In vitro cytotoxicity testing of airborne formaldehyde collected in serum-free culture media

Abstract

The purpose of this study was to identify a suitable sampling model for on-site toxicity assessment of soluble air contaminants such as formaldehyde, a well known industrial and indoor air contaminant. The in vitro cytotoxicity of formaldehyde, the selected model for soluble air contaminants, was studied using the MTS (tetrazolium salt) assay in two carcinoma cell lines, A549 epithelial lung and HepG2 hepatocarcinoma, and in skin fibroblasts. The cytotoxic effects of airborne formaldehyde were evaluated using test atmospheres in concentrations below 10 ppm (12.3 mg/m³), generated by a dynamic diffusion method and bubbled (0.3 L/min) through serum-free culture media for one or four hours. Human cells were treated with formaldehyde air samples, and cell viability was determined after four hours incubation. In parallel, the concentration of airborne formaldehyde was monitored, using the 3500 NIOSH method. Cell viability of the HepG2 cells exposed to formaldehyde air samples (8.75 ppm-4 h) was reduced to less than 50% (31.69/1.24%). The HepG2 cell lines were found to be more sensitive (IC₅₀=103.799/23.55 mg/L) to formaldehyde than both A549 cell lines (IC₅₀=198.369/9.54 mg/L) and skin fibroblasts (IC₅₀=196.689/36.73 mg/L) (PB/0.01). An average of 96.8% was determined for collection efficiency of formaldehyde in serum-free culture media. The results of this study suggest that absorption of soluble air contaminants, such as formaldehyde, in serum-free culture media can be used as a suitable sampling model for on-site toxicity assessments.

Keywords

air contaminants cytotoxicity formaldehyde in vitro toxicity MTS (tetrazolium salt) assay

Get full access to this article

View all access options for this article.

References

Alfaro Moreno, E. , Martinez, L. , Garcia Cuellar, C. et al. 2002: Biologic effects induced in vitro by PM 10 from three different zones of Mexico City. Environmental Health Perspectives 110, 715-720.

ATSDR , 1999: Toxicological profile for Formaldehyde. Atlanta: US Department of Health and Human Services, Public Health Service. Agency for Toxic Substances and Disease Registry.

Aufderheide, M. , Knebel, J. W. and Ritter, D. 2003: Novel approaches for studying pulmonary toxicity in vitro. Toxicology Letters 140-141, 205-211.

Blanquart, C. , Giuliani, I. , Houcine, O. et al. 1995: In vitro exposure of rabbit trachelium to SO2: effects on morphology and ciliarity beating. Toxicology In Vitro 9, 123-132.

Chen, L. C. , Fang, C. P. , Qu, Q. S. et al. 1993: A novel system for the in vitro exposure of pulmonary cells to acid sulfate aerosols. Fundamental and Applied Toxicology 20, 170-176.

Clemedson, C. and Ekwall, B. 1999: Overview of the final MEIC results: I. The in vitro-in vitro evaluation. Toxicology In Vitro 13, 657-663.

Conolly, R. B. , Kimbell, J. S. , Janszen, D. B. and Miller, F. J. 2002: Dose response for formaldehyde-induced cytotoxi-city in the human respiratory tract. Regulatory Toxicology and Pharmacology 35, 32-43.

Ekwall, B. 1999: Overview of the final MEIC results: II. The in vitro-in vivo evaluation, including the selection of a practical battery of cell tests for prediction of acute lethal blood concentrations in humans. Toxicology In Vitro 13, 665-673.

Glowala, M. , Mazurek, A. , Piddubnyak, V. et al. 2002: HSP70 over expression increases resistance of V79 cells to cytotoxicity of airborne pollutants, but does not protect the mitotic spindle against damage caused by airborne toxins. Toxicology 170, 211-219.

10.

Hamers, T. , Van Schaardenburg, M. D. , Felzel, E. C. et al. 2000: The application of reporter gene assay for the determination of the toxic potency of diffuse air pollution. The Science of the Total Environment 262, 159-174.

11.

IARC , 1995: International Agency for Research on Cancer - summaries & evaluations, formaldehyde, vol 62. Retrieved from http://www.inchem.org/documents/iarc/vol62/formal.html (accessed September 2003).

12.

IARC , 2004: IARC classifies formaldehyde as carcinogenic to humans. International Agency for Research on Cancer, Press Release, N 153.

