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A G Zapata

Publications and source records attributed to A G Zapata.

At least 37 records · Page 2Linked to original sources
PubMed1997

Expression of zebrafish rag genes during early development identifies the thymus.

Recent experiments have demonstrated that zebrafish is a vertebrate in which it is possible to carry out large-scale mutagenic screens to identify genes involved in specific developmental pathways. To follow development of the immune system in zebrafish, we have analyzed the expression of the recombination activating genes, rag1 and rag2, which we have previously isolated and characterized. These genes catalyze the rearrangement of immunoglobulin genes in immature B lymphocytes and of T cell receptor genes in immature T lymphocytes and are therefore appropriate markers to follow the development of organs containing these cells. By whole-mount in situ hybridization, we detected expression of both rag genes in a paired organ in the head, beginning on the fourth day after fertilization. Histological examination of this organ indicated that it corresponds to the thymus, as described for other fish, an organ that has not previously been identified in zebrafish. By histological analysis, the thymus primordium appears at 54 hr but does not enlarge significantly until 30 hr later. The thymus continues to enlarge and reaches its mature histological organization at 1 month. The pronephros, the major hematopoietic organ in the adult fish, begins to develop hematopoietic tissue about 2 weeks after fertilization. By 1 month, mature lymphocytes are distinguishable in the tissue surrounding renal tubules. Lymphocytes appear in the kidney too late for screening by whole-mount in situ hybridization; however, the pattern of rag1 expression in the thymus forms the basis of an assay for mutations affecting development of the thymus or its constituent lymphocytes.

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PubMed1997

Interleukin-7 treatment promotes the differentiation pathway of T-cell-receptor-alpha beta cells selectively to the CD8+ cell lineage.

In this report we have studied the influence of interleukin-7 (IL-7) on thymocyte differentiation by evaluating the effects of IL-7 on the generation of T-cell receptor-alpha beta (TCR-alpha beta) and TCR-gamma delta thymocyte subpopulations in rat fetal thymus organ culture. IL-7 enhanced the differentiation pathway of TCR alpha beta thymocytes, first increasing the numbers of immature CD8+ cells, and later those of both CD4+ CD8+ and mature thymocytes. The kinetics of thymocyte migration out of thymic lobes was also accelerated, and the average number of mature TCR-alpha beta phi emigrants per day was increased in the presence of IL-7. Moreover, mature CD4- CD8+ thymocytes were preferentially generated after IL-7 administration. This TCR-alpha beta hi cell population was not actively dividing, indicating that IL-7-promoted thymocyte differentiation was selective to the CD8 cell lineage. Distribution of some TCR-V alpha and TCR-V beta segments among mature thymocytes was also modified in IL-7-treated thymic lobes. On the contrary, the maturation of TCR-gamma delta was not affected by IL-7 addition during the first days of culture, but their numbers sharply increased by day 6 of culture. These results were confirmed with IL-7-treated cultures for 24 hr, showing that IL-7 responsiveness was acquired by TCR-gamma delta cells late in thymus ontogeny. The present results thus indicate a key role for IL-7 in the maturation of TCR-alpha beta thymocytes and the expansion of thymic TCR-gamma delta cells.

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PubMed1997

Role of IL-2 in rat fetal thymocyte development.

Early during rat thymus ontogeny, an important proportion of thymocytes expresses IL-2R and contains IL-2 mRNA. To investigate the role of the IL-2-IL-2R complex in rat T cell maturation, we supplied either recombinant rat IL-2 or blocking anti-CD25 mAb to rat fetal thymus organ cultures (FTOC) under several experimental conditions. The IL-2 treatment initially stimulated the growth of thymocytes and, as a result, induced T cell differentiation, but the continuous addition of IL-2 to rat FTOC, as well as the anti-CD25 administration, resulted in cell number decrease and inhibition of thymocyte maturation. These results indicate that immature rat thymocytes bear functional high-affinity IL-2R and that IL-2 promotes T cell differentiation as a consequence of its capacity to stimulate cell proliferation. Modifications in TCR alpha beta repertoire and increased numbers of NKR-P1+ cells, largely NK cells, were also observed in IL-2-treated FTOC. Furthermore, IL-2-responsiveness of different thymocyte subsets changed throughout thymic ontogeny. Immature CD4-CD8-cells responded to IL-2 in two stages, early in thymus development and around birth, in correlation with the maturation of two distinct waves of thymic cell progenitors. Mature CD8+ thymocytes maximally responded to IL-2 around birth, supporting a role for IL-2 in the increased proliferation of mature thymocytes observed in vivo in the perinatal period. Taken together, these findings support a role for IL-2 in rat T cell development.

