Effect of Exposure Mode on Amounts of Radiolabeled Cigarette Particles in Lungs and Gastrointestinal Tracts of F344 Rats

Abstract

This study was designed to compare the internal and external deposition of cigarette smoke particles in F344IN rats after nose-only or whole-body exposures and to provide information on how grooming affects the amount of smoke particles that pass through the gastrointestinal (GI) tract. Female rats were exposed to mainstream cigarette smoke by - four different modes: nose-only, tube-restrained (NOT); pelt-only, tube-restrained (POT); whole-body, tube-restrained (WBT); and whole-body, cage-housed (WBC). Croups of rats were exposed simultaneously for 40 min by 1 of the 4 modes to [¹⁴C]ldotriacontane (DTC) labeled cigarette smoke at a mean mass concentration of 327 mglm³. Half of the rats from each group were sacrificed immediately after exposure, and the others were sacrificed 24 h later. Head skin, a sample of subcutaneous fat, GI tract, trachea/lobar bronchi, lungs, depelted head, depelted carcass, and remaining pelt were analyzed to determine their ^UC content. About 60% of the ¹⁴C activity in the respiratory tract in the NOT and WBT groups was deposited in the pulmonary region, and about 40% was in the head airways and trachea. The radiolabeled DTC was cleared very slowly from the pulmonary region. The initial total body burdens of ¹⁴C in the rats exposed by the WBT and WBC modes were higher than those in the rats exposed by the NOT mode as a consequence of pelt contamination by the ¹⁴C-DTC. Crooming resulted in the ingestion of about 80-90% and 60% of the ¹⁴C activity originally deposited on the head skin and pelt, respectively, by 24 h after exposure. The ratio of the amount of smoke particles either contained within or passing through the CI tract to the amount in lung after 24 h was 2.6 for WBC-exposed rats and 1.3 for NOT-exposed rats. We concluded that compared to rats exposed using the NOT mode, WBC exposures increased the amount of smoke particles passing into the CI tract by about a factor of two.     

Fourteen-Day Inhalation Study in Rats, Using Aged and Diluted Sidestream Smoke from a Reference Cigarette: II. DNA Adducts and Alveolar Macrophage Cytogenetics

The chemical constituents of cigarette smoke are greatly dilutedin environmental tobacco smoke (ETS). In the typical indoorenvironment where cigarettes are smoked, the mean value of respirablesuspended particles is approximately 0.1 mg/m³. In this study,we used aged and diluted sidestream smoke (ADSS) of 1R4F Universityof Kentucky research cigarettes as a surrogate for ETS and exposedSprague-Dawley rats nose-only to 0, 0.1, 1.0, and 10 mg wettotal particulate matter (WTPM)/m³ for 6 hr per day for 14 consecutivedays. DNA from lung, heart, larynx, and liver was tested foradduct formation after 7 and 14 days of exposure and after 14days of recovery. In addition, alveolar macrophages from animalsexposed for 7 days were examined for chromosomal aberrations.Exposure-related DNA adducts were not observed in any of theanimals at 0.1 or 1.0 mg WTPM/ m³, which represent ambient and10-fold exaggerated ETS concentrations, respectively. Slightdiagonal radioactive zones, characteristic of adducts observedin human smokers and in animals exposed to mainstream smoke,were observed, but only in lung and heart DNA of animals exposedto the highest concentration of ADSS (10 mg WTPM/m³), a 100-foldexaggeration of typical field measurements of ETS. The meanrelative adduct labeling values (±SE) were 8.7 (±0.2)adducts per 10' nucleotides for lung DNA and 5.7 (±0.7)adducts per 10' nucleotides for heart DNA after 14 days of exposure.No elevation in chromosomal aberrations was observed in alveolarmacrophages. These results indicate a no-observed-effect-level(NOEL) of 1.0 mg/m³ for DNA adduct formation in lung and heartand a NOEL of at least 10 mg/m³ for the induction of chromosomeaberrations in alveolar macrophages under the conditions ofthis study.     

