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Biomedical subjects

A Coutinho

Publications and source records attributed to A Coutinho.

At least 271 records · Page 15Linked to original sources
PubMed1982

B-cell growth factor: distinction from T-cell growth factor and B-cell maturation factor.

A T-cell hybridoma was derived by somatic cell hybridization between concanavalin A-activated BALB/c spleen cells and the AKR thymoma BW 5147. Media conditioned by hybridoma cells, even at high dilutions (1:1,000) support the growth of lipopolysaccharide-stimulated B-cell blasts but not that of T-cell growth factor (TCGF)-reactive T-cells. This activity, herein designated B-cell growth factor (BCGF), has a Mr of approximately equal to 20,000 and it can readily be separated from TCGF (Mr approximately equal to 30,000) by gel filtration. BCGF is constitutively produced by the hybridoma cells, it is removed from conditioned media by incubation with target cells at +4 degrees C, and it is equally effective on B-cell blasts carrying different major histocompatibility complex and Ig haplotypes. BCGF shows no T-cell replacing factor (TRF) activity, and it is poor in supporting the development of Ig-secreting plaque-forming cells in B-cell blast cultures. Terminal maturation, however, can be induced in BCGF-dependent blasts by addition of conditioned media from normal helper T cell cultures, suggesting that two distinct factors are involved in the helper cell-dependent growth and maturation of B lymphocytes.

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PubMed1981

Very rapid decay of mature B lymphocytes in the spleen.

To determine the persistence of immunocompetent B lymphocytes at the periphery, regardless of cell division, we have followed the decay of lipopolysaccharide (LPS)-reactive B cells in LPS-nonresponder, histocompatible hosts. Both the numbers of transferred cells and of those persisting in the recipients could be determine with precision by limiting dilution analysis of the various cell populations. Decay rates of 50% per day were determined. Because we can exclude immune elimination of donor cells and the numbers of transferred cells were too low to result in gross alterations of the physiologic turnover rates, we conclude that the majority of LPS-reactive B lymphocytes and a large part of the whole B cell compartment show this astonishingly high rate of decay. The method introduced here might prove useful in a variety of clonal assays, as it can detect cells present at a very low frequency.

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PubMed1981

Immune networks. Frequencies of antibody- and idiotype-producing B cell clones in various steady states.

In limiting dilution analysis, the absolute frequencies of lipopolysaccharide-reactive B cell precursors producing anti-trinitrophenyl antibodies or the MOPC460 idiotype were studied in BALB/c mice, either normal, or immunized with antigen (Ab1), the idiotype (Ab2), or a monoclonal anti-idiotype antibody (Ab3). Anti-idiotype immunity results in the suppression of the B cell precursors for the relevant idiotype, and anti-(anti-idiotype) immunity leads to a 10-fold increase in precursor B cell frequencies, with a comparatively lower increase in antibody-producing precursors. The findings can only be explained by variations in the composition of the B cell compartment in the various immune states.

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PubMed1981

Hapten-specific helper T cells III. Fine specificity of the (4-hydroxy-3-nitro-phenyl)acetyl (NP)-specific response in Igh-1b mice.

Helper cells, with specificity for the haptens (4-hydroxy-3-nitro-phenyl)acetyl (NP) or (4-hydroxy-5-iodo-3-nitro-phenyl)acetyl (NIP), were raised in B10.BR mice by in vivo priming and in vitro long-term enrichment with hapten-derivatized syngeneic spleen cells. Upon co-culture with the homologous antigen (NP or NIP self), selected helper cells specifically responded by proliferation and by inducing large numbers of B cells to clonal expansion and immunoglobulin secretion. Criss-cross experiments demonstrated the nonheteroclitic nature of antigen recognition by helper cells, as the proliferative and helper cell activities were in every case one order of magnitude higher when confronted with the homologous hapten used for immunization.

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PubMed1981

Immunoglobulin isotype expression. II. Frequency analysis in mitogen-reactive B cells.

The frequency of lipopolysaccharide (LPS)-reactive B cells developing into clones that secrete the various immunoglobulin (Ig) classes has been determined in vitro, in cells from BALB/c mice, under culture conditions which detect all growth-inducible cells. Secretion of the different Ig classes was assessed in the protein A plaque assay for Ig-secreting, plaque-forming cells by using developing antisera specific for either IgM, IgG1, IgG2a, IgG2b, IgG3 or IgA. In all lymphoid organs tested (spleen, bone marrow, mesenteric lymph nodes and thoracic duct), a considerable proportion of all B cells (5-20%) was induced by LPS to yield a clone of IgM-secreting cells. Frequency determinations of LPS-reactive cells giving rise to descendants secreting other Ig isotypes revealed that, on an average, and irrespective of the origin of the cells, 7% of all IgM-secreting clones switched to the synthesis of IgG1, 39% to IgG2, 41% to IgG3 and 1% to IgA. Roughly the same frequencies of B cells switching CH gene expression were found among spleen cells of athymic nude mice. No correlation was found between the clonal frequencies of CH gene expression in polyclonally activated B cells and the in vivo "background" Ig-secreting cells suggesting that the CH gene expression in B cells is influenced by the quality of stimulation and other regulating influences.

