- Email: [email protected]
Effects of diesel exhaust particles on human airway epithelial cells in vitro
224
IOP4D87I
Poster Session 4D. Inhalation Toxicology NEWBIOSOLUBLE INSULATION FIBRES
P. Dachowski * 1, D. M. Bernstein-, C. Morscheidt", llsov er St. Gobain, Paris; 2Consultant in Toxicology, Geneva, Switzerland Over the past ten years, the Isover division of SI. Gobain has undertaken an extensive scientific program to understand what factors differentiate the potential of different fiber compositions to cause disease. In this context 12 fiber compositions have been evaluated directly and an additional 6 fibers as part of a joint program in studies ranging from acellular in-vitro solubility to inhalation biopersistence, intra-tracheal biopersistence, 28-day inhalation toxicity and intra-peritoneal injection. The se data and the relationship which has been developed and widely accepted of fiber biosolubility to potential health effects contributed significantly to the ability of the European Commission to develop the recent Directive 97/69IEC on fibers in December 1997 and have allowed SI. Gobain to develop a new generation of biosoluble glass and stone-wool insulation materials. The data across this wide range of studies has been found to be very coherent. They strongly indicate that it is the fibers which are longer than the macrophage can fully engulf which are most important in determining potential pathological response and that the bio-solubility of these fibers is closely related to actual response. These studies have also shown that fiber composition is directly related to bio-solubility, that the bio-solubility is the primary determinant in the ability of the fibers to persist in the lung, and that the number of fibers which persist is related to pathogenic response. The biopersistence of the new bio-soluble fibers developed by 51. Gobain is less than 10 days (weighted clearance half-time), indicating that these fibers are very rapidly dissolved after deposition in the lung without any pathogenic response. This is in contrast to durable fibers which can persist for years or crocidolite/amosite asbestos fibers which can persist for the human life-time without any degradation both of which have been shown to produce tumours in chronic inhalation and IF studies. These studies have been essential to the ability of Saint Gobain to put into commercial use biosoluble fibers which according to the EC directive are completely exempt from carcinogenicity classification. These fibers have been thoroughly tested for efficacy to assure that they meet all industrial and building requirements including high fire resistance and good ageing properties.
IOP4D88I
PARTICLES OF RESIDUALOIL FLYASH (ROFA) INDUCETOXICITY AND MUCINHYPERSECRETION IN RODENTAIRWAY EPITHELIALCELLS IN VITRO VIA AN OXIDANT-MEDIATED MECHANISM
K.B. Adler* , N.F. liang, I.A. Dye, K.L. Dreher. Col. Vet. Med., North Carolina State University, Raleigh, NC & U.S. Environmental Protection Agency, Research Triangle Park, NC, USA Inhalation of ambient air particulate matter (PM's) is associated with lung injury and inflammation. We have developed a technique to culture guinea pig (GPTE) and rat tracheal epithelial (RTE) cells in airlliquid interface, which maintains their differentiated structure and function in vitro. We exposed these cells to suspensions of 6 different urban ambient air PM samples, as well as dust from the Mt. SI. Helens' volcanic eruption of May 1979 (negative control). Effects of these particulates on cell viability (assessed via LDH assay) and secretion of mucin (measured by a monoclonal antibody-based ELISA) were determined . ROFA was the most toxic of the dusts, significantly increasing LDH release by 24 hours at relatively low concentrations (lOOJiglml). ROFA also enhanced mucin secretion after 8 hrs. exposure. ROFA exposure induced production of intracellular oxidants by the cells, as determined by dihydrorhodamine oxidation, and also depleted the cells of the antioxidant, glutathione. The oxidant scavenger, dimethylthiourea (DMTU), added to the cells
at 0.4 to 40 mM, attenuated ROFA-induced cytotoxicity and mucin secretion in a concentration-dependent manner. These results suggest that ROFA-induced cytotoxicity and mucin hypersecretion involve intracellularly-generated oxidants, and the delay in induction of cell injury and mucin secretion in response to ROFA may reflect the time necessary to deplete cellular antioxidant defenses. Supported by Grant # CR824355 from U.S. EPA, and HL 36982 from NIH