Short-Term Effects of Sidestream Smoke on Respiratory Epithelium in Mice: Cell Kinetics

Male strain A/J and C57BL/6 mice were exposed on five consecutivedays, for 6 hr a day, to sidestream smoke generated from Kentucky1R4F reference cigarettes. Chamber concentrations were 1 mg/m³of total suspended particulate matter and 528 to 549 µg/m³of nicotine. Cumulative labeling indices in the airways andin the pulmonary parenchyma were measured following 1, 3, or5 days exposure to unfiltered or filtered sides tream smoke.A significantly increased labeling index was found in A/J micein the epithelium lining large intrapulmonary airways and terminalbronchioles after 3 and 5 days exposure to unfiltered smoke,whereas following exposure to filtered smoke labeling indicesremained normal. The alveolar labeling index was not increasedfollowing smoke exposure. In C57BL/6 mice, sidestream smokedid not produce signs of increased cell proliferation in therespiratory tract. It is concluded that the response to sidestreamsmoke inhalation in mice may depend upon the strain of miceexamined.     

Proceedings. Radioactivity in diet.

Male and female Syrian golden hamsters were exposed to the smoke of [¹⁴C]dotriacontane-16,17 ([¹⁴C]DOT)-labelled cigarettes in 2 different exposure systems. These 2 systems differed in terms of the smoke concentration drawn into the exposure chamber. To compare the effectiveness of the exposure systems, the inhaled dose of ¹⁴C-labelled cigarette smoke was determined in the different parts of the hamster respiratory tract by liquid scintillation counting. About 6 times more smoke particles were deposited in the respiratory tract after exposure to high smoke concentration with intermittent puffs of fresh air (closed system) than after exposure to smoke diluted with air (open system). About 80% of the diluted smoke reached the bronchi and lung compared to approximately 60% of the concentrated smoke. The remainder of the dose was trapped in the upper respiratory tract, mainly by the nose and larynx. Additionally, in the open system the total dose of inhaled smoke was dependent upon the position in the exposure chamber. The results are discussed with respect to the use of the exposure systems for chronic cigarette smoke inhalation studies in experimental respiratory tract carcinogenesis.     

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.     

Deposition and Fate of Inhaled Ethylene Glycol Vapor and Condensation Aerosol in the Rat

Deposition and Fate of Inhaled Ethylene Glycol Vapor and CondensationAerosol in the Rat. Marshall, T.C. and Cheng, Y.S. (1983). Fundam.Appl. Toxicol. 3:175-181. Fischer-344 rats were exposed by nose-onlyinhalation for 30 min to ethylene glycol-¹⁴C vapor at a concentrationof 32 mg/m³. A second group of rats was exposed for 17 min toan ethylene glycol-¹⁴C aerosol formed by condensation of theglycol on 0.1 m ⁶⁷Ga₂O₃ particles. The mass median aerodynamicdiameter (MMAD) of the condensation aerosol was 2.3 µm (g = 1.8) with an ethylene glycol concentration of 184 mg/m³. Initial radiocarbon body burdens were at glycol levels of0.7 mg/kg for the vapor exposure and 2.4 mg/kg for the aerosolexposure. Estimates indicated that at least 60% of the ethyleneglycol inhaled for both exposures was deposited in rats withthe highest concentration of radioactivity being in the nasalcavities. Distribution from the site of deposition was rapidsince 75 to 80% of the initial body burden was found throughoutthe bodies of rats sacrificed immediately after exposure. Thepredominant routes of elimination during the first 4 days followingexposure were via the expired air as ¹⁴CO₂ (63% for the vaporexposure and 70% for the aerosol exposure) and urine as theunchanged glycol (20% and 11%). The proportion of the dosesexcreted as ¹⁴CO₂ suggested more complete metabolism of ethyleneglycol following inhalation relative to that observed afterintravenous injections reported previously. The half-time forclearance of plasma radiocarbon was 39 and 34 hr for the vaporexposure and aerosol exposure, respectively. The studies suggestedthat ethylene glycol plasma levels following acute exposureto high airborne concentrations of ethylene glycol vapor and/oraerosol would not approach the toxic concentrations associatedwith known dose-dependent changes in the glycol's metabolism.     

