Faculty Bibliography

A surprising number of cases of hypersensitivity pneumonitis have been observed at work sites employing automotive machinists. Because hypersensitivity pneumonitis is not typically associated with exposure to metalworking fluid aerosols, this study examined whether Mycobacterium immunogenum (M. immunogenum), a rapidly growing mycobacterium isolated from several affected work sites, could induce hypersensitivity pneumonitis in mice. Hypersensitivity pneumonitis-like histologic changes occurred in mice treated with heat-killed and lysed M. immunogenum. These lung lesions were characterized by peribronchial and perivascular lymphohistiocytic inflammation and noncaseating granulomas in the parenchyma. The pathologic changes observed in mice instilled with M. immunogenum-contaminated used metalworking fluid were indistinguishable from those observed with M. immunogenum alone. The role of genetic factors in M. immunogenum-induced lung lesions was examined by comparison of the response of eight inbred strains of mice. The observed immunologic changes in the lung were significantly greater in C57Bl/6, 129, and BALB/c mice than in the other strains, suggesting that genetic factor(s) contribute to the susceptibility of workers exposed to M. immunogenum-contaminated metalworking fluid aerosols. Thus, these studies provide indirect evidence that M. immunogenum is an unrecognized class of microorganisms capable of causing hypersensitivity pneumonitis and plays a role in the outbreaks of hypersensitivity pneumonitis in automotive plants

BACKGROUND: In the US cotton industry, airborne cotton dust levels are regulated, and other countries are moving to specify safety limits for airborne endotoxins. There is concern about potential respiratory health hazards associated with agricultural and other organic dusts. In laboratories, ranking which samples have high and low levels of endotoxin is usually in good agreement between laboratories. When different laboratories assay identical samples, the levels differ. The objective of this research was to evaluate the intra- and inter-laboratory variability for 13 laboratories measuring endotoxin in cotton dust. METHOD: Two inter-laboratory round robin endotoxin assay studies were conducted using cotton dust. In the first round robin, each laboratory used their normal in-house assay method and then used a common extraction protocol. In the second round robin, a common extraction protocol and endotoxin assay kit was used. RESULTS: The intra-laboratory results had small variations but inter-laboratory results had very high variations. The inter-laboratory results using a common extraction protocol showed reduced differences. Using the same extraction protocol and endotoxin assay kit, the intra-laboratory variation was small and inter-laboratory variation was reduced but not enough for inter-laboratory agreement. Most of the laboratories were able to discern between the high and low endotoxin concentration dusts. CONCLUSIONS: Standardization has reduced the differences in results between laboratories and possibly further standardization may bring closer inter-laboratory agreement

A workshop that brought together representatives of most of the laboratories that have conducted animal and/or human inhalation exposure studies with concentrated ambient air particles (CAPs) was convened by the Health Effects Institute in Boston on May 5, 2004. Participants agreed that CAPs researchers need to make serious efforts to harmonize their experimental and analytical protocols to permit the sharing of lessons learned, questions raised, and opportunities for more definitive studies. Standardized outcome measures based on spirometry and response markers in lung bronchoalveolar lavage (BAL) cells and fluids exist, including the appropriate times after exposure to collect samples and measurements. However, for the emerging focus on cardiac system responses, there are many different electrocardiographic (ECG) endpoints being examined, and little standardization on markers that are most informative about adverse effects; on when the measurements need to be made; and on how to make comparable measurements. The workshop focused on two aspects of dealing with these complexities: sorting out influential particulate matter (PM) components responsible for observed effects, and searching for time-varying responses in continuous outcome data. The need for more complete analyses of PM samples from the CAPs studies was also emphasized, as was obtaining a consistent set of parameters characterizing exposure atmospheres and the ambient PM from which the CAPs are sampled. CAPs studies have already had a significant impact within the air pollution health effects community, especially in regard to cardiovascular system effects, and a follow-up meeting with a greater focus on means to harmonize data collection and analysis is needed