IOP4D89 I
ASSESSING TOXICOLOGICAL EFFECTSOF COMPOUNDS ADSORBEDTO PARTICULATE MATTERUSINGTHE NEUTRAL RED UPTAKE BIOASSAY WITHHUMAN BRONCHIALEPITHELIAL CELLS
L. Maldonado-Baez *, DR Reyes, B.D. Jimenez, J.F. Rodriguez. U
P R/Medical Sciences Campus, San Juan, PR, USA Epidemiological studies performed throughout various regions of the world alludes to the fact that airborne particulate matter is one of the principal causes for respiratory illnesses, acute bronchitis, and asthmatic attacks. However, a direct correlation between the toxicity of these particulate matter extracts and specific components within the extracts have not been established. Toxicity testing of many urban particulate matter have depended greatly on the use of the traditional mutagenic Salmonella/microsome bioassay (Ames test) for establishing the mutagenicity of these extracts. Thi s research intends to bridge the gap between toxicity testing in humans and conventional mutagenicity assays. We have recently used normal human epidermal keratynocytes (NHEK) to test the toxic effect of adsorbed compounds from particulate, matter. In order to expand our analyses of toxic effects on airborne particulate, matter and respiratory tract diseases, we have incorporated an additional Neutral Red Bioassay using bronchial epithelial cells. Extracts from a urban dust standard reference material (5RM 1649), as well as extracts obtained from total suspended particles (T5P), and particulate matter of less than 10 Jim (PM IO) were obtained using the EPA standard sampling and Soxhlet extraction methodology. Three different solvents of distinct polarities were employed in the extraction procedure. The toxicity of each extract was evaluated using a human cell line. Comparisons between the NRSO obtained using both types of cells the human keratinocytes and the bronchial tracheal cells were found to be within the same range. The three SRM extracts ranged from I to 8 /LglmL, which is in agreement with previous results obtained using NHEK. None of the control extracts from a non industrialized area exhibited any detectable level of toxicity during winter. However, extracts from an industrialized area which were concurrently monitored during the same period exhibited toxic characteristics. There NRSO values fluctuated from 10 to 30 /Lg/mL.
IOP4D90 I EFFECTSOF DIESEL EXHAUSTPARTICLES ON HUMAN AIRWAY EPITHELIALCELLS IN VITRO
Sonja Boland *1 , Armelle Baeza1 , Thierry Foumiec2, Michel Aubier-'. Francelyne Marano 1 • J Laboratoire de
Cytophysiologie et Toxicologie Cellulaire. Universite Paris \Ill. 2 place Jussieu, 75005 Paris; 2°lnserm U408, BP 416, 75870 Paris cedex 18, France A diesel automobile emits some 30 to 100 times more particles than a comparable gasoline vehicle and about 90% of diesel exhaust particles (DEP) are of respirable size. DEP contain carbon nuclei which absorb a large variety of compounds among which are polycyclic aromatic hydrocarbons that are suspected to be responsible for the toxicity of DEP. The potential toxicity of DEP on the respiratory epithelium was studied by comparison with carbon black particles using cell culture systems of nasal and bronchial human epithelial
PosterSession 4D. Inhalation Toxicology cells. Viability assays (trypan blue exclusion and lactico deshydrogenase release) have shown that DEP from I to 10 J-Lg/cm 2 induce a dose dependent membrane damage after 5 days of treatment (but not after 24 hours) without a significant mortality. Moreover from 4 hours of treatment an endocytosis of DEP can be observed that increases with time treatment. The particles were confined in large vacuoles and we have demonstrated the existence of transcytosis. However the treated cells presenting phagocytosed particles exhibit no ultrastructural lesions. Phagocytosis of DEP, quantified by cytometric measurements of right angle light scatter, is a time and dose dependent event but not a specific event to DEP. On the other hand, DEP led to an increased release of the proinflamrnatory cytokins GM-CSF and IL8 after 24 and 48 hours of exposure whereas carbon black particles have a clearly lower effect. These data indicate that DEP are largely endocytosed by respiratory epithelial cells without producing an immediate cytotoxic effect. In addition they induce an inflammatory response which seems specific.