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PubMed1997

Immunity in fish larvae.

The route of immunization, dose and nature of antigens are relevant for induction of specific immune responses but they are particularly influenced by the state of immunomaturation of fish. It is difficult, however, to determine when the fish immune system has matured and, accordingly, when the fry can be successfully vaccinated. To establish some general parameters which can correlate the development of protective immunity with maturation of the immune system we review: (i) the appearance of first lympho-haemopoietic cell precursors; (ii) the histological and functional maturation of lymphoid organs; (iii) the relevance of phagocytosis and transport of maternal Ig to eggs for early vaccination; (iv) the appearance of both T-like and B-like cells and their correlation with the development of the cell-mediated and humoral immune responses and (v) the mechanisms which may be involved in the induction of tolerance following early immunization of fry. These data rather than a general rule for vaccination of larval fish indicate that the earliest age (size) that a fish can be vaccinated differs between species and vaccines.

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PubMed1997

Bronchial epithelium associated to lymphoid tissue does not selectively express vimentin.

The existence of a lymphoepithelium containing M cells in the bronchus-associated lymphoid tissue (BALT) of several species has repeatedly been questioned. In previous electron microscopical studies we failed to ultrastructurally identify these cells in the epithelium covering bronchial lymphoid tissue of adult rats. In the present study, we analyze immunohistochemically the expression of vimentin, an intermediate filament, reported to be a sensitive marker for rabbit M cells, in both BALT and Peyer's patches. Our results demonstrate, however, the absence of vimentin expression in the epithelium covering the bronchial lymphoid aggregates as well as in the lymphoepithelium of the Peyer's patches. On the contrary, both epithelia are strongly cytokeratin positive. Furthermore, numerous vimentin-positive lymphocytes appear in both lymphoid organs. Results are discussed from a view of the possible relationship between BALT and the so-called mucosae-associated lymphoid tissue (MALT).

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PubMed1996

Histogenesis of the epithelial component of rat thymus: an ultrastructural and immunohistological analysis.

BACKGROUND: Despite the assumed importance of thymic cell microenvironments for governing T-cell maturation, little is known about the ontogeny of their cell components. A few studies have analyzed previously the ontogenetical development of rat thymic epithelium (Bogojevic et al. 1990. Period. Biol., 92:126; Kampinga and Aspinall 1990 Harwood Acad. Pub., London, pp. 149-186; Micic et al., 1991 Dev. Comp. Immunol., 15:443-450) and recently we have reported the development of both interdigitating/dendritic cells and macrophages (Vicente et al., 1994 Immunology, 82:75-81, 1995 Immunology, 85:99-105). METHODS: In the present work we analyze in situ ultrastructural, immunohistochemical, and histoenzymatically the appearance and development of the thymic epithelial cell component in both embryonic and neonatal Wistar rats with special emphasis on the origin of the different epithelial cell types, the occurrence or absence of a common precursor for these, and the expression of MHC molecules. RESULTS: The thymic primordium of 13-day-old embryos is formed by a homogeneous population of primitive epithelial cells differentiating gradually into various epithelial cell subtypes of both the cortex and the medulla. In the cortex, subcapsular and stroma-supporting epithelial cells appear at days 14-15 as two structurally different cell entities. At the same time, stroma-supporting, keratinized, and vacuolated epithelial cells occur in the thymic medulla. These last two cell types differentiate subsequently into Hassall's bodies and hypertrophied cells. Lympho-epithelial cell complexes are identified in the deep cortex around birth, when the cortical parenchyma houses a transitional erythropoiesis. mAbs (His-39, RMC-20) which recognize medullary epithelial cells in the adult thymus stain positively cells of the thymic primordium as early as day 16 of embryonic life. Cortical epithelial cell markers (His-37, RMC-17) appear, however, slightly later and the subcapsulary region is not established until postnatal life. MHC class I and class II molecules can be identified on epithelial cells in the thymus of 15-day-old embryonic rats although they reach the highest expression around birth. CONCLUSIONS: Our results confirm the heterogeneity of the thymic epithelial component, the persistence of primitive, non-differentiated epithelial cells morphologically similar to those occurring in the early thymic primordium in adult thymus, and the mutual relevance of epithelial cells and thymocytes for an adequate development of rat thymus gland.

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PubMed1996

Expression of functionally active alpha 4 beta 1 integrin by thymic epithelial cells.

