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F Sanchez-Madrid

Publications and source records attributed to F Sanchez-Madrid.

At least 37 records · Page 2Linked to original sources
PubMed1989

Function of CD44(Pgp-1) homing receptor in human T cell precursors.

T cell precursors migrate from extrathymic hematopoietic tissues and differentiate after encountering the thymic microenvironment. We asked whether human T cell precursors express the CD44(Pgp-1/gp90HR) class of homing receptors that have been implicated in the traffic of hematopoietic cells, such as lymphocyte entry to peripheral lymphoid organs. Flow cytometry and immunoprecipitation studies demonstrate that CD7+34+, CD1-2-3-4-8-14-16-20- cells in bone marrow and thymus, which have been shown to exhibit features of T cell precursors, bear CD44. Immunohistological studies show that clusters of thymocytes in the subcapsular and the inner cortex and most medullary thymocytes are clearly CD44+, whereas the expression of CD44 is selectively downregulated in CD3- and CD3low functionally incompetent cortical thymocytes. The expression of CD44 is not restricted to T cell precursors but also occurs in thymic stroma, which bear a different molecular species of CD44. CD44-specific antibodies exert stimulatory effects on T cell precursors, a process that is dependent on stromal cells. We postulate that CD44 might be an adhesion molecule for precursor homing to thymus and that it participates in cell-to-cell interactions within the thymic environment.

Cell Movement↗
PubMed1988

Co-expression of Mac-1 and p150,95 on CD5+ B cells. Structural and functional characterization in a human chronic lymphocytic leukemia.

The leukocyte adhesion receptors (LAR) Mac-1 and p150,95 were thought to occur only on myeloid cells, but recently, mouse Ly-1+ (CD5+) B and pro-B cells were shown to bind M1/70 monoclonal antibody, an anti-Mac-1 antibody. Using immunoprecipitation, sodium dodecyl sulfate gel electrophoresis, immunofluorescence and flow cytometry, we have studied the expression of LAR in human CD5+ B cells from chronic lymphocytic leukemia (B-CLL) patients. 170,95 kDa Mac-1 and p150,95, as well as 180,95 kDa LFA-1 heterodimers are shown to be co-expressed on CD5+ mu + kappa + B cells from a patient where they retain their known function in homotypic cell-cell adhesion. Therefore, functional Mac-1 and p150,95 LAR are not restricted to myeloid cells.

Antibodies, Monoclonal↗
PubMed1988

Characterization of a CD11c-reactive monoclonal antibody (HC1/1) obtained by immunizing with phorbol ester differentiated U937 cells.

A mouse monoclonal antibody (HC1/1) specific for a differentiation marker of human monocytes and granulocytes has been generated by using as immunogen the monocytic cell line U937 differentiated with phorbol esters. This differentiation antigen has been characterized as the p150/95 member of the CD11 family of glycoproteins by cellular distribution studies, immunoprecipitation and competition experiments with MAb 3.9 specific for the CD11c antigen. Immuno-alkaline phosphatase staining of normal tissue sections with the HC1/1 MAb demonstrated that the CD11c antigen is a useful macrophage marker. Furthermore, the MAb HC1/1 stains specifically populations of macrophages on skin and lymph node sections from different pathologies such as Sarcoidosis, Granuloma annulare, Lepromatous leprosy and Toxoplasmosis.

Antibodies, Monoclonal↗
PubMed1987

Involvement of the CD4 molecule in a post-activation event on T cell proliferation.

A monoclonal antibody (mAb) directed to a human leukocyte 55-kDa cell surface molecule with identical cellular distribution and biochemical properties to the CD4 was able to inhibit T cell proliferation induced either in a mixed lymphocyte culture or by activation with mAb anti-CD3, anti-CD2 or phytohemagglutinin. The inhibitory effect of anti-CD4 was observed in the absence of monocytes and was directly exerted on T4+ cells. This effect on cellular proliferation appears to be due to an inhibition of a postactivation event since the rise of cytoplasmic Ca2+ after activation with anti-CD3 mAb is not affected by the presence of anti-CD4 and the proliferation that occurs after an activation pulse of 3 h with ionophore and phorbol myristate acetate can be inhibited when the anti-CD4 is added after the pulse period. Kinetic studies demonstrated that the inhibition of cellular proliferation by anti-CD4 mAb was observed even if the antibody was added as late as 18-24 h after the initiation of the culture. The effect of this blocking anti-CD4 mAb on the interleukin (IL) 2/IL 2 receptor signalling pathways was also examined. The presence of anti-CD4 slightly affected the production of IL2. In fact, addition of exogenous recombinant IL2 at the initiation of the cultures did not restore the proliferative response. However, anti-CD4 had a strong inhibitory effect on the expression of IL2 receptors as analyzed by direct immunofluorescence cytometry. Taken together, these results indicate that the binding of the anti-CD4 mAb to T cells interferes with a late metabolic step being capable of abolishing the proliferative activity of fully activated cells.

