Biomedical subjects

C Martinez-A

Publications and source records attributed to C Martinez-A.

At least 37 records · Page 2Linked to original sources

PubMed1997

Lck is necessary and sufficient for Fas-ligand expression and apoptotic cell death in mature cycling T cells.

Apoptotic cell death is induced in mature cycling T cells upon ligation of the Ag-specific TCR. This process is essential for the maintenance of homeostasis in the immune system, as it is capable of down-regulating ongoing immune responses. The analysis of the mechanism underlying TCR-induced programmed cell death has focused the attention of many scientists recently. In this regard, several recent reports have implicated Fas/Fas-ligand molecules as the final mediators of this process. Several other gene products have been implicated in the control of apoptosis (as Bcl-2, p53, and c-Myc); however, no information was available in the early signaling molecules that trigger this phenomena. The results presented in this work indicate that pp56(lck) src family kinase is actually required for the TCR to trigger cell death in mature cycling T cells. In fact, while inhibition of pp56(lck) expression with antisense oligonucleotides blocked TCR-induced apoptosis, pharmacologic inhibition of phosphatidylinositol 3-kinase activity had no effect. Accordingly, ligation of the Ag receptor in a cell line defective for pp56(lck) expression was unable to induce apoptosis, although it induced cellular stimulation, as measured by the expression of CD69. In addition, we show in this work that expression of constitutively active pp56(lck) mutants, but not pp59(fyn) mutants, in the absence of any other TCR-derived signal, is sufficient to induce apoptosis not only in transformed, but also in normal cycling T cells. Finally, evidence is presented indicating that a mechanism through which pp56(lck) regulates TCR-induced apoptosis in mature cycling T cells is by controlling Fas-ligand expression.

Apoptosis↗

PubMed1997

Intermediate affinity interleukin-2 receptor mediates survival via a phosphatidylinositol 3-kinase-dependent pathway.

Peripheral blood T lymphocytes require two signals to enter and progress along the cell cycle from their natural quiescent state. The first activation signal is provided by the stimulation through the T cell receptor, which induces the synthesis of cyclins and the expression of the high affinity interleukin-2 receptor. The second signal, required to enter the S phase, is generated upon binding of interleukin-2 to the high affinity alphabetagamma interleukin-2 receptor. However, resting T cells already express intermediate affinity betagamma interleukin-2 receptors. As shown here, T cell stimulation through intermediate affinity receptors is capable of inducing cell rescue from the apoptosis suffered in the absence of stimulation. Characterization of the signaling pathways utilized by betagamma interleukin-2 receptors in resting T cells, indicated that pp56(lck), but not Jak1 or Jak3, is activated upon receptor triggering. Compelling evidence is presented indicating that phosphatidylinositol 3-kinase associates with the intermediate affinity interleukin-2 receptor and is activated upon interleukin-2 addition. Bcl-xL gene was also found to be induced upon betagamma interleukin-2 receptor stimulation. Finally, pharmacological inhibition of phosphatidylinositol 3-kinase blocked both interleukin-2-mediated bcl-xL induction and cell survival. We conclude that betagamma interleukin-2 receptor mediates T-cell survival via a phosphatidylinositol 3-kinase-dependent pathway, possibly involving pp56(lck) and bcl-xL as upstream and downstream effectors, respectively.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1997

Conformational changes required in the human growth hormone receptor for growth hormone signaling.