Inhalation experiments with 14C-labelled cigarette smoke. II. The distribution of cigarette smoke particles in the hamster respiratory tract after exposure in two different smoking systems.

Male and female Syrian golden hamsters were exposed to the smoke of [14C]dotriacontane-16,17 ([14C]DOT)-labelled cigarettes in 2 different exposure systems. These 2 systems differed in terms of the smoke concentration drawn into the exposure chamber. To compare the effectiveness of the exposure systems, the inhaled dose of 14C-labelled cigarette smoke was determined in the different parts of the hamster respiratory tract by liquid scintillation counting. About 6 times more smoke particles were deposited in the respiratory tract after exposure to high smoke concentration with intermittent puffs of fresh air (closed system) than after exposure to smoke diluted with air (open system). About 80% of the diluted smoke reached the bronchi and lung compared to approximately 60% of the concentrated smoke. The remainder of the dose was trapped in the upper respiratory tract, mainly by the nose and larynx. Additionally, in the open system the total dose of inhaled smoke was dependent upon the position in the exposure chamber. The results are discussed with respect to the use of the exposure systems for chronic cigarette smoke inhalation studies in experimental respiratory tract carcinogenesis.     

Comparative Inhalation Toxicity of Nickel Subsulfide to F344/N Rats and B6C3F1 Mice Exposed for 12 Days

Comparative Inhalation Toxicity of Nickel Subsulfide to F344/NRats and B6C3F₁ Mice Exposed for 12 Days. BENSON, J. M., CARPENTER,R. L., HAHN, F. F., HALEY, P. J. HANSON, R. L., HOBBS, C. H.,PICKRELL, J. A., AND DUNNICK, J. K. (1987). Fundam Appl. Toxicol.9, 251–265. Groups of F344/N rats and B6C3F₁ mice wereexposed to aerosols of nickel subsulfide (Ni₃S₂) 6 hr/day for12 days not including weekends. Actual exposure concentrationswere within 3% of target (target 10.0, 5.0, 2.5, 1.2, 0.6,and 0.0 mg Ni₃S₂/m³). Nickel lung burdens of exposed rats andmice increased linearly with exposure concentration. Two malerats and all mice exposed to 10.0 mg Ni₃S₂/m³ died before theend of the exposures. Exposure to Ni₃S₂ had no elfect on thenatural killer cell activity of mouse spleen cells. Lesionsin rats and mice related to inhalation of Ni₃S₂ were found inthe nasal epithelium, lung, and bronchial lymph nodes. The mostextensive lesions were found in the lung and included necrotizingpneumonia. Emphysema developed in rats exposed to 5.0 or 10.0mg Ni₃S₂/m³ while fibrosis developed in mice exposed to 5.0mg Ni₃S₂/m³ Degeneration of the respiratory epithelium and atrophyof the olfactory epithelium of the nose occurred in rats exposedto as low as 0.6 mg Ni₃S₂/m³ and mice exposed to 1.2 mg/m³ Resultsindicate that inhalation exposure of rats and mice to Ni₃S₂/aerosol concentrations near the current threshold limit value(TLV) for nickel compounds (1 mg/m³ for Ni metal and roastingfume and dust and 0.1 mg/m³ as Ni for soluble compounds) canproduce lesions in the respiratory tract. Atrophy of lymphoidtissues (spleen, thymus, and bronchial lymph nodes) was foundin animals of the highest exposure concentration. Degenerationof the testicular germinal epithelium was also observed in miceand rats that survived 5.0 or 10.0 mg/m³ exposure concentrations.     

