Evaluation of the Developmental Toxicity of Ethylene Glycol Aerosol in CD-1 Mice by Nose-Only Exposure

Ethylene glycol (EG; CAS No. 107-21-1) is teratogenic to miceby whole-body (WB) exposure to aerosol (1000–2500 mg/m³The WB results were confounded by possible exposure from ingestionafter grooming andior from percutaneous absorption. Therefore,CD-1 mice were exposed to EG aerosol (MIMAD 2.6 ± 1.7µm) on Gestational Days (GD) 6 through 15, 6 hr/day, bynose-only (NO) (0, 500, 1000, or 2500 mg/rn or WB exposures(0 or 2100 mg/m³ as positive control), 30/group. Five additional"satellite" females each at 2500 mg/m³ NO and 2100mg/m³ WB wereexposed on GD 6 for measurement of EG on fur. Control environmentswere water aerosol (4200 mg/rn for NO; 2700 mg/rn for WB). Femaleswere weighed and evaluated for clinical signs and water consumptionthroughout gestation. On GD 18, maternal uterus, liver, andkidneys (2) were weighed, with kidneys examined microscopically.Corpora lutea and implantation sites were recorded. Live fetuseswere weighed, sexed, and examined for structural alterations.For NO dams, kidney weights were increased at 1000 and 2500mg/m³ no renal lesions and no other treatment-related maternaltoxicity were observed. There were no effects on pre- or postimplantationloss; fetal body weights/litter were reduced at 2500 mg/m³ At2500 mg/m³ incidences of fused ribs and skeletal variationswere increased. The 2500 mg/rn NO satellite animals had {smalltilde}330 mg/kg extractable EG. The WB group exhibited maternaland developmental toxicity including increased fetal skeletalmalformations and variations, confirming previous results, with1390 mg/kg extractable EG on fur. Therefore, exposure of CD-1mice to a respirable EG aerosol during organogenesis by NO inhalationresulted in minimal maternal toxicity at 1000 and 2500 mg/m³and developmental toxicity at 2500 mg/m³ The NOAEL was 500 mg/m³NO for maternal and 1000 mg/m³ NO for develop mental toxicity.This study supports the interpretation of the initial EG WBresults as due to systemic exposure from noninhala tion routessince limiting noninhalation routes prevented almost all ofthe effects (including teratogenicity) observed in mice afterWB exposure.     

The developmental toxicity of ethylene glycol in rats and mice

Timed-pregnant CD rats and CD-1 mice were dosed by gavage with ethylene glycol (EG) in distilled water on gestational days (gd) 6 through 15 (0, 1250, 2500, or 5000 mg kg-1 day-1 for rats; and 0, 750, 1500, or 3000 mg kg-1 day-1 for mice). Females were observed daily during treatment, but no maternal deaths or distinctive clinical signs were noted. Dose-related decreases in maternal weight gain during treatment were significant at all doses in rats and at the mid and high doses in mice. Gravid uterine weight was reduced in both species at the mid and high doses, and corrected maternal gestational weight gain showed a significant decreasing trend. At termination (gd 20, rats; gd 17, mice), the status of uterine implantation sites was recorded, and live fetuses were weighed and examined for external, visceral, and skeletal malformations. Dose-related increases in postimplantation loss per litter were observed in both species with the high dose significantly above controls only in rats. Fetal body weight per litter was significantly reduced at the mid and high doses in rats and at all doses in mice. The percentage of malformed live fetuses per litter and/or the percentage of litters with malformed fetuses was significantly elevated in all EG dose groups and greater than 95% of litters were affected at the high dose for each species. A wide variety of malformations were observed; the most common in both species were craniofacial and neural tube closure defects and axial skeletal dysplasia. EG produced severe developmental toxicity in two rodent species at doses that apparently failed to produce any serious maternal effects.     

Decreased Fetal Weights in Rats Exposed to Sidestream Cigarette Smoke

Pregnant Sprague-Dawley rats were exposed to sidestream Cigarettesmoke (SS) for 6 hr a day, at a concentration of 1 mg/m³ ofrespirable total suspended particulate material (TSP) on Days3, 6–10, and 13–17 of pregnancy. Controls were keptin an identical chamber without smoke exposure. The animalswere killed on Day 20 of gestation. No differences were foundin maternal body weight gain or average daily food consumptionbetween the smoke-exposed and control groups. The numbers offetuses and of implantation sites per litter were comparableamong the groups. None of the pups showed any gross malformationsand no difference was found between controls and SS-exposedpups when examined for reduced skeletal ossifications. However,there was a small but significant reduction in mean pup weight.We conclude that intermittent exposure of rats to sidestreamcigarette smoke at concentrations severalfold greater than thoseencountered in smokey public indoor environments causes intrauterinegrowth retardation.     

