* CIIT Centers for Health Research, P.O. Box 12137, 6 Davis Drive, Research Triangle Park, North Carolina 27709-2137;
Syngenta, Alderley Park, Macclesfield, Cheshire, SK104TJ, United Kingdom;
DuPont Haskell Laboratories, Newark, Delaware 19714
Received May 14, 2002; accepted July 29, 2002
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key Words: pigmentary titanium dioxide; lung response; rats; mice; hamsters; inhalation; bronchoalveolar lavage fluid; pulmonary lesions.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The pulmonary toxicological effects of chronic exposure by inhalation to TiO2 particles have been examined in rodents. Continuous exposure for 2 years to 250 mg/m3 pigmentary TiO2 (p-TiO2) led to the development of lung tumors in male and female rats with a greater incidence found in females (Lee et al., 1985). Exposure to other PSP, such as diesel exhaust, carbon black, and talc particles, has resulted in pulmonary tumors in rats but not in mice or hamsters following chronic exposure (Heinrich et al., 1986
; Hext, 1994
; Mauderly et al., 1987
, 1994
; Muhle et al., 1998
). Findings of fibrosis, bronchoalveolar hyperplasia, metaplasia, and pulmonary tumorigenesis following exposures to high doses of PSP for long periods of time appear to be unique to the rat relative to other rodent species. These PSP-induced effects occur under, and appear limited to, conditions of substantial particle lung burdens and corresponding impairment of alveolar macrophage-mediated lung clearance (pulmonary overload).
Subchronic inhalation toxicity studies of rats to PSP have demonstrated that at concentrations under which chronic exposure conditions would lead to overload, there is a persistent inflammatory response and an attendant impairment of particle clearance from the lung (Cullen et al., 2000; Warheit et al., 1997
). Although particle overload conditions can be achieved in rats as well as mice, the specific pathway leading to tumor development only in rats remains unclear. The nonneoplastic lung responses induced in rodents by exposure to PSP have recently been reviewed, and the experimental evidence supports the hypothesis that there is a link between chronic inflammation and the epithelial changes leading to pulmonary cancer in rats (Donaldson, 2000
). It is believed that the rat is, in some way, hyper responsive in terms of its tendency to mount an inflammatory response to particles.
There have been several recent reviews concerning the topic of the species-specificity of nonneoplastic lung responses to PSP (Donaldson, 2000; Levy, 1994
; Mauderly et al., 1994
; Snipes, 1996
). Despite the importance of the topic, there have been relatively few comparative studies of PSP that have employed multiple species in the same laboratory utilizing an identical exposure regimen; moreover, to date there have been no systematic species comparison studies with p-TiO2 particles. The objective of the present study was to test the hypothesis that the rat lung responds to high particulate loading in a manner that differs from that of mice and hamsters under identical exposure conditions to PSP. Selected pulmonary pathobiological responses of female rats, mice, and hamsters were compared following subchronic inhalation exposure to comparable concentrations of p-TiO2 particles.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Aerosol generation and monitoring.
P-TiO2 (rutile type) was obtained from the DuPont Company (Wilmington, DE). A single modified-rotating brush dust-feeding mechanism served as the generator. Inhalation exposures were conducted in 1-m3 H-1000 stainless-steel chambers (Lab Products, Maywood, NJ). During the exposures, particle concentrations were continuously monitored using light scatter (Model RAM-S, Monitoring Instruments for the Environment, Inc., Bedford, MA), and the time-averaged concentration was recorded at least six times over the 6-h exposure period. The particle size distribution of the aerosol was measured at least twice per exposure level (excluding the control chambers) during the course of the study. Measurements were made using a MOUDI impactor (micro-orifice uniform deposit impactor, model 100, MSP Corporation, Minneapolis, MN). Mean mass-median aerodynamic diameter was 1.40 µm (see Results).
Experimental design.