Growth hormone (GH) plays a significant role in normal growth and development. Signaling to the cell is believed to require growth hormone receptor (GHR) dimerization, which occurs following binding of a single growth hormone molecule to each of two receptors. We have developed human growth hormone receptor-specific monoclonal antibodies, one of which was used here to characterize hormone/receptor interactions. This antibody, GHR05, is directed against the hinge spanning subdomains I and II of the receptor's extracellular region. Antibody binding to the cell surface receptor increases upon receptor binding to growth hormone, but not when it binds a mutant form, hGHG120R, which does not trigger receptor activation. Growth hormone binding thus appears to lead to a conformational change in the receptor epitope recognized by GHR05, giving rise to the active dimer configuration, necessary for signal transduction. Using a chimeric receptor-expressing, growth hormone-dependent murine cell line, we find that GHR05 binds to the receptor in the absence of human GH and delivers a signal leading to cell proliferation. Finally, GHR05 treatment of IM-9 cells, a human cell line expressing a functional human GHR, leads to cell proliferation mediated by the generation of GH-specific signals, including phosphorylation of the JAK2 tyrosine kinase and activation of STAT5.

Animals↗

PubMed1996

Eosinophil recruitment to the lung in a murine model of allergic inflammation. The role of T cells, chemokines, and adhesion receptors.

Eosinophil accumulation is a distinctive feature of lung allergic inflammation. Here, we have used a mouse model of OVA (ovalbumin)-induced pulmonary eosinophilia to study the cellular and molecular mechanisms for this selective recruitment of eosinophils to the airways. In this model there was an early accumulation of infiltrating monocytes/macrophages in the lung during the OVA treatment, whereas the increase in infiltrating T-lymphocytes paralleled the accumulation of eosinophils. The kinetics of accumulation of these three leukocyte subtypes correlated with the levels of mRNA expression of the chemokines monocyte chemotactic peptide-1/JE, eotaxin, and RANTES (regulated upon activation in normal T cells expressed and secreted), suggesting their involvement in the recruitment of these leukocytes. Furthermore, blockade of eotaxin with specific antibodies in vivo reduced the accumulation of eosinophils in the lung in response to OVA by half. Mature CD4+ T-lymphocytes were absolutely required for OVA-induced eosinophil accumulation since lung eosinophilia was prevented in CD4+-deficient mice. However, these cells were neither the main producers of the major eosinophilic chemokines eotaxin, RANTES, or MIP-1alpha, nor did they regulate the expression of these chemokines. Rather, the presence of CD4+ T cells was necessary for enhancement of VCAM-1 (vascular cell adhesion molecule-1) expression in the lung during allergic inflammation induced by the OVA treatment. In support of this, mice genetically deficient for VCAM-1 and intercellular adhesion molecule-1 failed to develop pulmonary eosinophilia. Selective eosinophilic recruitment during lung allergic inflammation results from a sequential accumulation of certain leukocyte types, particularly T cells, and relies on the presence of both eosinophilic chemoattractants and adhesion receptors.

Animals↗

PubMed1996

Distinct expression and function of the novel mouse chemokine monocyte chemotactic protein-5 in lung allergic inflammation.

We have cloned a novel mouse CC chemokine cDNA from the lung during an allergic inflammatory reaction. The protein encoded by this cDNA is chemotactic for eosinophils, monocytes, and lymphocytes in vitro and in vivo. Based on its similarities in sequence and function with other CC chemokines, we have named it mouse monocyte chemotactic protein-5 (mMCP-5). Under noninflammatory conditions, expression of mMCP-5 in the lymph nodes and thymus is constitutive and is generally restricted to stromal cells. Neutralization of mMCP-5 protein with specific antibodies during an allergic inflammatory reaction in vivo resulted in a reduction in the number of eosinophils that accumulated in the lung. Moreover, mMCP-5 mRNA expression in vivo is regulated differently from that of other major CC chemokines in the lung during the allergic reaction, including Eotaxin. The presence of lymphocytes is essential for expression of mMCP-5 by alveolar macrophages and smooth muscle cells in the lung, and the induction of mMCP-5 RNA occurs earlier than that of the eosinophil chemokine Eotaxin during allergic inflammation. In contrast to Eotaxin, mRNA for mMCP-5 can be produced by mast cells. From these results, we postulate that mMCP-5 plays a pivotal role during the early stages of allergic lung inflammation.

Amino Acid Sequence↗

PubMed1996

The catalytic domain of pp56(lck), but not its regulatory domain, is sufficient for inducing IL-2 production.

