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Biomedical subjects

R Scollay

Publications and source records attributed to R Scollay.

At least 37 records · Page 2Linked to original sources
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

Perturbed development of T and B cells in mice expressing an Hlx homeobox transgene.

Within the lymphoid compartment of mice, the Hlx homeobox gene is expressed only at early stages of B-lymphoid differentiation. To determine whether Hlx influences lymphopoiesis, transgenic mice were developed to enforce Hlx expression throughout the B and T cell lineages. The strain with the highest transgene expression in both cell types (Hlx-94) exhibited marked perturbations in both B and T cell development. In these mice, the thymus lacked almost all mature CD4+8- and CD8+4- cells and the medulla was greatly shrunken, whereas nearly one-half the T cells in the periphery were CD4+8+, a cell type normally confined to the thymus. The peripheral CD4+8+ cells had some features of mature T cells, including responsiveness to mitogens. Presumably these cells had emigrated prematurely from the thymus and generated mature T cells in the periphery. Bone marrow transplantation experiments indicated that the defects was intrinsic to the Hlx-94 hematopoietic cells rather than support cells. Although thymocyte development in Hlx-94 mice was blocked at the stage when selection normally occurs, analysis of lymphocyte populations in the progeny of crosses with mice transgenic for an anti-HY T cell receptor indicated that neither positive nor negative selection of T cells was markedly affected. In addition to T cell defects, Hlx-94 mice had subnormal numbers of B lymphoid cells in the bone marrow and spleen, and their surface phenotype suggested that B cell development after the pro-B stage was impeded. Furthermore, the B cell response to stimulation with LPS was impaired. These striking developmental defects suggest that the Hlx gene may help to govern lymphoid maturation.

Animals↗
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

The gamma delta T lymphocytes of the perinatal murine thymus.

We have previously shown that the adult thymus contains three subsets of gamma delta T cells that can be defined by the expression of THY-1 and heat-stable antigen (HSA). In this study, the number of cells in each of these thymic gamma delta populations was investigated at different stages throughout life. In adult mice, the populations stay relatively constant, however, in contrast, there were major variations in them early in development. It was shown that only two of the gamma delta populations were present in the prenatal thymus, a major population of Thy-1+ HSA - cells, and a smaller population of Thy-1+ HSA+ cells. However, after birth, most of the Thy-1+ HSA - cells appear to loose the Thy-1 antigen, forming the third population of HSA - Thy-1 - cells. The adult configuration of populations appeared to be established within the first week after birth. Therefore, whereas the gamma delta populations stayed relatively constant from this time point onwards, there were major variations early in development. Throughout life, most gamma delta thymocytes are CD4- CD8-, however, in the neonatal thymus, there are some CD+ and CD+ gamma delta thymocytes, and these are contained in the Thy-1+ HSA - population.

Animals↗
PubMed1994

Thymic stem cells in mouse bone marrow.

There is still controversy concerning the nature of the stem cells from bone marrow that colonize the thymus during embryogenesis and continually throughout life. To identify the bone marrow stem cells that home to and populate the thymus, we screened murine bone marrow cells for the presence of a population of surface phenotype similar to the earliest known intrathymic precursor. We have identified a population characterized by expression of an intermediate level of heat stable antigen, a very low level of Thy-1, and high levels of CD44 and class I major histocompatibility complex antigens. It is negative for B-cell, granulocyte, macrophage, and erythrocyte markers (B220, Gr-1, Mac-1, and TER 119). All these markers are common to the intrathymic precursors and bone marrow stem cells. However, this new bone marrow population is Sca-2+, similar to the intrathymic precursor, which makes a clear distinction from the Sca-2- bone marrow hematopoietic stem cells previously characterized. The frequency of the new population in the normal mouse bone marrow is about 0.25%. When transferred intrathymically or intravenously to lethally irradiated mice, it has a higher expansion potential (2 x 10(5)) than has been found for the intrathymic precursors (10(3)), but less than was found for the Sca-2- multipotent stem cell (10(7)). These transfer studies also showed that it was pluripotent, in that its precursor activity was not restricted to the production of T or B lymphocytes. However, it gave a reduced spleen colony number and smaller colonies (day-12 colony-forming unit spleen) when compared with multipotent stem cells. Thus, the cell we have identified appears to be the latest pluripotent cells so far identified in bone marrow and is therefore a good candidate for a bone marrow prothymocyte, but it appears not to be T-cell-committed.