Animals were exposed to 10, 50, or 250 mg/m3 p-TiO2 for 6 h/day, 5 days/week, for 13 weeks. Controls were exposed to filtered air only. Hamsters were exposed separate from the mice and rats due to health consideration. Group size for rats was 65 animals and for mice and hamsters 73 animals. The difference in group size was to account for expected animal losses in the course of the study. Animals were sacrificed immediately following completion of the 13-week exposure and additional recovery groups were held for postexposure periods of 4, 13, 26, or 52 (46 for hamster) weeks in clean air. Following exposure and at each recovery time, the p-TiO2 burdens in the combined right lung lobes and the lung-associated lymph nodes were determined. The left lungs were used to assess lung cell proliferation and histopathology. A separate group of animals was used at each time point to assess the inflammatory response in bronchoalveolar lavage fluid (BALF) by assaying lactate dehydrogenase (LDH) and total protein levels and conducting cytologic characterization of BALF cells.
Titanium burden analysis.
Burden analysis was performed as described by Levine and colleagues (Levine et al., submitted manuscript). At necropsy, right lung and lymph node tissue samples were collected and frozen until ready for separate titanium burden analysis. Prior to burden analysis the tissues were thawed, weighed, dried overnight in a muffle furnace at 37°C, dessicated, and weighed again. Dry tissue samples were digested overnight in a mixture of nitric acid and hydrofluoric acid and then further digested in a microwave oven. The sample digests were diluted with deionized water to 25 ml, and samples were analyzed using an inductively coupled plasma optical emission spectrometer (ICP-OES; Thermo Jarrell Ash AtomScan 16,Thermo Elemental, Franklin, MA). Data for lung tissue are expressed as milligrams of p-TiO2 per gram dry lung. For lymph node tissues, the data are expressed as the total micrograms of p-TiO2 present because the harvest of lymph node tissues was subject to collection of extraneous tissue to assure collection of the nodes. The method quantitation limit (MQL) was 0.05 µg Ti/ml of sample digest. The minimum detectable concentrations (MDC) of p-TiO2 in pulmonary tissues were calculated using the MQL in combination with the average control lung weight, and they were calculated to be for lung 0.22, 0.03, and 0.03 mg p-TiO2/g dry weight for mouse, rat, and hamster, respectively, and for lymph nodes 2.09 µg p-TiO2/sample. Occasional values above the MDC were observed in control tissues (e.g., burden values for control hamsters at the end of the exposures), and the values reflect an increase in the baseline for that run of the assay rather than the presence of p-TiO2 in control tissues. Retention half-times for p-TiO2 in lung were calculated using the best-fit equations to the lung burden data at all time points.
Bronchoalveolar lavage.
Lungs were lavaged five times with equal volumes of phosphate-buffered saline. Fluid from the first two lavages was recovered, pooled, and placed on ice. The subsequent three lavages were pooled and also placed on ice. Recovered cells from all lavages were collected by centrifugation, resuspended in cell culture medium, and counted using an automated cell counter (Model ZM, Coulter, Marietta, GA). Cell differential counts were performed on Wright-Giemsa-stained cytocentrifuge slide preparations. LDH and total protein levels in cell-free fluid from the first two pooled lavages were quantitated spectrophotometrically using a COBAS FARA II automated analyzer (Roche Diagnostic Systems Inc., Montclair, NJ).
Lung cell proliferation.
Five days prior to euthanasia, animals were subcutaneously implanted with osmotic pumps (Alza, Palo Alto, CA) containing bromodeoxyuridine (BrdU; Sigma Chemical Co., St. Louis, MO). Rats and hamsters were implanted with model 2ML1 (10 µl/h) pumps containing 5 mg/ml BrdU. Mice were implanted with model 2001 (1 µl/h) pumps containing 16 mg/ml BrdU. At necropsy, left lungs were pressure-infused intratracheally (20 cm H2O for mice; 30 cm H2O for rats and hamsters) with 10% neutral-buffered formalin. Lungs were fixed for approximately 48 h and then changed to 70% ethanol. Subsequently, the lungs were embedded in paraffin, sectioned at 5 µm, and stained for BrdU by established methods (Rutten et al., 1994). Terminal bronchiolar and alveolar cell labeling indices were determined for each animal, and the mean labeling index was calculated for each group of five animals.
