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Pokeweed mitogen-induced B cell differentiation: Enhancement by phorbol myristic acetate (PMA) and the hypoxanthine analogue NPT 15392
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POKEWEED MITOGEN°INDUCED B CELL DIFFERENTIATION: ENHANCEMENT BY PHORBOL MYRISTIC ACETATE (PMA) AND THE HYPOXANTHINE ANALOGUE NPT 15392. A. Pompidou, B. Mace, N. Duchet, J. Lehmeyer and M. D. Cooper Hopital Saint Vincent de Paul, Paris, France and University of Alabama in Birmingham, USA Blood mononuclear cells from 6 normal adults were p a r t i a l l y depleted of macrophages by plastic adherence and cultured (lOO/ml/culture) in the presence of pokeweed mitogen extract (PWM, 1/200 di1.), phorbol myristic acetate (PMA, 100 rig.) and/or NPT 15392~(100 ng). The culture medium was RPMI plus 10% fetal calf serum, gentamycin and 2xlO-~M 2-ME. The cells and supernatants were harvested at i week, and B cell differentiation was assessed by immunofluorescence enumeration of plasmablasts and mature plasma cells and by radioimmunoassay of secreted IgM. PMA, a known inducer of B cell differentiation, enhanced plasmablast but not terminal plasma cell d i f f e r e n t i a t i o n , and this was reflected by an insignificant effect on IgM secretion. Pie4 induced both the development of plasmablasts and IgM plasma cell maturation. PI~ s i g n i f i c a n t l y enhanced PWM-induced B cell differentiation and IgM secretion (>2-fold increase). Similarly, NPT alone enhanced the development of plasmablasts, but not plasma cell differentiation, and also enhanced PWMinduced differentiation of IgM secreting cells. The results suggest that PMA and NPT act primarily as inducers of plasmablasts, which require additional stimulation for terminal plasma cell differentiation.
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DTC, AN IMMUNOPOTENTIATOR SPECIFIC FOR T CELLS. M. Roumiantzeff, C. Charbonnier, J. Armand, M. Renoux, G. Renoux. I n s t i t u t Merieux, 17 rue Bourgelat, 69002, Lyon, France. Sodium diethyldithiocarbamate, DTC, comply with the requirements for an agent of known pharmacology, and a c t i v i t i e s on the immune system. DTC is devoid of carcinogenicity, does not induce lymphoproliferative disorders, and is of low t o x i c i t y . As a biological response modifier, DTC demonstrates a unique a c t i v i t y on the T-cell lineage, as well as a regulatory influence inducing a restoration to normal levels of either excessive or impaired responses. These effects are mediated through an increased production of hormone-like factors specific for T cells. Phase I - I I studies confirm in man both the non-toxicity of DTC and i t s effectiveness as a biological and regulatory agent. Since DTC possesses also a degree of antibiotic a c t i v i t y , and a protective effect against chemically-induced cancers, one might expect i t w i l l be a useful drug to treat primary immunodeficiencies (at the example of i t s a c t i v i t y on nude mice), secondary immunodeficiencies associated with aging, infections, cancer and immunosuppressive therapies, and hopefully, autoimmune diseases due to a faltering T-cell population.
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ACTIVATION MECHANISMS OF HUMAN LYMYHOCYTES BY THE INOSINE DERIVATIVE, ISOPRINOSINE(R)(IP) A. Morin*, D. Tello**, J.J. Ballet**, *I~,munothdrapie Expdrimen~ale, Institut Pasteur, 28, rue du Dr Roux, 75015 Paris, France. **INSERM U 108, Hopital St-Louis, 75010 Paris, France. Enhancement of T cell-proliferative response to mitogens and antigens is indeed the most consistent effect of IF. This may be mediated in part by IL2, as we have found that IP can enhance IL2 production. On the other hand, we have observed that IP is able to augment the growth of a variety of lymphoblastoid cell lines including non T-cell. Since IF contains an inosine molecule, it can be envisaged that the stimulatory activity may involve its utilization in the purine pathway of the cell. It was therefore interesting to study with radiolabelled material the role of uncomplexed inosine compared co its complexed form IP. We found that the uptake of I~C inoslne by normal human mononuclear cells was increased when DIP-PAcBA (the other portion of the complex) was added. The uptake of uncomplexed I~C inosine was inhibited by IP and vice-versa, suggesting a cow,on mechanism of uptake. The uptake of inosine was relatively specific as suggested by its inhibition with inosine, hypoxanthine and to a lesser extent adenosine, but not by cytosine, thymidine, histamine or deoxyadenosine. These data suggest that an increased availability of inosine is provided by its complexed form IF. This might account, at least partially, for the stimulating effects of the immunomedulator.