Animals↗
PubMed1994

Assessment of CD4 expression by early T precursor cells and by dendritic cells in the human thymus.

The adult mouse thymus contains a minute population of early lymphoid precursor cells that express moderate levels of CD4. We searched for a corresponding population of early T precursors in the infant human thymus, by first depleting the majority of more mature thymocytes, then using immunofluorescence and flow cytometry to analyze cells bearing a range of early T lineage markers. No discrete population of early T precursors expressing CD4 was observed, in contrast to the murine thymus. Most putative very early human thymocytes were CD4-8-3-1-2lo44+34+7hi class I MHChi class II MHC-. However, a distinct population of human thymic dendritic cells expressing high levels of CD4 was isolated. These were CD4hi8-3-1-2-44+34-7- class I MHChi class II MHChi, and lacked markers of B cells, NK cells, or myeloid cells. They were large cells that exhibited dendritic morphology after brief periods of culture, and they were efficient stimulators of allogeneic T cells. The biologic implications of CD4 expression by thymic dendritic cells are discussed.

Adolescent↗
PubMed1993

Intrathymic lymphoid precursor cells during fetal thymus development.

Our previous studies have demonstrated the presence, in the adult mouse thymus, of a population of early precursor cells able to give rise to T and B lymphocytes but not myeloid cells. This population of cells expresses low levels of CD4 and has been termed the "low CD4 precursors." All these precursors were found to be c-kit positive, and they precede the better known CD4-CD8- precursor stage. In this study, embryonic and neonatal thymuses were examined to see whether a similar low CD4 precursor was part of the pathway of T cell development during ontogeny. A population with the phenotypic characteristics of the adult low CD4 precursor was found from day 15 of embryonic development, although the expression of low levels of CD4 was apparent only from embryonic day 17. Functional tests of these putative precursors showed they had no thymus-reconstituting ability when isolated from thymuses at any time during embryonic life, and very low reconstituting ability even 24 days after birth. These results raise questions about the adult low CD4 precursor as an obligatory stage in the development of T cells in the thymus.

Animals↗
PubMed1993

CD2-CD4-CD8- lymph node T lymphocytes in MRL lpr/lpr mice are derived from a CD2+CD4+CD8+ thymic precursor.

MRL lpr/lpr (lymphoproliferative, lpr) mice demonstrate an age-dependent lymphoproliferation and development of autoimmunity. Characteristic of the lymphoproliferation in these mice is the accumulation of large numbers of CD4-CD8-(CD4-8-),CD3+ T lymphocytes in their lymph nodes. The development of the CD4-8- cells, which also aberrantly express B220 and CD44 (Pgp-1) but are CD2-, has been shown to be thymus dependent. An unusual feature of lpr CD4-8-T lymphocytes is that although they appear unresponsive to stimulation, as defined by proliferation and IL-2 production, they have undergone thymic negative selection. As thymic deletion normally occurs at the CD4+CD8+ (CD4+8+) stage, this raises the dilemma that lpr CD4-8- T lymphocytes have either previously been CD4+8+, or they are able to undergo thymic selection as CD4-8- cells. We have addressed this question by examining the methylation status of the CD8 gene in MRL lpr CD4-8- lymph node cells. Demethylation of the CD8 gene has been shown to be an indicator of previous CD8 expression. We find that the CD8 gene in lpr CD4-8- lymph node cells, as well as in the abnormal B220+ CD4-8- lpr thymocytes, is demethylated, suggesting that these cells have previously expressed CD8. In addition, we find that the lpr CD4+8+ thymocyte population contains an increased percentage of atypical B220+, CD44+ cells that are virtually all CD2+. Taken together, these data are consistent with the lpr CD2-CD4-8- population of LNC having arisen from a CD2+ CD4+8+ thymic stage of differentiation.

