Tobacco Documents Online

CHRONIC EXPOSURE TO DIESEL EXHAUST AND COAL DUST FIG. 4. Sites of particle retention and tissue responses in rats. (A) DE rat. Soot within macrophages in alveolar septa (~rrows) and a small amount in the perivascular interstitium (arrowheads). In rats, particulate material was found in these interstitial compartments only in conjunction with particulate material in alveoli. ×200. (B) CD rat. Particulate material within macrophages in the pleura (arrowheads). In rats, particulate material was located in the pleura in foei where particle-laden macrophages were located in adjacent alveolar lumens (arrow). Note the locally intense alveolar epithelial hyperplasia in conjunction with the maerophage aggregation. ×240. (p < 0.001), but it was not possible to estimate an odds ratio because none of the controls had particles scored in the interstitium. There were no significant differences due to type of particle exposure in rats (p = 1.0). In monkeys, there was significantly more interstitial particulate material in particle-exposed thaa in C monkeys (p < 0.001) with an odds ratio of 84 (12, 570). There were no significant differ- ences due to type of particle exposure in monkeys (p = 0.3). Morphometry In the rats, the me-an volume fraction of alveolar paren- chyma (alveolar air and alveolar septa including calSillaries) was 89% of the total lung, the mean volume fraction of air was 79% of the parenchyma, and there were no differences among controls or any of the exposure groups. These values are similar to those reported for F344 rats of 82 and 85% for the volume fractions of alveolar parenchyma in the lung and air in the parenchyma, respectively (Pinkerton et al., 1982). Although these results suggest that there was slightly greater sampling of the parenchymia than the hilus, and that the lungs were slightly less inflated than the lungs examined by Pinkerton and co-workers (1982), the differences were small. Because most of the particulate material is located in the parenchyma of rats, the sampling and inflation of these lung sections would tend to increase the value for the volume density of particulate material (volume of particulate mate- rial/volume of lung). However, any error would be small, and it would be uniform across all exposure groups. In monkeys, the mean volume fraction of alveolar paren- chyma was 81% of the total lung, the mean volume fraction of air was 75% of the parenchyma, and there were no differ- ences among control or any of the exposure groups. Volume fraction of alveolar parenchyma has not been reported for cynomolgus or other macaque monkeys, but the volume frac- tion of alveolar parenchyma is consistently 80-90% of the total lung fixed under physiologic conditions for a large range of mammals including rats (82%), dogs (85%), pigs (86%), baboons (82%), and humans (85-90%) (reviewed

6~6889890~

CHRONIC EXPOSURE TO DIESEL EXHAUST AND COAL DUST ~.9 TABLE 5 Distribution of Histopathologic Severity Scores for Particles in the Interstitium among Groups of Rats and Monkeys Seve~tyscore" Rats Monkeys Exposu~ gmu~ 0 1 2 3 0 1 2 C 15c 0 0 0 4 7 3 DE 0 14 I 0 0 ! 14 CD 0 14 1 0 0 I 12 DECD 0 14 1 0 0 0 1.5 m ~ Severity scores: 0, lesion not observed; 1, slight; 2, minimal; and 3, mild. ~ Animals were exposed to filtered air as controls (C) or to diesel exhaust (DE), coal dust (CD), or a combination of diesel exhaust and coal dust (DECD) as described earlier. 15 rats examined in each group. 