We have investigated the expression and function of the VLA-4 heterodimer alpha 4 beta 1, a member of the beta 1 integrin subfamily, on human thymic epithelial cells (TEC) derived from cortical epithelium. The expression of the alpha 4 integrin chain was studied in four different cloned TEC lines derived from either fetal or post-natal human thymus by both flow cytometry and immunoprecipitation techniques with anti-alpha 4 MoAbs. All different cell lines assayed expressed significant levels of alpha 4, as revealed by their reactivity with MoAbs specific for distinct alpha 4 epitopes. The alpha 4 subunit expressed by TEC was associated to beta 1 but not to beta 1 chain, and displayed the characteristic 80/ 70 kD pattern of proteolytic cleavage. The VLA-4 integrin in these cells was constitutively active in terms of adhesiveness to both fibronectin and vascular cell adhesion molecule-1 (VCAM-1). In addition, this heterodimer localized to punctate regions of the cell in the area of contact with the substratum, named point contacts assessed by staining with the anti-beta 1 activation epitope 15/7 MoAb. According to the cortical origin of the TEC lines expressing VLA-4, human thymus sections stained with different anti-alpha 4 antibodies revealed the presence of cortical, and in smaller numbers medullary epithelial cells bearing alpha 4 integrin. The expression of alpha 4 in the thymus was also found in both adult and fetal rats, in which epithelial cells were also specifically stained. Altogether, our data show that VLA-4 is an additional component of the integrin repertoire of TEC, and suggest that it could have an important role in thymus epithelial cell-thymocyte interactions.

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PubMed1996

In vitro and in situ characterization of fish thymic nurse cells.

We present an enzyme- and immuno-cytochemical, and ultrastructural characterization of trout thymic nurse cells (TNCs). Our data suggest that isolated trout thymic multicellular complexes are epithelial cells with acidic compartments that may be involved in the processing of antigens and in the generation of the MHC-II proteins that these cell express, and also that isolated TNCs are the in vitro equivalent of the pale and intermediate electronlucent epithelial cells located in the inner zone of the trout thymus, constituting indirect evidence of the phylogenetical relationships of the inner zone of the teleost thymus with the thymic cortex of higher vertebrates.

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PubMed1996

Increased numbers of CD5+ B cells in the thymus of estradiol benzoate-treated rats.

In the present work we combine both flow cytometry and in situ immunohistochemical techniques to study the changes affecting a minor B cell population described within the normal rat thymus, after treatment with estradiol benzoate (EB). Our results, in agreement with previous data, show that the vast majority of these intrathymic B cells are CD5+. The existence of CD5+ B cells was confirmed flow cytometrically in both cervical lymph nodes and spleen of control, adult Wistar rats. Moreover, after EB administration intrathymic B cells increased significantly especially in those rats receiving 500 micrograms of EB, constituting cell masses around the blood vessels of cortico-medullary area and in the thymic medulla. We discuss the significance of this increased number of intrathymic CD5+ B cells, which is probably due to a selective cell migration from the periphery into the thymus, from the view of the effects of estradiol on the thymic vascular permeability.

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PubMed1995

T-dependent areas in the chicken bursa of Fabricius: an immunohistological study.

BACKGROUND: The so-called diffusely infiltrated lymphoid tissue of the chicken bursa of Fabricius was previously described as a T-dependent bursal area. METHODS: We have analyzed immunohistologically its postnatal development by using a battery of mAbs, most of them raised specifically to chicken T cells, in order to characterize phenotypically the nature of its cell components, appearance, development, and possible functional significance. RESULTS: Our results demonstrate that this tissue represents poorly developed lymphoid aggregates, the cell content reminiscent of that found in other lymphoid tissues occurring throughout the avian gut. The first lymphoid elements appear in this bursal area only after hatching, growing rapidly to reach the adult condition in the second week of postnatal life. They consist mainly of T lymphocytes, including principally. CD4+ TcR alpha V beta 1+ cells, which form cell groups and CD8+ cells, and TcR gamma delta+ lymphocytes, which predominate in the subepithelial area and within the epithelium. MHC Class II molecule-expressing cells and IgM-, IgG-, and IgA-positive B lymphocytes also occupy the subepithelial region. CONCLUSIONS: We propose that the bursal diffusely infiltrated lymphoid tissue of the chicken represents gut-associated lymphoid tissue involved in mounting immune responses to antigens that reach the bursal lumen via the cloaca.

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PubMed1995

Macrophage-lymphocyte cell clusters in the hypothalamic ventricle of some elasmobranch fish: ultrastructural analysis and possible functional significance.

