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No production in a undifferentiated and differentiated human intestinal cell line
112
Poster Session 2C. Nitric Oxide
LPS (10 IlglmJ) repressed CYP2Bl mRNA after 3 hours. 'The effect was maximalafter 12 hours and persisted at 24 hours. When a nitric oxide synthase inhibitor was added to the culture media (30 to 300 IlM) . no difference was noted. To determine the efficiency of the inhibitorswe measured NO production. which was blocked when an inhibitor was in the culture medium. We concluded that NO was not involved in the repression of CYP2BI mRNA by LPS in rat hepatocytes,
I
P2C74
1 STUDIES ON THE EFFECT OF CARBON DISULFIDE ON NITRIC OXIDE AND LIPID PEROXIDATION
Zhang Wen-chang " Tu Li-qing. Department of Occupational Health. Fujian Medical University Fuzhou 350004. P.R. China Studies on the effect of CS2 on the rat nitric oxide/endothelium-derived relaxing factor (NOIEDRF) and it, mechanism were carried out, The results showed that serum and cardiac NO were reducedand were significantly lower in Csj-exposed rats (320.160 mg/kg) than in the controls, and serum LPO (lipid peroxidation) higher in rats exposed to CS2than in controls (p < O.O\), but changes cardiac NOS (nitric oxide synthases) activities had not been found (p > 0.05). The results of stepwise regression analysis (serum NO (y). CSz dose (x l ). serum LPO (x2), cardiac NOS (x S) and cardiac NO (x4) showed that there was a close correlation between serum ~O and LPO or the dose of CS2. When F critical value was 5, the serum LPO became the only factor in the stepwise regression equation (R = -0.6665, p < 0.01). We also found that the serum LPO content was significantly reduced, and NO level was significantly increased when some anti-oxidation substances (Sc, Zn, Vit E and Vit C) were taken by rats which were exposed to CS2 at the same time. So, it was suggested that the lipid peroxidation resulted from CS2 would accelerate the oxidation and decomposition of NO. and the NO level in the body was reduced which resulted in the toxic effect on rnycardiovascular system.
IP2C7S1
STREPTOMYCES ANULATUS INDUCED PRODUCTION OF NITRIC OXIDE AND CYTOTOXICITY IN HUMAN ALVEOLAR TYPE 11 EPITHEUAL CELLS (A549)
M.-R. Hirvoncn .1 ,J. Jussila1 , M. Ruotsalainen 1 • K. Savolainerr", A. Nevalainen 1 , J Division ofEnvironmental Health. National Public Healrh Institute. P.D. Box 95. FIN-7070I Kuopio; 2Department of Pharmacology and Toxicology, University of Kuopio, P.D. Box 1627. FIN-702// Kuopio, Finland Dampness and mold growth in buildings cause spore generation into indoor air. which is associated with respiratory tract disorders. Specific agents or cellular mechanisms of diseases have not yet been identified. A likely contributingfactor in the etiological mechanism of these respiratory symptoms could be inflammatory response towards specific organic materials in the microbes. NO plays an important role as an immune defense molecule. A variety of cells in the airways can be stimulated to produce large amounts of NO for long periods of time through stimulation of inducible NO synthase (iNOS). The ability of Streptomyces anulatus, gram positive bacteria which was isolated from moldy buildings. and lipopolysaccharide (LPS) or interferon gamma (IFN) to induce NO production and to cause cytotoxicity in human alveolar type II epithelial cells (A549) were studied. Here were report that Streptomyces anulatus or IFN induce a significant dose-, and time dependent increase in l'O-production and decrease in the cell viability of A549 cells. The effects induced by Streptomyces anulatus alone. were further increased by IFN. LPS was not able to induce NO-production in these cells or to cause cytotoxicity. The present results suggests that Streptomyces anulatus, initiates a cascade of events in human alveolar A549 cells leading to cytotoxicity and production of NO, which is believed to
he an important inflammatory mediator in the pathogenesisof many respiratory diseases.
IP2C76!
ARECA NUT EXTRACTS STIMULATE DNA STRAND BREAKS BY GENERATION OF NITRIC OXIDE
T.y. Liu ", C.W.Chi. Veterans General Hospital-Taipei. Taipei, Taiwan. Republic of China International Agency for Research on Cancer (!ARC) clearly stated that chewingbetel quid links to the development of oral cancer. Areca nut is the majorcomponentof betelquid. Previously, we havedemonstrated that areca nut extract (ANE) can generate reactive oxygen species (ROS) and cause oxidative DNA damage through hydrogen peroxide formation and Fenton-type reaction, In this study, we have investigated the possibility that nitric oxide is also involved in the ANE-mediated DNAdamages. By using alkaline single cell gel electrophoresis (SCGE), we have shown that Al'E-induced DNA strand breaks dose-dependently in bovine endothelial aortic cells (BEAC). S-methyl-L-thiocitrulline and Nw-nitro-L-arginine methyl ester, inhibitors of nitric oxide synthase, could suppress the ANE-induced DNA strand breaks. Increase of nitroryrosine, a stable product of nitric oxide, was also detected in ANE-treated cells. These results suggest that ANE treatment may generate nitric oxide to damage DNA and consequently may be important in the pathogenesis of oral cancer.