225
TNF, Interleukin-6 IL-6) in AM supernatants was measured with isotopic labling method and MIT colorimetric assay respectively. AM supernatants with WI-38 fibroblast were cultured in vitro, then the fibroblast proliferation and collagen production were examined by 3H-TdR and 14C-Proline incorporation The hydroxy-proline of fibroblast was analysed with chloramine-T and inhibiting assay of fibroblast proliferation and collagen synthesis were carried out by using anti-TNF antibody and IFNy. The results show that mineral dusts can induce AM to release TNF and IL-6. The level of TNF were 852, 1369 and 1198U/rnl, and the level of IL-6 were 1335, 1336 and 1511U/ml respectively, which were higher than those in the Ti02 control. The lung fibroblast proliferation and collagen synthesis can be increased by the supernatant of the mineral dust treated AM. The cpm in 3H_TdR were 3201, 4750 and 2669U/ml respectively. The cpm in 14C-Proline were 18538, 13416 and 27952U/ml (200J-Lg/ml) respectively. The HOP content in WI-38 cell were 21.3, 22.4 and 24.0J-Lg/ml. Both anti-TNF antibody and IFN y can inhibit the fibroblast proliferation and decrease collagen synthesis.
This study was supported by Ademe and Renault (DIMAT).
I OP4D91 119EADJUVANT ACTIVITY OF POLYSTYRENE
IOP4D93! TOXICOLOGICAL DIFFERENCES BASED ON EXTRACTION PROCEDURES FOR ATMOSPHERIC PARTICULATE MATTER
PARTICLES IN FOURSTRAINSOF MICE
B. Granum *1 , P.I. Gaarder", M. Levik1 • I Department of
Environmental Medicine, National Institute of Public Health 2DepartmentofImmunology and Transfusion Medicine, Ullevdl University Hospital, Oslo, Norway Diesel exhaust particles are reported to increase the immune response to the model allergen ovalbumin. There is evidence that the non-extractable particle core is responsible for part of this adjuvant activity. The aim of our study was to investigate the activity of particles free from adsorbed chemical substances - that is, polystyrene particles - in different strains of mice. Four different strains of mice were used: Hsd:NIHS (outbred), NIH/ala (inbred), BALB/c (inbred) and C3H1HeJ (inbred). Particles and ovalbumin (P-ovalbumin) or ovalbumin alone were given intraperitoneally to different groups of eight mice each on days 1 and 16. All groups were reinjected with ovalbumin alone on day 43. Blood samples were drawn on days 0, 41 and 48. The serum levels of IgE anti-ovalbumin, total IgE, IgG2a anti-ovalbumin and total IgG2a were measured. In outbred. and inbred NIH mice given P-ovalbumin, the concentration of IgE anti-ovalbumin, total IgE and IgG2a anti-ovalbumin was increased compared to mice given ovalbumin alone. The concentration of total IgG2a remained unchanged. The production of total IgG2a was suppressed in BALB/c and C3H1HeJ mice given P-ovalbumin compared to mice given ovalbumin alone, while the production of total IgE was increased. By contrast, P-ovalbumin had no significant effects on the production of IgE anti-ovalbumin or IgG2a anti-ovalbumin in BALB/c or C3H1HeJ mice. Additional experiments are being performed, both with intraperitoneal immunisation and aerosol exposure, to elucidate these findings. Our results indicate that polystyrene particles had an adjuvant effect on the specific inunune response, including IgE, in outbred and inbred NIH mice. Our results further indicate that the genetic background is an important factor for the adjuvant effect of particles on allergy related immune responses.
