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Bone marrow antimitogenic factor and its regulation by histamine
Cell Biology
International
Reports,
Vol. 14, Abstracts
Supplement
BONE MARROWANTIMITOGENIC FACTOR AND ITS REGULATION BY HISTAMINE Nicolai N Belyaev, Gulnara K Zakiryanova. Ajtkhozhin'e Institute of Molecular Biology and Biochemistry,Alma-Ata,480012,USSR. Bone marrow contains maatocytee and bacapable to releaee sophyl-like celle, autocoid histamine under the action of IL3. In this work modulating influence of histamine on syntheeie of antimitogenic factor by bone marrow cells (B&K) in vitro ie studied. Supernatant of 480hour long BMC culture nom BALB/c mice inhibited inclusion of C-timidine by ConA-induced splenocytea, tymoma EL-4 and hybrito ataphylococcua enterodomas, specific toxine and horiogonadotropine. Antibodies synthesis by hybridoma increased. Supernatant effect decreaeed at eolution to 1:128 Preliminary 3-hour long treatment of BMC with IO@! histamine reliably weekened antimitogenic effect of supernatant. Analoic effect of histamine waa noted at stu%y of supernatant on intact BMC. In the BMC culture histamine induced almost depressing of spontanic proliferation.Addion of 25mM N-acetyl-B-galactosamine to BMC culture stabilized effects of eupernatants intact onea a6 well as hietamine-stimulated BMC. It'8 euppoeed, that hi&amine induces releaee of factor, inhibiting grcwth of bone marrow natural suppressors and elaboration of antimitogenic factor by them.
P330
THE INTRACELLULAR DISTRIBUTION OF IL-Id.: CHANGES IN DIFFERENTIATING MONOCYTES. Gillian R. Bullock, Amanda E. Semper, Janis K. Lazdins. Ciba-Geigy AG, Postfach, CH-4002, Easel. The cytokine interleukin 1 (IL-l) exists in two forms, IL-I& and IL-S, which show only 25% homology at the protein level. IL-l is produced by monocytes and macrophages following stimulation with lipopolysaccharide (LPS). Using antibodies specific for either IL-la? or IL-16 and an indirect alkaline phosphatase immunocytochemical technique, we were able to demonstrate intracellular IL-l. In freshly-isolated human monocytes cultured for 8-12hrs in the presence of LPS (300ng/ml), 90-952 of monocytes showed immunoreactivity for IL-ltf, or IL-B which was distributed uniformly throughout the cytoplasm. Following 2 weeks in culture most monocytes differentiate into macrophages. Subseguent stimulation of these cells with LPS, using identical conditions, gave the following results: 1) The number of cells showing IL-l&immunoreactivity was significantly reduced (eg: from 95.8 t1.22 to 68.1t1.12 for cells from the same donor "IL" ) . Reductions in the number of IL-18 positive cells were less apparent. 2) Many of the macrophages stained positively for IL-W showed reaction product restricted to or in a predominantly perinuclear location (7421.7% for donor "x"). Distribution of IL-1B remained virtually unchanged. Thus, after maturation, fewer cells produce IL-15 the location of which changes from general cytoplasmic to perinuclear. The distihution of IL-l/3 remains unchanged. The precise subcellular location and the effect of "in viva" rather than "in vitro" maturation of macrophages are now being investigated
pa32
P329
P331
Siegfried
1990
161
ALTERATIONS IN Krauw,
OF I’UIMCYTE FUNCTIONS HYPERCHOLE STEROLEHIA Carmen Pohl, Andreas Wolfgang Liische. Institute Biochemistry, Medical Academy, Erfurt, Serman Dem. Rep.
Pohl , of pathological DDR-5010 There are evidences that monocytes play a primary role in the development of atherosclerosis. In the present study we investigate the influence of experimental and clinical hypercholesterolemia (HC) on variPUS functions of circulating monocytes. Monocytes were isolated by centrifugation on Ficoll-Paque from (i) pigs fed an atherogenic diet containing 2 X cholesterol and 20 X beef tallow for 16 weeks and (ii) patients with primary HC. It was determined the monocyte adhesion on artificial surf aces, aggregat i on of mononucl ear cells, migration belong a chemotactic gradient, phagocytosis and chemi1uminescence. In blood from pigs fed an atherogenic diet the monocyte count W8S significantly higher compared to the control group, but in general the monocyte reactivity (adhesion and phrgocytosisl is decreased. tlonocytes from patients with HC showed a depressed chemotaxis, but in contrast the adhesiveness of the monocytes is higher than in controls. The results indicate that HC may influence monocyte behaviour. Hypaerctivity in isolated monocytes may result from depletion of highly reactive monocytes in tha circulation or cell preparation. The during discrepancy between the results that were obtained in experimental and clinical HC requires further examination.
