Biomedical subjects

P Hugo

Publications and source records attributed to P Hugo.

At least 55 records · Page 3Linked to original sources

PubMed1992

Thymic shared antigen-1. A novel thymocyte marker discriminating immature from mature thymocyte subsets.

In a previous study, we raised a mAb (MTS 35) reacting with a plasma membrane Ag expressed on both cortical thymocytes and a subset of thymic medullary epithelial cells. In view of the shared expression of this molecule, we have defined it as thymic shared Ag-1 (TSA-1). Considering its selective reactivity with cortical, but not medullary thymocytes, the relevance of TSA-1 as a marker of immature T cells was investigated in detail in this study, using multicolor flow cytometric analysis. TSA-1 was found on all immature thymocyte subsets (CD3-4-8-, CD3-4+8-, CD3-4-8+, CD3-4+8+, CD3low4+8+). Conversely, CD3high4+8- and CD3high4-8+ thymocytes, early thymic migrants and peripheral T cells were TSA-1-. More refined gating and analysis of the transitional CD3intermediate/high4+8+ thymocytes, proposed candidates for negative selection, demonstrated that approximately one half were TSA-1-. In fact, there was a directly inverse relationship between TSA-1 and CD3 expression on thymocytes. In the periphery, TSA-1 was detected on B lymphocytes. TSA-1 is PI-linked and has a molecular mass of 17 kDa nonreduced, or 12 to 13 kDa reduced. Through cross-correlation analysis, this molecule was distinct from H-2K, PNA-R, CD5, CD11a/18, Thy-1, HSA, Ly6A/E, Ly6C, ThB, CD25, CD44. Hence TSA-1 appears to be a unique marker which exquisitely separates mature from immature thymocytes.

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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

Towards an integrated view of thymopoiesis.

One prediction from the complex series of steps in intrathymic T-cell differentiation is that to regulate it the stroma controlling the process must be equally complex: the attraction of precursors, commitment to the T-cell lineage, induction of T-cell receptor (TCR) gene rearrangement, accessory molecule expression, repertoire expansion, major histocompatibility complex (MHC) molecule-based selection (positive and negative), acquisition of functional maturity and migratory capacity must all be controlled. In this review, Richard Boyd and Patrice Hugo combine knowledge of T-cell differentiation with thymic stromal cell heterogeneity to offer an integrated view of thymopoiesis within the thymic microenvironment.

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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

Selection of CD4+CD8+ thymocytes by complex formation with medulla-derived epithelial cells.

The role of lymphostromal complexes in T-cell differentiation is far from elucidated, mainly because a clear association of a particular stromal cell type with a distinct thymocyte subset has never been identified. Using an in vitro system, detecting the adherence of thymocytes to a thymic medullary epithelial cell line (E-5), we showed that the phenotype of these thymocytes was that of cortical type: Thy-1hi, LFA-1+, PNAhi, CD4+CD8+, MEL-14-/lo, IL-2R-, CD3-/lo, and TcR V beta 8-/lo. They were enriched in cells in G2/M at the time of complex formation, showed a higher basal proliferation in culture, and did not respond to PHA, IL-2 and only marginally to Con A. These data show that complex formation with mouse thymic medullary epithelium selects for CD4+CD8+ thymocytes, as shown by the marked decrease in CD4+CD8-/CD4-CD8+ thymocytes, and the incapacity of CD4-CD8- thymocytes to adhere.

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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.

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PubMed1989

Thymic lymphoma cells as a model for complex-formation between thymocytes and thymic medullary epithelial cells.

The formation of complexes between thymocytes and thymic stromal elements is known to be involved in T cell differentiation. We have previously described one type of lympho-stromal interaction involving CD4+ CD8+ thymocytes and a medullary epithelial cell line (E-5). In this study we report the potential for complex formation of two different thymic lymphoma cell lines (Ti-6 and RDM-4). Ti-6 cells were shown to adhere to the E-5 cells, while RDM-4 cells were totally incompetent. Phenotypic characterization of these cell lines suggests that immature thymocytes are not capable of forming complexes with medullary epithelium and that a certain level of differentiation is required to do so. Comparison of their phenotypes showed that the possibility of some classical T cell surface markers being the receptor for the E-5 ligand can be dismissed.

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PubMed1989

Dynamics of complex formation between thymocytes and thymic medullary epithelial cells.