IOP4D92I
EFECTOF CYTOKINE REDUCED BY MINERAL DUSTSON FIBROBLASTIN VITRO
Li Ren Zhou *, jian Hua Zhou, Ji Cheng Young. Department of
Radiation Medicine, Suzhon Medical College, P.R. China Rabbit's alveolar macrophages obtained by lavage were cultured with mineral dust in vitro. The level of cytokine (Tumor necrosis factor
D.R. Reyes *, O. Rosario, J.E Rodriguez, B.D. Jimenez. UPR, Departmentsof Chemistry and Biochemistry, San Juan, Puerto Rico, USA Respiratory illneses have been associated with levels of particulate matter in many epidemiological studies. An Industrialized area (Catano) in Puerto Rico has been declared by the Environmental Quality Board as a municipality with a history exceeding the National Air Quality Standards. The Public Health Department has also reported the highest cancer rates of all types for the same municipality. Total suspended particles (TSP) and particulate matter of less than 10 J-Lm (PM I 0) were collected throughout the year in the industrialized and a non-industrialized area from the East Coast and Puerto Rico. The particulate matter from both experimental and reference sites as well as a standard reference material (SRM 1649) was processed using Soxhlet and microwave assisted extraction (MAE) for comparison purposes. The extracts obtained with three solvents of different polarities were evaluated for their relative cytotoxicity on human keratinocytes using the Neutral Red Bioassay (NRB). The NR50 obtained using the Soxhlet extraction procedure ranged from 5 to 160 J-Lg/mL with most of the moderately-polar extracts displaying no detectable toxicity. However, the microwave extraction method generated higher toxicity values for the extracts obtained with our conditions. The NR50s for the SRM extracts obtained using the MAE ranged from 0.5 to 2 J-Lg/mL for the non-polar extract, 10 to 15 J-Lg/mL for the moderately-polar and from 30 to 36 J-Lg/mL for the polar solvents. Parallel Soxhlet extraction performed on the same samples generated NR50 of 6, 43 and 42 J-Lg/mL with ascending polarities. Chemical characterization of the various extract are on their way for both extraction methodologies in order to aid in the identification of responsible toxicological constituents.
IOP4D94!
MUTAGENICITY AND CHEMICALANALYSIS OF AIRBORNE PARTICULATE IN A RUBBER MANUFACTURING PLANT
M.E. Fracasso *, G. Perbellini, P. Franceschetti, A. Piubelli, L. Romeo. Institute of Pharmacology, and OccupationalHealth,
University of Verona, Italy Epidemiological studies conducted in the '80s revealed that people working in the rubber manufacturing industry have an increased risk of cancer and to despite the measures adopted to improve the situation, workers employed in rubber processing are still at risk. The
IOP4D87I
Poster Session 4D. Inhalation Toxicology NEWBIOSOLUBLE INSULATION FIBRES
P. Dachowski * 1, D. M. Bernstein-, C. Morscheidt", llsov er St. Gobain, Paris; 2Consultant in Toxicology, Geneva, Switzerland Over the past ten years, the Isover division of SI. Gobain has undertaken an extensive scientific program to understand what factors differentiate the potential of different fiber compositions to cause disease. In this context 12 fiber compositions have been evaluated directly and an additional 6 fibers as part of a joint program in studies ranging from acellular in-vitro solubility to inhalation biopersistence, intra-tracheal biopersistence, 28-day inhalation toxicity and intra-peritoneal injection. The se data and the relationship which has been developed and widely accepted of fiber biosolubility to potential health effects contributed significantly to the ability of the European Commission to develop the recent Directive 97/69IEC on fibers in December 1997 and have allowed SI. Gobain to develop a new generation of biosoluble glass and stone-wool insulation materials. The data across this wide range of studies has been found to be very coherent. They strongly indicate that it is the fibers which are longer than the macrophage can fully engulf which are most important in determining potential pathological response and that the bio-solubility of these fibers is closely related to actual response. These studies have also shown that fiber composition is directly related to bio-solubility, that the bio-solubility is the primary determinant in the ability of the fibers to persist in the lung, and that the number of fibers which persist is related to pathogenic response. The biopersistence of the new bio-soluble fibers developed by 51. Gobain is less than 10 days (weighted clearance half-time), indicating that these fibers are very rapidly dissolved after deposition in the lung without any pathogenic response. This is in contrast to durable fibers which can persist for years or crocidolite/amosite asbestos fibers which can persist for the human life-time without any degradation both of which have been shown to produce tumours in chronic inhalation and IF studies. These studies have been essential to the ability of Saint Gobain to put into commercial use biosoluble fibers which according to the EC directive are completely exempt from carcinogenicity classification. These fibers have been thoroughly tested for efficacy to assure that they meet all industrial and building requirements including high fire resistance and good ageing properties.