BACKGROUND: Previous studies have demonstrated the existence of lympho-haemopoietic tissue in the meninges and choroid plexuses of various primitive vertebrates, including the stingray Dasyatis akajei and in early human embryos. In the present study, we extend these results analyzing macrophage-lymphocyte cell clusters found in the floor of the hypothalamic ventricle of several specimens of elasmobranchs. METHODS: After aseptical isolation of the brain from several specimens of smooth dogfish Triakis scyllia, cloudy dogfish Scyliorhinus torazame, gummy shark Mustelus manazo, and stingray Dasyatis akajei their hypothalamic regions were processed routinely by light, scanning, and transmission electron microscopy. RESULTS: The study of serial histological sections demonstrated that the macrophage-lymphocyte cell clusters proceeded from the meningeal lymphohaemopoietic tissue, reaching the ventricular lumen along large blood vessels. In this tissue, macrophages, different sized lymphocytes, lymphoblasts, granulocytes, monocytes, and developing and mature plasma cells were closely packed among a meshwork of fibroblastic reticular cell processes. It never invaded the brain parenchyma. A cell layer of glial elements and a continuous basement membrane interposed between the lymphoid tissue and the neural elements although some macrophages had migrated across the ependymal cell layer. In the ventricular lumen very irregular macrophages with long cell processes and containing abundant engulfed material of unknown origin formed big cell clusters with neighboring lymphocytes, lymphoblasts, and plasma cells, similar to those described during the immune response. Moreover, electron lucent cells which resembled the antigen-presenting cells of higher vertebrates established intimate surface cell contacts with the surrounding lymphocytes. In the third ventricle of several specimens of gummy shark, Mustelus manazo, morphologically similar cell clusters appeared but these were not connected to the meningeal lympho-haemopoietic tissue. No intraventricular cell aggregates were found in the stingray brain. CONCLUSIONS: Although we cannot rule out that these macrophage-lymphocyte cell clusters represent a permanent structure in the elasmobranch brain they rather seem to be only established after specific stimulation for preventing the entrance of noxious, foreign materials into the elasmobranch brain parenchyma.

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PubMed1995

Changes in the blood-thymus barrier of adult rats after estradiol-treatment.

The accessibility of the thymus parenchyma for relatively large Mw (+/- 150 Kd) proteins has been studied by the intravenous injection of monoclonal antibodies (mAbs) specific either for all T cells (His-17) or MHC class II molecules (His-19) in control and estradiol benzoate (EB)-treated adult Wistar rats. In controls, the transcapsular route rather than cortical capillaries seems to be involved in the entry of molecules into the thymus. By contrast, a specific staining for either T cells (His-17) or MHC class II molecules (His-19 positive cells) disappears almost completely from the thymic cortex of EB-treated rats except in the immediate subcapsular epithelial cell layer. In these rats, T cells and epithelial cells intimately associated to blood vessels from both inner cortex and corticomedullary border showed additional staining with the respective mAbs confirmed by electron microscopy. The disappearance of the transcapsular route together with the increased vascular permeability of cortical blood vessels would be related to the reinforcement of the subcapsular epithelial cell layer and to direct effects of EB on vascular endothelia, respectively. These results are discussed in relationship to the cell migration into and out of adult thymus, as suggested by the changes in intrathymic T cell subsets evaluated by flow cytometry.

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PubMed1995

T-cell development in early partially decapitated chicken embryos.

We have evaluated the immunohistological and cytofluorometric changes that occur in the thymus of chicken embryos partially decapitated at 33-38 hr of incubation (DCx embryos) in an attempt to analyze possible neuroendocrinological influences on T-cell differentiation and, indirectly, the ontogeny of the so-called neuroendocrine-immune network. The thymus of DCx embryos shows important variations that profoundly and selectively affect different T-cell subsets, but not the nonlymphoid cell components of thymic stroma. These modifications include the accumulation of cell precursors, mainly DN (CD4-CD8-) cells and immature CD8lowCD4- cells, which expand but do not differentiate, resulting in an extreme decline of both DP (CD4+ CD8+) cells and TcR alpha beta-expressing cells. Accordingly, both subcapsulary and outer cortex increase in size, whereas the deep cortex and principally the thymic medulla almost disappear in DCx embryos. In contrast, other T-cell subsets of DCx embryos, largely CD8highCD4- cells and TcR gamma delta-expressing cells do not undergo significant variations throughout thymic ontogeny.

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PubMed1995

The relevance of cell microenvironments for the appearance of lympho-haemopoietic tissues in primitive vertebrates.