Developmental Toxicity Evaluation of Ethylene Glycol by Gavage in New Zealand White Rabbits

Artificially inseminated New Zealand white (NZW) rabbits wereadministered ethylene glycol (EG) by gavage on Gestational Days(GD) 6 through 19 at doses of 0, 100, 500, 1000, or 2000 mg/kg/day,with 23–24 inseminated animals per group. Clinical signswere recorded and water consumption was measured daily; doeswere weighed on GD 0, 6–19, 25, and 30. At necropsy (GD30), maternal liver, kidney, and gravid uterine weights wererecorded. Histopathologic examination was performed on kidneysfrom 10 does/dose and for all unscheduled deaths. Ovarian corporalutea were counted and uterine implantation sites (total sites,resorptions, dead and live fetuses) were recorded. All livefetuses were weighed, sexed, and examined for external, visceral,and skeletal malformations and variations. EG resulted in profoundmaternal toxicity at 2000 mg/kg/day (42% mortality; three earlydeliveries and one spontaneous abortion) associated with renalpathology and unaccompanied by any other indicators of maternaltoxicity. Renal lesions at 2000 mg/kg/day involved the corticalrenal tubules and included intraluminal oxalate crystals, epithelialnecrosis, and tubular dilatation and degeneration. No dose-relatedmaternal toxicity occurred at 100–1000 mg/kg/day. Therewas no indication of developmental toxicity at any dose tested,including no effects on pre- or postimplantation loss, numberof fetuses, fetal body weight, or sex ratio (% male fetuses)per litter, and no evidence of teratogenicity. The "no observableadverse effect level" (NOAEL) for maternal toxicity was therefore1000 mg/kg/day and the NOAEL for developmental toxicity wasat least 2000 mg/kg/day in this study. The sensitivity of NZWrabbits relative to that of Sprague—Dawley rats and Swissmice for maternal and developmental toxicity from gavage administrationof EG during organogenesis can be determined for maternal toxicity:rabbits>mice>rats, and for developmental toxicity, mice>>rats >> rabbits.     

The Pulmonary Response and Clearance of Ludox Colloidal Silica after a 4-Week Inhalation Exposure in Rats

Rats were exposed to Ludox colloidal silica (CS) at concentrationsof 0, 10, 50, and 150 mg/m³ for 6 hr/day, 5 days/week for 4weeks. Rats were killed after 4 weeks of exposure and 10 daysor 3 months post exposure (PE). The exposure concentration of10 mg/m³ Ludox CS is considered to be the no-effect concentration.There were no exposure-related clinical signs in any group.After 4 weeks exposure, lung weights were increased significantlyin rats exposed to 50 and 150 mg/m³ Ludox CS, but lung weightswere similar to those of controls at 3 months PE. After 4 weeksexposure to 50 mg/m³ Ludox CS, a slight alveolar macrophageresponse, polymorphonuclear leukocytic infiltration, and TypeII pneumocyte hyperplasia in alveolar duct regions were present.After 3 months PE, these pulmonary lesions had almost disappearedwith removal of most dust-laden alveolar macrophages (AMs).The pulmonary response to 150 mg/m³ Ludox CS was similar incharacter but increased in magnitude from that seen at 50 mg/m³At 3 months PE, most particleladen AMs had disappeared and theremaining AMs were aggregated and sharply demarcated. A fewaggregates of particle-laden AMs appeared to transform intosilicotic nodules comprising macrophages, epithelioid cells,and lymphocytic infiltration in some animals. Some silicoticnodules showed reticular fiber networks with minute collagenfiber deposition. Tracheobronchial lymph nodes were enlargedwith aggregates of particle-laden AMs and hyperplastic histiocyticcells. Lung-deposited Ludox cleared rapidly from the lungs withhalf-times of approximately 40 and 50 days for the 50 and 150mg/m³ groups, respectively.     

Determination of a No-Observed-Effect Level for Developmental Toxicity of Ethylene Glycol Administered by Gavage to CD Rats and CD-1 Mice

Previous studies have indicated that ethylene glycol (EG) isa developmental toxicant in rats and mice primarily when ingested.This study was designed to establish no-observed-effect levels(NOELs) for developmental toxicity of EG administered by gavagein both rodent species. Dams were administered EG on GestationDays 6–15; rats were given 0, 150, 500, 1000, or 2500mg EG/kg/ day; mice were dosed with 0, 50, 150, 500, or 1500mg EG/kg/day. In rat dams given 2500 mg EG/kg/day, water consumptionwas increased during treatment and body weights were reducedthroughout gestation; liver and kidney weights were increasedat euthanization (Gestation Day 21). Relative liver weightswere also increased at 1000 mg/kg/day. Effects observed in ratfetuses at 2500 mg/kg/day included the following hydrocephaly;gastroschisis; umbilical hernia; fused, duplicated, or missingarches, centra, and ribs; poor ossification in thoracic andlumbar regions; and reduced body weights. Reduced body weights,duplicated or missing ribs, centra, and arches, and poor ossificationwere also ob served in rat fetuses at 1000 mg/kg/day. In mice,there was no apparent treatment-related maternal toxicity. Inmouse fetuses (Gestation Day 18), effects were observed at 1500mg/kg/day and included reduced body weights, fused ribs andarches, poor ossification in thoracic and lumbar centra, andincreased occurrence of an extra 14th rib. At 500 mg/kg/day,slight reductions in fetal body weight and increased incidencesof extra ribs were observed. Under conditions of these studies,NOELs for developmental toxicity were 500 mg/kg/day for ratsand 150 mg/kg/day for mice, indicating that mice were more susceptiblethan rats to the teratogenic effects of EG.     