Histopathology.
Paraffin-embedded left lung tissues were sectioned at 5 µm and stained with Massons trichrome. The trichrome-stained lung sections were evaluated for particle-induced histopathologic changes.
Statistical methods.
All data were tested for normality and homogeneity of variance. If the hypotheses for these assumptions were rejected (p < 0.01), common transformations (e.g., log, square root, arc sine) were applied and the data retested. Comparisons to controls were made using Dunnetts test. The software package JMP (SAS Institute, Cary, NC) was used for the statistical analysis.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Body Weights
Body weights were collected weekly for the exposure phase of the study and biweekly from week 17 through the end of the postexposure period. A depression in body weight was noted in all species and in all groups following the end of the exposure period (45% in mice, 23% in rats, and 511% in hamsters) with recovery occurring over the next three to four weeks in mice and rats (data not shown). This recovery was slower in hamsters, occurring within approximately six weeks. A likely reason for the depression in body weight in the three species is the need for a period of acclimation to changes in environmental conditions experienced by the animals during the postexposure recovery period when they were no longer housed in inhalation exposure chambers and were moved to new housing.
Mice exposed to the high concentration of p-TiO2 had a consistent depression, not exceeding 10% of controls, in body weight during the recovery period (data not shown). Although the cause for this depression in body weight is unclear, it is probably reflective of the high retained-p-TiO2 lung burdens found in these animals. In contrast, rats exposed to the high concentration of p-TiO2 demonstrated a consistent pattern of elevated weights (less than 10%), compared to controls, during the latter half of the recovery period (data not shown). No obvious cause for this weight elevation was evident.
Changes in hamster body weights were more frequent and abrupt than in mice or rats (data not shown). Weight loss during the exposure was attributed to housing conditions and was reversed following changes in housing from suspended-wire to solid-bottom shoebox caging during the postexposure periods. The observed drop in body weights near the end of the study probably reflected age-related changes as well as clinical symptoms consistent with kidney disease, as noted above.
P-TiO2 Burdens
There were dose-related changes in p-TiO2 lung burdens for all three species. Following 13 weeks of exposure, mice exhibited the greatest (170 mg/g dry lung) p-TiO2 lung burdens followed by rats (120 mg/g dry lung) then hamsters (114 mg/g dry lung; Fig. 1). Particle burdens decreased in the lung with time postexposure in mice, rats, and hamsters (Fig. 1
). Burdens in the lymph nodes increased with time postexposure in rats of all dose groups, in mice of the low and mid-dose groups, and in hamsters of the high-dose group (Fig. 1
). Notably, the lymph node p-TiO2 burdens of the animals exposed to 250 mg/m3 demonstrated the greatest increase between 4 and 13 weeks after the exposure ended. Decreases in lung burdens with time after exposure were sharply different between exposure concentration groups in both rats and mice; high- and mid-dose burdens decreased slowly to approximately 75% of the initial burden, whereas low-dose burdens decreased to approximately 15%. There were very significant differences between species in the pulmonary clearance kinetics of p-TiO2; rats and mice of the high-dose group retained more (approximately 75%) of the initial burden after 52 weeks of recovery than did hamsters after 46 weeks of recovery (approximately 10%).
|
The number of recovered macrophages in rats, mice, and hamsters of the high-dose group was significantly elevated, relative to corresponding controls, following exposure. The absolute increase in this cell population was greatest in mice and approximately equivalent in rats and hamsters (Fig. 2). Postexposure recovery resulted in a decline in the number of macrophages recovered by lavage in all species; although these values remained significantly elevated over concurrent controls in mice of the mid- and high-dose groups and rats of the high-dose group at 52 weeks postexposure, hamsters had returned to control levels by 26 weeks postexposure (Fig. 2
).
|
Lymphocytes were minimal in control rats and mice (0.2 and 0.3% respectively), whereas these cells comprised from 0.2 to 2.0% of the recovered cell population in control hamsters (data not shown). Animals exposed to p-TiO2 (mid- and high-dose rats and mice and high-dose hamsters) demonstrated significantly elevated levels of lymphocytes, which remained so for rats (2.2%) and mice (1.4 and 1.0%) at the end of the 52-week postexposure period but had returned to control levels in hamsters by 46 weeks postexposure (data not shown).
