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M Sasportes

Publications and source records attributed to M Sasportes.

At least 37 records · Page 2Linked to original sources
PubMed1991

Perforin and granzyme B: predictive markers for acute GVHD or cardiac rejection after bone marrow or heart transplantation.

Monitoring of human allografts requires to use histological, immunohistochemical and functional techniques to characterize graft infiltrating cells. Granzyme B and perforin gene expression is of major importance in functional studies. Those proteins are present in the cytoplasmic granules of cytotoxic T lymphocytes and are secreted during granule exocytosis at the effector/target cell interface. Gene expression of both proteins has been studied by in situ hybridization using specific riboprobes on serial sections of biopsies in two pathological models. Our results show that cells infiltrating early skin lesions of patients with acute GVHD after bone marrow graft are exclusively composed of T cells, among which some of them express granzyme B and perforin genes. Similarly the presence of granzyme B and perforine-expressing cells in endomyocardial biopsies of heart transplanted patients has been associated to early and severe crisis of rejection. In contrast, the absence of functional markers in lymphoid infiltrates was coinciding with less aggressive and late episodes of rejection. Taken together, our data indicate that granzyme B and perforin gene expression in skin infiltrating lymphocytes during GVH or within heart infiltrating cells during crisis of rejection are in favor of severe processes. The study has allowed to predict during heart transplantation the apparition of a rejection crisis and to show the necessity for treating the patient with immunsuppresive drugs. This is also the case for patients with GVHD at the time of the first skin rash.

Acute Disease↗
PubMed1990

Structural organization of the hCTLA-1 gene encoding human granzyme B.

Cytotoxic T lymphocytes (CTLs) and natural killer/lymphokine-activated cells produce granzymes, a family of serine esterase proteins located in cytoplasmic granules. These might be involved in different cytotoxic pathways. We report the structural organization of the human gene encoding granzyme B (hCTLA-1). A 4.75-kb genomic DNA fragment containing all the sequences of granzyme B-encoding cDNA clones has been sequenced. The gene is composed of five exons and four introns. A comparison with the genomic organization of murine CCP1/CTLA-1 showed very similar structure and a 76% nucleotide homology in the coding sequences. This suggests that both genes may have a common ancestor. No typical regulatory element was detected in the 1160 bp upstream from the ATG start codon. The detection of a second locus related to hCTLA-1 is also described.

Amino Acid Sequence↗
PubMed1990

Granzyme B-gene expression: a marker of human lymphocytes "activated" in vitro or in renal allografts.

Human killer cells contain cytoplasmic granules in which serine esterases, or granzymes, and perforin have been identified. We have studied the induction of granzyme B-gene expression in peripheral blood lymphocytes and large granular lymphocytes activated by various stimuli in vitro as well as in cellular infiltrates at the site of renal allografts in patients with and without rejection. Using in situ hybridization, kinetic experiments have shown that granzyme B mRNA is an early marker of in vitro cell activation. Data obtained in vivo also indicate the presence of granzyme B mRNA-bearing cells, which argues in favor of the induction of granzyme B gene in cells activated in vivo.

Gene Expression Regulation, Enzymologic↗
PubMed1990

Involvement of granzyme B and perforin gene expression in the lytic potential of human natural killer cells.

Natural killer (NK) cells (CD3- LGL) spontaneously kill K562 targets but are unable to kill Daudi cells in the absence of IL-2 stimulation. IL-4 is reported to prevent or inhibit the IL-2 driven lymphokine activated killer (LAK) generation in NK cells. Therefore, we asked whether the antagonistic effect of IL-4 on the IL-2 induced LAK activity might regulate the expression of genes encoding proteins involved in lysis, like perforin the pore-forming protein or associated to lysis like granzymes A and B. By using in situ hybridization we show that, besides inducing LAK activity, IL-2 stimulation increases the amount of perforin and granzyme B mRNA at the single cell level in 40 to 100% of the total CD3- LGL population, which suggests the preferential implication of a cell subset within the LGL population. In addition, our results indicate that the stimulatory effect of IL-2 can be down-regulated by IL-4 with respect to both LAK activity and granzyme B and perforin gene expression. Here again one can notice a decrease in the amount of specific mRNA per cell. These findings suggest that the modulation of the lytic machinery via lymphokines might be associated to the regulation of the lytic potentiality of NK/LAK cells.

Cell Survival↗
PubMed1988

Proximity of the CTLA-1 serine esterase and Tcr alpha loci in mouse and man.

