Continuous exposure to chrysotile asbestos can cause transformation of human mesothelial cells via HMGB1 and TNF-? signaling.
ABSTRACT: Malignant mesothelioma is strongly associated with asbestos exposure. Among asbestos fibers, crocidolite is considered the most and chrysotile the least oncogenic. Chrysotile accounts for more than 90% of the asbestos used worldwide, but its capacity to induce malignant mesothelioma is still debated. We found that chrysotile and crocidolite exposures have similar effects on human mesothelial cells. Morphological and molecular alterations suggestive of epithelial-mesenchymal transition, such as E-cadherin down-regulation and ?-catenin phosphorylation followed by nuclear translocation, were induced by both chrysotile and crocidolite. Gene expression profiling revealed high-mobility group box-1 protein (HMGB1) as a key regulator of the transcriptional alterations induced by both types of asbestos. Crocidolite and chrysotile induced differential expression of 438 out of 28,869 genes interrogated by oligonucleotide microarrays. Out of these 438 genes, 57 were associated with inflammatory and immune response and cancer, and 14 were HMGB1 targeted genes. Crocidolite-induced gene alterations were sustained, whereas chrysotile-induced gene alterations returned to background levels within 5 weeks. Similarly, HMGB1 release in vivo progressively increased for 10 or more weeks after crocidolite exposure, but returned to background levels within 8 weeks after chrysotile exposure. Continuous administration of chrysotile was required for sustained high serum levels of HMGB1. These data support the hypothesis that differences in biopersistence influence the biological activities of these two asbestos fibers.
Project description:Quantifying the asbestos-related lung cancer burden is difficult in the presence of this disease's multiple causes. We explore two methods to estimate this burden using mesothelioma deaths as a proxy for asbestos exposure.From the follow-up of 55 asbestos cohorts, we estimated ratios of (i) absolute number of asbestos-related lung cancers to mesothelioma deaths; (ii) excess lung cancer relative risk (%) to mesothelioma mortality per 1000 non-asbestos-related deaths.Ratios varied by asbestos type; there were a mean 0.7 (95% confidence interval 0.5, 1.0) asbestos-related lung cancers per mesothelioma death in crocidolite cohorts (n=6 estimates), 6.1 (3.6, 10.5) in chrysotile (n=16), 4.0 (2.8, 5.9) in amosite (n=4) and 1.9 (1.4, 2.6) in mixed asbestos fibre cohorts (n=31). In a population with 2 mesothelioma deaths per 1000 deaths at ages 40-84 years (e.g., US men), the estimated lung cancer population attributable fraction due to mixed asbestos was estimated to be 4.0%.All types of asbestos fibres kill at least twice as many people through lung cancer than through mesothelioma, except for crocidolite. For chrysotile, widely consumed today, asbestos-related lung cancers cannot be robustly estimated from few mesothelioma deaths and the latter cannot be used to infer no excess risk of lung or other cancers.
Project description:BACKGROUND:We have conducted a population-based study of pleural mesothelioma patients with occupational histories and measured asbestos lung burdens in occupationally exposed workers and in the general population. The relationship between lung burden and risk, particularly at environmental exposure levels, will enable future mesothelioma rates in people born after 1965 who never installed asbestos to be predicted from their asbestos lung burdens. METHODS:Following personal interview asbestos fibres longer than 5?µm were counted by transmission electron microscopy in lung samples obtained from 133 patients with mesothelioma and 262 patients with lung cancer. ORs for mesothelioma were converted to lifetime risks. RESULTS:Lifetime mesothelioma risk is approximately 0.02% per 1000 amphibole fibres per gram of dry lung tissue over a more than 100-fold range, from 1 to 4 in the most heavily exposed building workers to less than 1 in 500 in most of the population. The asbestos fibres counted were amosite (75%), crocidolite (18%), other amphiboles (5%) and chrysotile (2%). CONCLUSIONS:The approximate linearity of the dose-response together with lung burden measurements in younger people will provide reasonably reliable predictions of future mesothelioma rates in those born since 1965 whose risks cannot yet be seen in national rates. Burdens in those born more recently will indicate the continuing occupational and environmental hazards under current asbestos control regulations. Our results confirm the major contribution of amosite to UK mesothelioma incidence and the substantial contribution of non-occupational exposure, particularly in women.