Animals↗
PubMed1985

Isolation of the major IgE-binding protein from Parietaria judaica pollen using monoclonal antibodies.

Allergen molecules from Parietaria judaica pollen, a widely distributed allergy inducer in Southern and Western Europe, have been studied using specific monoclonal antibodies (MAbs). MAbs against IgE-binding components were selected in a 4-step radioimmunoassay. Three different MAbs (AC/1.1, AC/7.1 and AC/15.1) were obtained which recognized epitope(s) located on a polypeptide of 10 Kd (Pj10). This polypeptide displayed the highest IgE-binding ability under either native or SDS-denatured conditions, as determined by immunoadsorption and immunodetection after SDS-PAGE, respectively. The Pj10-containing allergen, purified on an AC/1.1 MAb-Sepharose column, was able to inhibit most of the binding of specific IgE to the pollen extract coupled to paper discs in an inhibition radioallergosorbent test (RAST). The affinity-purified allergen exhibited the same immunoelectrophoretic behaviour as the native allergen.

Allergens↗
PubMed1985

Identification of a novel allergen molecule from Dermatophagoides by monoclonal antibodies.

Six different monoclonal antibodies (MAb) have been obtained which bind to components contained in extracts from Dermatophagoides. One out of the six MAb recognized molecules displaying IgE binding ability. This MAb (MADP-1) immunoprecipitated allergenic polypeptides of 42 kDa and 30 kDa from 125I-labeled extracts of D. pteronyssinus and D. farinae respectively. Purified allergen preparations from both extracts have been obtained by affinity chromatography using a column of purified MADP-1 coupled to Sepharose. The purified fractions partially compete the binding of specific IgE contained in sera from sensitized patients to the whole extracts, in a radioallergosorbent inhibition test.

Allergens↗
PubMed1985

Cell surface molecular changes on the activation of human thymocytes.

The changes in the expression of antigen molecules on the cell surface membranes of uncultured (nonactivated) and activated human thymocytes have been studied by flow cytometry and immunoprecipitation techniques. Nonactivated thymocytes do not have the phenotypic profile of a resting population because they express cell proliferation molecules such as the transferrin receptor and the 4F2 antigens (mainly the 100,000 dalton subunit). After activation with IL 2-containing supernatants, mature T3+, T6- thymocytes proliferate and are able to nonspecifically kill different target cells. The activated thymocytes are T3+, T11+, T6-, OKM1- and bear T4 or T8 antigens in mutually exclusive cell subpopulations. They also "de novo" express the IL 2 receptor, and the 210.000/130.000 molecular complex defined by the TS2/7 MAb. Activated human thymocytes express higher amounts of class I and class II MHC antigens, equal T3 and LFA-1, and lower quantities of T11 and T4 molecules than nonactivated thymocytes. Furthermore, activated thymocytes only express the T8 34,000 dalton polypeptide subunit, whereas the nonactivated thymocyte population expressed the T8 34,000 dalton associated with a 46,000 glycoprotein. We have demonstrated that this structural change in the T8 molecule from a complex of two polypeptide subunits of 46,000 and 32,000 does not indeed occur in the activation process but rather in the maturation from T6+ to T6- thymocytes.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1984

Human lymphocyte function-associated antigens.