[P2C'7!] NO PRODUCTION IN A UNDIFFERENTIATED AND DIFFERENTIATED HUMAN INTESTINAL CELL LINE
A.L. Vignoli, F. Zucco v" , A. Stammati, R.C. Srivastava". Istituto Superiore di Sanaa, Roma; J Istituto di Tecnologie Biomediche. CNR. Roma, Italy: 2 Industrial Toxicology Research Centre, Lucknow; India Nitric oxide (NO) may play an important role in intestinal inflammation by acting on the barrier integrity. thus reducing its selective permeability. Few studies have been performed in vitro on human intestinal cell lines. An inducible NO synthase has been shown in HT29 and OLD-I cells following proinflammatory cytokine exposure. On Caco-2 cells, able to spontaneously differentiate in vitro to small intestinal enterocytes, only indirect evidence has been obtained of NO production. By the use of different biochemical methods. we have been able to showthat Caco-2cells are producing ~O following PMA + yll'F exposure. In undifferentiated cells (7 days of culture) the production is very low, while at the 21'1 day of culture. when the cells are completely differentiated. a consisted production is detected. This effect is inhibited by nitro-D-arginine methyl ester (D-NAME). but N-monomethyl-L-arginine (NMLA) is more effective, suggesting that an inducible NO synthase (NOS) is involved. Indeed. the NOS activity has been detected by 3H-argininclcitrulline conversion method. Further studies concerning thc induction of the enzyme are in progress,as well as its correlation with the cytoskeletal modification and barrier integrity.
[P2Ctii] PROSTAGLANDIN E
2 SYNTHESIS AFFECTED BY NITRIC OXIDE IN MACROPHAGES
Cecilia Guastadisegni .1. Maria Balduzzi'", Luisa Minghetti2 • Alessia Nicolini", Elisabetta Polazzi2 . JLaboratory of EnvironmentalHygiene; 2Neurobiology Section. Laboratory of Pathophysiology Istituto Superiore di Sanita; .I Biomedical and Toxicological Department. ENEA. Rome. Italy Prostaglandins (PGs). the arachidonic acid (AA) metabolites of
Poster Session 2C. Nitric Oxide
LPS (10 IlglmJ) repressed CYP2Bl mRNA after 3 hours. 'The effect was maximalafter 12 hours and persisted at 24 hours. When a nitric oxide synthase inhibitor was added to the culture media (30 to 300 IlM) . no difference was noted. To determine the efficiency of the inhibitorswe measured NO production. which was blocked when an inhibitor was in the culture medium. We concluded that NO was not involved in the repression of CYP2BI mRNA by LPS in rat hepatocytes,
I
P2C74
1 STUDIES ON THE EFFECT OF CARBON DISULFIDE ON NITRIC OXIDE AND LIPID PEROXIDATION
Zhang Wen-chang " Tu Li-qing. Department of Occupational Health. Fujian Medical University Fuzhou 350004. P.R. China Studies on the effect of CS2 on the rat nitric oxide/endothelium-derived relaxing factor (NOIEDRF) and it, mechanism were carried out, The results showed that serum and cardiac NO were reducedand were significantly lower in Csj-exposed rats (320.160 mg/kg) than in the controls, and serum LPO (lipid peroxidation) higher in rats exposed to CS2than in controls (p < O.O\), but changes cardiac NOS (nitric oxide synthases) activities had not been found (p > 0.05). The results of stepwise regression analysis (serum NO (y). CSz dose (x l ). serum LPO (x2), cardiac NOS (x S) and cardiac NO (x4) showed that there was a close correlation between serum ~O and LPO or the dose of CS2. When F critical value was 5, the serum LPO became the only factor in the stepwise regression equation (R = -0.6665, p < 0.01). We also found that the serum LPO content was significantly reduced, and NO level was significantly increased when some anti-oxidation substances (Sc, Zn, Vit E and Vit C) were taken by rats which were exposed to CS2 at the same time. So, it was suggested that the lipid peroxidation resulted from CS2 would accelerate the oxidation and decomposition of NO. and the NO level in the body was reduced which resulted in the toxic effect on rnycardiovascular system.