14 C, 15 DE, 14 CD, and 15 DECD monkeys were examined. ¢ Number of animals with each severity score. by Pinkerton et al., 1992). The volume fraction of air in the parenehyma is 78% in rhesus macaques (Kapanci et aL, 1969). These results suggest that the sampling was represen- tative of the total monkey lung. The inflation may have been slightly less than that of the rhesus monkeys examined by Kapanci and co-workers (1969), but that the difference was minimal. Overall, these results suggest that the values for volume density of particulate material in the monkey lungs can be accepted with confidence. The relative volume density of particulate material was greater in the DE than in CD and DECD rats (Fig. 6). These differences in relative volume density were statistically sig- nificant between the DE and DECD rats (p = 0.003), but were not statistically significant between the DE and CD rats (p = 0.065) or the CD and DECD rats (p = 0.16). In the monkeys, the relative volume density of particles was less in the DE than the CD and DECD monkeys (Fig. 6). These differences in relative volume density were statisti- cally significant between the DE and CD monkeys (p = 0.026), but not between the DE and DECD monkeys (p = 0.29), or the CD and DECD monkeys (p = 0.19). The rela- tive volume density was also compared between rats and monkeys for each type of exposure after adjustment for dif- ferences between controls for each species. The CD and Rat Monkey \\\ \\\ \\\ \\\ 0 ~ Air DE CD OECD Air DE CO DECD Exposure Materials FIG. 6. R~lativ~ volum~ d~nsiti~ of p~iculate mate~al in rat and monkey lungs ~ ~e me~ volum~ percentage (+SE) for animals exposed to air, diesel exhaust (DE), coal dust (CD), or diesel exhaust and coal dust combined (DECD). DECD monkeys had significantly greater relative volume densities of particulate material than the CD and DECD rats, respectively (p < 0.001). There w.as no significant difference in the relative volume density of particulate material between DE monkeys and rats (p = 0.024). As explained earlier, interspecies comparisons of the relative volume density of particulate material must be interpreted cautiously. However, data for parenchymal and air volume fractions suggest that sampling and fixation were adequate; if errors were intro- duced, they would be small and would slightly increase the values for volume density of particulate material in rat lungs. These data correspond to the histopathologic observations that (1) the monkey lungs contained more total particulate material than the rat lungs, and (2) DE rat lungs seemed to contain more particulate material, while DE monkey lungs contained less particulate material, than the other exposure groups within each species. The particulate material in the control monkeys consisted of endogenous pigments, mite debris in some monkeys, and miscellaneous particles. The rare particulate material in control rats, which was seen at the 1280× magnification used for morphometry but not in FIG. 5. Sites of particle retention and tissue responses in monkeys. (A) DECD monkey. Particulate material in alveolar septa (arrowheads). Contrast O'~ the scarcity of particulate material in alveolar lumens and the lack of epithelial hyperplasia or other tissue responses with the photomicrographs from CO rats. AD, lumen of alveolar duct. ×200. (B) DE monkey. Soot in alveolar maerophages in the pleural connective tissue and lymphatics. Note the scarcity of particulate material in adjacent alveoli and the lack of tissue reaction in contrast with 4B. ×240. (C) DECD monkey. Particulate material in the interstitium of a respiratory bronchitic (RB). Macrophages containing particulate material are aggregated at bronchiolar branching sites (arrows). ×60. (D) DE monkey. Soot in the perivascular adventitia and lymphatics surrounding a pulmonary vein. Note the lack of soot in the surrounding alveoli in contrast with Fig. 4A. xl70.