Direct cell contact is an intrinsic part of several differentiation processes. A case in point is the formation of complexes between thymic lymphocytes and stromal cells, which are essential for T-cell maturation. The objective of the present work was to gain an insight into the mechanisms underlying the formation and dissociation of such lymphostromal complexes. Using an in vitro system, we show that the number of thymocytes adhering to a thymic medullary epithelial cell line (E-5) increases with time and reaches a plateau, after which some thymocytes spontaneously detach, while the rest remain attached to epithelium. The detached and forcibly removed thymocytes were analysed for their expression of L3T4 and Ly-2 antigens. The detached thymocytes showed a markedly lowered expression of both antigens. Both of these subtypes were shown to be totally refractory to form further complexes after a first encounter with E-5 cells. We also show that, once a mean adherence level of one thymocyte per E-5 cell was reached, (a) the level of further adherence increased exponentially, and (b) during this phase, binding of thymocytes to E-5 cells occurred in clusters, indicating pre-existing polarity or contact-induced polarization of the receptors at the E-5 cell surface.

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PubMed1989

Binding of cortical thymocytes to a medullary epithelial cell line: a brief review.

In this review, we bring together data regarding complexes formed between thymocytes and a line of thymic epithelial cells, E-5. We show that E-5 has all the markers which identify it as of epithelial and medullary origin. We show that thymocytes which form complexes with it belong to the double positive, CD4 + CD8 + population, and that the receptor responsible for binding to epithelial cells is likely to be an as yet unidentified glycoprotein. The ligand on E-5 cells is also a glycosylated protein. Adhering thymocytes are in a state of mitotic activity, and their incubation with E-5 cells does not induce increased responsiveness to polyclonal mitogens. Non-adhered thymocytes, on the other hand coincubated With E-5 cells increase their responsiveness to ConA and initiate responsiveness to PHA. Differentiation of adhering thymocytes is evidenced by their acquisition of refractoriness to a secondary adherence, and by the modification of their surface phenotype.

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PubMed1988

Interactions between lymphoid cells and a thymic stromal cell line in vitro.

In the present study, we have optimalized the adherence assay to allow monitoring of the level of contact between thymocytes and a thymic epithelial cell line, E-5, in vitro. This type of interaction is not MHC-restricted, thus is unlikely to participate in the education of thymocytes to self. It was also shown that adherence does not vary from strain to strain, except for B6lpr/lpr immunodeficient mice which showed a markedly decreased adherence. This might be caused by the high level of L3T4-, Lyt-2- thymocytes in these mice (Davignon et al., 1985), since enriched double negative cells were shown not to adhere to E-5 cells. Preliminary characterization of adhering thymocytes suggests an heterogeneous mature phenotype. These cells appear around day 16 of fetal life and increase gradually until birth to remain constant throughout life. On the basis of contact duration, two populations of adhering thymocytes exist: one spontaneously detached after 1.5 hr, refractive to further adherence and the other which adheres for up to 14 hr. Contact between lymphoid and E-5 cells was shown to induce PHA responsiveness.

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PubMed1988

Transient inhibition of mixed lymphocyte reactivity by dieldrin in mice.

Dieldrin, a non-aromatic organochlorinated pesticide, was shown to be a potent modulator of the immune system. We had earlier demonstrated that mice treated with a single sublethal dose of dieldrin showed an impaired antibody response and reduced viral restriction mediated by macrophages. These dieldrin-induced immunosuppressive effects were shown to be dose dependent, when administered by the oral or intraperitoneal (i.p.) route. This study was undertaken to investigate the effect of dieldrin on the T-cell immune response. Lymphoid cells from mice injected i.p. 7 days earlier with 36 mg/kg body weight (0.6 LD50) dieldrin were assessed for their ability to recognize a foreign antigen and to proliferate in a mixed lymphocyte reaction (MLR) at 4, 7, 14 and 24 days post-treatment. We have demonstrated a strong but transient inhibition of MLR at 7 days after pesticide exposure. This effect was reversible and could not be attributed to a direct cell cytotoxicity, nor to the modulation of the T-cell ratio in lymphoid organs. Since the mitogen response was not impaired at this time point, we suggest that T-cell ability to recognize a foreign antigen can be altered by dieldrin, but not the proliferative potential of the cells.

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PubMed1988

Abrogation of graft-versus-host reaction by dieldrin in mice.

Sublethal exposure to the organochlorine pesticide, dieldrin, decreased the T-cell immune response in mice. Indeed, a transient inhibition of the mixed lymphocyte reactivity (MLR) was noted at 7 days after intraperitoneal exposure to 0.6 LD50 dieldrin. The present study was undertaken to further investigate the effects of dieldrin on the T-cell immune response, using the graft-versus-host reaction (GVHR) as a model, in order to assess T-cell subset efficiency. Lymphoid cells of A/J mice injected intraperitoneally 7 days earlier with 36 mg/kg body weight dieldrin were transferred into H-2-incompatible F1 hybrids. With this model, known to induce a marked GVHR, we have observed that dieldrin inhibited the potential of parental cells to induce a GVHR in hybrid mice. This effect could not be attributed to a direct cell cytotoxicity, nor to the modulation of major T-cell subsets as shown by thymic and peripheral T-cell subpopulation analysis. Collaboration processes between these cellular subsets seem to represent a potential site for the dieldrin-induced suppression.

