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R Scollay

Publications and source records attributed to R Scollay.

At least 55 records · Page 3Linked to original sources
PubMed1991

CD4 expressed on earliest T-lineage precursor cells in the adult murine thymus.

A continuous but low input of stem cells or 'prothymocytes' is necessary to maintain T-cell development in the adult thymus, but the colonizing cell has not been characterized. Precursors of T cells have been found in the minor CD4-8- population of thymocytes, but even the earliest cells of this population already have partially rearranged T-cell antigen receptor (TCR) genes. We now demonstrate that the thymus contains a minute population of lymphoid cells similar in some but not all respects to bone marrow-derived haemopoietic stem cells. This population has TCR genes in a germline state. It gives a slow but extensive reconstitution of both alpha beta and gamma delta lineages on transfer into an irradiated thymus, with kinetics indicating that it includes the earliest intrathymic precursor cells so far isolated. Surprisingly, these cells express low surface levels of the mature T-cell marker CD4.

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PubMed1991

The effect of peripheral immunization with Mls-1a on the emigration of antigen-specific cells from the thymus.

Mature T cells found in the lymph nodes and spleen have the capacity to become activated and to proliferate in response to foreign antigens. The response of the thymus to such immunization is less well understood. We have examined one aspect of the thymic response by determining the effect of peripheral immunization upon cell emigration from the thymus. BALB/c (Mls-1b) mice were injected with spleen cells from DBA/2 (Mls-1a) mice, and V beta 6+ (Mls-1a-reactive) thymic emigrants were identified 3-30 days after immunization. Neither the rate of total cell migration from the thymus nor the proportion of V beta 6+ cells was altered, even though the immunizing spleen cells elicited an immune response in the draining (parathymic) lymph nodes. The same immunogen caused deletion of V beta 6+ cells in both the thymus and lymph nodes after intraperitoneal injection into the neonate. The inability of DBA/2 splenocytes to modify the development of adult thymocytes after intrathymic injection of the cells precluded the lack of entry into the thymus as the reason for the lack of any observed effect in the adult. Our results, therefore, indicate that the development of adult thymocytes is not modified by immunization, and suggest that the differing thymic response of mice injected as adults or neonates is related to changes in the intrathymic antigen presentation capacity associated with age.

Age Factors↗
PubMed1991

Characterization of immature CD4+CD8-CD3- thymocytes.

Previously we have described (Hugo, P. et al., Int. Immunol. 1990. 2: 209) an immature CD4+CD8-CD3- thymocyte subset which is thought to be the counterpart of the CD4-CD8+CD3- subset. In this study we show that the ontogeny of these two subsets is parallel in fetal thymic organ culture. Extensive phenotypic characterization of CD4+CD8-CD3- cells reveals that they closely resemble CD4-CD8+CD3- thymocytes being: HSAhigh, Thy-1high, interleukin 2 receptor alpha chain negative, CD44-, H-2K+/-, CD5low, MEL-14low/intermediate, CD2+, LFA-1+ and MTS 35+. Finally, we show that the proportion of CD4+CD8-CD3- thymocytes is highly variable between mouse strains.

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PubMed1991

CD4+CD8+CD3high thymocytes appear transiently during ontogeny: evidence from phenotypic and functional studies.

During T cell development thymocyte subsets emerge in a defined order, reflective of their maturational stage. In this study we determined the timing of appearance of CD4+CD8+CD3high thymocytes during in vivo and in vitro embryonic development, and thymic reconstitution after cortisone treatment. In these models, CD4+CD8+CD3high cells followed CD4+CD8+CD3low and preceded mature CD4+CD8-CD3high/CD4-CD8+CD3high thymocytes, while cortisone resistance was first seen among CD4+CD8+CD3high cells. CD4+CD8+CD3high thymocytes were also shown to display a pattern of antigen receptor-mediated calcium influx intermediate between that induced in other CD4+CD8+ cells and mature thymocytes. These results are consistent with a precursor-progeny relationship between CD4+CD8+CD3low and CD4+CD8+CD3high thymocytes, the latter developing to mature thymocytes (Hugo, P. et al., Int. Immunol. 1991. 3: 265).