50 NIKULA ET AL. Rat Monkey AI. IP AL. IP DE CD AL IP AL, IP AL IP AI. IP DECD DE CD OECD Exposure Materials FIG. 7. Volume percentages of particulate material in lumens of alveolar ducts and alveoli (AL) versus interstitium and pleura (IP) as the mean percentages of the total particulate material (+SE) for animals exposed to diesel exhaust (DE), coal dust (CD), or diesel exhaust and coal dust combined (DECD). The interstitial compartments with the greatest portion of the retained particulate material in each species are shown. Key to interstitial compartments: • alveolar septa, [] pleura, • perivascular interstitium, • interstitium of respiratory bronchioles (monkeys only), [] other interstitial compartments. the histopathologic examination, consisted of endogenous pigments. Approximately 73% of the particulate material in exposed rats was in the lumens of alveolar ducts and alveoli (Fig. 7). A much lower portion, approximately 43%, of the particulate material was in the lumens of alveolar ducts and alvcoli of exposed monkeys (Fig. 7). Approximately 27 and 52% of the particulate material in exposed rats and monkeys, respec- tively, was in the interstitium. As shown in the figure, parti- cles in the pleural lymphatics and connective tissue were grouped with the interstitial particles for this analysis. Inter- species comparison showed a significantly greater volume percentage of the total particulate material in the lumens of alveolar ducts and alveoli in the exposed rats than in the exposed monkeys (p < 0.001). Conversely, a significantly greater volume percentage of the total particulate material was in the interstitium of exposed monkeys than in exposed rats (p < 0.001). Within each species, there were no statisti- cal differences between DE, CD, and DECD animals for the volume percentage of the total particulate material in lumens of alveolar ducts and alveoli or for the volume percentage of the total particulate material in the interstitium. These data correspond well to the histopathologic observations of the relative distribution of the retained particles. In the rats, the interstitial compartments with the greatest portion of the retained particulate material, in descending order of amount, were the alveolar and alveolar duct septa, the pleura, and the perivascular interstitium (Figs. 4 and 7). These compartments comprise 21% of the volume fraction of the lung and contained 2 i% of the particulate material in exposed rats. In monkeys, the interstitial compartments with the greatest portion of the retained particulate material, in descending order of amount, were the alveolar and alveolar duct septa, the pleura, the interstitium of respiratory bronchi- olcs, and the perivascular interstitium (Figs. 5 and 7). These compartments comprised 27% of the volume fraction of the lung and contained 47% of the particulate material in ex- posed monkeys. DISCUSSION Both the histopathologic examination and the morphomet- ric determination of the relative volume density of particulate material suggested that a slightly, though not significantly, greater amount of diesel soot than coal dust was retained in the rat lungs. Although it was not possible to directly com- pare the amount of material visible by light microscopy to the amount of material determined by chemical analyses, these results are consistent with particle type differences in the lung burden data of Lewis et al. (1989). They determined that the particulate content of rat lungs, expressed as a per- centage of lung dry weight and corrected for analytib effi- ciency of recovery, was 1.13 and 0.92% for diesel soot and coal dust, respectively, at 24 months of exposure. In the monkeys, the histopathologic examination and the morpho- merry suggested that less diesel soot than coal dust was retained. Lewis et al. (1989) did not report lung burden data for the monkeys. Overall, both the histopathologic and morphometric data suggested that relatively more particulate material, regardless of type of exposure, was retained in