Project description:The onset of malignant mesothelioma (MM) is linked to exposure to asbestos fibers. Asbestos fibers are classified as serpentine (chrysotile) or amphibole, which includes the crocidolite, amosite, anthophyllite, tremolite, and actinolite types. Although few studies have been undertaken, anthophyllite has been shown to be associated with mesothelioma, and tremolite, a contaminant in talc and chrysotile, is a risk factor for carcinogenicity. Here, after characterizing the length and width of these fibers by scanning electron microscopy, we explored the cytotoxicity induced by tremolite and anthophyllite in cells from an immortalized human mesothelial cell line (MeT5A), murine macrophages (RAW264.7), and in a rat model. Tremolite and short anthophyllite fibers were phagocytosed and localized to vacuoles, whereas the long anthophyllite fibers were caught on the pseudopod of the MeT5A and Raw 264.7 cells, according to transmission electron microscopy. The results from a 2-day time-lapse study revealed that tremolite was engulfed and damaged the MeT5A and RAW264.7 cells, but anthophyllite was not cytotoxic to these cells. Intraperitoneal injection of tremolite in rats induced diffuse serosal thickening, whereas anthophyllite formed focal fibrosis and granulomas on peritoneal serosal surfaces. Furthermore, the loss of Cdkn2a/2b, which are the most frequently lost foci in human MM, were observed in 8 cases of rat MM (homozygous deletion [5/8] and loss of heterozygosity [3/8]) by array-based comparative genomic hybridization techniques. These results indicate that tremolite initiates mesothelial injury and persistently frustrates phagocytes, causing subsequent peritoneal fibrosis and MM. The possible mechanisms of carcinogenicity based on fiber diameter/length are discussed.
Project description:Chrysotile asbestos is closely associated with excess mortality from pulmonary diseases such as lung cancer, mesothelioma, and asbestosis. Although multiple mechanisms in which chrysotile asbestos fibers induce pulmonary disease have been identified, the role of autophagy in human lung epithelial cells has not been examined. In this study, we evaluated whether chrysotile asbestos induces autophagy in A549 human lung epithelial cells and then analyzed the possible underlying molecular mechanism. Chrysotile asbestos induced autophagy in A549 cells based on a series of biochemical and microscopic autophagy markers. We observed that asbestos increased expression of A549 cell microtubule-associated protein 1 light chain 3 (LC3-II), an autophagy marker, in conjunction with dephosphorylation of phospho-AKT, phospho-mTOR, and phospho-p70S6K. Notably, AKT1/AKT2 double-knockout murine embryonic fibroblasts (MEFs) had negligible asbestos-induced LC3-II expression, supporting a crucial role for AKT signaling. Chrysotile asbestos also led to the phosphorylation/activation of Jun N-terminal kinase (JNK) and p38 MAPK. Pharmacologic inhibition of JNK, but not p38 MAPK, dramatically inhibited the protein expression of LC3-II. Moreover, JNK2(-/-) MEFs but not JNK1(-/-) MEFs blocked LC3-II levels induced by chrysotile asbestos. In addition, N-acetylcysteine, an antioxidant, attenuated chrysotile asbestos-induced dephosphorylation of P-AKT and completely abolished phosphorylation/activation of JNK. Finally, we demonstrated that chrysotile asbestos-induced apoptosis was not affected by the presence of the autophagy inhibitor 3-methyladenine or autophagy-related gene 5 siRNA, indicating that the chrysotile asbestos-induced autophagy may be adaptive rather than prosurvival. Our findings demonstrate that AKT/mTOR and JNK2 signaling pathways are required for chrysotile asbestos-induced autophagy. These data provide a mechanistic basis for possible future clinical applications targeting these signaling pathways in the management of asbestos-induced lung disease.
Project description:<h4>Background</h4>While asbestos has long been known to cause mesothelioma, quantitative exposure-response data on the relation of mesothelioma risk and exposure to chrysotile asbestos are sparse.<h4>Methods</h4>Quantitative relationships of mortality from mesothelioma and pleural cancer were investigated in an established cohort of 5397 asbestos textile manufacturing workers in North Carolina, USA. Eligible workers were those employed between 1950 and 1973 with mortality follow-up through 2003. Individual exposure to chrysotile fibres was estimated on the basis of 3420 air samples covering the entire study period linked to work history records. Exposure coefficients adjusted for age, race, and time-related covariates were estimated by Poisson regression.<h4>Results</h4>Positive, statistically significant associations were observed between mortality from all pleural cancer (including mesothelioma) and time since first exposure (TSFE) to asbestos (rate ratio [RR], 1.19; 95% confidence interval [CI], 1.06-1.34 per year), duration of exposure, and cumulative asbestos fibre exposure (RR, 1.15; 95% CI, 1.04-1.28 per 100?f-years/mL; 10-year lag). Analyses of the shape of exposure-response functions suggested a linear relationship with TSFE and a less-than-linear relationship with cumulative exposure. Restricting the analysis to years when mesothelioma was coded as a unique cause of death yielded stronger but less precise associations.<h4>Conclusions</h4>These observations support with quantitative data the conclusion that chrysotile causes mesothelioma and encourage exposure-response analyses of mesothelioma in other cohorts exposed to chrysotile.