CEARACTBgIBATION FOR TECB P53 GAG
OF A HOAB SPECIFIC PBOTEIN OFTNENIV.
Mastraiicasa, N.Luisa Nolli, EdoardO Sari&i, narco Luigi Cwenaghi,*Febio Nalavasi 6 Mauririo Dmaro. Lepetit Research Center ,Germtmo, Varese, Italy. Wpnrtimento di Genetica, 8iolosir e Chimica Media MvcrsitA di Torino, Italy. Like other retroviruses, the gof the innnoddiciency virus (HIV) contains gag, pal and env gmes &ich code for the viral core proteins, vim1 enand the viral -lopa mpectively. These pmteim am translated as protein, pl-ewrsor polypeptides that we Past-translationally mcdified thrwgh proteolytic pwcwses md other modifications. In the case of the sag gene, its prc&ct is the p 53, a precursor polypcpride, that is cleaved, by the viral protease, into emrg which there is p 24, the major smiler pdypptides, internal core protein of NIV. Uith the aim of studying the protease activity in cells md tissues as nwker for viral infectivity, we proc*ced nmoclaal mtibcdics spcific for the p 53 of HIV. The deals with the stub/ P-t characterization of anab lC12 obtained using as imnnogm m antigen cmstitutd by c prtim of the p 53, cantainiw the ctewage pmtww. fused to the Ixtasire for the gslactosidase of E.&i. The lGl2 lnoab is reactive exclusively with the HIV part of the fusion Protein that is recognized before and not after clewape b# the NIV-1 protease. A further characterization dzmmstrated that nod, lGl2 birds only to the gag precursor p 53 and not to either p 24 or P 18, two of the pm&&s of the activation of p 53 by the protease, which are partially in&Ad in the fusiml protein. These results confirm that the qHtape nxos?ized by llDab 1Glt is in the gas portion of the fusim protein and that it is dcstruyed after the NIV-l that cleavage protease. was-tin) bv the recognitim site of the meb a? p 53 could be the cleavage site for the protease.
International
Reports,
Vol. 14, Abstracts
Supplement
BONE MARROWANTIMITOGENIC FACTOR AND ITS REGULATION BY HISTAMINE Nicolai N Belyaev, Gulnara K Zakiryanova. Ajtkhozhin'e Institute of Molecular Biology and Biochemistry,Alma-Ata,480012,USSR. Bone marrow contains maatocytee and bacapable to releaee sophyl-like celle, autocoid histamine under the action of IL3. In this work modulating influence of histamine on syntheeie of antimitogenic factor by bone marrow cells (B&K) in vitro ie studied. Supernatant of 480hour long BMC culture nom BALB/c mice inhibited inclusion of C-timidine by ConA-induced splenocytea, tymoma EL-4 and hybrito ataphylococcua enterodomas, specific toxine and horiogonadotropine. Antibodies synthesis by hybridoma increased. Supernatant effect decreaeed at eolution to 1:128 Preliminary 3-hour long treatment of BMC with IO@! histamine reliably weekened antimitogenic effect of supernatant. Analoic effect of histamine waa noted at stu%y of supernatant on intact BMC. In the BMC culture histamine induced almost depressing of spontanic proliferation.Addion of 25mM N-acetyl-B-galactosamine to BMC culture stabilized effects of eupernatants intact onea a6 well as hietamine-stimulated BMC. It'8 euppoeed, that hi&amine induces releaee of factor, inhibiting grcwth of bone marrow natural suppressors and elaboration of antimitogenic factor by them.