CHRONIC EXPOSURE TO DIESEL EXHAUST AND COAL DUST 51 the monkey lungs. The difference between the DE rats and monkeys in amount of particulate material visible by light microscopy was small and not statistically significant after correcting for the particulate material in controls. The larger interspeeies difference in the CD and DECD animals was statistically significant after correcting for particulate mate- rial in controls. Both the histopathologie evaluation and the morphometry showed clear differences between the rats and monkeys in the predominant sites of particle retention. In rats, almost three-fourths of the particulate material was retained in the lumens of alveolar ducts and alveoli. In monkeys, the partic- ulate material was almost equally divided between the lu- mens of alveolar ducts and alveoli, and the interstitium, but more of the material tended to be in the interstitium, and the amount in the interstitium was disproportionate to the size of the interstitial compartment. Within each species, the predominant site of particle retention did not vary by expo- sure material. The tendency for monkeys to retain particles in the interstitium may be related to structural features of the primate lung. For example, monkeys have respiratory bronchioles, and particulate material collected in the intersti- tium of the respiratory bronchioles. In primates, lymphatic vessels exist at the alveolar level adjacent to the respiratory bronehioles (Leak, 1977; Lauweryns and Baert, 1974). Blockage of these lymphatics has been proposed as the pri- mary event in formation of coal maeules in coal workers' lungs (Heppleston, 1954). In rats, which lack respiratory bronchioles, lymphatic vessels are located adjacent to the terminal bronchioles, but they do not exist at the alveolar level. Tl:le present findings, combined with the known ana- tomical differences and data showing that primates clear deposited particles more slowly than rats (Snipes, 1989, 1996), might suggest that a greater proportion of particles or particle-laden macrophages penetrate the airway epithe- lium and enter the interstitium in primates than in rats. Alter- natively, during chronic exposure, particle-laden macro- phages aggregated within alveoli of respiratory bronchioles may become incorporated into the interstitium as these alve- eli are obliterated (Heppleston, 1989; Green and Laquer, 1980). The particles in the interstitium may be cleared more slowly than those cleared via mucociliary action. If the pres- ence of respiratory bronchioles, the amount of interstitial and pleurat tissue, and the thickness of the alveolar septa are important determinants of the sites of particle retention, the differences between rats and humans might be even greater than the differences between rats and monkeys be- cause human:.!.ungs have more extensive interlobular septa, thicker pleura, and wider interstitial spaces in the alveolar septa than monkeys (McLaughlin et aL, 1966; Weibel, 1970, 1979). The response to particles, including alveolar epithelial hy- perplasia, inflammation, and focal septal fibrosis, was sig- nificantly greater in rats than monkeys. This difference was due to two factors: (1) the particles in the alveoli elicited much more of a response in the rats than monkeys, and (2) the particles in the interstitium, the retention site of over 50% of the particulate material in the monkeys, did not elicit proliferative or inflammatory responses in the monkeys at this exposure concentration and resultant lung burden. Thus, at equivalent exposure concentrations, both diesel soot and coal dust elicited less response in monkeys than in rats, despite the greater relative volume density of particulate material, especially coal dust, in the monkeys. This finding suggests that at equivalent lung burdens of coal dust, the greater response of rats than of monkeys would have been even more striking. This is not the only case where chronic inhalation of par- tieulate material resulted in different histopathology in rats and monkeys. Sprague-Dawley rats exposed to aerosols of petroleum coke dust developed accumulations of pigmented macrophages, chronic pulmonary inflammation, bronchioli- zation and adenomatous hyperplasia, sclerosis, squamous metaplasia of alveolar epithelium, and keratin cysts. Identi- cally exposed cynomolgus monkeys accumulated coke within pulmonary macrophages, but ~they did not develop