Animals↗
PubMed1993

Development of gamma delta T cells in the adult murine thymus.

The development of T cells belonging to the gamma delta lineage is not well understood. We have analyzed the cells in the adult murine thymus which express the gamma delta TcR on the surface in order to learn more about this process. Our data demonstrate a number of clear subpopulations of gamma delta expressing cells in the thymus based on the expression of Thy-1 and HSA (heat-stable antigen). Only one of these subpopulations, the one expressing both Thy-1 and HSA, contains dividing cells or has a significant rate of turnover. Together with the fact that emigrant gamma delta cells are HSA+Thy-1+, this suggests that this thymic subpopulation is the sole, or major, source of exported cells. However, the turnover of cells from this population is 5 x 10(4) - 10 x 10(4) cells per day, while previous estimates of the rate of export of gamma delta cells are in the order of 10(4) cells per day. Furthermore the V gamma profile of recent gamma delta+ emigrants differs from that of the thymic HSA+Thy-1+ cells. This raises the possibility that only a selected subpopulation of the thymic gamma delta+HSA+Thy-1+ population is exported, and that some gamma delta cells may die in situ in the thymus. The function of the other gamma delta thymic subpopulations, which are turning over very slowly or not at all, (i.e. the HSA-Thy-1- and HSA-Thy-1+ subpopulations) remains unclear.

Animals↗
PubMed1993

Regulation of T cell production in T cell receptor transgenic mice.

The thymus produces many more cells than it releases into the periphery. According to generally accepted models of T cell development most of this loss occurs in the thymic cortex, among CD4+8+ thymocytes. An interesting situation arises in the case of T cell receptor (TcR) transgenic mice in which all cells can potentially be positively selected, leading to a theoretical increase of about 30-fold in the survival rate of CD4+8+ cells and in their transition to mature CD4+8- or CD4-8+ thymocytes. This in turn should lead to a 30-fold increase in the size of the thymic medulla, in the emigration rate and in the size of the peripheral T cell pool. Increases in medullary or peripheral pool sizes of this magnitude are not seen in TcR transgenic mice. The question was therefore asked whether some form of homeostatic process regulated the size of the mature T cell pool and at what level it might operate. In this report we demonstrate that the increased rate of double-positive to single-positive transition in the TcR transgenic mice is directly reflected in an increased emigration rate, and that the medulla seems to be relatively efficient regardless of the number of cells passing through it. However, the potential increases in emigrant numbers in TcR transgenic mice are offset by the reduced size of the CD4+8+ thymocyte pool. It would appear then that regulation of T cell production, if it occurs, probably does so through regulation of the size of the CD4+8+ thymocyte pool. Mechanisms for regulation of this kind are not yet known.

Animals↗
PubMed1993

TCR beta and TCR alpha gene enhancers confer tissue- and stage-specificity on V(D)J recombination events.

We describe transgenic mice carrying germline variable gene segments associated with either the T cell receptor (TCR) beta or alpha gene enhancers (E beta or E alpha). Transgenic constructs underwent high rates of site-specific rearrangements predominantly in T cells from independent mice. Rearrangements of the E beta-containing transgenes began at different stages of T cell differentiation in embryonic and adult thymus than did the E alpha-containing ones, with a pattern superimposable upon the patterns of TCR beta or TCR alpha gene expression, respectively. We demonstrate that sequences within the TCR beta and TCR alpha gene enhancers confer tissue- and stage-specificity upon the V(D)J recombination events affecting adjacent gene segments. The patterns of transgene expression also gave information on developmental events and lineage relationships (gamma delta versus alpha beta) during T cell development.