Three cell surface molecules, designated LFA-1, LFA-2, and LFA-3 were identified by mAbs selected for their ability to block cytolysis by an OKT4+, HLA-DR-specific CTL line. The LFA mAbs block all CTL and proliferative functions studied. In addition, anti-LFA-1 mAbs inhibit NK-mediated cytolysis. By analogy with murine LFA-1, human LFA-1 may be involved in the adhesion stage of cellular interactions. LFA-2, the SRBC receptor molecule, appears to be a T cell function-specific molecule. We have not yet established whether LFA-2 participates in antigen recognition or whether it is involved in antigen-non-specific interactions. The anti-LFA-3 mAb specifically blocks function by binding to the target cells, implying that LFA-3 may be a target ligand for an effector-specific receptor. The CTL-target interaction involves a number of steps, including antigen recognition, cell adhesion, and delivery of the lethal hit [22]. The LFA antigens show the complexity of this process at the molecular level. The anti-LFA monoclonal antibodies will be useful probes into the T cell immune response and may prove clinically relevant, both diagnostically and therapeutically.

Antibodies, Monoclonal↗
PubMed1984

Glycoproteins of 210,000 and 130,000 m.w. on activated T cells: cell distribution and antigenic relation to components on resting cells and T cell lines.

A glycoprotein complex of 210,000 and 130,000 m.w., found on mitogen or alloantigen-stimulated human T cells and not on other hematopoietic cells, has been defined by a monoclonal antibody (Mab). The components of this complex are a subset of a larger family of proteins (210,000, 165,000 and 130,000 m.w.) defined by a second Mab. In a panel of hematopoietic cell lines and cell types, only activated T cells (including the cell line HUT-102) express the 210,000/130,000 complex and these cells also express the IL 2 receptor, a characteristic marker for activated T cells. The 210,000/130,000 m.w. complex (reactive with the Mab TS2/7) is present on all long-term activated T cells, including both the OKT4 and OKT8 subsets. The 210,000 m.w. subunit is expressed only on activated T cells. Other lymphoid cells express either the 130,000 m.w. subunit alone (unactivated lymphocytes, thymocytes, HUT-78) or the 130,000 subunit together with a 165,000 subunit (MOLT-4, HSB, and other leukemic T cell lines). The 210,000/130,000 m.w., 165,000/130,000 m.w. and 130,000 m.w. complexes are antigenically related in that all share reactivity with the Mab A- 1A5 . Among non-lymphoid hematopoietic cells and cell lines, none express the 210,000 m.w. chain; adherent cells (monocytes) and myeloid cell lines each express single proteins of 130,000 to 155,000 m.w. Granulocytes and red blood cells are negative and platelets express multiple bands (165,000 and 140,000 m.w.). Immunoperoxidase staining of tissue sections showed that a broad range of tissues and cell types had material cross-reactive with the lymphoid 130,000 m.w. protein. However, only a discrete subset of those tissues and cells including blood vessel walls, connective tissue, smooth muscle, kidney mesangial cells, and some non-cellular matrix tissue, had material cross-reactive with the 210,000 m.w. protein on activated T lymphocytes.

Animals↗
PubMed1983

A human leukocyte differentiation antigen family with distinct alpha-subunits and a common beta-subunit: the lymphocyte function-associated antigen (LFA-1), the C3bi complement receptor (OKM1/Mac-1), and the p150,95 molecule.

The human lymphocyte function-associated antigen-1 (LFA-1), the complement receptor-associated OKM1 molecule, and a previously undescribed molecule termed p150,95, have been found to be structurally and antigenically related. Each antigen contains an alpha- and beta-subunit noncovalently associated in an alpha 1 beta 1-structure as shown by cross-linking experiments. LFA-1, OKM1, and p150,95 alpha-subunit designations and their molecular weights are alpha L = 177,000 Mr, alpha M = 165,000 Mr, and alpha X = 150,000 Mr, respectively. The beta-subunits are all = 95,000 Mr. Some MAb precipitated only LFA-1, others only OKM1, and another precipitates all three antigens. The specificity of these MAb for particular subunits was examined after subunit dissociation by high pH. MAb specific for LFA-1 or OKM1 bind to the alpha L- or alpha M-subunits, respectively, while the cross-reactive MAb binds to the beta-subunits. Coprecipitation experiments with intact alpha 1 beta 1-complexes showed anti-alpha and anti-beta MAb can precipitate the same molecules. In two-dimensional (2D) isoelectric focusing-SDS-PAGE, the alpha subunits of the three antigens are distinct, while the beta-subunits are identical. Biosynthesis experiments showed alpha L, alpha M, and alpha X are synthesized from distinct precursors, as is beta. The three antigens differ in expression on lymphocytes, granulocytes, and monocytes. During maturation of the monoblast-like U937 line, alpha M and alpha X are upregulated and alpha L is downregulated. Some MAb to the alpha subunit of OKM1 inhibited the complement receptor type three. LFA-1, OKM1, and p150,95 constitute a novel family of functionally important human leukocyte antigens that share a common beta-subunit.