The lymphoid src kinase pp56(lck) has been shown to be essential for the induction of different T lymphocyte responses, including CD4-mediated enhancement of Ag-induced T cell activation, early T cell differentiation, induction of IL-2 production, and cytotoxicity. It is assumed that pp56(lck) acts on these processes by phosphorylating substrates. However, it has been recently reported that the NH2 regulatory domain is sufficient to mediate CD4 accessory function. In this report we address the contribution of the regulatory and catalytic domains of pp56(lck) to another function of this enzyme independent of CD4: TCR-induced IL-2 production. Two pp56(lck) mutants lacking either the entire catalytic domain or the entire NH2 regulatory domain were generated, and their abilities to trigger transactivation of the TCR-regulated nuclear factor of activated T cells (NF-AT) region of the IL-2 promoter were compared. Only the catalytic, but not the NH2 regulatory, domain of pp56(lck) was able to induce NF-AT region transactivation on its own and to cooperate with other intracellular signals to trigger this response. Moreover, the catalytic domain of pp56(lck) was able to induce IL-2 cytokine production to an extent similar to that of wild-type pp56(lck). We conclude that different domains of the pp56(lck) molecule contribute to regulate distinct biologic functions. In fact, while the NH2 regulatory domain is sufficient to mediate CD4 accessory function, we show here that the catalytic domain of pp56(lck) is sufficient for induction of IL-2 production, mimicking TCR ligation.

Allosteric Site↗

PubMed1996

Enterotoxin septic shock protection and deficient T helper 2 cytokine production in growth hormone transgenic mice.

Neuroendocrine hormones have long been thought to play a role in lymphoid development and function. In particular, growth hormone has been shown to mediate thymic development as well as to promote T cell engraftment in severe combined immunodeficiency mice. Murine T helper cells are classified into two subsets based on their cytokine production pattern. Here, we report that transgenic mice for bovine growth hormone show significant alterations in T cell function and decreased capability for cytokine production, an effect that is more acute in T helper cells as measured by their inability to produce IL-4 upon in vivo injection with Staphylococcus aureus enterotoxin B. Furthermore, upon immunization with conventional Ags, growth hormone transgenic mice produce an altered Ig isotype pattern characterized by a response shift from IgG1 in nontransgenic mice to IgG2 in transgenic mice. The impaired T cell responses correlated with survival from septic shock mediated by bacterial enterotoxins. We conclude that growth hormone may have the potential of regulating immune responses in pathologic processes associated with hyperactivation of T cells or with massive cytokine production.

Animals↗

PubMed1996

Linomide prevents the lethal effect of anti-Fas antibody and reduces Fas-mediated ceramide production in mouse hepatocytes.

Fas is an apoptosis-signaling receptor molecule expressed in vivo on thymocytes, liver, heart, and ovary. In vivo administration of the anti-Fas Jo2 antibody in mice induces severe apoptotic liver damage leading to fulminant hepatitis and death. Linomide, a quinoline 3-carboxamide, inhibits apoptosis of B and T cells induced by various stimuli including viruses, superantigens, and glucocorticoids. Mice treated with linomide survived the lethal effect of anti-Fas antibody, did not accumulate ceramide in hepatocytes, and recovered liver structure and function within 96 h of anti-Fas injection, as confirmed by histology and glutamic oxalacetic transaminase, glutamic pyruvic transaminase, and lactate dehydrogenase levels. Surviving mice showed severe depletion of cortical thymocytes, but medullar thymic cells expressing high CD3 and Fas levels also survived the treatment with anti-Fas in the presence of linomide. Heart, lung, and ovary showed no signs of apoptosis promoted by Fas ligation. These results suggest that linomide prevents cell death triggered by Fas ligation and can be useful for therapeutic intervention in fulminant hepatitis.

Animals↗

PubMed1996

CD2 ligation abrogates antigen-independent apoptosis in B cells.