the other lesions (Klonne et aL, 1987). MacFarland and co- workers (1982) identically exposed F344 rats and cynomol- gus monkeys to raw or processed shale dusts. All of the rats developed proliferative bronchiolitis and alveolitis (i.e., inflammation with epithelial hyperplasia), and most devel- oped chronic inflammation with nonprogressive fibrosis, cholesterol clefts, and microgranulomas. The monkeys accu- mulated pigment-laden macrophages in the bronchiolar and alveolar walls more than in the alveolar lumens. The majority of monkeys had no reaction to the accumulated material; a minority had occasional foei of subacute inflammation. None of the monkeys developed epithelial hy~erplasia or a fibrotic reaction. Squirrel monkeys and two strains of rats (Charles River-Caesarean derived and Greenacres Controlled Flora) were identically exposed to bertrandite or beryl ore by Wagner and co-workers (1969). Alveolar epithelial hyper- plasia and chronic inflammation with granulomas were pres- ent in rats exposed to both materials, and the incidence of neoplasms increased in the beryl-exposed rats. No lesions other than accumulations of particle-containing macrophages and mononuclear cells around respiratory bronchioles and blood vessels were present in monkeys. • Epithelial hyperplasia concomitant with the aggregation of particle-laden macrophages in alveolar lumens is a charac- teristic response to many poorly soluble particles in the rat lung, both at exposure concentrations that result in lung tumors (Nikula et aL, 1995; NTP, 1993, 1994a,b) and at exposure concentrations below those resulting in lung tu- mors (Maudedy et al., 1987; NTP, 1993, 1994a,b). Hyper- plasia of the surrounding epithelium in response to accumu-

NIKULA ET AL. lation of particulate material in focal aggregatss of alveolar macrophages was not characteristic of the response to diesel soot or coal dust in monkeys in this investigation, nor is it characteristic of coal workers' pneumoconiosis (Green and Laqueur, 1980; Merchant et al., 1986; Kleinerman et al., 1979), silicosis (Peters, 1986) or talc pneumoconiosis (Gam- ble, 1986) in humans. If human lungs respond to particles more like monkey lungs than rat lungs, this investigation suggests that the pul- monary response of rats to particles may not be predictive of the response in human lungs at concentrations represent- ing highoccupational exposures. Epidemiological studies suggest that diesel exhaust may increase lung cancer risk in heavily exposed humans (reviewed in Mauderly, 1992; California EPA, 1994; Health Effects Institute, 1995), and it is assumed that diesel soot-associated organic carcinogens may be important to this response. However, it has been shown that diesel-soot-associated organic carcinogens play little role in the carcinogenicity of diesel soot in rats (Hein- rich et al., 1992; Nikula et al., 1995). Therefore, the mecha- nism of carcinogenicity in rats exposed at high concentra- tions may differ from the potential mechanism in humans exposed at lower concentrations. The present findings sug- gest that perhaps the carcinogenicity data from rats exposed to high concentrations of diesel exhaust, which greatly ceed expected human exposure concentrations, should not be used to quantitatively predict responses in humans posed at lower rates because the inflammatory and epithelial proliferative responses that seem critical to the rat response to high concentrations of particles may not occur in primate lungs exposed at environmental or occupational concentra- tions. ACKNOWLEDGMENTS The substantial efforts of all the individuals who conducted the original study, especially the authors, T. R. Lewis (decreased), F. H. Y. Green, W. J. Moorman, J. R. Burg, and D. W. Lynch. arc gratefully acknowledged. The authors express their appreciation to Drs. Val Vallyathan, Francis H. Y. Green, and Frank Salomon of NIOSH, who facilitated our use of these slides and provided additional information concerning the original study. The authors thank Drs. Fletcher Hahn and M. Burton Shills of ITRI for helpful discussions concerning pulmonary pathology and particle deposition and clearance. The authors also thank the ITRI Technical Communications staff