Project description:The importance of the role of fibroblasts in cancer microenvironment is well-recognized. However, the relationship between fibroblasts and asbestos-induced lung cancer remains underexplored. To investigate the effect of the asbestos-related microenvironment on lung cancer progression, lung cancer cells (NCI-H358, Calu-3, and A549) were cultured in media derived from IMR-90 lung fibroblasts exposed to 50 mg/L asbestos (chrysotile, amosite, and crocidolite) for 24 h. The kinetics and migration of lung cancer cells in the presence of asbestos-exposed lung fibroblast media were monitored using a real-time cell analysis system. Proliferation and migration of A549 cells increased in the presence of media derived from asbestos-exposed lung fibroblasts than in the presence of media derived from normal lung fibroblasts. We observed no increase in proliferation and migration in lung cancer cells cultured in asbestos-exposed lung cancer cell medium. In contrast, increased proliferation and migration in lung cancer cells exposed to media from asbestos-exposed lung fibroblasts was observed for all types of asbestos. Media derived from lung fibroblasts exposed to other stressors, such as hydrogen peroxide and UV radiation didn't show as similar effect as asbestos exposure. An enzyme-linked immunosorbent assay (ELISA)-based cytokine array identified interleukin (IL)-6 and IL-8, which show pleiotropic regulatory effects on lung cancer cells, to be specifically produced in higher amounts by the three types of asbestos-exposed lung fibroblasts than normal lung fibroblasts. Thus, the present study demonstrated that interaction of lung fibroblasts with asbestos may support the growth and metastasis of lung cancer cells and that chrysotile exposure can lead to lung cancer similar to that caused by amphibole asbestos (amosite and crocidolite).
Project description:The six natural silicates known as asbestos may induce fatal lung diseases <i>via</i> inhalation, with a latency period of decades. The five amphibole asbestos species are assumed to be biopersistent in the lungs, and for this reason they are considered much more toxic than serpentine asbestos (chrysotile). Here, we refined the atomic structure of an amosite amphibole asbestos fibre that had remained in a human lung for ?40 years, in order to verify the stability <i>in vivo</i>. The subject was originally exposed to a blend of chrysotile, amosite and crocidolite, which remained in his parietal pleura for ?40 years. We found a few relicts of chrysotile fibres that were amorphous and magnesium depleted. Amphibole fibres that were recovered were undamaged and suitable for synchrotron X-ray micro-diffraction experiments. Our crystal structure refinement from a recovered amosite fibre demonstrates that the original atomic distribution in the crystal is intact and, consequently, that the atomic structure of amphibole asbestos fibres remains stable in the lungs for a lifetime; during which time they can cause chronic inflammation and other adverse effects that are responsible for carcinogenesis. The amosite fibres are not iron depleted proving that the iron pool for the formation of the asbestos bodies is biological (haemoglobin/plasma derived) and that it does not come from the asbestos fibres themselves.
Project description:Asbestos is still a social burden worldwide as a carcinogen causing malignant mesothelioma. Whereas recent studies suggest that local iron reduction is a preventive strategy against carcinogenesis, little is known regarding the cellular and molecular mechanisms surrounding excess iron. Here by differentially using high-risk and low-risk asbestos fibers (crocidolite and anthophyllite, respectively), we identified asbestos-induced mutagenic milieu for mesothelial cells. Rat and cell experiments revealed that phagocytosis of asbestos by macrophages results in their distinctive necrotic death; initially lysosome-depenent cell death and later ferroptosis, which increase intra- and extra-cellular catalytic Fe(II). DNA damage in mesothelial cells, as assessed by 8-hydroxy-2'-deoxyguanosine and ?-H2AX, increased after crocidolite exposure during regeneration accompanied by ?-catenin activation. Conversely, ?-catenin overexpression in mesothelial cells induced higher intracellular catalytic Fe(II) with increased G2/M cell-cycle fraction, when p16INK4A genomic loci localized more peripherally in the nucleus. Mesothelial cells after challenge of H2O2 under ?-catenin overexpression presented low p16INK4A expression with a high incidence of deletion in p16INK4A locus. Thus, crocidolite generated catalytic Fe(II)-rich mutagenic environment for mesothelial cells by necrotizing macrophages with lysosomal cell death and ferroptosis. These results suggest novel molecular strategies to prevent mesothelial carcinogenesis after asbestos exposure.
Project description:Human malignant mesothelioma (MM) is an aggressive tumor strongly associated with asbestos exposure. SM patients generally have poorer prognosis compared to EM patients. To identify potential genes accounting for the differential prognosis between these two subtypes, we compared the microarray gene expression profiles of rat SM and EM tissues induced by intraperitoneal injections of 3 types of asbestos (chrysotile,crocidolite and amosite). Carcinogenesis protocol was performed using specific pathogen-free male and female F1 hybrid rats between Fischer344 and Brown-Norway strains. A total of 28 microarrays (Whole Rat Genome Microarray) were used for screening purpose: The 2 arrays were used for knife-scraped peritoneal mesothelial cells, 2 arrays for cultured peritoneal mesothelial cells and 24 arrays for MM samples.