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PubMed1991

T-cell subset relationships in thymocyte development.

During the past couple of years there has been significant progress in our understanding of the development of different lineages of T cells within the thymus. Pathways, subpopulations and cellular dynamics are all becoming clearer. Signal transduction through primary and accessory receptors is also beginning to be understood. However, the exact nature of the events that lead uncommitted cells to choose a particular lineage (either alpha beta/gamma delta or CD4+/CD8+) has still not been determined.

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PubMed1991

Timing of deletion of autoreactive V beta 6+ cells and down-modulation of either CD4 or CD8 on phenotypically distinct CD4+8+ subsets of thymocytes expressing intermediate or high levels of T cell receptor.

In this paper we describe a differentiation sequence amongst adult murine thymocytes which goes from CD4+8+3lo(low) to CD4+8+3int(intermediate) to CD4+8+3hi(high) and then to mature single positive CD3hi thymocytes. Phenotypic characterization of CD4+8+3int/hi cells for a number of other surface markers is consistent with them being in transition from CD4+8+3lo phenotype to mature phenotype. The same observation was made for sensitivity towards ionomycin-mediated apoptosis. In the thymus of Mls-1a mice, where autoreactive TCR-V beta 6+ cells are negatively selected, deletion of TCR-V beta 6+ cells was first detected in the CD4+8+3int subset, and was complete by the CD4+8+3hi stage, suggesting that up-regulation of the TCR/CD3 complex is required for deletion of Mls-1a autoreactive thymocytes. No sign of apoptosis was detected among any fresh thymocyte subsets suggesting that apoptotic cells are rapidly cleared from the thymus. The CD4+8+3int/CD4+8+3hi cells are therefore populations in transit from the typical cortical thymocytes to the mature T-cells.

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PubMed1990

Lineage relationships and developmental kinetics of immature thymocytes: CD3, CD4, and CD8 acquisition in vivo and in vitro.

T lymphocytes develop in the thymus from immunologically naive bone marrow precursors. Based on T cell receptor rearrangement and transcription, and thymic reconstitution potential, we have deduced a developmental sequence among immature thymocytes, before the acquisition of the lineage markers CD3, CD4, and CD8. In the current study, we have followed the ontogenic progression of the latter stages in this sequence, using two different systems: (a) in vivo, by direct injection into the thymus of nonirradiated, congenic recipients; and (b) in vitro, using culture medium without mitogens or cytokines. In vivo, the less mature Pgp-1- interleukin 2 receptor alpha-positive (IL-2R alpha+) CD3-4-8- subset (also heat-stable antigen high) requires 3 d before becoming predominantly IL-2R alpha- CD3lo4+ 8+ typical cortical-type cells, and at least 5 d before the appearance of any mature single-positive cells (CD3hi4+ 8- or CD3hi4-8+). However, these Pgp-1- IL-2R alpha+ precursors do not differentiate further in unstimulated culture. The more mature Pgp-1- IL-2R alpha- CD3-4-8- subset becomes primarily CD3lo4+ 8+ within 1 d after transplantation, and some mature single-positive progeny are evident by day 3. By 5 d, most of these Pgp-1-IL-2R alpha- precursor cells have become CD3hi, and have lost or are downregulating either CD4 or CD8. In culture, these Pgp-1- IL-2R alpha- cells also acquire high levels of CD4 and CD8 within 1 d, and low levels of CD3 by 2 d. However, they do not progress further to mature single positives in vitro, and most of them die by day 3. These experiments directly confirm our previously proposed developmental sequence, and demonstrate the kinetics of T lymphocyte production in a low-stress, steady-state environment.

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PubMed1990

Differentiation of hematopoietic stem cells in irradiated mouse thymic lobes. Kinetics and phenotype of progeny.

