Permeation of roxarsone and its metabolites increases caco-2 cell proliferation
Author(s)
Gladys S. Bayse,
Latanya P. Hammonds-Odie*,
Kimberly M. Jackson,
Deidre K. Tucker,
Ward G. Kirlin
Affiliation(s)
Department of Chemistry, Spelman College, Atlanta, USA. Department of Biology, Spelman College, Atlanta, USA. Department of Pharmacology and Toxicology, Morehouse School of Medicine, Atlanta, USA.
Department of Chemistry, Spelman College, Atlanta, USA. Department of Biology, Spelman College, Atlanta, USA. Department of Pharmacology and Toxicology, Morehouse School of Medicine, Atlanta, USA.
ABSTRACT
The benzenearsonate, Roxarsone, has been used since 1944 as an antimicrobial, growth-promoting poultry feed additive. USGS and EPA report that Roxarsone (4-hydroxy-3-nitrobenzenearsonate) and metabolites, including AHBA (3-amino-4-hydroxybenzenearsonate), contaminate waterways at greater than 1100 tons annually. To assess human impact of these organic arsenic water contaminants, it was important to study their potential absorption. The human adenocarcinoma cell line, Caco-2, is a model for intestinal absorption. We found proliferative effects on Caco-2 cells at micromolar levels of these compounds, as monitored by [3H]-thymidine incorporation into DNA. Flow cytometry cell cycle analysis confirmed accumulation in S phase from 21% (control) to 36% (24 hour exposure to 10 μM AHBA). Confluent Caco-2 cells grown on collagen-coated Transwell plates were dosed on the apical side. After exposure, media from apical and basolateral sides were collected separately. Following removal of FBS by 30K centrifugal filtration, the benzenearsonates in the collected media were analyzed by HPLC. Analyses were at wavelengths in the ultraviolet/visible range where the absorbance values were linear with respect to concentration. Concentrations were calculated by comparison with analytically-prepared commercial standards. Results from cells dosed at 10 μM for 24 hours with AHBA, Roxarsone, or Acetarsone indicated 6%-29% permeation occurring from apical to basolateral side, modeling absorption across intestinal epithelium to the circulatory system. Benzenearsonate feed additives are frequently applied in combination with antibiotics, raising additional health concerns. We conclude that micromolar levels of these benzenearsonates are adequate to stimulate Caco-2 cell proliferation.
The benzenearsonate, Roxarsone, has been used since 1944 as an antimicrobial, growth-promoting poultry feed additive. USGS and EPA report that Roxarsone (4-hydroxy-3-nitrobenzenearsonate) and metabolites, including AHBA (3-amino-4-hydroxybenzenearsonate), contaminate waterways at greater than 1100 tons annually. To assess human impact of these organic arsenic water contaminants, it was important to study their potential absorption. The human adenocarcinoma cell line, Caco-2, is a model for intestinal absorption. We found proliferative effects on Caco-2 cells at micromolar levels of these compounds, as monitored by [3H]-thymidine incorporation into DNA. Flow cytometry cell cycle analysis confirmed accumulation in S phase from 21% (control) to 36% (24 hour exposure to 10 μM AHBA). Confluent Caco-2 cells grown on collagen-coated Transwell plates were dosed on the apical side. After exposure, media from apical and basolateral sides were collected separately. Following removal of FBS by 30K centrifugal filtration, the benzenearsonates in the collected media were analyzed by HPLC. Analyses were at wavelengths in the ultraviolet/visible range where the absorbance values were linear with respect to concentration. Concentrations were calculated by comparison with analytically-prepared commercial standards. Results from cells dosed at 10 μM for 24 hours with AHBA, Roxarsone, or Acetarsone indicated 6%-29% permeation occurring from apical to basolateral side, modeling absorption across intestinal epithelium to the circulatory system. Benzenearsonate feed additives are frequently applied in combination with antibiotics, raising additional health concerns. We conclude that micromolar levels of these benzenearsonates are adequate to stimulate Caco-2 cell proliferation.