Pulmonary Toxicity End Points
Bronchoalveolar lavage was performed on one group of five animals for each time point and dose group. Pooled fluids (BALF) from the first and second lavages were assayed for LDH and total protein as markers of lung injury.
LDH levels were significantly elevated in mouse BALF in the mid- and high-dose animals and remained so through 26 weeks postexposure for the mid-dose animals and 52 weeks postexposure for the high-dose animals. Similarly, rats in the high- and mid-dose groups had persistent elevation of LDH in BALF; however those in the mid-dose group had returned to control levels by 26 weeks postexposure. Although hamsters exposed to the high dose of p-TiO2 had elevated LDH levels, the elevations were significantly different from control only through four weeks postexposure (Fig. 3).
|
|
Increased terminal bronchiolar cell replication was measured in mice and rats of the high-dose group following the 13-week exposure interval; however, the labeling index of these cells in both species had returned to control values by four weeks postexposure. No significant increases in the labeling index of bronchiolar cells of the hamster were observed (Fig. 5).
|
|
With the counting scheme used, no attempt was made to quantify cell replication in focal lesions associated with particle and particle-laden macrophage accumulation; however, it was apparent that labeled cells were often associated with these sites.
Lung Histopathology
The microscopic pattern of particle retention and lesions varied by species, by exposure concentration level, and with time postexposure. A characteristic finding in all three species was the intraluminal aggregation of particle-laden macrophages filled with opaque particles of p-TiO2. In each of the three concentration groups, the majority of the particles were found in macrophages within alveolar and alveolar duct lumens, although some of the intraluminal particulate material was extracellular. Extracellular particles were most prominent immediately following the exposure and were by far most numerous in the high-concentration group. Over time postexposure, aggregations of particle-laden macrophages coalesced and extracellular particulate material diminished. In all three species, the low concentration exposure groups had only a minimal diffuse increase in alveolar macrophages, some of which contained p-TiO2 particles. No associated pathology was present in these animals.
Rats.
The mid- and high-dose rats had a variety of lung lesions associated with retained p-TiO2 particles. Particles and particle-laden alveolar macrophages were most numerous in centriacinar lung regions, although in the high-dose animals there was a more diffuse panacinar distribution at the end of the 13-week exposure period. While the nature of the epithelial lesions was similar in mid- and high-dose animals, there were significant differences in lesion severity. The number of particle-laden macrophages observed individually and in aggregates was much greater in the high-dose animals. Although most particles were retained intraluminally, a minimal to mild interstitial accumulation of particle-laden macrophages was present in mid- and high-dose rats and remained over the course of the 52-week recovery. Immediately postexposure, both mid- and high-dose rats had alveolar hypertrophy and hyperplasia of type II epithelial cells surrounding aggregations of particle-laden macrophages (Fig. 7B). These lesions were generally minimal to mild in mid-dose and mild to moderate in high-dose animals. In both of these exposure concentration groups at this time point, histological evidence of chronic active inflammation was noted by the infiltration of neutrophils. Histological recognition of neutrophil infiltrates diminished in mid-dose rats by four weeks recovery although they persisted throughout the 52-week recovery period in the high-dose rats.
|
|
|
Hamsters.
The particle retention pattern in hamsters reflected the rapid clearance of p-TiO2. Similar to findings in mice, particles and particle-laden macrophages in exposed hamsters appeared to be concentrated in central portions of the lung lobes, although there were occasional subpleural focal accumulations of heavily particle-laden macrophages, similar to those noted in rats. Minimal type II epithelial cell hypertrophy and hyperplasia were observed associated with free particles and particle-laden macrophages, in both mid- and high-dose animals. These effects were reduced in incidence and severity with increasing time postexposure, correlating with the rapid removal of p-TiO2 particles from the hamster lung. Alveolar metaplasia and fibroproliferative changes were not present in hamster lungs (Fig. 8F). Polymorphonuclear leukocytes, both neutrophils and eosinophils, were found in all high-dose hamster lungs at each postexposure time point with the exception of the 46-week final time point. The severity of inflammation subsided with increasing time postexposure and with the associated diminution of particle burden.