Effects of Inhaled Chromium Dioxide Dust on Rats Exposed for Two Years

Effects of Inhaled Chromium Dioxide Dust on Rats Exposed forTwo Years. LEE, K. P., ULRICH, C. E., GEIL, R. G., AND TROCHIMOWICZ,H. J. (1988). Fundam. Appl. Toxicol 10, 125–145. Ratswere exposed by inhalation to chromium dioxide (C1O2) dust atdesign concentrations of 0, 0.5 mg/m³(stabilized and unstabilized,respectively), or 25 mg/m³ (stabilized) for 6 hr/day, 5 days/weekfor 2 years. No dust-exposure-related pathological changes otherthan lung lesions were observed in all exposed rats. There wereno significant differences in pulmonary responses between unstabilizedand stabilized CrC>2 at the 0.5 mg/m³ exposure level. Thelungs showed minute dust deposition in the alveoli adjacentto the alveolar ducts, but maintained an intact general architecture.The pulmonary responses satisfied the biological criteria fora nuisance dust. At 25 mg/m³, dust deposition was sharply confinedto the alveoli in the alveolar duct region. Alveolar walls enclosingdust-laden macrophage (dust cell) aggregates were thickenedwith hyperplastic Type II pneumocytes and slightly collagenizedfibrosis. Alveoli adjacent to the terminal bronchioles werelined with bronchiolar epithelium (alveolar bronchiolar-ization).In addition, lungs showed foamy macrophage response, cholesterolgranulomas, alveolar protoeinosis, and minute fibrotic pleurisy.These pulmonary lesions occurred predominantly in female rats.Of 108 female rats, 6 developed keratin cysts and 2 had cystickeratinizing squa-mous cell carcinoma (CKSCC). None of 106 malerats had either a keratin cyst or a CKSCC. The lung tumors developedfrom metaplastic squamous cells in the areas of alveolar bronchio-larizationat the alveolar duct region. The lung tumors were well differentiatedand devoid of characteristics of true malignancy. The CKSCCis an experimentally induced, unique tumor type and is differentfrom the type of spontaneous lung tumor seen in man or animals.The relevance to man of this type of lung tumor appears to benegligible.     

Ninety-Day Inhalation Study in Rats, Using Aged and Diluted Sidestream Smoke from a Reference Cigarette: DNA Adducts and Alveolar Macrophage Cytogenetics

To study the genotoxic effects of subchronic exposure to environmentaltobacco smoke, Sprague-Dawley rats were exposed to 0, 0.1, 1.0,and 10 mg total particulate matter (TPM)/m³ of aged and dilutedsidestream smoke (ADSS) from 1R4F reference cigarettes 6 hrper day, 5 days a week for 13 weeks. DNA from lung, heart, larynx,bladder, and liver was tested for adduct formation by the ³²P-postlabelingassay after 28 (except bladder) and 90 days of exposure and90 days after cessation of exposure. In addition, alveolar macrophagesfrom animals exposed for 28 or 90 days were examined for chromosomalaberrations. Exposure-related DNA adducts were not observedin any tissue in any of the animals exposed to 0.1 or 1.0 mgTPM/m³. However, increased levels of DNA adducts with diagonalradioactive zones were observed in lung, heart, and larynx DNAof animals exposed to the highest concentration of ADSS (10mg TPM/m³). Adduct analyses with varying amounts of DNA fromlungs of mid-and high-exposure animals clearly indicate thatthe dose-response for DNA adduct formation is nonlinear. Theadduct levels were highest after 90 days of exposure and weresignificantly reduced in all target tissues 90 days after cessationof exposure. Chromosomal aberrations in alveolar macrophageswere not elevated in any group after 28 or 90 days of exposure.These results indicate a no-observed-effect-level (NOEL) ofat least 1.0 mg/m³ for DNA adduct formation in lung, heart,and larynx, and a NOEL of at least 10 mg/m³ for the inductionof chromosome aberrations in alveolar macrophages, under theconditions of this study.     