BACKGROUND: Individuals may develop tolerance to the induction of adverse pulmonary effects following repeated exposures to inhaled toxicants. Previously, we demonstrated that genetic background plays an important role in the development of pulmonary tolerance to inhaled zinc oxide (ZnO) in inbred mouse strains, as assessed by polymorphonuclear leukocytes (PMNs), macrophages, and total protein in bronchoalveolar lavage (BAL) phenotypes. The BALB/cByJ (CBy) and DBA/2J (D2) strains were identified as tolerant and non-tolerant, respectively. The present study was designed to identify candidate genes that control the development of pulmonary tolerance to inhaled ZnO. METHODS: Genome-wide linkage analyses were performed on a CByD2F2 mouse cohort phenotyped for BAL protein, PMNs, and macrophages following 5 consecutive days of exposure to 1.0 mg/m3 inhaled ZnO for 3 hours/day. A haplotype analysis was carried out to determine the contribution of each quantitative trait locus (QTL) and QTL combination to the overall BAL protein phenotype. Candidate genes were identified within each QTL interval using the positional candidate gene approach. RESULTS: A significant quantitative trait locus (QTL) on chromosome 1, as well as suggestive QTLs on chromosomes 4 and 5, for the BAL protein phenotype, was established. Suggestive QTLs for the BAL PMN and macrophage phenotypes were also identified on chromosomes 1 and 5, respectively. Analysis of specific haplotypes supports the combined effect of three QTLs in the overall protein phenotype. Toll-like receptor 5 (Tlr5) was identified as an interesting candidate gene within the significant QTL for BAL protein on chromosome 1. Wild-derived Tlr5-mutant MOLF/Ei mice were tolerant to BAL protein following repeated ZnO exposure. CONCLUSION: Genetic background is an important influence in the acquisition of pulmonary tolerance to BAL protein, PMNs, and macrophages following ZnO exposure. Promising candidate genes exist within the identified QTL intervals that would be good targets for additional studies, including Tlr5. The implications of tolerance to health risks in humans are numerous, and this study furthers the understanding of gene-environment interactions that are likely to be important factors from person-to-person in regulating the development of pulmonary tolerance to inhaled toxicants

The goal of the Complex Trait Consortium is to promote the development of resources that can be used to understand, treat and ultimately prevent pervasive human diseases. Existing and proposed mouse resources that are optimized to study the actions of isolated genetic loci on a fixed background are less effective for studying intact polygenic networks and interactions among genes, environments, pathogens and other factors. The Collaborative Cross will provide a common reference panel specifically designed for the integrative analysis of complex systems and will change the way we approach human health and disease

Large volumes of metalworking fluids (MWFs) are used in manufacturing industries for cooling and lubrication of metal pieces and tools during machining. MWFs accumulate microbial growth through continuous recirculation and reuse. We studied the progression of microbial contamination for 6 months after dumping, cleaning and recharging (DCR) of a large semi-synthetic MWF system managed with several biocides. Fresh, uncontaminated fluid was added to the system after extensive cleaning. The following samples were collected and analyzed: pre-DCR fluid (before system cleaning); neat fluid diluted to 6% with water; in use MWF 12 h and 1, 3 and 6 months post-DCR. Samples were analyzed for total microorganism concentrations by direct counting using fluorescence microscopy and by plate counting on various media (R2A, BHI, Middlebrooks and rose bengal under aerobic conditions). In addition, PCR was performed for the detection of mycobacteria. There was a rapid progression in the total bacterial counts as determined by fluorescence microscopy: 5.7 x 10(7) cells/ml in the pre-DCR used fluid, no measurable bacteria in the neat fluid, 6.9 x 10(6) cells/ml after 12 h and 2.2 x 10(6), 3.6 x 10(8) and 6.1 x 10(8) cells/ml after 1, 3 and 6 months, respectively. On average, only 0.2% of the direct count organisms were quantified on R2A cultures. PCR showed the presence of mycobacteria in the used MWF at 3 and 6 months. Mycobacteria were also identified from cultures on Middlebrooks and R2A. This study demonstrates that standard methods for cleaning MWF systems are inadequate since residual bacteria in the system can rapidly repopulate the newly charged MWF