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The retained lung p-TiO2 particle burdens in this study indicated that mid- and high-dose rats and mice developed significant impairment of alveolar macrophage-mediated clearance and thus overloaded lungs. Animals in the mid- and high exposure concentration groups had calculated particle retention half-times of 417 and 621 days (mice) or 324 and 838 days (rats) respectively. This contrasts with retention half-times of approximately 50 days for mice and 100 days for rats in the low concentration group and less than 110 days for all hamsters. The prolongation of p-TiO2 particle retention in exposed rats in the present study is consistent with the retention half-times of 68, 110, and 330 days previously reported for rats exposed for four weeks by inhalation to aerosol concentrations of 5, 50, and 250 mg/m3 p-TiO2 particles, respectively (Warheit et al., 1997).
In addition to significant differences in total retained lung and LALN p-TiO2 burdens, all three rodent species had distinctive particle microdosimetry retention patterns. Despite these differences, the three species had primarily airspace aggregation of particles and particle-laden macrophages, as opposed to the interstitial patterns of particle retention that have been reported in people and in nonhuman primate models (Nikula et al., 2001). Although particle distribution has been proposed to account for marked differences in rat and human responses to inhaled particles (Nikula et al., 2001
), the marked interspecies differences in pulmonary toxicity and pathology findings in the present rodent study cannot be explained by dose and anatomic site of particle distribution alone.
Although lung overload can be achieved in hamsters it requires prolonged exposure and higher lung burdens than are necessary to achieve this condition in rats (Muhle et al., 1990). The retained p-TiO2 lung particle burdens indicated that under the conditions of the present study, hamsters were better able to clear particles than were similarly exposed rats and mice. In part, the efficient lung clearance and corresponding decreased retention of p-TiO2 particles may explain the relative lack of long-term pulmonary toxicity and pathology noted in the heavily exposed hamsters. The lung burden findings in hamsters following the subchronic p-TiO2 exposure were somewhat surprising since we did not anticipate such dramatic differences between rats and hamsters. There have been relatively few clearance studies of PSP in Syrian golden hamsters, although a previous chronic study had demonstrated significant differences between the response of rats and Syrian golden hamsters to the inhalation of particles including p-TiO2 (Muhle et al., 1990
). Creutzenberg and colleagues noted the stimulation of clearance of radiolabeled polystyrene tracer particles in female Syrian golden hamsters following PSP particle exposure and speculated that hamsters as burrowing animals might have a well-developed adaptive mechanism due to living in a dry, dusty environment (Creutzenberg et al., 1998
).
The most striking findings of the present study are the interspecies differences noted in inflammation and lung lesions found in highly exposed animals. These experiments demonstrate that rats, mice, and Syrian golden hamsters vary significantly in their lung responses to inhaled p-TiO2 particles, confirming the generally held view that rats respond differently to PSP inhalation at overload exposure concentrations (Donaldson, 2000). The inflammatory changes, epithelial proliferative changes, and fibroproliferative lung lesions noted in rats in this study are identical to findings previously noted in subchronic (Warheit et al., 1997
) and chronic (Lee et al., 1985
) p-TiO2 studies that have used similar particles and exposure concentrations. Our findings differ from those of Baggs and colleagues (Baggs et al., 1997
) who observed the regression of pulmonary lesions with time postexposure, whereas in the present study, animals with equivalent initial lung burdens did not exhibit fibroproliferative lung lesions. A major difference between these two studies is the gender of the rats used. Since the incidence of pulmonary tumors in female rats has been shown to be greater than in male rats (Lee et al., 1985
), and gender differences in defense mechanisms (e.g., phagocytosis and neutrophil function) are known to exist (Spitzer, 1999
) it is possible that this would result in the observed differences.