Studies on the deposition and distribution of catechol from whole cigarette smoke in BC3F1Cum mice

Tritiated catechol has been used to follow the pharmacokinetics and metabolic fate of inhaled catechol in cigarette smoke in $\text{BC3F1}\text{Cum}$ mice. The presence of [³H]catechol in the smoke was verified by silica gel chromatography, high-performance liquid chromatography, and gas chromatography/mass spectrometry. Mice were exposed to 10% ($\text{v}\text{v}$) 2R1 cigarette smoke on the Walton Horizontal Smoking Machine under standard conditions of 35 ml puff volume, 2 sec/puff, 10 puffs/cigarette. The deposition and distribution of inhaled catechol were determined in all internal tissues, urine, and feces. Data showed that clearance was occurring during the 10-min smoke exposure period. Immediately after exposure, over 50% of the radioactivity was found in the blood, with 10% found in the lung, and approximately 12% in the respiratory tract. Over 91% of the inhaled radioactivity was found in the urine 120 min after exposure. Less than 0.5% of the total dose was found in the lung at this time. We conclude that catechol in smoke is rapidly absorbed, redistributed, and excreted from mice exposed to whole cigarette smoke.     

Deposition and distribution of the total particulate matter of cigarette smoke in mice using a large-capacity smoke exposure system

A newly developed automatic smoke exposure machine (SEM II) was used to generate [¹⁴C]dotriacontane-labeled University of Kentucky reference 2A1 or 2R1 cigarette smoke. The SEM II is a large-capacity (480 mice) dynamic smoke exposure system in which smoke is routed through the animal containment system as a continuously flowing stream. Mice are restrained about the neck in stock-like holders for “nose-only” exposure. Using standard smoke exposure conditions, the deposition and internal distribution of the total particulate matter (TPM) from cigarette smoke was determined in BC3Fl/Cum male and female mice. Results show: (a) smoke exposure conditions can be varied so that deposition from 30 to 200 μg TPM/lung can be obtained, (b) 80–90% of the TPM deposition was found in the respiratory tissues, (c) the mouse-to-mouse variation for TPM deposition in pulmonary tissue was ～20%, (d) similar deposition and distribution of TPM was observed in male and female mice, and (e) deposition and distribution of TPM was not altered in mice exposed to smoke on a daily basis over a 6-month period of time.     

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.     

Inhalation experiments with 14C-labelled cigarette smoke III. Body size dependent distribution of particulate matter in small rodents during cigarette smoke inhalation

Syrian golden and European hamsters were exposed to ¹⁴C-labelled cigarette smoke. The deposition of the ¹⁴C activity in the respiratory and digestive tracts was determined immediately after the exposure. From the results obtained it can be seen that the distribution of particulate matter between the upper and lower respiratory tract, as well as, between the respiratory and digestive tracts is related to the body weight of the experimental animals.     

Pulmonary Retention of Inhaled Diesel Particles after Prolonged Exposures to Diesel Exhaust

Pulmonary Retention of Inhaled Diesel Particles after ProlongedExposures to Diesel Exhaust CHAN, T. L., LEE, P. S., AND HERING,W. E. (1984). Fundam. Appl. Toxicol. 4, 624–631. The effectof continuous exposure to diluted diesel exhaust on the pulmonaryretention of inhaled diesel particles was studied in male Fischer344 rats. Test animals were first exposed to clean air or diluteddiesel exhaust in exposure chambers at nominal particulate concentrationsof 250 µg/m³ or 6 mg/m³ for 20 hr/day, 7 days/week, forperiods lasting from 7 to 112 days, followed by a nose-onlyexposure to ^l4C-tagged diesel particles for 45 min. At preselectedtime intervals after the radioactive exposure, the ¹⁴C-activitiesin the lungs of groups of four animals were measured to determinethe clearance of the ^l4C-diesel particles up to 1 year. Thepulmonary retention of the radioactive diesel particles wasgreater in animals which had been preexposed to diesel exhaustThe slower alveolar clearance of particle-laden macrophagesand leukocytes can be described by a normal Diphasic clearancemodel. Since some of the macrophages were found sequesteredas aggregates in the pulmonary region, a slow-clearing residualcomponent was included in a modified lung retention model. Whenthese residual fractions were determined and excluded from theactive particulate transport within the lungs, normal alveolarclearance rates were calculated for the animals with a preexposurediesel particulate dose less than 0.8 mg. Slower clearance wasobserved at a dose of 6.5 mg and no clearance was evident ata dose of 11.8 mg in their lungs. In effect, the greater retentionof inhaled particles can be interpreted as-a sign of impairedlung clearance attributable to the prolonged exposures to highconcentrations of diesel exhaust gases and/or the presence ofaccumulated carbonaceous particles residing in sequestered macrophageaggregates in the lungs.     

