4-CD8- triple negative CD44+CD25+ pro-T cells in combination with stem cell factor. We show that both the p35 and p40 chains of IL-12 are produced in the fetal and adult mouse thymus and that thymic stromal cells are a potential source of this cytokine.

Animals↗
PubMed1994

Monoclonal antibody 2D10 recognizes a novel T cell costimulatory molecule on activated murine B lymphocytes.

We have developed a panel of rat mAbs against dibutyryl cAMP-activated 5C2 cells. In this panel, one mAb, 1G10, recognized murine B7. Another mAb designated 2D10 did not bind to murine B7 but could recognize a surface molecule expressed only on dibutyryl cAMP-activated 5C2 mouse B lymphoma cells or on LPS-stimulated splenic B cells. This new molecule is referred to as early T cell costimulatory molecule-1 (ETC-1). From both activated 5C2 cells and splenic B cells, mAb 2D10 immunoprecipitated a 59- to 60-kDa protein, which was different from the 47- to 55-kDa murine B7 protein precipitated from the same cell populations. FACS analysis showed that, in contrast to B7, the expression of ETC-1 on 5C2 cells was induced by lower concentrations of dibutyryl cAMP and displayed a faster kinetics. LPS-stimulated splenic B cells expressed relatively low levels of B7 and much higher levels of ETC-1. Importantly, in an Ag presentation assay using activated 5C2 cells as APC, the secretion of IL-2 by C8A3 T hybrids was partially inhibited by mAb 2D10 alone and completely blocked by combination use of mAbs 2D10 and 1G10 in a dose-dependent and synergistic fashion. In a one-way primary MLR, mAb 2D10 alone at 0.1 to 1 microgram/ml inhibited T cell proliferation by 19 to 56%. However, an additive blocking effect (up to 76%) was observed when two mAbs were added in combination. Thus, our data have demonstrated that a novel T cell costimulatory molecule is present on activated murine B cells, which, in cooperation with B7, may play a critical role in optimal T cell activation.

Animals↗
PubMed1994

Thymic epithelial cell lines that mediate positive selection can also induce thymocyte clonal deletion.

Negative selection of potentially autoreactive thymocytes occurs mainly in the thymus and is thought to be induced primarily by interaction with bone marrow-derived cells. However, some studies have also reported a role for radioresistant thymic cells, which are probably epithelial in origin, in the deletion of thymocytes reacting to endogenous superantigens. We have previously demonstrated that thymic epithelial cell lines could induce thymocyte-positive selection in vivo. In this study, we assessed the potential of these cells to delete thymocytes reacting to the staphylococcal enterotoxin A or B superantigens in vitro. In the presence of staphylococcal enterotoxin A or B we found that all thymic epithelial cell lines used in this study were capable of activating T cell hybrids or deleting CD4+CD8+ thymocytes expressing an appropriate TCR. The extent of superantigen-mediated thymocyte deletion mediated by thymic epithelial cell lines was comparable to that mediated by a thymic macrophage cell line. Similar results were obtained with three phenotypically distinct thymic cell lines, suggesting that the ability to induce thymocyte deletion might be a general feature of various subsets of thymic epithelium. The observations provided in this study, combined with our previous demonstration that the same thymic epithelial cell lines can participate in positive selection, suggest that a given stromal cell population might be capable of taking part both in positive and negative selection of thymocytes.

Animals↗
PubMed1994

Expression and functional analysis of murine B7 delineated by a novel monoclonal antibody.

The B cell surface molecule designated B7 has been shown to be expressed by activated human B cells and monocytes and to be a ligand for the CD28 and CTLA-4 molecules on T cells. B7/CD28 interactions can provide a second signal to T cells (in addition to occupancy of the T cell antigen receptor) that is needed for T cell activation. COS cells transfected with the mouse homologue of B7 have been demonstrated to provide a stimulatory signal to murine and human T cells. In this report we describe a rat anti-mouse B7 mAb designated 1G10. Scatchard and/or FACS analyses utilizing 1G10 demonstrated that B7 was not expressed on resting splenic T cells or B cells, but could be induced at high levels on B cells cocultured with a syngeneic I-Ak-restricted autoreactive T cell hybridoma. Furthermore, activation of B cells with dibutyryl-cAMP (db-cAMP), a second messenger for class II MHC signaling, or with LPS induced the expression of B7 and the two agents showed additive effects. In contrast to B cells, freshly isolated mouse peritoneal macrophages constitutively expressed B7. Antibody-blocking experiments indicated that anti-B7 antibody partially inhibited T cell proliferative responses to primary antigenic stimulation but had no effect on the responses of previously activated T cells to antigenic restimulation.

Animals↗
PubMed1994

Flow cytometric analysis reveals unexpected shared antigens between histologically defined populations of thymic stromal cells.