We have described recently the prevention of apoptosis by CD2-soluble CD48 interaction on antigen B cell receptor occupancy. Here, we show that CD2 ligation is also able to interfere with B cell receptor-independent apoptosis pathways such as spontaneous death in spleen B cells or serum deprivation and hydrogen peroxide exposure in the BAL-17 cell line. In all cases, CD2 ligation induces a signal that prevents the downregulation of Bcl-2 expression. The specific CD2 signal pathway involved in this phenomenon is still unknown. As reported, CD2 did not appear to induce Ca2+ mobilization, phosphatidylinositol turnover, or PKC translocation in B cells. Nevertheless, we show that CD2 receptor ligation is coupled to the tyrosine phosphorylation pathway in B cells. These observations indicate that CD2 is functionally able to trigger at least an early signal that could play a role in apoptosis blockage B cells in addition to the adhesion one. The results suggest the participation of cellular membrane receptors other that CD40 in apoptosis rescue, not only in the antigen-dependent but also in the antigen-independent phases of B cell lymphopoiesis.

Animals↗

PubMed1996

Phosphatidic acid generation through interleukin 2 (IL-2)-induced alpha-diacylglycerol kinase activation is an essential step in IL-2-mediated lymphocyte proliferation.

Proliferation of T lymphocytes is triggered by the interaction of interleukin 2 (IL-2) with its high affinity specific receptor that is expressed on the cell surface following T lymphocyte activation. Significant advances have recently been made in identifying the multiple signals that follow IL-2 receptor occupancy, although the exact mechanism responsible for IL-2-induced proliferation remains an enigma. It has been shown previously that unique species of phosphatidic acid are rapidly produced in vivo following IL-2 binding. It was then suggested that, in contrast to other eukaryotic growth factor systems, phosphatidic acid was at least in part generated through IL-2-induced diacylglycerol (DG) kinase activation. In the present study we demonstrate IL-2-dependent activation of the alpha isoform of DG kinase. Confocal microscopy studies reveal that the enzyme is located in the cytosol and nuclei of resting T cells. Interleukin 2 stimulation induces translocation of the enzyme to the perinuclear region. Furthermore, our results indicate that inhibition of the alpha isoform of DG kinase has a profound effect on IL-2-induced T cell growth. Studies on cell cycle distribution demonstrate that the inhibition of IL-2-induced phosphatidic acid production induces arrest in late G1 phase of IL-2 dependent cells. Altogether, these results link previous observations of interleukin 2 and phosphatidic acid production to activation of an specific isoform of DG kinase and suggest that activation of this enzyme is part of a novel signaling cascade that utilizes phosphatidic acid as an effector molecule.

Cell Compartmentation↗

PubMed1996

Decrease in cAMP levels promoted by CD48-CD2 interaction correlates with inhibition of apoptosis in B cells.

The authors recently reported that CD2 ligation rescues B cells from antigen-induced apoptosis by upregulation of intracellular Bcl-2 levels. However, the characterization of the early signals involved in apoptosis rescue by CD2 ligation has not been well established. In this context, CD2 does not promote either phosphatidylinositol turnover or CA2+ mobilization in B cells. In this paper the authors show that CD2 interaction with its ligand CD48 also reduces the apoptosis induced by forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine and, to a much lesser extent, the apoptosis induced by cholera toxin in murine B splenocytes. Using a cAMP detection system sensitive to the picomolar range, the authors demonstrate that CD2-CD48 interaction decreases the intracellular cAMP concentrations induced by forskolin but not by cholera toxin. In comparison with the CD2-CD48 interaction, CD40-CD40 ligand interaction completely inhibits the apoptosis induced by cAMP increases without affecting the intracellular cAMP levels promoted by forskolin or cholera toxin. These results indicate that CD2 can also control the apoptosis at the very early steps after receptor signalling, such as the adenylate cyclase activity. Given that heterotrimeric G-proteins can mediate the adenylate cyclase activity the authors suggest that CD2 signalling could act through these small proteins, which would explain the inability of CD2 signalling to rescue from the apoptosis induced by cholera toxin, a Gs-protein activator. Conversely, CD40 seems to control apoptosis further downstream of the cAMP-PKA pathway where the survival and apoptotic signals are confluent, which might therefore render it a more efficient system to block apoptosis.