To define cell populations which participate in the very early stages of T cell development in the mouse thymus, we enriched hematopoietic stem cells from mouse bone marrow and injected them into thymic lobes of irradiated Ly-5 congenic recipients. The progeny of the stem cells were identified and their phenotypes were determined by two-color flow cytometry for the expression of various cell surface differentiation Ag during the course of their subsequent intrathymic development. The majority of the differentiation which occurred in the first 10 days after intrathymic cell transfer was myeloid in nature; hence, this study demonstrates that the irradiated thymus is not strictly selective for T cell development. Further, the maximum rate of T cell development was observed after intrathymic injection of 200 stem cells. Donor-derived cells which did not express Ag characteristic of the myeloid lineage could be detected and their phenotypes could be determined by flow cytometry as early as 7 days after intrathymic injection. At this time, the cells were still very similar phenotypically to the bone marrow hematopoietic stem cells. Exceptions to this were the expression of stem cell Ag 2 and a decrease in the level of MHC class I Ag expression. After 9 days, the donor-derived cells expressed high levels of the Thy-1 Ag and proceeded to change in cell surface phenotype as differentiation continued. These cell phenotypes are described for the time frame ending 18 days after injection, when most donor-derived cells were phenotypically small CD4+ CD8+ (double-positive) thymocytes.

Animals↗
PubMed1990

Selection of the T-cell repertoire in transgenic mice expressing a transplantation antigen in distinct thymus subsets.

Transgenic mice that expressed a transplantation antigen, H-2Kb, in an unusual tissue distribution have been developed. Gene-regulatory elements from the immunoglobulin heavy-chain locus (Emu enhancer and heavy chain promoter) were linked to the class I Kb gene and the construct microinjected into fertilized mouse eggs of a different haplotype. It was expected that such gene-regulatory elements would direct expression of the foreign class I molecules only to B and T lymphocytes. However, expression was also detected in a subset of thymus medullary epithelium. The Kb molecules expressed on this thymic subset were unable to positively select T cells for passage to the periphery. The mice were, however, tolerant of the cell types expressing the foreign Kb molecules and were also tolerant of Kb presented as skin grafts. These results suggest that not all components of thymic epithelium are involved in positive selection of T cells and that transplantation antigens expressed on non-dendritic cells can induce tolerance.

Animals↗
PubMed1990

Development of immature thymocytes: initiation of CD3, CD4, and CD8 acquisition parallels down-regulation of the interleukin 2 receptor alpha chain.

We have previously identified a developmental sequence among immature thymocytes, prior to their expression of the lineage markers CD3, CD4, and CD8. This sequence is marked by transient expression of the interleukin 2 receptor alpha chain (IL 2R alpha). The most mature cells in this sequence (surface phenotype heat-stable antigen (HSA)++ Pgp-1- IL 2R alpha-) are the immediate precursors to CD4+CD8+ small cortical thymocytes, and have by definition been considered to be CD4-CD8-. We now show that these cells display low levels of surface CD4 and CD8, but not CD3. This low-level expression begins to appear immediately after the loss of IL 2R alpha expression. Northern blot analysis for mRNA expression confirms that these IL 2R alpha- cells are transcribing CD4 and CD8 mRNA, in contrast to their immediate (IL 2R alpha+) precursor. Upon unstimulated culture, these IL 2R alpha- cells gradually acquire high levels of CD4 and CD8, as well as low levels of CD3, whereas IL 2R alpha+ cells do not. These findings suggest that the IL 2R alpha+ subset is the end of the true CD3-CD4-CD8- phase, and that the intracellular signals for CD3, CD4, and CD8 acquisition occur simultaneously with, or immediately prior to, the signal for down-regulation of IL 2R alpha.

Animals↗
PubMed1990

Kinetics of mature T-cell development in the thymus.