The serine esterase CTLA-1 gene was shown by in situ hybridization to map to the D segment of mouse chromosome 14, the same localization as a member of the immunoglobulin superfamily, Tcr alpha. To further demonstrate the proximity of CTLA-1 and Tcr alpha, genetic linkage was tested in mouse using restriction fragment length polymorphisms and a backcross progeny, and no recombination was observed in the 100 backcross products studied. Recombination events between Tcr alpha/CTLA-1 and the markers Gdh-X and NP-1 show that the most probable order of these loci in the mouse 14D region is NP-1-Tcr alpha/Ctla-1-Gdh-X. In man, the human homologue of CTLA-1 was shown by in situ hybridization to map on chromosome 14, at 14q11-q12, where Tcr alpha also maps. Using the human cell line SUP-T1, bearing the inversion inv(14) (q11;q32), we further demonstrated the loci order in man to be centromere-NP-1-Tcr alpha-CTLA-1. To complement the cytogenetic and genetic mapping data, we tried to determine the physical distance between the two genes by pulsed field gel electrophoresis (PFGE). DNA prepared from various cell types, both mouse and human, were digested with a panel of rare cutter enzymes and hybridized first with CTLA-1, then with Tcr alpha probes. None of the bands identified hybridized with both Tcr alpha and CTLA-1 probes for either mouse or human cells. Although the physical mapping by PFGE is inconclusive, the cytogenetic and genetic data support close linkage of the Tcr alpha and CTLA-1 genes in both mouse and man, suggesting homology between the D region of mouse chromosome 14 and the q11-q12 region of human chromosome 14, encompassing the Tcr alpha and CTLA-1 loci. These findings also provide another example of proximity of genes coding for a member of the Ig super-family and a serine esterase.

Animals↗
PubMed1986

Interferon: a cytotoxic T lymphocyte differentiation signal.

Human T cell clones which were able to proliferate in response to specific stimuli but could not kill even in the presence of lectins were found to acquire the specific lytic function when interferon alpha or gamma was added on day 1 of the 7-day restimulation culture. These results demonstrate that interferon may act as a cytotoxic T lymphocyte differentiation signal. This signal can be blocked by the monoclonal antibody LeoA1 which recognizes a 70-kDa cell surface structure, involved in cytotoxic T lymphocyte differentiation.

Antibodies, Monoclonal↗
PubMed1986

Allospecific proliferative human T-cell clones acquire the cytotoxic effector function after three months in culture, in IL-2 conditioned medium.

Allostimulated T lymphocytes were cloned by micromanipulation and expanded in IL-2 conditioned medium. Three T3+,T4+,T8-, clones called BJ1, BJ4, and BJ37, were extensively studied. The BJ1 cells were able to proliferate and kill the specific target. The BJ4 and BJ37 cells were able to proliferate with the specific restimulator but could not kill even in lectin-dependent cell-mediated cytotoxic assay; however, they acquired the specific cytolytic activity in the 6-day culture when fresh irradiated autologous peripheral blood mononuclear cells as feeder cells were added to the specific irradiated Epstein-Barr virus transformed cell line, in the presence of recombinant IL-2. This observation strongly suggested that the culture conditions could be involved in the differentiation of proliferative clones into cytotoxic T lymphocyte (CTL) clones, by the lymphokines, either present in the IL-2 conditioned medium or secreted by the mixed allogeneic irradiated feeder cells. Moreover, it was shown that the acquisition of the cytolytic function could be blocked by the monoclonal antibody LeoA1, previously described and which recognized the TLiSA1 structure involved in the CTL differentiation.

Antibodies, Monoclonal↗
PubMed1986

In vitro allostimulation of peripheral blood mononuclear cells generates autologous reactive T lymphocytes: analysis at the clonal level.

Cells from two siblings within family PER with a maternal crossing-over were used as responder and stimulator cells; they differed in class II MHC gene products. The responder cells were cloned and studied for their specificity. All the clones were T4+, most of them, were allospecific cytotoxic T-cell clones, some were only proliferative. Among the 62 clones that have been screened for their phenotype and function, we could expand, in our cloning system, two autoreactive T-cell clones specific for the self-class II MHC products. The results indicated that this autoreactivity was also observed in serum-free medium, which excludes any serum components susceptible to alter the self-class II MHC products. For the autorecognition, the two clones used the T3-Ti complex, and only one of them, the T4 structure.

Antibodies, Monoclonal↗
PubMed1986

What is a T-cell clone? Effect of rIFN on T-cell clone function and T-cell receptor gene rearrangement.

A T4+ proliferative, noncytotoxic cloned line acquires specific lytic function by treatment with recombinant interferon alpha or gamma. Simultaneous with the acquisition of this new cell function, a rearrangement of the T-cell receptor alpha gene occurs. These changes necessitate a revised concept of a T-cell clone regarding its T-cell receptor gene configuration and cell function.

Cell Line↗
PubMed1986

Recombinant interferon alpha can induce rearrangement of T-cell antigen receptor alpha-chain genes and maturation to cytotoxicity in T-lymphocyte clones in vitro.