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PubMed1981

Effect of anti-delta antibodies on clonal expansion and maturation of B lymphocytes.

The effect of a monoclonal anti-delta antibody on the in vitro B cell response to the B cell mitogen lipopolysaccharide (LPS) has been studied. Titration of anti-delta antibody in mass cultures stimulated by LPS resulted in a dose-dependent reduction of thymidine incorporation. IgM and IgG plaque forming cell (PFC) responses were poorly affected at high concentration, and slightly increased at low concentrations of anti-delta antibody. By limiting dilution analysis it was shown that anti-delta antibody inhibit 30-50% of LPS-reactive B cells to grow as IgM secreting clones, while increasing the average size of clones that grew in the presence of anti-delta as compared to control cultures. Anti-delta also results in increased frequencies of IgG secreting clones. By immunofluorescence it was possible to show the presence of a higher relative number of cells containing immunoglobulin as an effect of anti-delta antibodies. Observations made at early times of culture indicate that the cells that do not proliferate in the presence of anti-delta undergo an early maturation to secretion. Experiments performed on LPS blasts suggest that the effects of anti-delta on cell proliferation require the presence of antibodies at early times in the response, while the effects on maturation can be manifested during clonal development. The relevance of membrane IgD and of the IgM-to-IgD ratio in the maintenance versus exhaustion of the clone is discussed.

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PubMed1981

The enhancement of antibody response by IgM antibodies is dependent on antigen-specific T helper cells.

The enhancement of antibody responses by IgM antibodies administered with low doses of antigen has been studied in a T-dependent (SRBC) and an T-independent (alpha 1,6 dextran) system. It has been found that IgM anti-SRBC antibodies do not enhance a SRBC response in nude mice. The T-cell dependency was also directly demonstrated by showing the effect of IgM on T-cell priming in transfer experiments. The simultaneous injection of antigen and IgM antibody also induced a polyclonal increase of IgM, PFC, which was not due to a non-specific "adjuvant" effect of IgM, as we could not detect a similar effect on an ongoing response to HRBC in mice simultaneously given SRBC and IgM anti-SRBC antibodies. The specificity of the helper cell for either the antibody or the antigen was investigated in a response to alpha 1, 6 dextran, in which we could demonstrate antibody-specific helper T cells, but no antigen-specific help. We have found that IgM anti-dextran antibodies do not enhance and rather suppress the response of normal, high-responder mice, to dextran, suggesting that the T cells mediating the "19S enhancement" are antigen-specific. The magnitude of the enhancement response, as compared to the responses induced by either antigen or antibody alone, implies a synergistic mechanism, possibly involving antigen-specific and antibody(idiotype)-specific T helper cells.

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PubMed1981

Suppression of a "recurrent" idiotype results in profound alterations of the whole B-cell compartment.

The progeny of BALB/c female mice actively immunized with the trinitrophenyl-binding myeloma protein MOPC460 and producing anti-idiotypic antibodies during pregnancy were compared with mice born of normal mothers for several characteristics of B lymphocytes and their precursors. In all cases, maternal anti-idiotypic immunity resulted in the suppression of the expression of that idiotype by immunocompetent cells in the progeny, as shown by limiting-dilution analysis in single clones of mitogen-reactive IgM-secreting cells. At critical concentrations of circulating maternal antibodies, suppression of the antibody idiotype was found to be accompanied by a large increase in the total number of mature small B lymphocytes. This increase can be accounted for by the selective expansion of B cells bearing nonimmunoglobulin surface structures crossreactive with a MOPC460 idiotope recognized by a monoclonal antibody. In addition, the large majority of newly formed mature B lymphocytes, as well as a large fraction of immunoglobulin-negative cells in the bone marrow of suppressed mice, bear such nonimmunoglobulin MOPC460 crossreactive determinant(s). These results suggest that the suppression of a given "recurrent" idiotype has profound consequences for a large part of the immune system.

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PubMed1981

Allogeneic H-2 dose not cross-react, at the helper T-cell level, with Hapten-modified Self.