Antibodies, Monoclonal↗
PubMed1983

Mapping of antigenic and functional epitopes on the alpha- and beta-subunits of two related mouse glycoproteins involved in cell interactions, LFA-1 and Mac-1.

Mouse Mac-1, a complement receptor-associated surface structure on macrophages, and LFA-1, a function-associated structure on lymphocytes, comprise a novel family of leukocyte differentiation antigens participating in adhesive cell interactions. Mac-1 and LFA-1 contain alpha-subunits of 170,000 and 180,000 Mr, respectively, and beta-subunits of 95,000 Mr noncovalently associated in alpha 1 beta 1 complexes. The structural relation between the alpha- and between the beta-subunits, and the location of functionally important sites on the molecules, have been probed with antibodies. Both non-cross-reactive and cross-reactive monoclonal antibodies (MAb) and antisera prepared to the purified molecules or the LFA-1 alpha-subunits were used. Reactivity with individual subunits was studied by immunoprecipitation after dissociation induced by high pH treatment, or by immunoblotting after SDS-PAGE. Cross-reactive epitopes on Mac-1 and LFA-1 were found to be present on the beta-subunits, which were immunologically identical. Non-cross-reactive epitopes that are distinctive for Mac-1 or LFA-1 were localized to the alpha-subunits. MAb to LFA-1 alpha-subunit epitopes inhibited CTL-mediated killing. Two MAb to Mac-1 alpha-subunit epitopes but not a third MAb to a spatially distinct alpha-epitope inhibited complement receptor function. Neither function was inhibited by a MAb binding to a common beta-subunit epitope. Therefore, sites of Mac-1 and LFA-1 involved in their respective adhesion-related functions, as well as distinctive structural features, have been localized to the alpha-subunits.

Animals↗
PubMed1983

Human lymphocyte function associated antigen-1 (LFA-1): identification of multiple antigenic epitopes and their relationship to CTL-mediated cytotoxicity.

Human lymphocyte function-associated antigen (LFA)-1, a heterodimeric lymphocyte surface glycoprotein of 177,000 and 95,000 relative molecular weight has been implicated to function in the cytotoxic T lymphocyte (CTL) effector mechanism. Seven mouse hybridoma lines producing monoclonal antibodies (MAb) reactive with this structure were studied. Three unique and 3 partially over-lapping epitopes on human LFA-1 were defined by competitive cross inhibition binding assays using biosynthetically labeled anti-LFA-1 MAb. In contrast, of five rat antimouse LFA-1 MAb, all five recognized a common or shared epitope. An HLA-B7 specific human CTL line expressed 1.1 X 10(5) LFA-1 sites per cell with a direct saturation binding assay. Human CTL expressed two to four times more LFA-1 than peripheral blood lymphocytes or B and T lymphoblastoid cell lines. Titration of each of the anti-LFA-1 MAb in a 51chromium release cytolytic assay revealed quantitative differences in the ability of the different anti-LFA-1 MAb to block cytolysis indicating distinct functional and antigenic epitopes exist on the human LFA-1 molecule. Anti-LFA-1 MAb reversibly inhibited the CTL reaction by slowing the initial rate of cytolysis. These results suggest anti-LFA-1 MAb inhibit CTL function by specific blockade of a functionally relevant molecule.

Animals↗
PubMed1983

Cytotoxic T cells directed against HLA-DR antigens and their surface proteins.