Four-Week Inhalation Toxicity Study with Ludox Colloidal Silica in Rats: Pulmonary Cellular Responses

Four-Week Inhalation Toxicity Study with Ludox Colloidal Silicain Rats: Pulmonary Cellular Responses. WARHEIT, D. B., CARAKOSTAS,M. C, KELLY, D. P., AND HARTSKY, M. A. (1991). Fundam. Appl.Toxicol. 16, 590–601. This study was designed to complementa traditional subchronic inhalation toxicity study with Ludoxcolloidal silica. CD rats were exposed nose-only for 2 or 4weeks at concentrations of 0, 10, 50, and 150 mg/m³ Ludox (driedSiO₂). Additional groups of rats exposed for 4 weeks were givena 3-month recovery period. Following exposure and/or recovery,fluids and cells were recovered from the lungs by bronchoalveolarlavage (BAL) and measured for cellular and biochemical parameters.Additional groups of animals were processed for cell labelingstudies or lung deposition studies. Inhaled doses of Ludox colloidalsilica were measured after 4-week exposures and were found tobe 489 µg/lung (10 mg/m³ group), 2418 µg/lung (50mg/m³), and 7378 µg/lung (150 mg/m³), respectively. Resultsshowed that exposures to 150 mg/m³ Ludox for 2 or 4 weeks producedpulmonary inflammation along with increases in BAL protein,LDH, and alkaline phosphatase values (p<0.05) and reducedmacrophage phagocytosis. Inflammatory responses, evidenced byincreased numbers of neutrophils, were also measured in thelungs of the 50 mg/m³ group following 2 and/or 4 weeks of exposure.Most biochemical parameters for all groups returned to controlvalues following a 3-month recovery period. Autoradiographicstudies demonstrated that the labeling indices of terminal bronchiolarand lung parenchymal cells were generally increased in the 50and 150 mg/m³ groups after 2 and 4 weeks of exposure but, withone exception, returned to normal levels following a 3-monthpostexposure period. No significant alterations in any measuredparameters were detected in rats exposed to 10 mg/m³ Ludox atany time postexposure. The determination of a no-observable-effectlevel (NOEL) of 10 mg/m³ was consistent with results obtainedby conventional toxicology methods and affirms the utility ofthese biochemical, cellular, and autoradiographic techniquesfor providing a predictive screen to assess the toxicity ofinhaled particles.     

Deposition of Cigarette Smoke Particles in the Rat

The fractional deposition of cigarette smoke particles in therespiratory tracts of rats was studied. Male and female ratswere conditioned in nose-only exposure tubes 25 min/day for2 days, exposed to cigarette smoke at mass concentrations of95 or 341 mg/m³ 25 min/day for 3 days, and then exposed to smokeat mass concentrations of 212 and 657 mg/m³, 25 min/day for5 days. Mainstream cigarette smoke was generated by a modifiedWalton smoking machine from two 1R3 research cigarettes burnedsequentially for each exposure. Deposition studies were conductedby placing the rats in plethysmograph tubes to allow respiratoryminute volume measurements during exposure, then exposing themto [¹⁴C] cigarette smoke at mass concentrations of 202 or 624mg/m³ for 25 min, using the same smoking machine. Size distribution,real-time concentration, and ¹⁴C activity of the smoke particleswere determined using a multijet Mercer impactor, a real-timeaerosol monitor, and filter samples, respectively. Immediatelyafter the exposure, the rats were terminated to determine thedistribution of the ¹⁴C. Individual lung lobes, trachea andlobar bronchi, head, larynx, kidneys, liver, gastrointestinal(GI) tract, blood, and depelted carcass of each rat were analyzedfor ¹⁴C content. Results showed that the GI tract contained16–31% of the total activity, indicating significant clearancefrom the large airways and nose to the GI tract during the exposureand during the 10–15 min between the cessation of theexposure and the removal ofthe organs. Total deposition of theinhaled ¹⁴C activity was 20.1 ? 1.6% for both exposure concentrations.The intrapulmonary deposition fractions (lung lobes plus airwaysbelow the lobar bronchi) were 12.4 ? 0.9 and 15.9 ? 1.4% forconcentrations of 202 and 624 mg/m³ respectively, suggestinga slight enhancement in upper airway deposition for animalsexposed to the higher smoke concentration.     