Helper T cells raised in CBA mice and enriched in vitro with specificity for trinitrophenylmodified syngeneic spleen cells can activate hapten-modified B cells bearing Ik. These helper cells fail to activate B cells expressing any of six independent I-region haplotypes, indicating lack of cross-reactivity between hapten-self and allogeneic I-region products. Furthermore, hapten-derivatized allogeneic B cells are also not activated, demonstrating that the elements restricting T-cell function are poorly, if at all, cross-reactive.

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PubMed1981

Genetics of a non-H-2 antigen that stimulates "unrestricted" helper T cells and MLR.

The inheritance of antigens expressed by C3H/Tif B cells that stimulate MHC-unrestricted helper T cells from C3H/HeJ was investigated. F1 hybrids between C3H/HeJ and C3H/Tif and 39 C3H/HeJ X F1 backcross mice were characterized as to the ability of their spleen cells to stimulate a proliferative C3H/HeJ T helper cell response and to respond to helper cell activity by the development of polyclonal plaque-forming cell responses. Backcross progeny wee also typed for the following markers segregating in this cross: 1) Responsiveness to the B cell mitogen lipopolysaccharide (LPS); 2) LyM-1 allotype; 3) antigen(s) stimulating a primary non-H-2 MLR between these strains, previously ascribed to Mls locus differences, 4) expression of target antigens for cytotoxic T cells raised in the same strain combination. The antigen(s) recognized by helper cells and those stimulating primary MLR are controlled by autosomal gene(s) and segregate as a single trait. These antigens, however, are not encoded in genes linked to either the Lps or the Mls loci, and are not recognized by cytotoxic T cells raised in the same strain combination.

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PubMed1981

Regulation of T cell growth factor production: arrest of TCGF production after 18 hours in normal lectin-stimulated mouse spleen cell cultures.

The detailed kinetics of TCGF accumulation in Con A-stimulated spleen cell cultures shows a maximum at 24 hr, with a subsequent decrease in activity. This decrease is not due to the appearance of inhibitory substances "masking" TCGF activity. Pulse experiments show that the rate of TCGF production falls sharply after 18 hr and is completely arrested after 24 hr of Con A stimulation. The arrest in TCGF production is the result neither of culture depletion in medium components nor of limiting accessory cell function or inactivation of the lectin, and it thus seem to be the result of inactivation of TCGF-producing T cells. This regulation is not the result of a TCGF-mediated feedback mechanism but rather of lectin-induced suppressive cells that appear in culture after 24 hr and turn off de novo production of TCGF in fresh cultures.

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PubMed1981

Frequency analysis of immunoglobulin V-gene expression and functional reactivities in bone marrow B cells.

Frequency of immunocompetent B cells in bone marrow has been determined in vitro under culture conditions that allow the development in vitro under culture conditions that allow the development of every growth-inducible B cell into a clone of IgM-secreting PFC. Three limiting dilution culture systems were employed: a specific helper assay with SRBC as antigen and using activated T helper cells, a nonspecific helper assay using Con A-induced factors as a source of help, and polyclonal activation with LPS. From unseparated, normal C57BL/6J bone marrow 1 in 2200 to 1 in 2820 B cells were induced to form a clone of PFC to SRBC in each of the 3 systems. This corresponds to a frequency of 1 SRBC-specific clone in every 900 IgM-secreting LPS-reactive clones. The frequencies of specific plaque-forming B cell clones in terms of LPS-reactive B cells was 1 in 36 for NNP1-SRBC, 1 in 58 for TNP30-SRBC, 1 in 75 for NIP1-SRBC, and 1 in 230 for TNP3-SRBC. These frequencies of v-gene expression in bone marrow B cells are of the same magnitude as the corresponding frequencies for splenic B cells. Bone marrow B cells are also fully susceptible to stimulation by antigen in combination with either specific or nonspecific T cell help, as well as polyclonal activation by LPS, since every 3rd Ig-positive cells in marrow could be induced to form a clone of IgM-secreting cells. There is thus no difference in immunocompetence between surface Ig-bearing B cells from bone marrow and spleen.

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PubMed1981

Quantitative studies on concanavalin A-induced, TCGF-reactive T cells. I. Correlation between proliferation and lectin-dependent cytolytic activity.

A limiting dilution assay was used to determine the frequencies of T cells that could be induced to proliferate and kill by Con A and TCGF. The frequency of proliferating cells was 1 in 5 nylon wool-purified spleen cells and 1 in 7 normal lymph node cells. The frequencies in the same cell populations of lectin-dependent killer cell precursors were 1 in 5 and 1 in 9, respectively. Even at limiting numbers of T cells seeded per culture, very few cultures proliferated without showing cytotoxic activity. Thus, under these conditions Con A and TCGF, preferentially if not exclusively, activate T cells that will kill in the presence of PHA. This experimental system can be used for estimating the repertoires of specific CTL-precursors activated by Con A and expanded in TCGF.

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