Share This Article:

Stachybotrys chartarum (atra) spore extract alters surfactant protein expression and surfactant function in isolated fetal rat lung epithelial cells, fibroblasts and human A549 cells

Full-Text HTML XML Download Download as PDF (Size:4579KB) PP. 243-256
DOI: 10.4236/ojped.2013.33043    3,978 Downloads   5,853 Views   Citations

ABSTRACT

Moulds, notably Stachybotrys chartarum (atra), are constant contributors to air pollution particularly to air quality in buildings. The spores themselves or their volatile organic products are present in variable amounts in almost all environments, particularly in buildings affected by flooding. These moulds and products can account for the sick building syndrome and have been tied to such occurrences as the outbreak of pulmonary hemosiderosis and hemorrhage in infants in Cleveland, Ohio. The present study was designed to investigate the effects of S. chartarum extracts on surfactant protein expression, surfactant quality and cell survival in the developing lung. S. chartarum extracts were incubated with cultures of several cell types; isolated fetal lung type II cells and fetal lung fibroblasts, and human lung A549 cells, a continuously growing cell line derived from surfactant producing type II alveolar cells. MTT formazan assays were employed to test cell viability. The synthesis and release of the predominant surfactant protein A (SP-A), which is involved in the regulation of surfactant turnover and metabolism, and surfactant protein B (SP-B) involved in shuttling phospholipids between surfactant subcompartments was also assessed. Antibodies to these proteins and western blotting results were used to assess the quantity of protein produced by the various cell types. A novel approach utilizing captive bubble surfactometry was employed to investigate the quality of surfactant in terms of surface tension and bubble volume measurements. Electron microscopy was used to examine changes in cellular structure of control and S. chartarum-treated cells. Results of the study showed that exposure to the S. chartarum extracts had deleterious effects on fetal lung epithelial cell viability and their ability to produce pulmonary surfactant. S. chartarum extracts also induced deleterious changes to the developing fetal lung cells in terms of expression of SP-A and SP-B as well as to the surface tension reducing abilities of the pulmonary surfactant. Ultrastructurally, spore toxin associated changes were apparent in the isolated lung cells most notably in the lamellar bodies of fetal rat lung alveolar type II and human A549 cells. This study has demonstrated the potential damage to surfactant production and function which may be induced by inhaling S. chartarum toxins.

Conflicts of Interest

The authors declare no conflicts of interest.

Cite this paper

Pollard, G. , Shaw, A. , Sowa, M. , Rand, T. , Thliveris, J. and Scott, J. (2013) Stachybotrys chartarum (atra) spore extract alters surfactant protein expression and surfactant function in isolated fetal rat lung epithelial cells, fibroblasts and human A549 cells. Open Journal of Pediatrics, 3, 243-256. doi: 10.4236/ojped.2013.33043.