Pulmonary Fibrosis Produced in F-344 Rats by Subchronic Inhalation of Aerosols of a 4000 Molecular Weight Ethylene Oxide/Propylene Oxide Polymer

Pulmonary Fibrosis Produced in F-344 Rats by Subchronic Inhalationof Aerosols of a 4000 Molecular Weight Ethylene Oxide/PropyleneOxide Polymer. KLONNE, D. R., DODD, D. E., Losco, P. E., TROUP,C. M., AND TYLER, T. R. (1988), Fundam Appl. Toxicol 10, 682–690.Inhalation of aerosols of the ethylene oxide/propylene oxidepolymer (U-5100) evaluated in this study has previously beenshown in acute and 2-week studies to produce toxicologic effectson the lungs, with increased lung weights and microscopic findingsof congestion and hemorrhage of pulmonary alveolar capillariesand necrosis of alveolar epithelial cells (D. R. KLONNE, D.J. NACHREINER, D. E. DODD, P. E. Losco, AND T. R. TYLER, 1987,Fundam. Appl. Toxicol. 9, 7737–784). In the present studies,F-344 rats were exposed 6 hr/day, 5 day/week for 2 weeks toaerosols at mean concentrations of 0,0.9, or 5.0 mg/m³ or for13 weeks to mean concentrations of 0, 0.3, 1.1, or 5.2 mg/m³.Following the 2-week study, minimal multifocal hemorrhage andeosinophilic proteinaceous debris in alveoli were observed inthe 0.9 mg/m³ group; similar lesions plus alveolar cell necrosiswere found in the 5 mg/m³ group. In the 13-week study, the 5.2mg/m³ group had a slight decrease in body weight gain, whileincreases in absolute and/or relative lung weights occurredfor both the 1.1 and 5.2 mg/m³ groups at the end of the exposureregimen and at the end of a 5-week recovery period. Histologiclesions of the lungs occurred in all U-5100-exposed groups andconsisted of hemorrhage, alveolar histiocytosis, interstitialpneumonia, and multifocal fibrosis. The incidence and severityof the pulmonary lesions were concentration related. At theend of the 5-week recovery period, there was little change inthe severity or incidence of the pulmonary lesions in the 1and 5 mg/m³ groups when compared to rats killed the day afterthe termination of exposures. In conclusion, exposure to aerosolsof U-5100 for 13 weeks produced generally slight but biologicallysignificant pulmonary fibrosis in rats at all the exposure concentrationstested in this study.     

Developmental Toxicity Evaluation of Inhaled Methyl Isobutyl Ketone in Fischer 344 Rats and CD-1 Mice

Developmental Toxicity Evaluation of Inhaled Methyl IsobutylKetone in Fischer 344 Rats and CD-1 Mice. Tyl, R. W., FRANCE,K. A., FISHER, L. C., PRITTS, I. M., TYLER, T. R., PHILLIPS,R. D., and MORAN, E. J. (1987). Fundam. Appl. Toxicol. 8, 310–327.Pregnant Fischer 344 rats and CD-1 mice were exposed to methylisobutyl ketone vapor (CAS No. 108-10-1) by inhalation on GestationalDays 6 through 15 at concentrations of 0, 300, 1000, or 3000ppm (mean analytical values of 0, 305, 1012, and 2997 ppm, respectively).The animals were sacrificed on Gestational Day 21 (rats) or18 (mice), and live fetuses were examined for external, visceral,and skeletal alterations. In rats, exposure to 3000 ppm resultedin maternal toxicity expressed as clinical signs, decreasedbody weight and body weight gain, increased relative kidneyweight, and decreased food consumption, and in fetotoxicityexpressed as reduced fetal body weight per litter and reductionsin skeletal ossification. In mice, exposure to 3000 ppm resultedin maternal toxicity expressed as exposure-related increasesin deaths (12.0%, 3/25 dams), clinical signs, and increasedabsolute and relative liver weight, and in fetotoxicity expressedas increased incidence of dead fetuses, reduced fetal body weightper litter, and reductions in skeletal ossification. No treatment-relatedincreases in embryotoxicity or fetal malformations were seenin either species at any exposure concentration tested. Therewas no evidence of treatment-related maternal, embryo, or fetaltoxicity (including malformations) at 1000 or 300 ppm in eitherSpecies.     

Dose-dependent nonlinear pharmacokinetics of ethylene glycol metabolites in pregnant (GD 10) and nonpregnant Sprague-Dawley rats following oral administration of ethylene glycol.