for assistance in preparing this manuscript. This research was supported by Volkswagen AG under a Funds-In-Agreement with the U.S. Department of Energy under Contract No. DE-AC04-76EV01013. REFERENCES California Environmental Protection Agency (1994). Health Risk Assess- ment for Diesel Ex'haust, Office of Environmental Health and Hazard Assessment, School of Public Health, Berkeley, CA. Crapo, J. D.. Barry, B. E., Gehr, P., Bachofen. M., and Weibel, E. R. (1982). Cell numbers and cell characteristics of the normal human lung. Am. Rev. Respir. Dis. 126, 332-337. Elias, H., and Hyde, D. M. (1983). A Guide to Practical Steriology. Karger. New York. Gamble, J. F. (I 986). Silicate pneumoconiosis. In Occnpational Respiratoo' Disease (J. A. Merchant, B. A. Boehlecke, and G. Taylor, Eds.), pp. 243- 285, Department of Health and Human Services Publication, Centers for Disease Control, NIOSH, DHHS(NIOSH) Publication No. 86-102, U.S. Government Printing Office, Washington, DC. Glasar. U., Hoehrainer, D., Otto, F. J.. and Oldiges, H. (1990). Careinoge- nieity and toxicity of four cadmium compounds inhaled by rats. ToxicoL Environ. Chem. 27, 153-16.2. Green, F. H. Y., and Laqueur, W. A. (1980). Coal workers' pneumoeonio- sis. Pathol. Annu. 1980 Part 2 15, 333-410. Health Effects Institute (I 995). Diesel Exhaust: A Critical Analysis of Emis- sions, Exposures, and Health Effects, Cambridge, MA. Heinrich, U., Muhle, H., Takenaka, S., Ernst. H., Fuhst. R., Mohr, U.. Pott, F., and Strber, W. (1986). Chronic effects on the respiratory tract of hamsters, mice, and rats after long-term inhalation of high concentrations of filtered and unfiltered diesel engine emissions. J. AppL Toxicol. 6, 383-395. Heinrieh, U., Peters, L., Ernst, H., Rittinghansen, S., Dasenbmck, C., and KCSnig, H. (1989). Investigation on the carcinogenic effects of various cadmium compounds after inhalation exposure in hamsters and mice. Exp. Pathol. 37, 253-258. Heinrieh, U., puhst, R., and Mohr, U. (1992). Tierexperimentelle inhala- tionsstudien zur frage der tumorinduzierenden wirkung yon dieselmo- torabgasen and zwei testiluben. GSF-Forsehangszentrum fOr Umwelt and Gesundheit mbH, Neuherberg. In Ptojekttrager Umwelt- und Klima- forschung (L Ende, Ed.), pp. 21-30. Auswirkungen yon Dieselmotorab- gas auf die Gesundheit, Munich. Henderson, R.F., Piekrell, J. A., Jones, R. K., Sun, J. D., Benson, J. M.. Manderly, J.L., and McClellan, R. O. (1988). Response of rodents to inhaled diluted diesel exhaust: Biochemical and cytological changes in bmnchoalveolar lavage fluid and in lung tissue. Fundam. AppL Toxicol. ' 11, 546-567. Heppleston, A. G. (1954). The pathogenesis of simple pneumoconiosis in coal workers. J. Pathol. Bacteriol. 67, 51-63. Heppleston, A. G. (1989). Relationship of lipid secretion and particle size to diffuse interstitial change in pneumoeoniosis: A pathogenic perspec- tive. Am. J. Ind. Med. 15, 427-439. Kapanei, Y., Weibel, E.R., Kaplan, H.P., and Robinson. F.R. (1969). Pathogenesis and reversibility of the pulmonary lesions of oxygen toxicity in monkeys. II. Ultrastruetural and morphometrie studies. Lab. b~vest. 20, 101-118. Kapanei, Y., Tosco, R., Eggerman, J., and Gould, V. E. (1972). Oxygen pneumonitis in man: Light and electron microscopic morphometric stud- ies. Chest 62, 162-169. Kleinerman, J., Green, F., Harley, R.S., Lapp, L., Laqueur, W., Naeye, R.L., Pratt, P., Taylor, G., Wiot, J., and Wyatt, J. (1979). Pathology standards for coal workers' pneumoconiosis. Arch. PathoL Lab. Med. 103, 375-432. Klonne, D. R., Burns, J. M., Halder, ~. A., Holdsworth, C. E., and Ulrich, C. E. (1987). Two-year inhalation toxicity study of petroleum coke in rats and monkeys. Am. J. Ind. Med. 11, 375-389. Lauweryns, J. M.. and Baert, J. H. (1974). The role of the pulmonary lym- phatics in the defenses of the diseased lung: Morphological and experi- mental studies of the transport mechanisms of intratraeheally instilled particles. Ann. N.Y. Acad. Sci. 221, 244-275. Leak, L. V. (1977). Pulmonary lymphatics and their role in the removal of interstitial fluids and particulate matter. In Respirator3" Defense Mecha- nisms (J. D. Brain, D. F. Proctor, and L. M. Reid, Eds.). Part II, pp. 631 - 685, Dekker. New York.