73
POKEWEED MITOGEN°INDUCED B CELL DIFFERENTIATION: ENHANCEMENT BY PHORBOL MYRISTIC ACETATE (PMA) AND THE HYPOXANTHINE ANALOGUE NPT 15392. A. Pompidou, B. Mace, N. Duchet, J. Lehmeyer and M. D. Cooper Hopital Saint Vincent de Paul, Paris, France and University of Alabama in Birmingham, USA Blood mononuclear cells from 6 normal adults were p a r t i a l l y depleted of macrophages by plastic adherence and cultured (lOO/ml/culture) in the presence of pokeweed mitogen extract (PWM, 1/200 di1.), phorbol myristic acetate (PMA, 100 rig.) and/or NPT 15392~(100 ng). The culture medium was RPMI plus 10% fetal calf serum, gentamycin and 2xlO-~M 2-ME. The cells and supernatants were harvested at i week, and B cell differentiation was assessed by immunofluorescence enumeration of plasmablasts and mature plasma cells and by radioimmunoassay of secreted IgM. PMA, a known inducer of B cell differentiation, enhanced plasmablast but not terminal plasma cell d i f f e r e n t i a t i o n , and this was reflected by an insignificant effect on IgM secretion. Pie4 induced both the development of plasmablasts and IgM plasma cell maturation. PI~ s i g n i f i c a n t l y enhanced PWM-induced B cell differentiation and IgM secretion (>2-fold increase). Similarly, NPT alone enhanced the development of plasmablasts, but not plasma cell differentiation, and also enhanced PWMinduced differentiation of IgM secreting cells. The results suggest that PMA and NPT act primarily as inducers of plasmablasts, which require additional stimulation for terminal plasma cell differentiation.
74
DTC, AN IMMUNOPOTENTIATOR SPECIFIC FOR T CELLS. M. Roumiantzeff, C. Charbonnier, J. Armand, M. Renoux, G. Renoux. I n s t i t u t Merieux, 17 rue Bourgelat, 69002, Lyon, France. Sodium diethyldithiocarbamate, DTC, comply with the requirements for an agent of known pharmacology, and a c t i v i t i e s on the immune system. DTC is devoid of carcinogenicity, does not induce lymphoproliferative disorders, and is of low t o x i c i t y . As a biological response modifier, DTC demonstrates a unique a c t i v i t y on the T-cell lineage, as well as a regulatory influence inducing a restoration to normal levels of either excessive or impaired responses. These effects are mediated through an increased production of hormone-like factors specific for T cells. Phase I - I I studies confirm in man both the non-toxicity of DTC and i t s effectiveness as a biological and regulatory agent. Since DTC possesses also a degree of antibiotic a c t i v i t y , and a protective effect against chemically-induced cancers, one might expect i t w i l l be a useful drug to treat primary immunodeficiencies (at the example of i t s a c t i v i t y on nude mice), secondary immunodeficiencies associated with aging, infections, cancer and immunosuppressive therapies, and hopefully, autoimmune diseases due to a faltering T-cell population.
75
ACTIVATION MECHANISMS OF HUMAN LYMYHOCYTES BY THE INOSINE DERIVATIVE, ISOPRINOSINE(R)(IP) A. Morin*, D. Tello**, J.J. Ballet**, *I~,munothdrapie Expdrimen~ale, Institut Pasteur, 28, rue du Dr Roux, 75015 Paris, France. **INSERM U 108, Hopital St-Louis, 75010 Paris, France. Enhancement of T cell-proliferative response to mitogens and antigens is indeed the most consistent effect of IF. This may be mediated in part by IL2, as we have found that IP can enhance IL2 production. On the other hand, we have observed that IP is able to augment the growth of a variety of lymphoblastoid cell lines including non T-cell. Since IF contains an inosine molecule, it can be envisaged that the stimulatory activity may involve its utilization in the purine pathway of the cell. It was therefore interesting to study with radiolabelled material the role of uncomplexed inosine compared co its complexed form IP. We found that the uptake of I~C inoslne by normal human mononuclear cells was increased when DIP-PAcBA (the other portion of the complex) was added. The uptake of uncomplexed I~C inosine was inhibited by IP and vice-versa, suggesting a cow,on mechanism of uptake. The uptake of inosine was relatively specific as suggested by its inhibition with inosine, hypoxanthine and to a lesser extent adenosine, but not by cytosine, thymidine, histamine or deoxyadenosine. These data suggest that an increased availability of inosine is provided by its complexed form IF. This might account, at least partially, for the stimulating effects of the immunomedulator.