The kinetics of orally administered ethylene glycol (EG) and its major metabolites, glycolic acid (GA) and oxalic acid (OX), in pregnant (P; gestation day 10 at dosing, GD 10) rats were compared across doses, and between pregnant and nonpregnant (NP) rats. Groups of 4 jugular vein-cannulated female rats were administered 10 (P and NP), 150 (P), 500 (P), 1000 (P), or 2500 (P and NP) mg (13)C-labelled EG/kg body weight. Serial blood samples and urine were collected over 24-hr postdosing, and analyzed for EG, GA, and OX using GC/MS techniques. Pharmacokinetic parameters including Cmax, Tmax, AUC, and betat((1/2)) were determined for EG and GA. Pregnancy status (GD 10-11) had no impact on the pharmacokinetic parameters investigated. Blood levels of GA were roughly dose-proportional from 10 to 150 mg EG/kg, but increased disproportionately from 500 to 1000 mg EG/kg. EG and GA exhibited dose-dependent urinary elimination at doses > or = 500 mg EG/kg, probably due to saturation of metabolic conversion of EG to GA, and of GA to downstream metabolites. The shift to nonlinear kinetics encompassed the NOEL (500 mg EG/kg) and LOEL (1000 mg EG/kg) for developmental toxicity of EG in rats, providing additional evidence for the role of GA in EG developmental toxicity. The peak maternal blood concentration of GA associated with the LOEL for developmental toxicity in the rat was quite high (363 microg/g or 4.8 mM blood). OX was a very minor metabolite in both blood and urine at all dose levels, suggesting that OX is not important for EG developmental toxicity.     

Prechronic Inhalation Toxicity Studies of 2-Mercaptobenzimidazole (2-MBI)in F344/N Rats

2-Mercaptobenzimidazole (2-MBI), used in rubber processing,is a suspect carcinogen structurally related to ethylene thiourea.The inhalation toxicity of 2-MBI was evaluated in male and femaleF344/N rats exposed 6 hr/day, 5 days/week to respirable aerosolsgenerated by spray atomization of aqueous suspensions of the2-MBI powder and subsequent drying of the resulting aerosols.Twelve exposures at target concentrations of 0, 6.3, 12.5, 25.0,50.0, or 100 mg/m³ of 2-MBI produced a dose-related reductionin body weight gains, thyroid follicular cell hyperplasia, adrenalcortex fatty change, and pituitary atrophy. Sub-chronic exposureswere conducted at target concentrations of 0, 3.1, 6.2, 12.5,25.0, and 50.0 mg/m³ of 2-MBI. Rats at 25 mg/m³ displayed hunchedposture, hypoactivity, and reduced body weight gain, with compoundrelated mortality at the highest exposure level. Anemia; increasedSGPT, SGOT, alkaline phosphatase, sorbitol dehydrogenase, BUN,and cholesterol; and reduced free fatty acid were seen in ratsat 25 mg/m³. Increased thyroid weight and thyroid follicularcell hyperplasia were noted in both sexes at 6.2 mg/m³, withreduced triiodothyronine and thyroxine levels in both sexesat > 12.5 mg/m³. Thyroid follicular cell hyperplasia wasalso seen in rats at 3.1 mg/m³. Thymus weights were significantlyreduced in both sexes at all exposure levels with liver weightincreases at 6.2 mg/m³. Exposure-related histopathologic changesincluded pituitary cytoplasmic vacuolization, adrenal cortexnecrosis, lymphoid depletion, thymic atrophy, liver cell hypertrophy,renal mineralization and tubular atrophy, and hypocellularityof the bone marrow.     

A 90-Day Inhalation Toxicity Study with Benomyl in Rats

A 90-Day Inhalation Toxiaty Study with Benomyl in Rats. WARHEIT,D. B., KELLY, D. P., CARAKOSTAS, M. C., AND SINGER, A. W. (1989).Fundam Appl Toxicol./ 12, 333-345. Benomyl [methyl 1-(butylcarbamoyl)-2-benzimidazolecarbamate,CAS Registry No. 17804-35-2] is a fungicide and the possibilityfor inhalation exposure exists for field workers. To assessthe toxicity of benomyl, groups of 20 male and 20 female CDrats were exposed nose-only 6 hr a day, 5 days a week, to concentrationsof 0, 10, 50 or 200 mg/m³ of a benomyl atmosphere. At the midpoint(approximately 45 days on test) and at the end of the exposureperiod, blood and urine samples for clinical evaluation werecollected from 10 rats/group/sex, and these animals were sacrificedfor pathological examination. Similar evaluations were performadon all remaining rats at the end of the 90-day test period.After approximately 45 days on test, compoundrelated degenerationof the olfactory epithelium was observed in all males and in8 of 10 female rats exposed to 200 mg/m³ benomyl. Two male ratsexposed to 50 mg/m³ had similar, although less severe, areasof olfactory epithelial degeneration. After approximately 90days of exposure, the remaining 10 rats/group/sex were sacrificedand examined. Of these rats, all of the males and females exposedto 200 mg/m³ had olfactory degeneration, along with 3 malesexposed to 50 mg/m³ of benomyl. No other observed lesions wereinterpreted to have been caused by the benomyl exposure. Inaddition, male rats exposed to 200 mg/m³ benomyl had depressedmean body weights compared to controls and this finding correlatedwith a reduction in food consumption. Based on pathologicalobservations, 10 mg/m³ represents the no-observable-effect level(NOEL) for the male rats, and 50 mg/m³ is the NOEL for the femalerats.     