CHRONIC EXPOSURE TO DIESEL EXHAUST AND COAL DUST 53 Leak, L. V., and Jamuar, M. P. (1983). Ultrastructur~ of pulmonary lym- phatic vessels. Am. Rev. Respir. Dis. 128, $59-$65. Lewis, T. R., Green, F. H. Y., Moorman, W.J., Burg, J. R., and Lynch, D. W. (1989). A chronic inhalation toxicity study of diesel engine emis- sions and coal dust, alone and combined. J. Am. Coil. Toxicol. 8, 345- 375. MacFarland, H. N., Coate, W. B., Disbennett, D. B., and Ackerman, L.J. (1982). Long-term inhalation studies with raw and processed shale dusts. Am~ Occnp. Hyg. 26, 213-226. Manderly, J. L. (1992). Diesel exhaust. In Environmental Toxicants--Hu- man Exposures and Their Health Effects (M. Lippmann, Ed.), Chap. 5, p. 119, Van Nostrand Reinhold, New York. Mauderly, J. L. (1994). Contribution of inhalation bioassays to the assess- ment of human health risks from solid ~irborne particles. In Toxic and Carcinogenic Effects of Solid Particles in the Respirator. Tract (U. Mohr, D. L. Dungworth, J. L., Mauderiy, and G. Oberd6rster, F, ds.), pp. 355- 365, International Life Sciences Institute (ILSI) Press, Washington, DC. Mauderly, J. L. (1995). Current assessment of the carcinogenicity hazard of diesel exhaust. Toxicol. Environ. Chem. 49, 167-180. Mauderly, J. L., Jones, R. K., Griffith, W. C., Henderson, R. F., and McClel- lan, R. O. (1987). Diesel exhaust is a pulmonary carcinogen in rats ex- posed chronically. Fundam. AppL Toxicol. 9, 208-21 I. Mauderly, J. L., Banns, D. A., Griffith, W. C., Hahn, F. F., Henderson, R. F., and McClellan, R. O. (1996). Diesel exhaust is not a pulmonary carcino- gen in CD-I mice exposed under conditions carcinogenic to F344 rats. Fundam. AppL Toxicol. 30, 233-242. McCuilagh, P., and Nelder, J. A. (1983). Generalized Linem" Models, Chap- man and Hall, London. McLaughlin, R. F., Tyler, W. S., and Canada, R. O. (1961). A study of the subgross pulmonary anatomy in various mammals. Am. J. Anat. 108, 149-165. McLaughlin, R. F., Jr., Tyler, W. S., and Canada, R. O..(1966). Subgross pulmonary anatomy of the rabbit, rat, and guinea pig, with additional notes on the human lung. Am. Rev. Respir. Dis. 94, 380-387. Mercer, R. R., and Crape, J. D. (1988). Structure of the gas exchange region of the lungs determined by three-dimensional reconstruction. In Toxicol- ogy of the ~mgs. pp. 43-70. Raven Press, New York. Merchant, J. A., Taylor, T. K., and Hodous, G. (1986). Coal worker's pneu- moconiosis and exposure to other carbonaceous dusts. In Occupational Respiratm3, Disease (J. A. Merchant, Boehlecke, B. A., and Taylor, G., Eds.), pp. 329-384. Department of Health and Human Services Publica- tion, Centers for Disease Control, NIOSH, DHHS(NIOSH) Publication No. 86-102, U.S. Government Printing Office, Washington, DC. National Toxicology Program (NTP) (1993). Toxicology and Carcinogene- sis Studies of Talc in F344/N Rats and B6C3FI Mice (b)halation Studies), NTP Technical Report 421, NIH Publication No. 92-3152, U.S. Depart- ment of Health and Human Services, Public Health Service, National Institutes of Health. National Toxicology Program (NTP) (1994a). Toxicology and Carcinogene- sis Stndies of Nickel Subsulfide in F344/N Rats and B6C3FI Mice (Inhala- tion Studies), NTP Technical Report 453, NIH Publication No. 94-3369, U.S. Department of