The lowest exposure concentration of p-TiO2 particles (10 mg/m3) employed in this study did not result in inflammation or lesions, confirming the low toxicity of this particle-type and giving credibility to the notion that these results can be extrapolated to PSPs in general. Clearly, particle size, physicochemical attributes, and airborne concentration are known to play a role in PSP-induced effects. Therefore other interspecies studies using additional particle compositions are warranted (Cullen et al., 2000).
Our results demonstrate that p-TiO2-induced epithelial and fibroproliferative lesions associated with pulmonary overload and high lung burdens in the rat are progressive even following cessation of particle exposure and the diminution of pulmonary inflammation. The time course of lesion development in the present study shows that alveolar cell metaplasia (bronchiolization) occurs prior to the development of severe interstitial fibrosis in the lesion regions in those animals with high lung burdens. Further, the present studies confirm that pulmonary overload occurs in mice but with less neutrophilic inflammation and without the histopathological lesions observed in rats. The epithelial metaplasia (bronchiolization), persistent alveolar cell hyperplasia, and septal fibrosis noted in p-TiO2 particle-overloaded rats was a concentration-dependent effect and unique to that species when compared to the mice and hamsters. These lesions appeared to be highly correlated with the severe neutrophilic inflammation that characterized the rat response to inhaled p-TiO2 particles at exposure concentrations that resulted in pulmonary overload. Pulmonary neutrophilia in the rat was much more severe and persistent than in mice or hamsters, and may in large part explain the genesis of the species-specific pulmonary parenchymal lesions that occur. These lesions are believed to be strongly associated with PSP-induced tumorigenesis occurring in chronic inhalation bioassays using rats (Bellmann et al., 1992; Heinrich et al., 1995
; Mauderly et al., 1987
, 1994
; Muhle et al., 1990
).
Utilizing multiple species exposed in the same experiment and incorporating a long-term postexposure component, we demonstrated significant species differences in pulmonary responses to inhaled p-TiO2 particles. Under conditions where the retained lung p-TiO2 burdens were similar and likely to induce pulmonary overload, rats demonstrated a more severe and persistent pulmonary inflammatory response than either mice or Syrian golden hamsters. Rats also were unique in the development of a progressive fibroproliferative lesion and alveolar epithelial metaplasia in response to a subchronic exposure to a high concentration of p-TiO2 particles.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
NOTES |
---|
2 Present address: Advanced Inhalation Research, Cambridge, MA 02139-3789.
3 Present address: Wyeth Research, Collegeville, PA 19087-4517.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bellmann, B., Muhle, H., Creutzenberg, O., and Mermelstein, R. (1992). Irreversible pulmonary changes induced in rat lung by dust overload. Environ. Health Perspect. 97, 189191.[ISI][Medline]
Bingham, E., Corhssen, B., and Powell, C. H., Eds. (2001). Pattys Toxicology. Wiley, New York.
Creutzenberg, O., Bellmann, B., Muhle, H., Dasenbrock, C., Morrow, P., and Mermelstein, R. (1998). Lung clearance and retention of toner, TiO2, and crystalline silica, utilizing a tracer technique during chronic inhalation exposure in Syrian golden hamsters. Inhal. Toxicol. 10, 731751.[ISI]
Cullen, R. T., Tran, C. L., Buchanan, D., Davis, J. M., Searl, A., Jones, A. D., and Donaldson, K. (2000). Inhalation of poorly soluble particles. I. Differences in inflammatory response and clearance during exposure. Inhal. Toxicol. 12, 10891111.[ISI][Medline]
Donaldson, K. (2000). Nonneoplastic lung responses induced in experimental animals by exposure to poorly soluble nonfibrous particles. Inhal. Toxicol. 12, 121139.[Medline]
Heinrich, U., Fuhst, R., Rittinghausen, S., Creutzenberg, O., Bellmann, B., Koch, W., and Levsen, K. (1995). Chronic inhalation exposure of Wistar rats and two different strains of mice to diesel engine exhaust, carbon black, and titanium dioxide. Inhal. Toxicol. 7, 533356.[ISI]
Heinrich, U., Muhle, H., Takenaka, S., Ernst, H., Fuhst, R., Mohr, U., Pott, F., and Stober, 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, 383395.[ISI][Medline]
Hext, P. M. (1994). Current perspectives on particulate induced pulmonary tumours. Hum. Exp. Toxicol. 13, 700715.[ISI][Medline]
Lee, K. P., Trochimowicz, H. J., and Reinhardt, C. F. (1985). Pulmonary response of rats exposed to titanium dioxide (TiO2) by inhalation for two years. Toxicol. Appl. Pharmacol. 79, 179192.[ISI][Medline]
Levy, L. S. (1994). Squamous lung lesions associated with chronic exposure by inhalation of p-aramid fibres (fine fibre dust) and to titanium dioxide: Findings of a pathology workshop. In Toxic and Carcinogenic Effects of Solid Particles in the Respiratory Tract (D. L. D. U. Mohr, J. L. Mauderly, and G. Oberdorster, Eds.), pp. 473478. ILSI Press, Washington, DC.