References

[1] Montana, E., Etzel, R.A., Allan, T., Horgan, T.E. and Dearborn, D.G. (1997) Environmental risk factors associated with pediatric idiopathic pulmonary hemorrhage and hemosiderosis in a Cleveland community. Pediatrics, 99, E5. doi:10.1542/peds.99.1.e5
[2] Etzel, R.A., Montana, E., Sorenson, W.G., Kullman, G.J., Allan, T.M., Dearborn, D.G., Olson, D.R., Jarvis, B.B. and Miller, J.D. (1998) Acute pulmonary hemorrhage in infants associated with exposure to Stachybotrys atra and other fungi. Archives of Pediatrics & Adolescent Medicine, 152, 757-762. doi:10.1001/archpedi.152.8.757
[3] Weiss, A. and Chidekel, A.S. (2002) Acute pulmonary hemorrhage in a Delaware infant after exposure to Stachybotrys atra. Delaware Medical Journal, 74, 363-368.
[4] CDC Update (2000) Pulmonary hemorrhage/hemosiderosis among infants—Cleveland, Ohio, 1993-1996. Morbidity and Mortality Weekly Report (MMWR), 49, 180-184.
[5] Nikulin, M., Reijula, K., Jarvis, B.B. and Hintikka, E.L. (1996) Experimental lung mycotoxicosis in mice induced by Stachybotrys atra. International Journal of Experimental Pathology, 77, 213-218. doi:10.1046/j.1365-2613.1996.9250323.x
[6] Nikulin, M., Reijula, K., Jarvis, B.B., Veijalainen, P. and Hintikka, E.L. (1997) Effects of intranasal exposure to spores of Stachybotrys atra in mice. Fundamental and Applied Toxicology, 35, 182-188. doi:10.1006/faat.1996.2274
[7] Yike, I., Miller, M.J., Sorenson, W.G., Walenga, R., Tomashefski Jr., J.F. and Dearborn, D.G. (2002) Infant animal model of pulmonary mycotoxicosis induced by Stachybotrys chartarum. Mycopathologia, 154, 139-152. doi:10.1023/A:1016098331428
[8] Rao, C.Y., Brain, J.D. and Burge, H.A. (2000) Reduction of pulmonary toxicity of Stachybotrys chartarum spores by methanol extraction of mycotoxins. Applied and Environmental Microbiology, 66, 2817-2821. doi:10.1128/AEM.66.7.2817-2821.2000
[9] Jarvis, B.B., Salemme, J. and Morais, A. (1995) Stachybotrys toxins. Natural Toxins, 3, 10-16. doi:10.1002/nt.2620030104
[10] Jarvis, B.B., Sorenson, W.G., Hintikka, E.L., Nikulin, M., Zhou, Y., Jiang, J., Wang, S., Hinkley, S., Etzel, R.A. and Dearborn, D. (1998) Study of toxin production by isolates of Stachybotrys chartarum and Memnoniella echinata isolated during a study of pulmonary hemosiderosis in infants. Applied and Environmental Microbiology, 64, 3620-3625.
[11] Feinburg, B. and Maclaughlin, C.S. (1989) Biochemical mechanisms of action of trichothecenes mycotoxins. In: Beasley, V.R., Ed., Trichothecene Mycotoxins: Pathophysiological Effects, Boca Raton, CRC Press, 27-36.
[12] Vesper, S., Dearborn, D.G., Yike, I., Allan, T., Sobolewski, J., Hinkley, S.F., Jarvis, B.B. and Haugland, R.A. (2000) Evaluation of Stachybotrys chartarum in the house of an infant with pulmonary hemorrhage: Quantitative assessment before, during, and after remediation. Journal of Urban Health, 77, 68-85. doi:10.1007/BF02350963
[13] Vesper, S.J., Magnuson, M.L., Dearborn, D.G., Yike, I. and Haugland, R.A. (2001) Initial characterization of the hemolysin stachylysin from Stachybotrys chartarum. Infection and Immunity, 69, 912-916.
[14] Hossain, M.A., Ahmed, M.S. and Ghannoum, M.A. (2004) Attributes of Stachybotrys chartarum and its association with human disease. Journal of Allergy and Clinical Immunology, 113, 200-208. doi:10.1016/j.jaci.2003.12.018
[15] Pestka, J.J., Yike, I., Dearborn, D.G., Ward, M.D. and Harkema, J.R. (2008) Stachybotrys chartarum, trichothecene mycotoxins, and damp building-related illness: New insights into a public health enigma. Toxicological Sciences, 104, 4-26. doi:10.1093/toxsci/kfm284