P330
THE INTRACELLULAR DISTRIBUTION OF IL-Id.: CHANGES IN DIFFERENTIATING MONOCYTES. Gillian R. Bullock, Amanda E. Semper, Janis K. Lazdins. Ciba-Geigy AG, Postfach, CH-4002, Easel. The cytokine interleukin 1 (IL-l) exists in two forms, IL-I& and IL-S, which show only 25% homology at the protein level. IL-l is produced by monocytes and macrophages following stimulation with lipopolysaccharide (LPS). Using antibodies specific for either IL-la? or IL-16 and an indirect alkaline phosphatase immunocytochemical technique, we were able to demonstrate intracellular IL-l. In freshly-isolated human monocytes cultured for 8-12hrs in the presence of LPS (300ng/ml), 90-952 of monocytes showed immunoreactivity for IL-ltf, or IL-B which was distributed uniformly throughout the cytoplasm. Following 2 weeks in culture most monocytes differentiate into macrophages. Subseguent stimulation of these cells with LPS, using identical conditions, gave the following results: 1) The number of cells showing IL-l&immunoreactivity was significantly reduced (eg: from 95.8 t1.22 to 68.1t1.12 for cells from the same donor "IL" ) . Reductions in the number of IL-18 positive cells were less apparent. 2) Many of the macrophages stained positively for IL-W showed reaction product restricted to or in a predominantly perinuclear location (7421.7% for donor "x"). Distribution of IL-1B remained virtually unchanged. Thus, after maturation, fewer cells produce IL-15 the location of which changes from general cytoplasmic to perinuclear. The distihution of IL-l/3 remains unchanged. The precise subcellular location and the effect of "in viva" rather than "in vitro" maturation of macrophages are now being investigated
pa32
P329
P331
Siegfried
1990
161
ALTERATIONS IN Krauw,
OF I’UIMCYTE FUNCTIONS HYPERCHOLE STEROLEHIA Carmen Pohl, Andreas Wolfgang Liische. Institute Biochemistry, Medical Academy, Erfurt, Serman Dem. Rep.
Pohl , of pathological DDR-5010 There are evidences that monocytes play a primary role in the development of atherosclerosis. In the present study we investigate the influence of experimental and clinical hypercholesterolemia (HC) on variPUS functions of circulating monocytes. Monocytes were isolated by centrifugation on Ficoll-Paque from (i) pigs fed an atherogenic diet containing 2 X cholesterol and 20 X beef tallow for 16 weeks and (ii) patients with primary HC. It was determined the monocyte adhesion on artificial surf aces, aggregat i on of mononucl ear cells, migration belong a chemotactic gradient, phagocytosis and chemi1uminescence. In blood from pigs fed an atherogenic diet the monocyte count W8S significantly higher compared to the control group, but in general the monocyte reactivity (adhesion and phrgocytosisl is decreased. tlonocytes from patients with HC showed a depressed chemotaxis, but in contrast the adhesiveness of the monocytes is higher than in controls. The results indicate that HC may influence monocyte behaviour. Hypaerctivity in isolated monocytes may result from depletion of highly reactive monocytes in tha circulation or cell preparation. The during discrepancy between the results that were obtained in experimental and clinical HC requires further examination.
CEARACTBgIBATION FOR TECB P53 GAG
OF A HOAB SPECIFIC PBOTEIN OFTNENIV.
Mastraiicasa, N.Luisa Nolli, EdoardO Sari&i, narco Luigi Cwenaghi,*Febio Nalavasi 6 Mauririo Dmaro. Lepetit Research Center ,Germtmo, Varese, Italy. Wpnrtimento di Genetica, 8iolosir e Chimica Media MvcrsitA di Torino, Italy. Like other retroviruses, the gof the innnoddiciency virus (HIV) contains gag, pal and env gmes &ich code for the viral core proteins, vim1 enand the viral -lopa mpectively. These pmteim am translated as protein, pl-ewrsor polypeptides that we Past-translationally mcdified thrwgh proteolytic pwcwses md other modifications. In the case of the sag gene, its prc&ct is the p 53, a precursor polypcpride, that is cleaved, by the viral protease, into emrg which there is p 24, the major smiler pdypptides, internal core protein of NIV. Uith the aim of studying the protease activity in cells md tissues as nwker for viral infectivity, we proc*ced nmoclaal mtibcdics spcific for the p 53 of HIV. The deals with the stub/ P-t characterization of anab lC12 obtained using as imnnogm m antigen cmstitutd by c prtim of the p 53, cantainiw the ctewage pmtww. fused to the Ixtasire for the gslactosidase of E.&i. The lGl2 lnoab is reactive exclusively with the HIV part of the fusion Protein that is recognized before and not after clewape b# the NIV-1 protease. A further characterization dzmmstrated that nod, lGl2 birds only to the gag precursor p 53 and not to either p 24 or P 18, two of the pm&&s of the activation of p 53 by the protease, which are partially in&Ad in the fusiml protein. These results confirm that the qHtape nxos?ized by llDab 1Glt is in the gas portion of the fusim protein and that it is dcstruyed after the NIV-l that cleavage protease. was-tin) bv the recognitim site of the meb a? p 53 could be the cleavage site for the protease.