Mauderly, J. L., Jones, R. K., Griffith, W. C., Henderson, R. F., and McClellan, R. O. (1987). Diesel exhaust is a pulmonary carcinogen in rats exposed chronically by inhalation. Fundam. Appl. Toxicol. 9, 208221.[ISI][Medline]
Mauderly, J. L., and McCunney, R. J., Eds. (1996). Particle Overload in the Rat Lung and Lung Cancer: Implications for Human Risk Assessment. Taylor and Francis, Washington, D.C.
Mauderly, J. L., Snipes, M. B., Barr, E. B., Belinsky, S. A., Bond, J. A., Brooks, A. L., Chang, I. Y., Cheng, Y. S., Gillett, N. A., Griffith, W. C., Henderson, R. F., Mitchell, C. E., Nikula, K. J., and Thomassen, D. G. (1994). Pulmonary toxicity of inhaled diesel exhaust and carbon black in chronically exposed rats. Part I: Neoplastic and nonneoplastic lung lesions. Res. Rep. Health Eff. Inst. 6875.
Muhle, H., Bellmann, B., Creutzenberg, O., Koch, W., Dasenbrock, C., Ernst, H., Mohr, U., Morrow, P., and Mermelstein, R. (1998). Pulmonary response to toner, TiO2 and crystalline silica upon chronic inhalation exposure in Syrian golden hamsters. Inhal. Toxicol. 10, 699729.[ISI]
Muhle, H., Creutzenberg, O., Bellmann, B., Heinrich, U., and Mermelstein, R. (1990). Dust overloading of lungsinvestigations of various materials, species-differences, and irreversibility of effects. J. Aerosol Med.-Depos. Clear. Eff. Lung 3, S111S128.
Nikula, K. J., Vallyathan, V., Green, F. H., and Hahn, F. F. (2001). Influence of exposure concentration or dose on the distribution of particulate material in rat and human lungs. Environ. Health Perspect. 109, 311318.[ISI][Medline]
Rutten, A. A., Bermudez, E., Mangum, J. B., Wong, B. A., Moss, O. R., and Everitt, J. I. (1994). Mesothelial cell proliferation induced by intrapleural instillation of man-made fibers in rats and hamsters. Fundam. Appl. Toxicol. 23, 107116.[ISI][Medline]
Snipes, M. B. (1996). Current information on lung overload by nonrodent mammals: Contrast with rats. Inhal. Toxicol. 8(Suppl.), 91109.
Spitzer, J. A. (1999). Gender differences in some host defense mechanisms. Lupus 8, 380383.[ISI][Medline]
Warheit, D. B., Hansen, J. F., Yuen, I. S., Kelly, D. P., Snajdr, S. I., and Hartsky, M. A. (1997). Inhalation of high concentrations of low toxicity dusts in rats results in impaired pulmonary clearance mechanisms and persistent inflammation. Toxicol. Appl. Pharmacol. 145, 1022.[ISI][Medline]