Health and Human Services, Public Health Service, National Institutes of Health. National Toxicology Program (NTP) (1994b). Toxicology and Carcinogen- esis Studies of Nickel Oxide in F344/N Rats m~d B6C3F~ Mice (b~halation Studies), NTP Technical Report 45 I, NIH Publication No. 94-3363. U.S. Department of Health and Human Services, Public Health Service, Na- tional Institutes of Health. Nikula, K: L. Snipes, M. B., Ban', E. B., Griffith, W. C., Henderson, R. F., and Mauderly, J. L. (1995). Comparative pulmonary toxicities and carci- nogenicities of chronically inhaled diesel exhaust and carbon black in F344 rats. Fundam. AppL ToxicoL 25, 80-94. Peters, J. M. (1986). Silicosis. In Occupational Respirator3, Disease (J. A. Merchant, B. A. Boehl~cke, and G. Taylor, Eds.), pp. 219-237. Depart- ment of Health and Human Services Publication, Centers for 'Disease Control, NIOSH, DHHS(NIOSH) Publication No. 86-102, U.S. Govern- ment Printing Office, Washington, DC. Phalen, R. F., and Oldham, M. J. (1983). Airway structures: Tracheobron- chial airway stractur~ as revealed by casting techniques. Am. Rev. Respi~: Di~: 128, S 1 -$4. Pinkerton, K. E., Barry, B. E., O'Neii, J.J., Raub, J. A., Pratt, P. C., and Crape, J, D. 0982). Morphologic changes in the lung during the lifespan of Fischer 344 rats. Am. J. Anat. 164, 155-174. Pinkerton, K. E., Gehr, P., and Crape, D. (1992). Architectur~ and cellular composition of the aft-blood barrier. In Comparative Biology of the Normal lamg (R. A. Parent, Ed.), pp. 121 -128. CRC Press, Boca Raton, FL. Saffiotti, U. (1995). Carcinogenesis by crystallin~ silica: Animal, cellular, and molecular studies. In Silica mtd Silica-b~duced Lung Diseases (V. Castranova, V. Vailyathan, and W. E. Wallace, Eds.), pp. 345-381. CRC Press, Inc., Boca Raton, FL. Schrcider, J. P., and Raabe, O. G. (1981). Structure of the human acinus. Am. J. Anat. 162, 221-232. Schultz, M. (1996). Comparative pathology of dust-induced pulmonary le- sions: Significance of animal studies to humans. InhaL Toxicol. 8, 433- 456, 1996. Snipes, M. B. (1989). Long-term retention and clearance of particles inhaled by mammalian species. Crit. Rev. ToxicoL 20, 175-21 I. Snipes, M. B. (1996). Cun.cnt information on lung overload in nonrodent mammals: Contrast with rats. b,t~al. ToxicoL 8, 91-109. Tyler, W. S. 0983). Small airways and terminal units: Comparative sub- gross anatomy of lungs. Ant. Rev. Respir. Dis. 128, $32-$36. Tyler, W.S., and Julian, M. D. (1992). Gross and subgross anatomy of lungs, pleura, connective tissue septa, distal airways, and structural units. In Comparative Biology of the Normal Lung (R. A. Parent, Ed.), Vol. 1, pp. 37-48. CRC Press, Boca Raton, FL. Wagner, W. D., Groth, D. H., Holtz, J. L., Madden, G. E., and Stokinger, H. E. (1969). Comparative chronic inhalation toxicity of beryllium ores, bertrandite and beryl, with production of pulmonary tumors by beryl. ToxicoL Appl. PharmacoL 15, 10-29. Weibel, E. R. (1970). Morphometric estimation of pulmonary diffusing ca- pacity. I. Model and method. Respir. Physiol. 11, 54-75. Weibel, E. R. (1979). Oxygen demand and the size of respiratory structures in mammals. In The Evoh,tion of Respiratoo, Processes (S. C. Wood and C. Lenfant, Eds.), Vol. 13, pp. 289-346. Dekker, New York. 0 / O~ O~