13.

IPCS , 1989: Formaldehyde. Environmental health criteria, No 89. International Programme on Chemical Safety. Geneva: World Health Organization.

14.

Jabbour, A. J. , Altman, L. C. , Wight, T. N. and Luchtel, D. L. 1998: Ozone alters the distribution of beta1 integrins in cultured primate bronchial epithelial cells. American Journal of Respiratory Cell and Molecular Biology 19, 357-365.

15.

Knebel, J. W. , Ritter, D. and Aufderheide, M. 1998: Development of an in vitro system for studying effects of native and photochemically transformed gaseous compounds using an air/liquid culture technique. Toxicology Letters 96 -97, 1-11.

16.

Lieber, M. , Smith, B. , Szakal, A. et al. 1976: A continuous tumour-cell line from a human lung carcinoma with properties of type II alveolar epithelial cells. International Journal of Cancer 17, 62-70.

17.

Lovschall, H. , Eiskjaer, M. and Arenholt-Bindslev, D. 2002: Formaldehyde cytotoxicity in three human cell types assessed in three different assays. Toxicology In Vitro 16, 63-69.

18.

Malich, G. , Markovic, B. and Winder, C. 1997: The sensitivity and specificity of the MTS tetrazolium assay for detecting the in vitro cytotoxicity of 20 chemicals using human cell lines. Toxicology 124, 179-192.

19.

Monticello, T. M. , Miller, F. J. and Morgan, K. T. 1991: Regional increases in rat nasal epithelial cell proliferation following acute and subchronic inhalation of formalde-hyde. Toxicology & Applied Pharmacology 111, 409-421.

20.

Muckter, H. , Zwing, M. , Bader, S. et al. 1998: A novel apparatus for the exposure of cultured cells to volatile agents. Journal of Pharmacological and Toxicological Methods 40, 63-69.

21.

Newell, G. W. 1983: Overview of formaldehyde. In Gibson, J. E. , editor, Formaldehyde toxicity. Washington, DC: Hemisphere Publishing Corporation, 3-15.

22.

NIOSH , 1994: NIOSH manual of analytical methods (NMAM), fourth edition. National Institute for Occupational Safety and Health. Retrieved from http://www.cdc.gov/niosh/nmam/nmammenu.html (accessed March 2003).

23.

NIOSH , 2002: The registry of toxic effects of chemical substances, formaldehyde, RTECS NO LP8925000. National Institute for Occupational Safety and Health. Retrieved from http://www.cdc.gov/niosh/rtecs/lp882f48.html (accessed September 2003).

24.

Promega, CellTiter 96® Aqueous Non-Radioactive Cell Proliferation Assay: Technical Bulletin No 169. Madison, USA: Promega Corporation

25.

Ritter, D. , Knebel, J. W. and Aufderheide, M. 2001: In vitro exposure of isolated cells to native gaseous compounds - development and validation of an optimized system for human lung cells. Experimental and Toxicologic Pathology 53, 373-386.

26.

Rusznak, C. , Devalia, J. L. , Sapsford, R. J. and Davies, R. J. , 1996: Ozone-induced mediator release from human bronchial epithelial cells in vitro and the influence of nedocromil sodium. European Respiratory Journal 9, 2298-2305.

27.

Saltzman, B. E. 1997: Preparation of known concentration of air contaminants. In Dinardi, S. R. , editor, The occupational environment - its evaluation and control. Fairfax, VA: American Industrial Hygienists Association, 131-154.

28.

Van Der Zee, J. , Dubbelman, T. M. A. R. , Raap, T. K. and Van Steveninck, J. 1987: Toxic effects of ozone on murine L929 fibroblasts. Enzyme inactivation and glutathion depletion. Biochemical Journal 242, 707-712.

29.

WHO, 2002: Formaldehyde, Concise International Chemical Assessment Document, No 40. Geneva: World Health Organization.

30.

Wilmer, J. W. , Woutersen, R. A. , Appelman, L. M. et al. 1989: Subchronic (13-week) inhalation toxicity study of formal-dehyde in male rats: 8-hour intermittent versus 8-hour continuous exposure. Toxicology Letters 47, 287-293.

31.

Yamaguchi, T. and Yamazaki, H. 2001: Cytotoxicity of airborne particulates sampled roadside in rodent and human lung fibroblasts. Journal of Health Science 47, 272-277.