Animals↗

PubMed1994

Age-dependent changes in the response to staphylococcal enterotoxin B.

In the present study we investigated the response of old mice to staphylococcal enterotoxin B (SEB) immunization. Old mice were susceptible to lethal toxic shock, probably mediated by tumor necrosis factor-alpha, although lethal toxic shock was not observed in young mice. Old mice were able to produce more IL-2 and IL-4 than young mice in response to in vivo immunization with SEB. V beta 8+CD4+ T cells of old mice expanded less in vivo and were not deleted in response to SEB. However, in spite of the absence of clonal deletion, SEB was found to induce energy of SEB reactive cells in old mice, as demonstrated by reduced in vitro T cell proliferation to SEB and reduced in vitro IL-2 and IL-4 production.

Aging↗

PubMed1990

Inside the thymus, Mls antigen is exclusively presented by B lymphocytes.

The ability to stimulate an Mls-1 mixed lymphocyte reaction (MLR) is predominantly expressed by low density B lymphocytes in the spleen and peritoneal cavity of normal adult mice, and is absent in splenic B cells 1 month after lethal irradiation and reconstitution from autologous bone marrow. Coreconstitution of these mice with normal syngeneic peritoneal cells restores the stimulatory potential of splenic B cells, but sorted CD5+ or CD5- IgM+ lymphocytes from peritoneum are equally good stimulators, suggesting that functional Mls-1 expression may require long life spans and selection. Bone-marrow-reconstituted DBA/2 mice that fail to express Mls-1 antigens in the periphery nevertheless maintain T-cell receptor V beta 6 and 8.1 deletions among the newly formed T cells. These findings led us to directly investigate the Mls stimulatory ability of purified antigen-presenting cell populations inside the thymus. We report here that thymic B lymphocytes seem to represent the only intrathymic cell population able to stimulate Mls-1 MLR.

Animals↗

PubMed1989

Further evidence for coelomic-associated B lymphocytes.

Previous observations indicate that "CD5 B lymphocytes" are preferentially clustered in gut-related mesenchymal areas, such as peritoneum, thymus and tonsils. We have now found that pleuropericardial spaces contain an homogeneous population of large-sized, noncycling, nonsecretory B cells, expressing very high levels of surface IgM, little or no IgD, Mac-1 and low levels of B220. This phenotype and the over-representation of some antibody clonotypes suggest that the pleuropericardial cavity contains a pure "CD5 B cell" population. In all mouse strains analyzed, however, many of these cells are CD5-. These findings, together with the common origin of peritoneum and pleural layers in the primitive coelomic cavity, suggest that such B cells differentiate locally from intraembryonic precursors; we propose to designate them as "coelomic", to distinguish them from "stromal", bone marrow-derived B cells.

Animals↗

PubMed1987

The role of I-A/E molecules in B-lymphocyte activation. II. Mechanism of inhibition of the responses to lipopolysaccharide by anti-I-A/E antibodies.

We have previously demonstrated that monoclonal anti-I-A/E antibodies inhibit B-cell responses to lipopolysaccharide (LPS). In the present report, the inhibitory effects were shown to be carried out directly on B cells, and to be totally independent of the LPS concentration used, thereby showing that antibodies do not mediate their effect through blocking of accessory cells or steric hindrance of LPS-receptors. Of the three different phases in B-cell activation/induction, proliferation, and maturation, induction was shown to be the most sensitive to inhibition by anti-I-A/E antibodies. Thus, kinetic studies showed that anti-I-A/E antibodies are only inhibitory for the first 16 h of LPS activation, after which B cells can no longer be inhibited by these antibodies. Class II MHC molecules appear, therefore, to be part of a membrane molecular complex which regulates delivery of activation signals to resting B cells. Since it was also shown that this time period corresponds approximately to the time required for B cells to express functional reactivity to growth factors, we suggest that anti-I-A/E antibodies act on resting B lymphocytes to inhibit mitogen-dependent induction of growth receptor expression.