Epidemiology studies suggest that exposure to air pollution increases the frequency of cardiac arrhythmias. A limitation of these studies is that it is difficult to link an increased risk of arrhythmias to a specific air pollutant. Animal exposure studies offer the opportunity to examine the effects of concentrated ambient fine particulate matter (PM), ultrafine PM, and copollutant gases separately. Male Fischer 344 rats, aged 18 mo, with implanted electrocardiograph (ECG) transmitters were used to determine the effects of PM on the frequency of arrhythmias. We found that old F344 rats had many spontaneous arrhythmias. An arrhythmia classification system was developed to quantify arrhythmia frequency. Arrhythmias were broadly grouped into two categories: premature beats and delayed beats. The rats were exposed to concentrated ambient PM (CAPS) or air for 4 h. The rats were exposed twice with a crossover design so each rat could serve as its own control. The CAPS concentrations were 160 microg/m(3) and 200 microg/m(3) for the first and second exposures, respectively. There was a significant increase in the frequency of irregular and delayed beats after exposure to CAPS. The same rats were subsequently exposed to laboratory-generated ultrafine carbon particles, to SO(2), or to air with a repeated crossover design. In these experiments there was no significant change in the frequency of any category of spontaneous arrhythmia following exposure to ultrafine carbon or SO(2). Thus, this study adds supporting evidence that acute exposure to elevated levels of ambient PM increases the frequency of cardiac arrhythmias

Various chemicals are used in the manufacture of cooling and lubricating fluids and fall into the classes of straight, soluble, semisynthetic, and synthetic metalworking fluids. The diversity of chemicals and in-use contaminants makes the risk assessment of metalworking fluids quite difficult. Toxicologists have used a number of methods to evaluate the component(s) responsible for the adverse pulmonary effects of metal working fluid aerosols encountered in the workplace. Although investigators have studied the adverse effects of metalworking fluid chemicals alone and in combination, the majority of evidence strongly suggests that the microbial changes that occur in fluid composition, during use and storage in the workplace, are responsible for the pulmonary effects reported for workers exposed to metalworking fluid aerosols. This review discusses the methodologies used to examine the toxicity of the complex nature of modern metalworking fluids and the findings that point toward bacterial endotoxin as a major contributor to their adverse effects

Beryllium (Be) has physical-chemical properties, including low density and high tensile strength, which make it useful in the manufacture of products ranging from space shuttles to golf clubs. Despite its utility, a number of standard setting agencies have determined that beryllium is a carcinogen. Only a limited number of studies, however, have addressed the underlying mechanisms of the carcinogenicity and mutagenicity of beryllium. Importantly, mutation and chromosomal aberration assays have yielded somewhat contradictory results for beryllium compounds and whereas bacterial tests were largely negative, mammalian test systems showed evidence of beryllium-induced mutations, chromosomal aberrations, and cell transformation. Although inter-laboratory differences may play a role in the variability observed in genotoxicity assays, it is more likely that the different chemical forms of beryllium have a significant effect on mutagenicity and carcinogenicity. Because workers are predominantly exposed to airborne particles which are generated during the machining of beryllium metal, ceramics, or alloys, testing of the mechanisms of the mutagenic and carcinogenic activity of beryllium should be performed with relevant chemical forms of beryllium

There are limited data on the efficacy of procedures for adapting rodents to restraint in nose-only holders. We examined: (1) What effect does restraint in nose-only holders have on heart rate and body temperature? (2) Does a gradual increase in the duration of restraint facilitate adaptation? (3) How long does it take for rodents to become fully adapted to nose-only holders? (4) Do rats and mice respond and adapt similarly to restraint in nose only holders? Heart rate and body temperature were monitored as measures of stress using electrocardiograph (ECG) transmitters in male C57Bl/6J mice and Sprague-Dawley rats. In naive animals during the first hour of restraint, heart rate increased by 58 beats per minute (BPM) (18.6%) in rats and by 174 BPM (32.3%) in mice as compared to cage controls. Temperature increased by 2 degrees C in mice and was unchanged in rats compared to cage controls. Heart rate and temperature values remained within normal physiologic values during restraint. In rats, the response to restraint in nose-only holders was the same after 4 days regardless of whether the duration of restraint was increased gradually to 4 h/day or kept constant at 4 h/day. In mice, the group that was gradually adapted had a statistically significant higher heart rate and temperature after 4 days than the fixed-duration adapted group. Rats and mice restrained for 4 h/day every day showed a gradual decrease in heart rate and temperature over time. Full adaptation to restraint required 14 days of fixed-duration daily restraint