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.     

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.     

Kinetics of Respiratory Tract Absorption and Plasma Clearance of Horseradish Peroxidase in Guinea Pigs

Kinetics of Respiratory Tract Absorption and Plasma Clearanceof Horseradish Peroxidase in Guinea Pigs. CONNER, M. W., CHAUDHURI,I., ROGERS, A. E., and AMDUR, M. O. (1985). Fundam. Appl. Toxicol.5, 99–104. Horseradish peroxidase (HRP) absorption acrossthe wall of the upper airway, monitored by the amount detectedin the blood, is used to measure epithelial damage by toxins.Full details of kinetics of absorption and blood clearance havenot been reported previously. We measured the kinetics underthe experimental conditions used in testing toxins. HRP wasadministered to guinea pigs either by intraarterial injectionof a 7.5 µg bolus (plasma clearance) or by intratrachealinstillation of 1 mg (respiratory tract absorption). Plasmaconcentrations were monitored for 60 min. Plasma concentrationsof HRP rose linearly with time after intratracheal instillation,reaching 236 ± 51 ng/ml (mean ± SE) at 60 minafter instillation. HRP was cleared from the plasma rapidlyafter bolus injection. The elimination coefficient, k2, determinedfrom the biphasic log normal plot, was 0.322 min^–1. Thesedata were used to estimate the kinetics of absorption acrossthe respiratory epithelium. A single 3-hr exposure to an atmospherecontaining 2.5 mg/m³ of submicrometer zinc oxide particles increasedplasma concentration of HRP after intratracheal deposition (407± 63 ng/ml at 60 min) and had no effect on plasma clearance(k² = 0.342 min^–1). Therefore plasma concentrations ofHRP measured after intratracheal deposition can be used as asensitive indicator to evaluate the effects of inhalation ofa test atmosphere on epithelial permeability, if plasma clearancekinetics are not altered by the exposure.     

Evaluation of the Developmental Toxicity of Ethylene Glycol Aerosol in the CD Rat and CD-1 Mouse by Whole-Body Exposure