T-cell receptor genes are found in the germ-line configuration as well as in rearranged forms, as detected in T lymphocytes by different patterns in Southern blot analysis. We have recently shown that cytotoxic T-lymphocyte (CTL) "precursor" clones, in which rearrangements are not detected in the gene encoding the T-cell receptor alpha-chain (TCRA), acquire specific lytic function induced by treatment with recombinant interferon alpha or gamma. We now report that, coincident with the acquisition of cytotoxic function by the precursor CTL clone; recombinant interferon alpha appears to induce a rearrangement of TCRA. In addition, in a mature CTL clone (i.e., one already showing lytic function) in which one TCRA allele is rearranged, treatment with recombinant interferon alpha appears to induce a new rearrangement of a TCRA gene.

Cell Differentiation↗
PubMed1985

Phorbol ester induces a differential effect on the effector function of human allospecific cytotoxic T lymphocyte and natural killer clones.

Six-day allosensitized human peripheral blood lymphocytes treated for 12-18 hr with phorbol 12-myristate 13-acetate (PMA) were found to lose the capacity to kill the specific target in a standard cell-mediated lympholysis (CML) assay, but they were still effective in their ability to kill the tumor cell line K562. We investigated which antigens on the cell membrane involved in alloimmune recognition might be modified by PMA, since it was found that in a lectin-dependent CML assay, the lytic mechanism was not impaired. For this purpose, T3+, T4+ allospecific cytotoxic T lymphocyte (CTL) clones and either T3+ or T3- natural killer (NK) clones were generated from 6-day allostimulated lymphocytes. The results indicated that PMA inhibited the cytolytic function of both alloimmune CTL and NK T3+ clones. In contrast, PMA did not modify the effector cell function of T3- NK clones. Phenotypic analysis of T-cell surface antigens from T3+ clones showed that T3 molecule expression on the cell membrane was reduced by 80-90% after PMA treatment, whereas expression of both accessory T4 molecules, involved in antigen recognition, and receptor for interleukin 2 was increased. Moreover, the loss of function was transitory and could be restored 4 days after PMA treatment when the T3 molecules were fully reexpressed at the cell surface. Taken together, these data strongly suggest that (i) PMA prevents cell-mediated cytotoxicity by modulating the disulfide-linked heterodimer associated to T3 and described as the receptor for antigen on the cell surface of major histocompatibility complex (MHC) and non-MHC specific CTL clones, without affecting the lytic mechanism per se, and (ii) the expression of the receptor for the antigen present on the tumor cell line K562 is not decreased on T3- NK clones after PMA treatment and must be different from that on T3+ T-cell clones.

Antibodies, Monoclonal↗
PubMed1985

[Phorbol diester induces phenotypic and functional changes in human T-lymphocyte clones].

Tumor promoting phorbol myristate acetate (PMA) induce to enhance the expression of IL2 Receptor and to decrease the antigen receptor expression on the cell surface. The same phenotypic changes are also observed when the T cell clones are stimulated by the specific ligand. In contrast to the IL2 Receptor induced by the specific antigen, the ones induced by PMA are less active.

Animals↗
PubMed1982

A five-generation family with sacral agenesis and spina bifida: possible similarities with the mouse T-locus.

In man, a malformation that recalls some of the defects associated with T/t mutants in the mouse is sacral agenesis. We report on a family with a high incidence of sacral malformation, ranging from a complete absence of the sacrum (SA), with or without spina bifida aperta, to a spina bifida occulta (SBO) that could only be detected by x-ray. The condition appeared in a man with four children who were all affect, and thereafter, to varying degrees, in 17 of his 28 descendants. Segregation analysis has been performed in this family, using the Elston and Stewart transmission probability model [1971]. The two traits (SA and SBO) were first studied separated and then together. A fully penetrant major dominant gene is show to cause SA. When the phenotypes SA and SBO are considered together, Mendelian transmission is rejected. This could be explained genetically by two alternative hypotheses: genetic heterogeneity or a dominant major gene transmitted in excess by heterozygotes (tau Aa A = 0.896), suggesting a segregation distortion property of an allele at a T-like locus.

Adult↗
PubMed1981

Regulation of the human allogeneic proliferative response in vitro.

We have investigated the glass-wool-adherent characteristics of human lymphocytes able to suppress allogeneic proliferation in vitro. We observed that: (1) Suppressor cells obtained after in vitro priming and their precursors are adherent to glass wool, whereas cytotoxic cells and their precursors are not; (2) primed cells adherent to glass wool are mainly HLA-DR positive, while nonadherent cells are HLA-DR negative or weakly positive; (3) the nonadherent, unprimed T cells used as responder cells in a primary MLR are more readily suppressed by suppressor cells than the same cells unfiltered. This may suggest an enrichment of 'acceptor' cells for suppression.

Cytotoxicity, Immunologic↗