Aging↗
PubMed1993

Emigration of selected subsets of gamma delta + T cells from the adult murine thymus.

Cells bearing the gamma delta form of the TCR make up only 1-3% of T cells in the adult murine thymus and peripheral lymphoid organs. Evidence from studies of nude mice suggests that the development of at least some gamma delta T cells is thymus dependent; however, until now it has not been directly demonstrated that gamma delta cells are exported from the thymus. In this paper we have used the technique of labelling thymocytes in vivo with FITC, followed by flow cytometric analysis to trace cells emigrating from the thymus to the spleen. Using this approach we have been able to demonstrate for the first time that gamma delta T cells are exported from the adult murine thymus to the spleen. We also demonstrate that the cells emigrating to the spleen are a selected subset of gamma delta thymocytes being heat stable antigen positive, Thy-1+, and expressing low levels of CD44 (Pgp-1). In addition, investigation of TCR V gamma gene usage among adult gamma delta + thymocytes, recent emigrants, and spleen cells, indicated a selective emigration of gamma delta cells expressing certain V gamma genes.

Animals↗
PubMed1992

The surface phenotype of dendritic cells purified from mouse thymus and spleen: investigation of the CD8 expression by a subpopulation of dendritic cells.

A new procedure for rapid isolation of dendritic cells (DC) was devised, involving collagenase digestion of tissues, dissociation of lymphoid-DC complexes, selection of light-density cells, then depletion of lymphocytes and other non-DC by treatment with a mixture of lineage-specific monoclonal antibodies (mAbs) and removal with anti-immunoglobulin-coupled magnetic beads. This enriched population (approximately 80% DC) was further purified when required by fluorescence-activated cell sorting for cells expressing high levels of class II major histocompatibility complex (MHC). The isolated DC were characterized by immunofluorescent staining using a panel of 30 mAbs. Thymic DC were surface positive for a number of markers characteristic of T cells, but they were distinct from T-lineage cells in expressing high levels of class II MHC, in lacking expression of the T cell receptor (TCR)-CD3 complex, and having TCR beta and gamma genes in germline state. Splenic DC shared many markers with thymic DC, but were negative for most T cell markers, with the exception of CD8. A substantial proportion of DC from both thymus and spleen expressed CD8 at high levels, comparable with that on T cells. This appeared to be authentic CD8, and was produced by the DC themselves, since they contained CD8 alpha mRNA. Thymic DC presented both the CD8 alpha and beta chains on the cell surface (Ly-2+3+), although the alpha chain was in excess; the splenic DC expressed only the CD8 alpha chain (Ly-2+3-). It is suggested that the expression of CD8 could endow certain antigen-presenting DC with a veto function.

Animals↗
PubMed1992

Seeding of neonatal lymph nodes by T cells and identification of a novel population of CD3-CD4+ cells.

Mature T cells first appear in the thymus of the mouse a few days before birth, about 7-8 days after the thymus was colonized by stem cells. These mature cells are exported to the peripheral lymphoid organs beginning at about the time of birth, but because the number is very small at this stage, little is known about the phenotype or function of these early emigrants. We have examined the cells that accumulate in the peripheral lymph nodes (LN) during the first week of life to understand better the initial seeding of the periphery by T cells. Our studies showed that a high proportion of neonatal LN cells were CD4+, but that the majority of these were CD3- during the first few days of life. The CD3- population did not increase greatly in number after birth and rapidly diminished in proportion as the number of CD3+ cells increased. These CD3-CD4+CD8- cells were found to be Thy-1loCD44+ and to lack surface expression of heat-stable antigen. B220 and Mac-1. They had lymphoid morphology, did not phagocytose latex, and did not exhibit any precursor activity for cells of hemopoietic lineages. Their origin (intra- or extrathymic) as well as their function and physiological role, therefore, remains unknown. CD3+ T cells, both CD4+CD8- and CD4-CD8+, were present in low numbers during the first 1-2 days of life, but at post-natal day 3, a sharp increase in the accumulation of these cells occurred in both LN and spleen. By day 3 the CD4:CD8 ratio in LN was about 2:1, as in the adult. Crude estimates of the rate of export from the thymus from day 3 onwards gave values around 1% of thymocytes per day, i.e. close to our previous estimates for young adult thymus. We found no evidence of particularly high levels of emigration from the thymus during the first week after birth. Both CD4+CD8- and CD4-CD8+ T cell subsets were present in the LN as early as 1 day post-natally with CD4-CD8+ predominating among LN T cells, even though CD3+CD4+CD8- cells predominated over CD3+CD4-CD8+ cells in the thymus. By day 3 the ratio had changed to 2:1 (as in the adult). T cells, therefore, appear to emigrate from the thymus from about the time of birth with a dramatic increase around day 3 after birth.