Ethylene glycol (EG) is a major industrial chemical, shown tobe teratogenic at high doses by gavage in rodents. Since oneroute of industrial exposure is to the aerosol at high concentrations,timed-pregnant CD rats and CD-1 mice were exposed, whole-body,to a respirable aerosol of EG (mass median aerodynamic diameter,2.3 µm) on Gestational Days (GD) 6 through 15 for 6 hrper day at target exposure concentrations of 0, 150, 1000, or2500 mg/m³ (analytical concentrations of 0, 119 ± 13,888 ± 149, and 2090 ± 244 mg/m³, respectively),with 25 plug-positive animals per species per group. Clinicalobservations and maternal body weights were documented throughoutgestation for both species. Maternal food and water consumptionwas measured in rats only throughout gestation. At schedulednecropsy (GD 21 for rats, GD 18 for mice), maternal animalswere evaluated for body weight, liver weight, kidney weight,gravid uterine weight, number of ovarian corpora lutea, andstatus of implantation sites, i.e., resorptions, dead fetuses,live fetuses. Fetuses were dissected from the uterus, counted,weighed, sexed, and examined for external, visceral, and skeletalmalformations and variations. All rat dams survived to scheduledtermination. Minimal maternal toxicity was indicated by a significantincrease in absolute and relative liver weight at 2500 mg/m³.Food and water consumption, maternal body weights and weightgain, and maternal organ weights (other than liver) were unaffectedby exposure. Gestational parameters were unaffected by exposure,including pre- and post-implantation loss, live fetuses/litter,sex ratio, and fetal body weight/litter. There was no treatment-relatedincrease in the incidence of any individual malformation, inthe incidence of pooled external, visceral, or skeletal malformations,or in the incidence of total malformations by fetus or by litter.There were no increases in the incidence of external or visceralvariations. Evidence of fetotoxicity, expressed as reduced ossificationin the humerus, the zygomatic arch, and the metatarsals andproximal phalanges of the hind-limb, was observed at 1000 and2500 mg/m³. All mouse dams survived to scheduled termination.One dam at 2500 mg/m³ was carrying a totally resorbed litterat termination. Maternal toxicity was observed at 1000 and 2500mg/m³, expressed as reduced body weight and weight gain duringand after the exposure period, and reduced gravid uterine weight.(Maternal effects may have been due, in part or in whole, toeffects on the conceptuses; see below.) Embryo/fetal toxicitywas also observed at 1000 and 2500 mg/m³, expressed as an increasein nonviable implantations/litter, a reduction in viable implantations/litter,and reduced fetal body weights (male, female, and total)/litter.The incidences of individual and pooled external, visceral,and skeletal malformations were increased at 1000 and 2500 mg/m³,as was the incidence of total malformations. Malformations werefound in the head (exencephaly), face (cleft palate, foreshortenedand abnormal face, and abnormal facial bones), and skeleton(vertebral fusions, and fused, forked, and missing ribs). Theincidences of many fetal variations were also increased at 1000and 2500 mg/m³ (and only a few at 150 mg/m³). The no observableadverse effect level (NOAEL) for maternal toxicity in rats was1000 mg/m³ (analytical concentration 888 mg/m³) and in micewas 150 mg/m³ (analytical concentration 119 mg/m³). The NOAELfor development toxicity in rats was 150 mg/m³ and in mice wasat or below 150 mg/m³, under the conditions of this study. Analysisof EG on the fur of rats and mice during and after the exposureperiod at 2500 mg/m³ indicated that much of the EG "dose" (65–95%)was potentially derived from ingestion after grooming and/orpercutaneous absorption. This contribution of the ingested and/orabsorbed chemical could have been sufficient, per se, to producethe teratogenic effects observed in mice. The definitive evaluationof the possible role of inhaled EG aerosol alone in teratogenesisrequires an exposure regimen which limits or precludes exposureby any other route.     

Histopathology,Urine Mutacenicity,and Bone Marrow Cytocenetics of Mice Exposed Nose-Only to Smoke from Cigarettes that Burn or Heat Tobacco

Abstract

Male and female B6C3F₁ mice were exposed nose-only to smoke from a test cigarette that heats tobacco, or from a reference cigarette that burns tobacco. Cigarette smoke was generated by a smoking machine, and the concentrations of wet total particulate matter (WTPM) were adjusted to 0, 0.16, 0.32, or 0.64 mg/l. Exposures were performed 1 h/day for 14 consecutive days. Urine mutagenicity was assessed by a modified Ames bacterial assay Clastogenesis (sister-chromatid exchanges, chromosome aberrations, and micronuclei) was evaluated in bone marrow cells. Respiratory rate was depressed significantly by exposure to smoke from the reference cigarette, but not the test. Blood carboxyhemoglobin, plasma nicotine, and plasma cotinine showed exposure-dependent increases in the smoke-exposed animals. Histopathological changes similar to those noted previously in smoke-exposed rats were noted, with fewer and less pronounced changes in the animals exposed to smoke from the test cigarette when compared with the reference. Positive urine mutagenicity and clastogenic responses were observed in the animals treated with positive control chemicals. However the urine mutagenicity and clastogenic responses of smoke-exposed animals (both cigarette types) were not different from those of sham-exposed animals, except for the micronucleus assay where animals exposed to high concentrations of reference cigarette smoke showed a significant increase over controls.