IOP4D88I
PARTICLES OF RESIDUALOIL FLYASH (ROFA) INDUCETOXICITY AND MUCINHYPERSECRETION IN RODENTAIRWAY EPITHELIALCELLS IN VITRO VIA AN OXIDANT-MEDIATED MECHANISM
K.B. Adler* , N.F. liang, I.A. Dye, K.L. Dreher. Col. Vet. Med., North Carolina State University, Raleigh, NC & U.S. Environmental Protection Agency, Research Triangle Park, NC, USA Inhalation of ambient air particulate matter (PM's) is associated with lung injury and inflammation. We have developed a technique to culture guinea pig (GPTE) and rat tracheal epithelial (RTE) cells in airlliquid interface, which maintains their differentiated structure and function in vitro. We exposed these cells to suspensions of 6 different urban ambient air PM samples, as well as dust from the Mt. SI. Helens' volcanic eruption of May 1979 (negative control). Effects of these particulates on cell viability (assessed via LDH assay) and secretion of mucin (measured by a monoclonal antibody-based ELISA) were determined . ROFA was the most toxic of the dusts, significantly increasing LDH release by 24 hours at relatively low concentrations (lOOJiglml). ROFA also enhanced mucin secretion after 8 hrs. exposure. ROFA exposure induced production of intracellular oxidants by the cells, as determined by dihydrorhodamine oxidation, and also depleted the cells of the antioxidant, glutathione. The oxidant scavenger, dimethylthiourea (DMTU), added to the cells
at 0.4 to 40 mM, attenuated ROFA-induced cytotoxicity and mucin secretion in a concentration-dependent manner. These results suggest that ROFA-induced cytotoxicity and mucin hypersecretion involve intracellularly-generated oxidants, and the delay in induction of cell injury and mucin secretion in response to ROFA may reflect the time necessary to deplete cellular antioxidant defenses. Supported by Grant # CR824355 from U.S. EPA, and HL 36982 from NIH
IOP4D89 I
ASSESSING TOXICOLOGICAL EFFECTSOF COMPOUNDS ADSORBEDTO PARTICULATE MATTERUSINGTHE NEUTRAL RED UPTAKE BIOASSAY WITHHUMAN BRONCHIALEPITHELIAL CELLS
L. Maldonado-Baez *, DR Reyes, B.D. Jimenez, J.F. Rodriguez. U
P R/Medical Sciences Campus, San Juan, PR, USA Epidemiological studies performed throughout various regions of the world alludes to the fact that airborne particulate matter is one of the principal causes for respiratory illnesses, acute bronchitis, and asthmatic attacks. However, a direct correlation between the toxicity of these particulate matter extracts and specific components within the extracts have not been established. Toxicity testing of many urban particulate matter have depended greatly on the use of the traditional mutagenic Salmonella/microsome bioassay (Ames test) for establishing the mutagenicity of these extracts. Thi s research intends to bridge the gap between toxicity testing in humans and conventional mutagenicity assays. We have recently used normal human epidermal keratynocytes (NHEK) to test the toxic effect of adsorbed compounds from particulate, matter. In order to expand our analyses of toxic effects on airborne particulate, matter and respiratory tract diseases, we have incorporated an additional Neutral Red Bioassay using bronchial epithelial cells. Extracts from a urban dust standard reference material (5RM 1649), as well as extracts obtained from total suspended particles (T5P), and particulate matter of less than 10 Jim (PM IO) were obtained using the EPA standard sampling and Soxhlet extraction methodology. Three different solvents of distinct polarities were employed in the extraction procedure. The toxicity of each extract was evaluated using a human cell line. Comparisons between the NRSO obtained using both types of cells the human keratinocytes and the bronchial tracheal cells were found to be within the same range. The three SRM extracts ranged from I to 8 /LglmL, which is in agreement with previous results obtained using NHEK. None of the control extracts from a non industrialized area exhibited any detectable level of toxicity during winter. However, extracts from an industrialized area which were concurrently monitored during the same period exhibited toxic characteristics. There NRSO values fluctuated from 10 to 30 /Lg/mL.