Animals↗

PubMed1987

Idiotypic multireactivity of 'natural' antibodies. 'Natural' anti-idiotypes also inhibit helper cells with cross-reactive clonotypes.

One hundred and twenty IgM-secreting hybridomas derived from unmanipulated 6-day-old BALB/c mice were screened for reactivity with the prototype idiotypic and anti-idiotypic monoclonal antibodies, defining three well established systems, namely TEPC 15:10/13-15, J558:CD3.2, and MOPC 460:F 6(51). Up to 25% of all IgM antibodies reacted with at least one of the six specific ligands, half of the latter being 'monospecific', the others reacting with two or more antibodies. A detailed analysis of the four most multi-reactive clones showed individually specific patterns of reactivity and revealed reactions of the same IgM molecule in idiotypic systems previously studied independently. Furthermore, when tested for functional interactions with syngeneic helper T cells expressing MOPC 460-like clonotypes, one of these antibodies was found to inhibit effector helper activity. The results show the existence of 'natural antibodies' with idiotypic reactivities related to recurrent clonotypes in the strain. They may be either 'specific' or 'multireactive', and might connect idiotypes on T and B cells and on antigenic systems so far studied independently.

Animals↗

PubMed1986

The basis for major histocompatibility complex (MHC) and immunoglobulin gene control of helper T cell idiotopes.

BALB/c helper T cells, prepared against 2,4,6-trinitrophenyl (TNP)-derivatized syngeneic spleen cells, fail to recognize other hapten modifications [(4-hydroxy-3-nitrophenyl)acetyl, fluorescein isothiocyanate] of syngeneic cells, as well as TNP-derivatized cells from major histocompatibility complex-congenic donors. T helper cell interactions with "presenting" cells and "target" B lymphocytes are inhibited by monoclonal antibodies directed either to the hapten or to I-A molecules on target cells. Helper activity is also inhibited by monoclonal anti-idiotypic antibodies directed to an idiotope expressed by the TNP-binding myeloma protein MOPC460, but not by soluble TNP-protein conjugates. The study of congenic mouse strains revealed that while the fine specificity of TNP recognition and the quantitative levels of M 460 idiotope expression are I-A linked, the expression of the M 460 idiotope by helper cells is controlled by IgCh-linked genes. Absence of anti-idiotope inhibition of helper cells prepared in anti-mu-suppressed mice suggests, however, that immunoglobulin idiotype expression by T cells results from network interactions selecting available lymphocyte repertoires which operate before antigenic stimulation.

Animals↗

PubMed1986

Differentiation of human mature thymocytes: existence of a T3+4-8- intermediate stage.

A T3 complex-bearing subpopulation was characterized within an in vivo cycling T4-8- early thymocyte compartment which contains cells constitutively expressing interleukin 2 and transferrin receptors. We show differentiation in vitro of both mature subsets of thymocytes (T3+4+8- and T3+4-8+) from the above T4-8- compartment, their appearance being preceded by cells in a T3+4-8- intermediate stage. Furthermore, those mature thymocytes generated in vitro contain functionally competent cells which use T3, T4 and T8 structures for their cytolytic activity. The finding of T3+4-8- thymocytes in vivo, together with the observation that T3 antigen expression precedes that of T4 or T8 molecules in vitro, shows that T3 (and presumably Ti) is present early in ontogeny, and suggests that T3+4-8- cells constitute an "intermediate" stage relevant to the connection between early precursors and mature thymocytes during T lymphocyte ontogeny.

Antigens, Differentiation, T-Lymphocyte↗