Animals↗
PubMed1992

Commitment to the T cell receptor-alpha beta or -gamma delta lineages can occur just prior to the onset of CD4 and CD8 expression among immature thymocytes.

Two types of T lymphocytes, distinguishable by their surface expression of either the gamma delta or the alpha beta T cell receptor (TcR) for antigen, populate the periphery in the adult. In addition, immature precursors of both T cell types can be found in the thymus. While it is generally accepted that these two cell types represent distinct lineages, it is not known at which developmental stage these lineages diverge. The most mature thymocyte precursor population not yet expressing T lineage-specific surface markers (i.e. CD3, CD4, and CD8) is known to be capable of generating TcR-alpha beta T cells, and has been thought to be preprogrammed into the TcR-alpha beta lineage at an earlier developmental stage. We now show that this late-stage precursor is capable of giving rise to cells of both the TcR-alpha beta and -gamma delta lineages, both in vitro after intrathymic transplantation, and in vitro in simple culture medium or medium with cytokines. Thus it appears that the divergence of TcR-alpha beta and -gamma delta cells can occur at a relatively late stage of intrathymic development, just prior to the onset of CD4 and CD8 expression in most cells.

Animals↗
PubMed1992

Regulation of TCR alpha and beta gene allelic exclusion during T-cell development.

Early in their development, most T cells become committed to the expression of one, and only one, TCR alpha beta combination. How do T cells achieve this TCR allelic exclusion? This article discusses the configuration and expression of TCR alpha and beta genes in mature T-cell lines and TCR alpha beta transgenic mice, and proposes three nonexclusive models to account for the significant occurrence of T cells with two productive alpha gene rearrangements.

Alleles↗
PubMed1991

Developmental potential of the earliest precursor cells from the adult mouse thymus.

A new, numerically minute population of cells representing the earliest T precursor cells in the adult mouse thymus has recently been isolated. This population has been shown to be similar to bone marrow hemopoietic stem cells in surface antigenic phenotype and to express moderate levels of CD4. We now show, by fluorescence-activated cell sorting and intrathymic transfer to irradiated mice, that this apparently homogeneous population differs from multipotent stem cells in expressing the surface stem cell antigen 2 (Sca-2), that it differs from most early B lineage cells in lacking B220 and class II major histocompatibility complex expression, and that it binds rhodamine 123 like an activated rather than a quiescent cell. Irradiated recipient mice differing at the Ly 5 locus were used to compare the developmental potential of these early intrathymic precursors with bone marrow stem cells. Only T lineage product cells were detected when the intrathymic precursor population was transferred back into an irradiated thymus. However, when the intrathymic precursor population was transferred intravenously, it displayed the capacity to develop into both B and T lymphoid cells in recipient bone marrow, spleen, and lymph nodes, but no donor-derived myeloid cells were detected. The absence of myeloid and erythroid precursor activity was confirmed by showing that the intrathymic precursor population was unable to develop into myeloid or erythroid spleen colonies on intravenous transfer or to form colonies in an agar culture. These findings indicate that this earliest intrathymic precursor population has become restricted (or strongly biased) to lymphoid lineage development, but not exclusively to T lymphocytes.

Agar↗