Utilizing flow cytometry, the expression of antigens recognized by six thymic stromal cell (TSC) reactive mAbs was investigated on fresh TSCs and TSC lines. It was found that some thymic epithelial cells and dendritic cells share antigenic phenotypes, and that most TSC reactive mAbs have a more extensive distribution than would have been predicted from immunohistology. While these findings illustrate the higher sensitivity of flow cytometric analysis, they more importantly emphasize the great complexity of TSC that direct T cell development. In order to identify the molecular parameters that define the various steps involved in T cell differentiation, TSC antigens (non-TCR/MHC/co-receptor) that are functional will have to be identified. This study represents the initial steps in characterizing such antigens.

Animals↗
PubMed1993

Isolation of a cDNA encoding thymic shared antigen-1. A new member of the Ly6 family with a possible role in T cell development.

We have previously characterized a novel mouse thymocyte marker, defined as thymic shared Ag-1 (TSA-1), present on both immature thymocytes and a subset of thymic medullary epithelial cells. MTS 35, a mAb specific for TSA-1, alters T cell differentiation when added to fetal thymic organ cultures, suggesting TSA-1 may be important for T cell development in the thymus. In this study, we describe the isolation of a cDNA encoding TSA-1 using transient expression of COS-7 cells and selection with MTS 35. The predicted amino acid sequence of this cDNA encodes a 15 to 17-kDa protein and the expressed protein is linked to the membrane via a phosphatidylinositol moiety. TSA-1 is transcriptionally active at various levels in all organs examined, suggesting that its role is not solely intrathymic. TSA-1 shares amino acid sequence homology to the mouse Ly6 multigene family, epidermal growth factor-like receptors, and to cobra venom neurotoxin. The Tsa-1 locus is located on chromosome 15 linked to Ly6 on the mouse genome. We also examined the effects of MTS 35 in fetal thymic organ cultures repopulated with two subsets of thymocytes representing defined stages of T cell development. Our results suggest that TSA-1 may play a role during positive selection and the transition from CD4+CD8+ thymocytes to the mature CD4+CD8- and CD4-CD8+ subsets.

Amino Acid Sequence↗
PubMed1993

A lymphostromal molecule, thymic shared Ag-1, regulates early thymocyte development in fetal thymus organ culture.

Previously, we detailed the characterization of thymic shared Ag-1, a unique marker of immature thymocytes and isolated thymic stromal cells, defined by the mAb MTS 35. In this study, the functional relevance of this molecule to thymopoiesis was investigated by the addition of purified MTS 35 to fetal thymus organ culture. It down-regulated thymic shared Ag-1 expression and dramatically reduced thymocyte cell yield through inhibition of alpha beta-TcR+ T cell differentiation, post CD3-CD4-CD8- triple negative thymocytes. These effects were specific for the mAb MTS 35, because controls, which include both isotype-matched and other lymphostromal mAb, showed no similar effects. These results demonstrate that thymic shared Ag-1 is a functionally important marker of early thymocyte differentiation, particularly with regard to the alpha beta-TcR lineage.

Animals↗
PubMed1993

A developmental pathway involving four phenotypically and functionally distinct subsets of CD3-CD4-CD8- triple-negative adult mouse thymocytes defined by CD44 and CD25 expression.

We have subdivided mouse CD4-CD8-CD3- triple-negative (TN) thymocytes into four subsets based upon expression of CD44 and CD25, including CD44+CD25-, CD44+CD25+, CD44-CD25+ and CD44-CD25-. Characterization of these cells revealed several features distinct to each subset, in particular the expression of high levels of c-kit (the receptor for stem cell factor) by CD44+CD25-TN and CD44+CD25+TN but not by CD44-CD25+TN and CD44-CD25-TN. The CD44+CD25+TN subset also included the IL-7 and stem cell factor-responsive cells, whereas only minimal responsiveness was observed by the CD44- populations. These subsets also showed differential cytokine production potential (CD44+CD25- > CD44+CD25+ > CD44-CD25+ > CD44-CD25-) after stimulation with calcium ionophore, PMA and IL-1. The repopulation potential of these subsets in 2-deoxyguanosine-treated fetal thymic lobes supports the following maturation sequence: CD44+CD25- -->CD44+CD25+ -->CD44-CD25+ -->CD44-CD25-. Furthermore, the sequence of progression from CD44+CD25+ to CD44-CD25+ cells was confirmed by their TCR beta-chain gene configuration. The former population exhibits germ-line TCR beta-chain configuration, whereas the latter subset shows a rearranged pattern.

Animals↗
PubMed1993

Control points in early T-cell development.

Intrathymic T-cell differentiation involves the generation, expansion and selection of distinct T-lymphocyte subsets. While positive and negative selection have been a focal point of T-cell development, these events represent the final stages in a complicated sequence of differentiation steps. Here, Dale Godfrey and Albert Zlotnik summarize recent advances in our understanding of early T-cell development and describe five 'control points' that identify key events in this sequence.

Animals↗