The authors review their recent research involving the generation of cytotoxic T lymphocytes (CTL) directed against HLA-DR antigens. A mouse anti-human xenogeneic system first suggested that HLA-DR antigens could be recognized by CTL. Human allogeneic CTL specific for HLA-DR6 were generated and found to be OKT4+. The fact that these CTL were OKT4+ while anti HLA-A,B CTL were OKT8+ suggested that these T cell surface antigens may be involved in MHC antigen recognition; ie, they may be part of the T cell receptor. These OKT4+, HLA-DR specific CTL were further used to generate monoclonal antibodies (1) which block cytolysis and define novel antigens involved in the CTL-target interaction and (2) which define an antigenic complex on alloantigen activated T cells.

Animals↗
PubMed1983

The functional significance, distribution, and structure of LFA-1, LFA-2, and LFA-3: cell surface antigens associated with CTL-target interactions.

Three cell surface antigens associated with the cytolytic T lymphocyte(CTL)-target cell interaction were identified by generation of monoclonal antibodies (MAb) against OKT4+, HLA-DR-specific CTL and selection for inhibition of cytolysis in a 51Cr-release assay. These MAb block cytolysis by both OKT4+ and OKT8+ CTL and the proliferative responses to PHA and the mixed lymphocyte response (MLR). LFA-1 is an antigen widely distributed on lymphoid tissues and is composed of two polypeptides of 177,000 and 95,000 Mr on all cell types studied. Anti-LFA-1 MAb block NK cell-mediated cytolysis in addition to T lymphocyte-mediated cytotoxicity and proliferation. LFA-2 (Mr = 55,000 to 47,000), a determinant on the sheep red blood cell receptor, is expressed by T cells but not B cells and appears specific for T cell functions. LFA-3 (Mr = 60,000) is a widely distributed antigen present on both hematopoietic and nonhematopoietic tissues and appears to only be involved in T cell functions. MAb to LFA-1 and LFA-2 inhibit function by binding to effector cell surface molecules, whereas anti-LFA-3 MAb appear to block by binding to the target cells. Together with previously described molecules, LFA-1, LFA-2, and LFA-3 demonstrate the complexity of CTL-mediated cytotoxicity at the molecular level.

Antibodies, Monoclonal↗
PubMed1983

Blocking of CTL-mediated killing by monoclonal antibodies to LFA-1 and LYT-2,3. I. Increased susceptibility to blocking after papain treatment of target cells.

It is now established that monoclonal antibodies (MAb) against LFA-1 and Lyt-2,3 antigens on cytolytic T lymphocytes (CTL) block killing function in the absence of C. It has been suggested that the blocking is inversely related to CTL-target affinity. In this report, we studied the effect of papain pretreatment of target cells, because papain is known to remove H-2 and to render target cells more resistant to allospecific CTL. CTL-target conjugate formation was weaker with papain-treated target cells (based on reduced post-dispersion lysis in dextran-containing medium). The concentration of MAb required to produce 40 to 60% inhibition of 51Cr release (2-hr assay) was reduced four to 29-fold for alpha LFA-1 and 64 to 114-fold for alpha Lyt-2,3. Papain, however, did not induce blocking by MAb to other CTL antigens such as Thy-1, H-2, and T200. Flow cytometric analysis confirmed that papain selectively removed more than 95% of H-2. In kinetic studies of removal and recovery, H-2 density and conjugate formation correlated well with each other. Sensitivity to blocking was not as well correlated, raising the possibility that an unidentified papain-sensitive target cell molecule other than H-2 plays an important role in CTL-target interaction.

Animals↗
PubMed1983

Stable hamster-mouse hybridomas producing IgG and IgM hamster monoclonal antibodies of defined specificity.

Specific antibody-secreting hybridomas have been obtained by fusing Syrian or Armenian hamster (Mesocricetus auratus or Cricetulus migratorius) spleen cells with mouse myeloma cells. The hamsters were immunized to mouse cytolytic T lymphocytes. Hybrids were selected either by an indirect binding assay using an 125I-monoclonal antibody (MAb) reactive with hamster kappa-chains or by their ability to block T cell-mediated cytolysis. Three hybridoma clones were obtained that secreted intact IgM-like and IgG-like hamster MAb as shown by SDS-PAGE. The clones were stable as shown by subcloning. Two MAb recognized antigens of wide tissue distribution; the third bound specifically to T lymphocytes, gave strong inhibition of T cell-mediated cytolysis, and immunoprecipitated the Lyt-2,3 molecule.

Animals↗