IP2C7S1
STREPTOMYCES ANULATUS INDUCED PRODUCTION OF NITRIC OXIDE AND CYTOTOXICITY IN HUMAN ALVEOLAR TYPE 11 EPITHEUAL CELLS (A549)
M.-R. Hirvoncn .1 ,J. Jussila1 , M. Ruotsalainen 1 • K. Savolainerr", A. Nevalainen 1 , J Division ofEnvironmental Health. National Public Healrh Institute. P.D. Box 95. FIN-7070I Kuopio; 2Department of Pharmacology and Toxicology, University of Kuopio, P.D. Box 1627. FIN-702// Kuopio, Finland Dampness and mold growth in buildings cause spore generation into indoor air. which is associated with respiratory tract disorders. Specific agents or cellular mechanisms of diseases have not yet been identified. A likely contributingfactor in the etiological mechanism of these respiratory symptoms could be inflammatory response towards specific organic materials in the microbes. NO plays an important role as an immune defense molecule. A variety of cells in the airways can be stimulated to produce large amounts of NO for long periods of time through stimulation of inducible NO synthase (iNOS). The ability of Streptomyces anulatus, gram positive bacteria which was isolated from moldy buildings. and lipopolysaccharide (LPS) or interferon gamma (IFN) to induce NO production and to cause cytotoxicity in human alveolar type II epithelial cells (A549) were studied. Here were report that Streptomyces anulatus or IFN induce a significant dose-, and time dependent increase in l'O-production and decrease in the cell viability of A549 cells. The effects induced by Streptomyces anulatus alone. were further increased by IFN. LPS was not able to induce NO-production in these cells or to cause cytotoxicity. The present results suggests that Streptomyces anulatus, initiates a cascade of events in human alveolar A549 cells leading to cytotoxicity and production of NO, which is believed to
he an important inflammatory mediator in the pathogenesisof many respiratory diseases.
IP2C76!
ARECA NUT EXTRACTS STIMULATE DNA STRAND BREAKS BY GENERATION OF NITRIC OXIDE
T.y. Liu ", C.W.Chi. Veterans General Hospital-Taipei. Taipei, Taiwan. Republic of China International Agency for Research on Cancer (!ARC) clearly stated that chewingbetel quid links to the development of oral cancer. Areca nut is the majorcomponentof betelquid. Previously, we havedemonstrated that areca nut extract (ANE) can generate reactive oxygen species (ROS) and cause oxidative DNA damage through hydrogen peroxide formation and Fenton-type reaction, In this study, we have investigated the possibility that nitric oxide is also involved in the ANE-mediated DNAdamages. By using alkaline single cell gel electrophoresis (SCGE), we have shown that Al'E-induced DNA strand breaks dose-dependently in bovine endothelial aortic cells (BEAC). S-methyl-L-thiocitrulline and Nw-nitro-L-arginine methyl ester, inhibitors of nitric oxide synthase, could suppress the ANE-induced DNA strand breaks. Increase of nitroryrosine, a stable product of nitric oxide, was also detected in ANE-treated cells. These results suggest that ANE treatment may generate nitric oxide to damage DNA and consequently may be important in the pathogenesis of oral cancer.
[P2C'7!] NO PRODUCTION IN A UNDIFFERENTIATED AND DIFFERENTIATED HUMAN INTESTINAL CELL LINE
A.L. Vignoli, F. Zucco v" , A. Stammati, R.C. Srivastava". Istituto Superiore di Sanaa, Roma; J Istituto di Tecnologie Biomediche. CNR. Roma, Italy: 2 Industrial Toxicology Research Centre, Lucknow; India Nitric oxide (NO) may play an important role in intestinal inflammation by acting on the barrier integrity. thus reducing its selective permeability. Few studies have been performed in vitro on human intestinal cell lines. An inducible NO synthase has been shown in HT29 and OLD-I cells following proinflammatory cytokine exposure. On Caco-2 cells, able to spontaneously differentiate in vitro to small intestinal enterocytes, only indirect evidence has been obtained of NO production. By the use of different biochemical methods. we have been able to showthat Caco-2cells are producing ~O following PMA + yll'F exposure. In undifferentiated cells (7 days of culture) the production is very low, while at the 21'1 day of culture. when the cells are completely differentiated. a consisted production is detected. This effect is inhibited by nitro-D-arginine methyl ester (D-NAME). but N-monomethyl-L-arginine (NMLA) is more effective, suggesting that an inducible NO synthase (NOS) is involved. Indeed. the NOS activity has been detected by 3H-argininclcitrulline conversion method. Further studies concerning thc induction of the enzyme are in progress,as well as its correlation with the cytoskeletal modification and barrier integrity.
[P2Ctii] PROSTAGLANDIN E
2 SYNTHESIS AFFECTED BY NITRIC OXIDE IN MACROPHAGES
Cecilia Guastadisegni .1. Maria Balduzzi'", Luisa Minghetti2 • Alessia Nicolini", Elisabetta Polazzi2 . JLaboratory of EnvironmentalHygiene; 2Neurobiology Section. Laboratory of Pathophysiology Istituto Superiore di Sanita; .I Biomedical and Toxicological Department. ENEA. Rome. Italy Prostaglandins (PGs). the arachidonic acid (AA) metabolites of