IOP4D90 I EFFECTSOF DIESEL EXHAUSTPARTICLES ON HUMAN AIRWAY EPITHELIALCELLS IN VITRO
Sonja Boland *1 , Armelle Baeza1 , Thierry Foumiec2, Michel Aubier-'. Francelyne Marano 1 • J Laboratoire de
Cytophysiologie et Toxicologie Cellulaire. Universite Paris \Ill. 2 place Jussieu, 75005 Paris; 2°lnserm U408, BP 416, 75870 Paris cedex 18, France A diesel automobile emits some 30 to 100 times more particles than a comparable gasoline vehicle and about 90% of diesel exhaust particles (DEP) are of respirable size. DEP contain carbon nuclei which absorb a large variety of compounds among which are polycyclic aromatic hydrocarbons that are suspected to be responsible for the toxicity of DEP. The potential toxicity of DEP on the respiratory epithelium was studied by comparison with carbon black particles using cell culture systems of nasal and bronchial human epithelial
PosterSession 4D. Inhalation Toxicology cells. Viability assays (trypan blue exclusion and lactico deshydrogenase release) have shown that DEP from I to 10 J-Lg/cm 2 induce a dose dependent membrane damage after 5 days of treatment (but not after 24 hours) without a significant mortality. Moreover from 4 hours of treatment an endocytosis of DEP can be observed that increases with time treatment. The particles were confined in large vacuoles and we have demonstrated the existence of transcytosis. However the treated cells presenting phagocytosed particles exhibit no ultrastructural lesions. Phagocytosis of DEP, quantified by cytometric measurements of right angle light scatter, is a time and dose dependent event but not a specific event to DEP. On the other hand, DEP led to an increased release of the proinflamrnatory cytokins GM-CSF and IL8 after 24 and 48 hours of exposure whereas carbon black particles have a clearly lower effect. These data indicate that DEP are largely endocytosed by respiratory epithelial cells without producing an immediate cytotoxic effect. In addition they induce an inflammatory response which seems specific.
225
TNF, Interleukin-6 IL-6) in AM supernatants was measured with isotopic labling method and MIT colorimetric assay respectively. AM supernatants with WI-38 fibroblast were cultured in vitro, then the fibroblast proliferation and collagen production were examined by 3H-TdR and 14C-Proline incorporation The hydroxy-proline of fibroblast was analysed with chloramine-T and inhibiting assay of fibroblast proliferation and collagen synthesis were carried out by using anti-TNF antibody and IFNy. The results show that mineral dusts can induce AM to release TNF and IL-6. The level of TNF were 852, 1369 and 1198U/rnl, and the level of IL-6 were 1335, 1336 and 1511U/ml respectively, which were higher than those in the Ti02 control. The lung fibroblast proliferation and collagen synthesis can be increased by the supernatant of the mineral dust treated AM. The cpm in 3H_TdR were 3201, 4750 and 2669U/ml respectively. The cpm in 14C-Proline were 18538, 13416 and 27952U/ml (200J-Lg/ml) respectively. The HOP content in WI-38 cell were 21.3, 22.4 and 24.0J-Lg/ml. Both anti-TNF antibody and IFN y can inhibit the fibroblast proliferation and decrease collagen synthesis.
This study was supported by Ademe and Renault (DIMAT).