[16] Andersen, B., Nielsen, K.F. and Jarvis, B.B. (2002) Characterization of Stachybotrys from water-damaged buildings based on morphology, growth, and metabolite production. Mycologia, 94, 392-403. doi:10.2307/3761773
[17] Jarvis, B.B. (2003) Stachybotrys chartarum: A fungus for our time. Phytochemistry, 64, 53-60. doi:10.1016/S0031-9422(03)00275-9
[18] Miller, J.D., Rand, T.G. and Jarvis, B.B. (2003) Stachybotrys chartarum: Cause of human disease or media darling? Medical Mycology, 41, 271-291. doi:10.1080/1369378031000137350
[19] Nielsen, K.F., Huttunen, K., Hyvarinen, A., Andersen, B., Jarvis, B.B. and Hirvonen, M.R. (2002) Metabolite profiles of Stachybotrys isolates from water-damaged buildings and their induction of inflammatory mediators and cytotoxicity in macrophages. Mycopathologia, 154, 201-205. doi:10.1023/A:1016383402963
[20] Flemming, J., Hudson, B. and Rand, T.G. (2004) Comparison of inflammatory and cytotoxic lung responses in mice after intratracheal exposure to spores of two different Stachybotrys chartarum strains. Toxicological Sciences, 78, 267-275. doi:10.1093/toxsci/kfh064
[21] Yike, I., Vesper, S., Tomashefski Jr., J.F. and Dearborn, D.G. (2003) Germination, viability and clearance of Stachybotrys chartarum in the lungs of infant rats. Mycopathologia, 156, 67-75. doi:10.1023/A:1022924306264
[22] Yike, I. and Dearborn, D.G. (2004) Pulmonary effects of Stachybotrys chartarum in animal studies. Advances in Applied Microbiology, 55, 241-273. doi:10.1016/S0065-2164(04)55009-8
[23] Shelton, B.G., Kirkland, K.H., Flanders, W.D. and Morris, G.K. (2002) Profiles of airborne fungi in buildings and outdoor environments in the United States. Applied and Environmental Microbiology, 68, 1743-1753. doi:10.1128/AEM.68.4.1743-1753.2002
[24] Rand, T.G., Mahoney, M., White, K. and Oulton, M. (2002) Microanatomical changes in alveolar type II cells in juvenile mice intratracheally exposed to Stachybotrys chartarum spores and toxin. Toxicological Sciences, 65, 239-245. doi:10.1093/toxsci/65.2.239
[25] McCrae, K.C., Rand, T., Shaw, R.A., Mason, C., Oulton, M.R., Hastings, C., Cherlet, T., Thliveris, J.A., Mantsch, H.H., MacDonald, J. and Scott, J.E. (2001) Analysis of pulmonary surfactant by Fourier-transform infrared spectroscopy following exposure to Stachybotrys chartarum (atra) spores. Chemistry and Physics of Lipids, 110, 1-10. doi:10.1016/S0009-3084(00)00199-7
[26] McCrae, K.C., Rand, T.G., Shaw, R.A., Mantsch, H.H., Sowa, M.G., Thliveris, J.A. and Scott, J.E. (2007) DNA fragmentation in developing lung fibroblasts exposed to Stachybotrys chartarum (atra) toxins. Pediatric Pulmonology, 42, 592-599. doi:10.1002/ppul.20608
[27] Scott, J.E., Possmayer, F., Quirie, M.A., Tanswell, A.K. and Harding, P.G. (1986) Alveolar pre-type II cells from the fetal rabbit lung: Effect of confluence on the production of disaturated phosphatidylcholine. Biochimica et Biophysica Acta, 879, 292-300. doi:10.1016/0005-2760(86)90218-3
[28] Mason, C.D., Rand, T.G., Oulton, M., MacDonald, J.M. and Scott, J.E. (1998) Effects of Stachybotrys chartarum (atra) conidia and isolated toxin on lung surfactant production and homeostasis. Natural Toxins, 6, 27-33. doi:10.1002/(SICI)1522-7189(199802)6:1<27::AID-NT6>3.0.CO;2-B
[29] Bradford, M.M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248-254. doi:10.1016/0003-2697(76)90527-3
[30] Bligh, E.G. and Dyer, W.J. (1959) A rapid method of total lipid extraction and purification. Canadian Journal of Biochemistry and Physiology, 37, 911-917. doi:10.1139/o59-099