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PubMed1988

Resistance to RadLV-induced leukemia: non-participation of splenic natural killer cells.

The phenotypic expression of genetically determined resistance to radiation leukemia virus (RadLV)-induced leukemia in mice has been shown to reside in the bone marrow. Because the bone marrow contains precursors of natural killer (NK) cells, known to play a role in retrovirally induced infections, and because these cells have been suggested as participating in resistance to radiation-induced leukemia, it was pertinent to establish whether their levels differed in strains of mice susceptible and resistant to leukemia. We therefore tested splenic NK cell levels in C57BL/Ka (susceptible) and B10.A(5R) (resistant) mice before viral inoculation, immediately after viral inoculation, and throughout the preleukemic period and showed that they were not different. This indicates that splenic NK cell levels have no bearing on the resistance to RadLV-induced leukemia and that other immune or non-immune mechanisms must be sought.

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PubMed1988

Characterization of the in vitro interaction between thymocytes and a medullary thymic epithelial cell line.

T-cell differentiation is known to be mediated by lympho-stromal interactions. However, the precise role of cellular complexes formed during this process is far from clear. We have previously established a thymic medullary epithelial cell-line, E-5, and have shown the adherence of thymocytes by a receptor-mediated-mechanism. We report here that the thymocytes able of complex-formation with E-5 cells appear at day 16 of gestation. Moreover, while the proportion of such thymocytes is constant throughout post-natal life, their absolute number decreases with thymic involution. We have also investigated the influence of the genetic background of thymocytes on adherence and found that polymorphic regions of H-2 genes were not involved in contact recognition. Therefore, this type of lympho-stromal interactions is unlikely to participate in the education to self MHC-restriction. However, thymocytes from B6/lpr mice, which spontaneously develop systemic lupus erythematosus (SLE) and have impaired T-cell differentiation, were shown incapable of adhering to E-5 cells. These results are interpreted as showing that interaction between thymocytes and medullary epithelial cells reflect a discrete stage of T-cell differentiation.

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PubMed1987

Suppression of humoral immunity in inbred mice by dieldrin.

The effect of single, sublethal i.p. injection of dieldrin on the primary antibody response to thymodependent (sheep red blood cells, SRBC) and T-cell-independent (lipopolysaccharide, LPS) antigens were investigated in inbred C57Bl/6 mice. Time-course studies showed significant suppression of the anti-SRBC IgM and anti-LPS IgM response at 7-24 days and at 4-14 days, respectively, after exposure to 0.6 LD50 of dieldrin. The anti-SRBC IgG response was also suppressed by dieldrin exposure, however, maximal suppressory effect was found at 48 days after the pesticide exposure. Similar patterns of the dieldrin-induced suppression of the primary IgM response to the thymodependent and T-cell-independent antigens, in addition to the overall control of cytotoxicity of lymphoid cell populations, suggest rather dysfunction of cellular cooperation during the inductory phase of the immune humoral response.

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PubMed1986

Suppression of MHV3 virus-activated macrophages by dieldrin.

Dieldrin (36 mg/kg body weight) administered intraperitoneally prolonged recovery from infection with mouse hepatitis virus 3 (MHV3) in the genetically-resistant A/J strain, affected the humoral anti-MHV3 IgG immune response, and inhibited the intrinsic antiviral activity of peritoneal macrophages upon in vitro rechallenge with the virus. Infection of untreated A/J animals and vehicle controls with MHV3 resulted in marked and reproducible activation of peritoneal macrophages, observed in vitro as resistance to MHV3-cytopathic effects 48 hr after rechallenge with the virus, whereas exposure to dieldrin resulted in apparent loss of the intrinsic capacity of cells to restrict replication of MHV3 and to protect them from cytolysis. In addition, in vitro treatment of MHV3 virus-activated macrophages with dieldrin, mitomycin C and X-irradiation, inhibited the intrinsic capacity of cells to restrict MHV3 replication. This mechanism of cellular restriction of the virus by MHV3-activated macrophages from the resistant A/J strain appeared to be one of the sensitive targets for the suppressive action of dieldrin on host resistance, as no major changes in macrophage cellular parameters were observed in in vitro studies of cell viability, adherence to plastic, and superoxide anion generation; the increased cell yield in the peritoneal exudates during MHV3 virus infection was not affected by dieldrin exposure; and the attachment and uptake of [3H]MHV3 by virus-activated macrophages was shown to be unchanged by dieldrin exposure.

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