In higher vertebrates, mainly in mammals, a role for the non-lymphoid components of lymphoid organs in governing the maturation and functioning of immune system has been largely demonstrated. In contrast, such a role in the evolution of the vertebrate immune system has only been evidenced indirectly. In the present review we summarize histophysiological results which emphasize the relevance of lympho-haemopoietic stromal elements in the emergence and evolution of vertebrate lymphoid organs. The most primitive vertebrates, the Agnatha, have no true lymphoid organs and, accordingly, their immune responses seem more related to the non-anticipatory defence mechanisms of invertebrates than to the immune responses of vertebrates. So, the appearance and evolution of vertebrate lymphoid organs seems closely related with the emergence of immune capacities. Thymus, spleen and gut-associated lymphoid organs appear early in phylogeny whereas lymph nodes and bone marrow are late phylogenetical adquisitions. However, bone marrowless vertebrates contain numerous organs (i.e., gonads, kidney, brain, etc...), the cell microenvironments of which support lympho-haemopoiesis mimicking the condition of higher vertebrate bone marrow. On the other hand, the lack of germinal centres, another feature of the lymphoid organs of ectothermic vertebrates which impedes the selection of B cells raised after somatic hypermutation, presumably reflects the absence of some of the elements necessary for this organization.

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PubMed1995

Ontogeny of rat thymic macrophages. Phenotypic characterization and possible relationships between different cell subsets.

In the present study we combined electron microscopy, immunohistology and primary stromal cell cultures to analyse the ontogeny of rat thymic macrophages (M phi) in an attempt to clarify the relationships between the different macrophage cell subsets described in adult rat thymus. Although phagocytic cells were observed in 15-day-old fetal thymus, monoclonal antibodies (mAb) which recognize different adult macrophage types were unable to identify positive cells until the end of embryonic life. However, our in vitro results from primary thymic stromal cell cultures of 16-day-old fetal rats, and the phenotyping of enriched thymic CD2- cell suspensions, demonstrated that monocyte-like cells which strongly expressed major histocompatibility complex (MHC) class II molecules colonized the embryonic thymus early, giving rise later to distinct macrophage subsets. During the process of maturation, macrophage precursors gradually lost their MHC class II expression, acquired other surface markers (CD45, Thy-1, CD25, CD4, etc.) and increased the acid phosphatase activity. In this respect, ED1+ macrophages, which appeared for the first time in the last stages of embryonic life, consisted of a MHC class II molecule-expressing phagocytic cell population, presumably involved in the elimination of non-selected cortical thymocytes, and of non-phagocytic cells which, in the thymic cortex, might differentiate to ED2+ macrophages throughout ED1+ED2lo/med and ED1+ ED2high intermediate cell stages, observed in vitro in 16-day-old fetal thymic stromal cell cultures. At the end of embryonic life and during the postnatal period the numbers of thymic macrophages increased, particularly in the medulla and corticomedullary border (CMZ), and more slowly in the thymic cortex. This increase was presumably due to the arrival, through perivascular spaces, of new macrophage progenitors, rather than in situ proliferation of pre-existent mature macrophages. The possible function of different thymic macrophage subsets, as well as the relationships between themselves and with their presumptive monocyte-like precursors, are discussed.

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PubMed1994

Ultrastructural changes in the adult rat thymus after estradiol benzoate treatment.

Although numerous authors have correlated high levels of circulating estrogens with thymic involution, a systematic analysis to date on the histological changes affecting the thymus gland in that situation is lacking. In the present study we report both histological and ultrastructural changes occurring in the thymus of adult Wistar rats which received a single dose either of 100 micrograms or 500 micrograms of estradiol benzoate. Both doses induced thymic involution which correlated well with histological changes observed in the lymphoid populations but also with profound modifications in the thymic epithelial component. Moreover, intrathymic erythro-and granulopoiesis, increased numbers of both macrophages and plasma cells, and important variations in the thymic vascular permeability occurred in estradiol benzoate treated rats. These results are discussed from the perspective that changes in both the non-lymphoid cell components of thymic microenvironments and vascular permeability are essential to understand the general effects of sex steroids on the immune system.

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PubMed1994

Prolactin and early T-cell development in embryonic chicken.

Currently, the information available on the ontogeny of the immune-neuroendocrine network is somewhat scant. However, increasing data suggest a role for prolactin, an important immunoregulatory molecule that resembles cytokines, in early T-cell development. In this article, Javier Moreno and colleagues present evidence for this role using the chicken embryo as a model system.

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