We have reexamined the balance between cell birth, cell maturation, and cell death in the thymus by labeling dividing thymocytes and their progeny in vivo with [3H]-thymidine, isolating clearly defined subpopulations by fluorescence-activated cell sorting, and determining the distribution of label by autoradiography. When mature thymocytes were precisely defined (as CD4+CD8- CD3+ or CD4-CD8+ CD3+) and separated from immature single positives (CD4+CD8- CD3- and CD4-CD8+ CD3-), a lag was observed in the rate of entry of [3H]thymidine into mature cells. Thus, many of the mature thymocytes appear to derive from a small nondividing cortical thymocyte pool, rather than originating directly from the earliest dividing CD4+CD8+ blasts. There was little evidence for cell division during or after mature thymocyte formation, suggesting a one-for-one differentiation from cortical cells rather than selective clonal expansion. The rate of production of mature single positive thymocytes agreed closely with estimates of the rate of export of mature T cells from the thymus and was only 3% of the rate of production of double-positive cortical thymocytes. This was compatible with a stringent selection process and extensive intrathymic cell death and suggested that no extensive negative selection occurred after the mature cells were formed.

Animals↗
PubMed1990

CD4-CD8- thymocytes that express the T cell receptor may have previously expressed CD8.

Amongst CD4-CD8- (double negative) thymocytes there is a sizeable population (variable from strain to strain) of cells expressing surface T cell receptor (TCR). These TCR+ double negatives are predominantly non-cycling, have very little precursor activity, and, unlike the TCR-CD4-CD8- thymocytes, appear not to be part of the mainstream of thymocyte development. A unique feature of this population is the biased V beta-gene region usage. In CBA mice, 60-70% of TCR+ CD4-CD8- cells express receptors that utilize V beta 8 gene products, compared with peripheral T cells from the same strain which are only 20-30% V beta 8+. This suggests that the high V beta 8 usage may be the result of some selective process. A growing body of experimental data suggests that TCR specificity selection occurs at the CD4+CD8+ stage of thymocyte development. In order to gain some insight into the previous history of the TCR+ double negatives, in particular whether or not they have previously expressed CD8 and therefore been eligible for selection, we have determined the methylation state of the CD8 gene and compared it to other thymocyte populations. We show that the TCR+ CD4-CD8- thymocytes are demethylated at some sites in the CD8 gene, consistent with previous CD8 expression. However, the demethylation pattern is distinct from that seen on typical peripheral T cells or on mature thymocytes, suggesting that the TCR+ CD4-CD8- thymocytes are not derived from mature thymocytes or peripheral T cells which have returned to the thymus and downregulated CD8 expression.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Intrathymic selection of murine TCR alpha beta+CD4-CD8- thymocytes.

The CD4-CD8- thymocyte population contains the precursors of all other thymocytes. However, it also contains a significant proportion of cells which express surface TCR alpha beta, and have little or no precursor activity. Like peripheral T cells, but unlike most other thymocytes, these TCR alpha beta+CD4-CD8- thymocytes do not express heat stable antigen. Both the origin and developmental status of these cells are unclear, and are the subject of this report. We have measured the proportion of V beta 8.1+ cells amongst TCR+HSA-CD4-CD8- thymocytes in MIs-1a versus MIs-1b mice, in order to determine whether they have undergone negative selection. The proportions were similar in both strains, in contrast to mature T cells, indicating that neither they nor their precursors had undergone clonal deletion. We also measured the accumulation of these cells over the early life of the animal and found that it was extremely slow. Our data also show that although TCR-V beta 8.1+ cells are reactive to MIs-1a in association with MHC class II, most mature TCR-V beta 8.1+ cells in MIs-1b mice are CD8+, suggesting an additional reactivity with MHC class I. We raise the possibility that TCR-V beta 8.1+CD4-CD8- thymocytes are derived from TCR-V beta 8.1+CD4+CD8+ thymocytes, and that the reactivity of TCR-V beta 8.1 with both MHC classes I and II has resulted in the down-regulation of both CD4 and CD8.

Animals↗
PubMed1990

Ontogeny of a novel CD4+CD8-CD3- thymocyte subpopulation: a comparison with CD4- CD8+ CD3- thymocytes.