Use of Lung Toxicity and Lung Particle Clearance to Estimate the Maximum Tolerated Dose (MTD) for a Fiber Glass Chronic Inhalation Study in the Rat

Short–term toxicity and lung clearance were assessed inrats exposed by inhalation to size-selected fibrous glass (FG)for 13 weeks. Results from this study and from a recent FG chronicinhalation study are presented here as guidelines for the selectionof a maximum tolerated dose (MTD) for chronic inhalation studiesof fibers. Fischer 344 rats were exposed using nose–onlyinhalation chambers, 6 hr/day, 5 days/week, for 13 weeks toone of five concentrations of FG (36, 206, 316, 552, or 714fibers/cc; expressed gravirnetrically, 3, 16, 30, 45, or 60mg/m³) or to filtered air. Rats were then held for an additional10 weeks of postexposure recovery. Test fiber was size–selectedfrom glass wool having a chemical composition representativeof building insulation. Rats were terminated at 7, 13, 19, and23 weeks after the onset of exposure to evaluate pulmonary pathology,lung epithelium cell proliferation, lung fiber burden, and lunglavage cells and chemistry. The effect of fiber inhalation onlung clearance of innocuous microspheres was also evaluated:following fiber exposure, six rats/group were exposed to ⁸⁵Sr–labeled3.0-µm polystyrene microspheres by intratracheal inhalationand then monitored for whole–body radioactivity duringthe 10–week recovery period. Data from the short–termstudy support the choice of 30 mg/m³ as the MTD for the previouschronic FG study and also provide indicators of long–termlung toxicity and functional impairment that can be used toestimate the MTD for future chronic fiber inhalation studies.     

Developmental Toxicity Evaluation of Inhaled 2-Ethoxyethanol Acetate in Ficher 344 Rats and New Zealand White Rabbits

Developmental Toxicity Evaluation of Inhaled 2–EthoxyethanolAcetate in Fischer 344 Rats and New Zealand White Rabbits. Tyl,R. W., Pritts, I. M, France, K. A., Fisher, L. C, and Tyler,T. R. (1988). Fundam. Appl. Toxicol. 10, 20–39. PregnantFischer 344 rats and New Zealand white rabbits were exposedto 2–ethoxyethanol acetate (EEA; CAS No. 111–15–9)vapor by inhalation on Gestational Days 6 through 15 (rats)or 6 through 18 (rabbits) at concentrations of 0, 50, 100,200,or 300 ppm, 6 hr/day. The animals were terminated on Gesta–tionalDay 21 (rats) or 29 (rabbits) and fetuses were examined forexternal, visceral, and skeletal malformations and variations.In rabbits, exposure to 100–300 ppm resulted in maternaltoxic–ity: decreased weight gain at 100–300 ppm,clinical signs at 200–300 ppm, alterations in hema–tologyat 100–300 ppm, reduced gravid uterine weight at terminationat 200–300 ppm, and elevated absolute liver weight at300 ppm. Developmental toxicity was observed at 100–300ppm: an increased incidence of totally resorbed litters at 200–300ppm, an increase in nonviable fetuses at 300 ppm, and a decreasein viable implants (live fetuses) per litter at 200–300ppm. The incidence of fetal malformations (external, visceral,and skeletal) was increased at 200–300 ppm. The incidenceof total malformations was 100% at 300 ppm and significantlyincreased at 200 ppm. Reduced fetal ossification was observedat 100–300 ppm. In rats, exposure to 100–300 ppmalso resulted in maternal toxicity: reduced weight gain andreduced food consumption at 200–300 ppm and elevated relativeliver weight and alterations in hematology at 100–300ppm. Absolute maternal liver weight was increased at all EEAexposure concentrations; relative liver weight was increasedat 100–300 ppm. Developmental toxicity was observed at100–300 ppm: increased nonviable implantations/litter(300 ppm), reduced fetal body weight/litter (200–300 ppm),and increased incidence of external (300 ppm), visceral, andskeletal (100–300 ppm) variations indicative of toxicity.The incidence of visceral, skeletal, and total malformationswas increased at 200–300 ppm. In conclusion, in both species,inhalation exposure to EEA during organogenesis produced maternaltoxicity at 100–300 ppm and developmental toxicity at100–300 ppm, including teratogenicity at 200–300ppm. At 50 ppm in both species, there was no evidence of maternalor developmental toxicity, including teratogenicity.     

