Blastic Transformation of Mouse Spleen Lymphocytes by a Water-Soluble Mitogen Extracted from Nocardia

Bona, Constantin A.; Damais, C.; Chedid, L

doi:10.1073/pnas.71.5.1602

Cited by 47 publications

(34 citation statements)

References 11 publications

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“…Thus, NWSM is able to stimulate human, rabbit, mouse, rat, and avian Bderived lymphocytes (16,23,26); lipoprotein from E. coli and peptidoglycan from gram-positive and gram-negative species can stimulate rabbit and mouse B-cells (16,22,28,29), whereas LPS and its lipid A moiety enterotoxin B, DxS, pneumococcal SIII polysaccharides fail to induce a high [ZH]thymidine incorporation and to stimulate nonspecifically human, monkey, guinea pig, and rabbit lymphocytes (30)(31)(32)(33)(34).…”

Section: Discussionmentioning

confidence: 99%

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Mitogenic analysis of murine B-cell heterogeneity.

Bona¹,

Yano²,

Dimitriu³

et al. 1978

The Journal of Experimental Medicine

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Murine B lymphocytes appear to be heterogeneous from a functional point of view. Thus, those B cells that give rise to antibody-producing cells can be divided into a subset that requires cooperation with T-derived lymphocytes and another subset that is T independent (1-6). Recently, it has been shown that the B-cell population also contains suppressor cells that regulate various functions of T cells (7)(8)(9)(10)(11) and that these suppressor cells seem to be different from antibody-producing cells (10, 11).There is contradictory evidence as to whether the response to different mitogens defines different B-cell subpopulations. In mice, Melchers and Andersson (12) concluded that PPD and lipepelysaccharide (LPS) I stimulate the same cell, whereas Gronowicz and Coutinho (13) The aim of this work was to test directly various alternative hypotheses that could explain the differences in the responsiveness of routine B lymphocytes with regard to various mitogens: (a) One cell possesses the receptors for various mitogens and can be stimulated by all known murine B-cell mitogens separately; (b) the ability to respond to various mitogens depends on the stage of maturation of the B cell; (c) there are different subsets of lymphocytes for different mitogens.We have chosen three well-known B-cell mitogens for this study; LPS (12), Nocardia water-soluble mitogen (NWSM) (18) and DxS (13). We have used the 5-bromo-deoxyuridine (BUdR) suicide technique of Zoschke and Bach (20). This should be a more critical method for ascertaining whether different cells respond to different mitogens by eliminating or inactivating those cells that respond to an initial mitogen.The specific killing by rabbit anti-mouse immunoglobulin serum and by antiIa alloantisera was used to investigate the presence of Ig and Ia on the surface of cells reactive to the above mitogens.1 Abbreviations used in this paper: BUdR, 5-bromo-deoxyuridine; CNRS, Centre National de la Recherche Scientifique; Con A, concanavalin A; DxS, dextran sulfate; LPS, lipopolysaccharide; NWSM, Nocardia water-soluble mitogen. 136J. ExP. MED.

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Section: Discussionmentioning

confidence: 99%

“…We have chosen three well-known B-cell mitogens for this study; LPS (12), Nocardia water-soluble mitogen (NWSM) (18) and DxS (13). We have used the 5-bromo-deoxyuridine (BUdR) suicide technique of Zoschke and Bach (20).…”

mentioning

confidence: 99%

Mitogenic analysis of murine B-cell heterogeneity.

Bona¹,

Yano²,

Dimitriu³

et al. 1978

The Journal of Experimental Medicine

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“…Single cell suspensions were obtained from spleens according to a previously described technique (7). Lymphocytes (1.5 x 10 e were cultured for 3 days in I ml RPML-1640 medium (Grand Island Biological Company, Grand Island, N.…”

Section: Methodsmentioning

confidence: 99%

“…[SH]thymidine (1 pCi) was added to each culture tube 12 h before harvesting the cells and incorporation of radioactivity was measured by liquid scintillation counting (7).…”

Section: Methodsmentioning

confidence: 99%

“…In rabbit in vitro systems, anti-allotypic antibodies induce blast transformation (3) and inhibit the synthesis of allotypes (4) whereas Nocardia water-soluble mitogen (NWSM) is a B-cell mitogen for several mammalian species (5,6) as polyclonal activator induces the differentiation of rabbit small lymphocytes to plasma cells and their polyclonal activation (7).…”

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Release from maternally-induced allotypic suppression in rabbit by Nocardia water-soluble mitogen.

Bona¹,

Cazenave²

1977

The Journal of Experimental Medicine

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In vivo allotypic suppression in rabbits has been described in newborn heterozygotes which were offspring of mothers immunized against the father's immunoglobulin aUotype (1) or which were injected at birth with antiserum directed against the father's allotype (2). This suppressed allotype is either entirely unexpressed or expressed in very low amounts (1).In rabbit in vitro systems, anti-allotypic antibodies induce blast transformation (3) and inhibit the synthesis of allotypes (4) whereas Nocardia water-soluble mitogen (NWSM) is a B-cell mitogen for several mammalian species (5, 6) as polyclonal activator induces the differentiation of rabbit small lymphocytes to plasma cells and their polyclonal activation (7).The aim of the present work was to study the synthesis of allotypes by lymphocytes from suppressed rabbits subsequent to in vitro stimulation by NWSM. Materials and MethodsAnimals. 4 day, 3, 6, and 12-wk-old offspring (b4/b5) were obtained from b4/b4 mothers immunized against the paternal allotype bS/b5 according to methods described by Dray (1), 12-too old offspring which had escaped from maternally allotypic suppression and 12-too old normal heterozygous b4/b5 rabbits (Pasteur Institute, Garches, France) were used in these experiments.Reagents. Rabbit antiallotypic sera directed against the a and b series allotypes were obtained according to Oudin (8) NWSM from Nocardia opaca was prepared according to Ciorbaru et al. (9) Concanavalin A (Con A) was obtained from Miles Yeda Ltd. and [aH]thyrnidine with 1 Ci/mM sp act was obtained from Commissariat a l'Energie Atomique, Saclay, France.Blast Transformation Assay. Single cell suspensions were obtained from spleens according to a previously described technique (7). Lymphocytes (1.5 x 10 e were cultured for 3 days in I ml RPML-1640 medium (Grand Island Biological Company, Grand Island, N. Y.) supplemented with 15% heat inactivated b6/b6 rabbit serum. The cultures were performed in plastic tubes which were incubated in an incubated model 1-H 100 Gallenkamp under a continuous flow mixture of 5% CO2 and 95% Air.[SH]thymidine (1 pCi) was added to each culture tube 12 h before harvesting the cells and incorporation of radioactivity was measured by liquid scintillation counting (7).Measure ofln Vitro Synthesis of AUotype. Splenic lymphocytes (25 x 10 e) were cultured for 72 h in 5 ml RPML-1640 medium supplemented with 10% fetal calf serum (Flow Laboratories, Inglewood, Calif.) in the same conditions described above.The amount of immunoglobulin in 72-h supernatant cultures was measured by a quantitative inhibition radioimmunoassay. IgG fractions were prepared from b4/b4 and b5/b5 homozygous rabbits by DEAE cellulose chromatography (10) and were labeled with ~5I, according to Green-* Present address:

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