We have studied the ontogeny of a novel thymocyte subset, CD4+CD8-CD3-. Three-colour flow cytometric analysis demonstrated that these cells constituted approximately 1% of the total thymocyte content in adult CBA mice, and were not present in lymph nodes. They were mainly blastic, cortisone-sensitive, and localized in the outer thymic cortex. During foetal life they were first observed at day 15 and reached a maximum (6%) at day 17, beyond which they decreased to the adult level. This kinetic profile was similar to that of the CD4-CD8+CD3- subpopulation, except that the CD4+CD8-CD3- cells appeared slightly earlier and their percentage was lower. Both these populations appeared after the CD4-CD8-CD3- cells but before the CD4+CD8+CD3- cells. Similar observations were made during thymic reconstitution following dexamethasone treatment. In this case, both CD4+CD8-CD3- and CD4-CD8+CD3- thymocytes disappeared 48 h after the treatment. While their absolute number increased up to 14 days post-treatment, their percentage was maximal at day 7 post-treatment and returned to normal values by day 10 post-treatment. These results argue strongly that not only the CD4-CD8+CD3- population but also the CD4+CD8-CD3- population can be considered an intermediate precursor in CBA thymuses.

Animals↗
PubMed1990

Analysis of recent thymic emigrants with subset- and maturity-related markers.

The thymus plays an integral role in the development and production of T lymphocytes. However, thymocytes differ markedly in their phenotypic characteristics from the T cells normally found in the peripheral lymphoid organs. We have examined the phenotypic characteristics of recent thymic emigrants and compared them with both mature phenotype thymocytes (CD4+ CD8-CD3+ and CD4-CD8+ CD3+) and lymph node T cells. Recent thymic emigrants were defined as those fluorescein-positive cells found in the lymph node up to 16 h after intrathymic injection of fluorescein. Most cells emigrating from the thymus expressed CD3 and either CD4 or CD8, indicating maturity. Recent thymic emigrants, like mature phenotype thymocytes, were slightly larger on average than peripheral T cells, but this differential was lost within 24 h of emigration. Also like mature thymocytes but unlike peripheral T cells, some recent emigrants expressed heat-stable antigen. This did not change within 24 h of emigration. The antigen CD44 (Pgp-1, Ly-24) was expressed on a proportion of mature thymocytes, recent thymic emigrants, and peripheral T cells, and its expression did not show any clear relationship to maturity. The antigen CD45R also did not show marked changes associated with maturity, but our data do not parallel the published data of the expression of CD45R in the human. We conclude that recent thymic emigrants are phenotypically mature with respect to some antigens but not others. None of the antigens we investigated could have been used to uniquely distinguish recent thymic emigrants from peripheral T cells or from mature thymocytes.

Animals↗
PubMed1990

The kinetics of immature murine thymocyte development in vivo.

The dynamics of cell generation and turnover in the young adult murine thymus has been studied by in vivo administration of [6-3H]deoxythymidine, isolation of thymocyte subpopulations by negative depletion and cell sorting procedures, and assessment of dividing cells and their products by autoradiography. The flow of label through subpopulations of CD4-CD8- thymocytes defined by the markers heat stable antigen (HSA), phagocyte glycoprotein 1 (Pgp-1), interleukin 2 receptor (p55) (IL-2R), and CD3 was determined, to check the developmental sequence deduced from intrathymic transfer and molecular approaches. In addition, the flow of label 'downstream' into the CD4+CD8+ cortical populations was followed to check if cells expressing CD8 alone were obligatory intermediates. The main findings were: (i) support for the following sequence within the CD4-CD8- group: HSA++Pgp-1+IL-2R(-)----HSA++Pgp-1-IL-2R(+)----HSA++Pgp-1-IL- 2R-; (ii) the majority of cell generation and cell turnover within the CD4-CD8- population was due to the HSA++IL-2R-Pgp-1- subpopulation; (iii) the rate of cell output from the proposed intermediate CD3-CD4-CD8+ subpopulation was equivalent to only 55% of the cell output from its proposed precursor, the most mature CD4-CD8- subpopulation, suggesting that many double negatives differentiate directly (or via CD3-CD4+CD8- intermediates) into double positives; and (iv) the CD4-CD8-HSA- (and CD3+) thymic subpopulation contained very few cycling cells and turned over extremely slowly, indicating that these slowly accumulating product cells are off the mainstream of T cell development.

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