[31] Bartlett, G.R. (1959) Phosphorus assay in column chromatography. The Journal of Biological Chemistry, 234, 466-468.
[32] Schoel, W.M., Schurch, S. and Goerke, J. (1994) The captive bubble method for the evaluation of pulmonary surfactant: Surface tension, area, and volume calculations. Biochimica et Biophysica Acta, 1200, 281-290. doi:10.1016/0304-4165(94)90169-4
[33] Luft, J.H. (1961) Improvements in epoxy resin embedding methods. Journal of Biophysical and Biochemical Cytology, 9, 409-414. doi:10.1083/jcb.9.2.409
[34] Ikegami, M. and Jobe, A.H. (1998) Surfactant protein-C in ventilated premature lamb lung. Pediatric Research, 44, 860-864. doi:10.1203/00006450-199812000-00006
[35] Hawgood, S., Derrick, M. and Poulain, F. (1998) Structure and properties of surfactant protein B. Biochimica et Biophysica Acta, 1408, 150-160. doi:10.1016/S0925-4439(98)00064-7
[36] Liu, Y., Peterson, D.A., Kimura, H. and Schubert, D. (1997) Mechanism of cellular 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. Journal of Neurochemistry, 69, 581-593. doi:10.1046/j.1471-4159.1997.69020581.x
[37] Gregory, L., Pestka, J.J., Dearborn, D.G. and Rand, T.G. (2004) Localization of satratoxin-G in Stachybotrys chartarum spores and spore-impacted mouse lung using immunocytochemistry. Journal of Toxicologic Pathology, 32, 26-34. doi:10.1080/01926230490260790
[38] Rehan, V.K., Sugano, S., Wang, Y., Santos, J., Romero, S., Dasgupta, C., Keane, M.P., Stahlman, M.T. and Torday, J.S. (2006) Evidence for the presence of lipofibroblasts in human lung. Experimental Lung Research, 32, 379-393. doi:10.1080/01902140600880257
[39] McGowan, S.E. and Torday, J.S. (1997) The pulmonary lipofibroblast (lipid interstitial cell) and its contributions to alveolar development. Annual Review of Physiology, 59, 43-62.doi:10.1146/annurev. physiol.59.1.43
[40] Brody, J.S. and Kaplan, N.B. (1983) Proliferation of alveolar interstitial cells during postnatal lung growth. Evidence for two distinct populations of pulmonary fibroblasts. American Review of Respiratory Disease, 127, 763-770.
[41] Hastings, C., Rand, T., Bergen, H.T., Thliveris, J.A., Shaw, A.R., Lombaert, G.A., Mantsch, H.H., Giles, B.L., Dakshinamurti, S. and Scott, J.E. (2005) Stachybotrys chartarum alters surfactant-related phospholipid synthesis and CTP: cholinephosphate cytidylyltransferase activity in isolated fetal rat type II cells. Toxicological Sciences, 84, 186-194. doi:10.1093/toxsci/kfi045
[42] Mason, C.D., Rand, T.G., Oulton, M., MacDonald, J. and Anthes, M. (2001) Effects of Stachybotrys chartarum on surfactant convertase activity in juvenile mice. Toxicology and Applied Pharmacology, 172, 21-28. doi:10.1006/taap.2001.9127
[43] Bi, X., Taneva, S., Keough, K.M., Mendelsohn, R. and Flach, C.R. (2001) Thermal stability and DPPC/Ca2+ interactions of pulmonary surfactant SP-A from bulk-phase and monolayer IR spectroscopy. Biochemistry, 40, 13659-13669. doi:10.1021/bi011188h
[44] Andreassen, S., Steimle, K.L., Mogensen, M.L., Bernardino de la Serna, J., Rees, S. and Karbing, D.S. (2010) The effect of tissue elastic properties and surfactant on alveolar stability. Journal of Applied Physiology, 109, 1369-1377. doi:10.1152/japplphysiol.00844.2009
[45] Curstedt, T. and Johansson, J. (2005) New synthetic surfactants—Basic science. Biology of the Neonate, 87, 332-337. doi:10.1159/000084881

  
comments powered by Disqus

Copyright © 2018 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.