Toxicity of Calcium Sodium Metaphosphate Fiber: II. Chronic Inhalation and Oncogenicity Study

Male and female Fischer 344 rats (80/sex/group) were exposedto CSM fiber 6 hr/day, 5 days/week at target-exposure levelsof 0, 1, 5, or 25 mg/m³ for 24 months, corresponding to 0, 27,80, and 513 fibers/cc, respectively. Number and size of theairborne fibers were determined during the course of the study.At 3 and 12 months, 10 rats/sex/group were euthanized and at18 and 24 months 5 rats/sex/group were euthanized. In addition,5 rats/ sex/group were removed from exposure at 18 months andmaintained for a 6-month recovery period. All animals survivingat the completion of the exposure period were maintained ina clean environment for up to 5 additional months. Clinicallaboratory examinations were performed on 10 animals/sex/groupat 3, 12, and 24 months. The number of fibers in the lung werealso determined at 3, 12, 18, and 24 months. Body weight andsurvival did not appear to be affected by treatment. There wereno biologically significant effects on clinical parameters.There was a dose-related increase in lung weight during theexposure period which was generally reversible during the recoveryperiods. There also was a dose-related increase in the numberof fibers/ milligram of lung, but no increase in lung fiberburden after the first 3 months. The number of fibers in thelungs of animals exposed to CSM fiber for 18 months and allowed6-month recovery period showed a decrease especially at thehigh dose. No increase in tumors (benign or malignant) was observedin this study. Microscopic changes considered reflective ofan irritant response were observed in the nasal turbinates notablyat the 5 and 25 mg/m³ levels. Histological changes were alsoobserved in the lungs at the 5 and 25 mg/m³ levels. The incidenceand/ or severity of histopathological changes in the 1 mg/m³group was considered to be essentially comparable to controls.     

Chronic Inhalation Toxicity and Carcinogenicity Study of Respirable Polymeric Methylene Diphenyl Diisocyanate (Polymeric MDI) Aerosol in Rats

Four groups of 60 Wistar rats of each sex were exposed by inhalationto 0, 0.2, 1.0, or 6.0 mg/m³ respirable polymeric methylenediphenyl diisocyanate (polymeric MDI) aerosol (93.5% < 4.2µm)for 6 hr a day, 5 days a week for up to 24 months. In addition,satellite groups of 10 rats/sex/group received the same treatmentfor 12 months. There was no adverse effect on general health,survival, body weight, or hematological or clinical chemistryparameters. Lung weights were increased in both males and femalesexposed to 6.0 mg polymeric MDI/m³ for 12 or 24 months. Grossexamination at autopsy of males exposed to 6.0 mg polymericMDI/m³ for 24 months revealed an increased incidence of spottedand discolored lungs. Increased incidences of degeneration andbasal cell hyperplasia of the nasal olfactory epithelium, oftenaccompanied by hyperplasia of Bowman's glands, were found inthe 1.0 and 6.0 mg/m³ groups. Light and electron microscopicstudies of the lungs revealed accumulations of alveolar macrophagescontaining polymeric MDI-associated refractile yellowish materialat the level of the alveolar duct in all exposed groups. Alveolarduct epithelialization as well as fibrosis of tissues surroundingthe macro phage accumulations occurred at the 1.0 and 6.0 mg/m³exposure levels. In addition, increased incidences of calcareousdeposits and localized alveolar bronchiolization were seen inthe 6.0 mg/m³ group. Moreover, eight pulmonary adenomas (sixin males and two in females) and one pulmonary adenocarcinoma(in a male) were observed in the 6.0 mg/m³ exposure group. Thetime sequence of the spectrum of pulmonary changes indicatesthat recurrent alveolar wall damage by polymeric MDI and/orpolymeric MDI-containing alveolar macrophages leads to alveolarbronchiolization and ultimately to bronchioloalveolar tumors.No lung tumors were found in the lower concentration groupsand in the control group. The incidence and distribution ofother types of tumors were not influenced by polymeric MDI.It was concluded that in the present study, the "no-observed-adverse-effectlevel" of polymeric MDI was 0.2 mg/m³ and that chronic exposureto polymeric MDI at a level of 6.0 mg/m³ was related to theoccurrence of pulmonary tumors. It was also concluded that exposureto polymeric MDI at concentrations not leading to recurrentlung tissue damage will not produce pulmonary tumors.     

Effect of Chronic Cigarette Smoke Exposure on Lung Clearance of Tracer Particles Inhaled by Rats

Cigarette smoking can influence the pulmonary disposition ofother inhaled materials in humans and laboratory animals. Thisstudy was undertaken to investigate the influence of cigarettesmoke exposures of rats on the pulmonary clearance of inhaled,relatively insoluble radioactive tracer particles. Following13 weeks of whole-body exposure to air or mainstream cigarettesmoke for 6 hr/day, 5 days/week at concentrations of 0, 100,or 250 mg total particulate matter (TPM)/m³, rats were acutelyexposed pernasally to ⁸⁵Sr-labeled fused aluminosilicate (⁸⁵Sr-FAP)tracer particles, then air or smoke exposures were resumed.A separate group of rats was exposed to the ⁸⁵Sr-FAP then seriallyeuthanized through 6 months after exposure to confirm the relativeinsolubility of the tracer particles. We observed decreasedtracer particle clearance from the lungs that was smoke concentration-dependent.By 180 days after exposure to the tracer aerosol, about 14,20, and 40% of the initial activity of tracer was present incontrol, 100 mg TPM/m³, and 250 mg TPM/m³ groups, respectively.Body weight gains were less in smoke-exposed rats than in controls.Smoke exposure produced lung lesions which included increasednumbers of pigmented alveolar macrophages distributed throughoutthe parenchyma and focal collections of enlarged alveolar macrophageswith concomitant alveolar epithelial hyperplasia and neutro-philicalveolitis. The severity of the lesions increased with smokeexposure duration and concentration to include interstitialaggregates of pigmented macrophages and interstitial fibrosis.Our data confirm previous findings that exposure to cigarettesmoke decreases the ability of the lungs to clear inhaled materials.We further demonstrate an exposure-concentration related magnitudeof effect, suggesting that the cigarette smoke-exposed rat constitutesa useful model for studies of the effects of cigarette smokeon the disposition of inhaled particles.     