I OP4D91 119EADJUVANT ACTIVITY OF POLYSTYRENE
IOP4D93! TOXICOLOGICAL DIFFERENCES BASED ON EXTRACTION PROCEDURES FOR ATMOSPHERIC PARTICULATE MATTER
PARTICLES IN FOURSTRAINSOF MICE
B. Granum *1 , P.I. Gaarder", M. Levik1 • I Department of
Environmental Medicine, National Institute of Public Health 2DepartmentofImmunology and Transfusion Medicine, Ullevdl University Hospital, Oslo, Norway Diesel exhaust particles are reported to increase the immune response to the model allergen ovalbumin. There is evidence that the non-extractable particle core is responsible for part of this adjuvant activity. The aim of our study was to investigate the activity of particles free from adsorbed chemical substances - that is, polystyrene particles - in different strains of mice. Four different strains of mice were used: Hsd:NIHS (outbred), NIH/ala (inbred), BALB/c (inbred) and C3H1HeJ (inbred). Particles and ovalbumin (P-ovalbumin) or ovalbumin alone were given intraperitoneally to different groups of eight mice each on days 1 and 16. All groups were reinjected with ovalbumin alone on day 43. Blood samples were drawn on days 0, 41 and 48. The serum levels of IgE anti-ovalbumin, total IgE, IgG2a anti-ovalbumin and total IgG2a were measured. In outbred. and inbred NIH mice given P-ovalbumin, the concentration of IgE anti-ovalbumin, total IgE and IgG2a anti-ovalbumin was increased compared to mice given ovalbumin alone. The concentration of total IgG2a remained unchanged. The production of total IgG2a was suppressed in BALB/c and C3H1HeJ mice given P-ovalbumin compared to mice given ovalbumin alone, while the production of total IgE was increased. By contrast, P-ovalbumin had no significant effects on the production of IgE anti-ovalbumin or IgG2a anti-ovalbumin in BALB/c or C3H1HeJ mice. Additional experiments are being performed, both with intraperitoneal immunisation and aerosol exposure, to elucidate these findings. Our results indicate that polystyrene particles had an adjuvant effect on the specific inunune response, including IgE, in outbred and inbred NIH mice. Our results further indicate that the genetic background is an important factor for the adjuvant effect of particles on allergy related immune responses.
IOP4D92I
EFECTOF CYTOKINE REDUCED BY MINERAL DUSTSON FIBROBLASTIN VITRO
Li Ren Zhou *, jian Hua Zhou, Ji Cheng Young. Department of
Radiation Medicine, Suzhon Medical College, P.R. China Rabbit's alveolar macrophages obtained by lavage were cultured with mineral dust in vitro. The level of cytokine (Tumor necrosis factor
D.R. Reyes *, O. Rosario, J.E Rodriguez, B.D. Jimenez. UPR, Departmentsof Chemistry and Biochemistry, San Juan, Puerto Rico, USA Respiratory illneses have been associated with levels of particulate matter in many epidemiological studies. An Industrialized area (Catano) in Puerto Rico has been declared by the Environmental Quality Board as a municipality with a history exceeding the National Air Quality Standards. The Public Health Department has also reported the highest cancer rates of all types for the same municipality. Total suspended particles (TSP) and particulate matter of less than 10 J-Lm (PM I 0) were collected throughout the year in the industrialized and a non-industrialized area from the East Coast and Puerto Rico. The particulate matter from both experimental and reference sites as well as a standard reference material (SRM 1649) was processed using Soxhlet and microwave assisted extraction (MAE) for comparison purposes. The extracts obtained with three solvents of different polarities were evaluated for their relative cytotoxicity on human keratinocytes using the Neutral Red Bioassay (NRB). The NR50 obtained using the Soxhlet extraction procedure ranged from 5 to 160 J-Lg/mL with most of the moderately-polar extracts displaying no detectable toxicity. However, the microwave extraction method generated higher toxicity values for the extracts obtained with our conditions. The NR50s for the SRM extracts obtained using the MAE ranged from 0.5 to 2 J-Lg/mL for the non-polar extract, 10 to 15 J-Lg/mL for the moderately-polar and from 30 to 36 J-Lg/mL for the polar solvents. Parallel Soxhlet extraction performed on the same samples generated NR50 of 6, 43 and 42 J-Lg/mL with ascending polarities. Chemical characterization of the various extract are on their way for both extraction methodologies in order to aid in the identification of responsible toxicological constituents.
IOP4D94!
MUTAGENICITY AND CHEMICALANALYSIS OF AIRBORNE PARTICULATE IN A RUBBER MANUFACTURING PLANT
M.E. Fracasso *, G. Perbellini, P. Franceschetti, A. Piubelli, L. Romeo. Institute of Pharmacology, and OccupationalHealth,
University of Verona, Italy Epidemiological studies conducted in the '80s revealed that people working in the rubber manufacturing industry have an increased risk of cancer and to despite the measures adopted to improve the situation, workers employed in rubber processing are still at risk. The