Acute, Subacute, and Subchronic Inhalation Toxicity Studies of Respirable Polymeric Methylene Diphenyl Diisocyanate (Polymeric MDI) Aerosol in Rats

Short-term inhalation toxicity studies with respirable polymericmethylene diphenyl diisocyanate (polymeric MDI) aerosol wereperformed in rats. The 4-hr LC50 was found to be 490 mg polymericMDI/m³ (95.5% <4.3 µm). Exposure of (4-week-old) ratsto 0, 2.2, 4.9, or 13.6 mg polymeric MDI/m³ (95% < 5 µm)for 2 weeks resulted in mortality, severe growth retardation,and elevated lung weights at 13.6 mg/m³ at 4.9 mg/m³ slightgrowth retardation and slightly elevated lung weights were observed.A 13-week study with 6-week-old rats exposed to 0.35, 1.4, or7.2 mg polymeric MDI/m³ (95% < 5 µm) revealed transientgrowth retardation and a slightly increased number of pulmonaryalveolar macrophages occasionally accompanied by increased numbersof mononuclear cells and fibroblasts in alveolar septa onlyat 7.2 mg/ m³ In a second 2-week study with 4 or 6-week-oldrats exposed to 14.1 mg polymeric MDI/m³ (95% < 5 µm),4-week-old rats died earlier and in greater numbers than 6-week-oldrats. In a second 13-week study with 6-week-old rats, usingexposure concentrations of 0, 4.1, 8.4, and 12.3 mg polymericMDI/ m³ (95% < 5 µm) and including a 4-week recoveryperiod, 12.3 mg/ m³ induced mortality, growth retardation, severerespiratory distress, increased lung weights, degeneration andhyperplasia of the nasal epithelium, accumulations of macrophagesin the lungs and mediastinal lymph nodes, and focal inflammatorychanges in the lungs. Rats exposed to 8.4 mg/ m³ showed respiratorydistress, lower body weights in males, increased lung weights,and similar, but much less severe, histopathological changesin the respiratory tract and mediastinal lymph nodes. Most ofthe histopathological changes seen at the higher concentrationswere also seen at 4.1 mg/m³ but to a very minor degree and ina few rats only. At the end of the 4-week posttreatment periodthe microscopical changes in nose, lungs, and mediastinal lymphnodes were still present but generally to a much less degreethan at the end of the exposure period. It was concluded thatthe dose-effect curve for repeated exposures of rats to respirablepolymeric MDI is very steep, and that the "no-observed-adverse-effectlevel" of polymeric MDI was 1.4 mg/m³ the actual no-adverse-effectlevel being lower than but most probably very close to 4.1 mg/m³.     

Oral developmental toxicity study of methylsulfonylmethane in rats.

Methylsulfonylmethane (MSM) is a metabolite of dimethyl sulfoxide, and occurs naturally at low levels in many foods. MSM has received wide attention as a dietary supplement to promote joint health. The objective of these studies was to determine the developmental toxicity potential of MSM when administered orally to pregnant rats during the period of major organogenesis and histogenesis. In a preliminary dose-finding study, distilled MSM microprill (i.e., microspherical pellets of MSM) was administered by oral gavage at dose levels of 0 (vehicle control), 50, 250, 500, and 1000 mg/kg/day to 8-9 sperm-positive female Sprague-Dawley rats/group/day on gestation days 6-20. No evidence of maternal or fetal toxicity was observed. For the definitive developmental study, four groups of 24-25 timed-bred primiparous female rats were administered 0, 50, 500, or 1000 mg MSM/kg/day via gavage on gestation days 6-20. Maternal feed consumption, body weight, body weight gain, uterus weight and corrected body weight/body weight gain were unaffected by treatment. No evidence of maternal toxicity, and no significant differences in litter viability, litter size, or litter body weight were detected. Fetal evaluations failed to show any biologically significant increase in the incidence of anomalies in the MSM treated groups, and no malformations were seen in any of the fetuses. No evidence of fetal mortality, alterations to growth, or structural alterations were observed in the fetuses of dams administered 50-1000 mg/kg/day. Therefore, under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for maternal and developmental toxicity was 1000 mg/kg/day.