Cite this paper
Bayse, G. , Hammonds-Odie, L. , Jackson, K. , Tucker, D. and Kirlin, W. (2013) Permeation of roxarsone and its metabolites increases caco-2 cell proliferation. Advances in Biological Chemistry, 3, 389-396. doi: 10.4236/abc.2013.34041.
Bayse, G. , Hammonds-Odie, L. , Jackson, K. , Tucker, D. and Kirlin, W. (2013) Permeation of roxarsone and its metabolites increases caco-2 cell proliferation. Advances in Biological Chemistry, 3, 389-396. doi: 10.4236/abc.2013.34041.
References
[1] Kolpin, D.W., et al. (2002) Pharmaceuticals, hormones, and other organic wastewater contaminants in US streams: A national reconnaissance. Environmental Science & Technology, 36, 1202-1211. doi:10.1021/es020136s [2] Osterberg, D. and Wallinga, D. (2004) Addressing externalities from swine production to reduce public health and environmental impacts. American Journal of Public Health, 94, 1703-1708. doi:10.2105/AJPH.94.10.1703 [3] US Food and Drug Administration (2010) 21 CFR Part 558.530. Food and drugs: new animal drugs for use in animal feeds. Electronic code of federal regulations. http://www.gpo.gov/fdsys/pkg/CFR-2010-title21-vol6/xml/CFR-2010-title21-vol6-part558.xml#seqnum558.530 [4] Calvert, C.C. (1975) Arsenicals in animal feeds and wastes. In: Woolson, E.A. Ed., Arsenical Pesticides, American Chemical Society, Washington DC, 70-79. doi:10.1021/bk-1975-0007.ch005 [5] Garbarino, J.R., Bednar, A.J., Rutherford, D.W., Beyer, R.S. and Wershaw, R.L. (2003) Environmental fate of roxarsone in poultry litter. I. Degradation of roxarsone during composting. Environmental Science & Technology, 37, 1509-1514. doi:10.1021/es026219q [6] Jackson, B.P. and Bertsch, P.M. (2001) Determination of arsenic speciation in poultry wastes by IC-ICP-MS. Environmental Science & Technology, 35, 4868-4873. doi:10.1021/es0107172 [7] Rutherford, D.W., et al. (2003) Environmental fate of roxarsone in poultry litter. Part II. Mobility of arsenic in soils amended with poultry litter. Environmental Science & Technology, 37, 1515-1520. doi:10.1021/es026222+ [8] Hileman, B. (2007) Arsenic in chicken production. Chemical and Engineering News, 85, 34-35. doi:10.1021/cen-v085n015.p034 [9] Cody, C. (2003) Siloam springs: Farmers told to stop spreading litter. Arkansas Democrat Gazette, 1. [10] Joyner, C. (2011) Feds warn of potential water contamination. The Atlanta Journal-Constitution, 14, SA1-SA12. [11] Nachman, K.E., Graham, J.P., Price, L.B. and Silbergeld, E.K. (2005) Arsenic: A roadblock to potential animal waste management solutions. Environ Health Perspectives, 113, 1123-1124. doi:10.1289/ehp.7834 [12] Moody, J.P. and Williams, R.T. (1964) The metabolism of 4-hydroxynitrophenylarsonic acid in hens. Food and Cosmetics Toxicology, 2, 707-710. doi:10.1016/S0015-6264(64)80422-3 [13] Cortinas, I., et al. (2006) Anaerobic biotransformation of roxarsone and related N-substituted phenylarsonic acids. Environmental Science & Technology, 40, 2951-2957. doi:10.1021/es051981o [14] Stoltz, J.F., et al. (2007) Biotransformation of 3-nitro-4-hydroxybenzene arsonic acid and release of inorganic arsenic by Clostridium species. Environmental Science & Technology, 41, 818-823. doi:10.1021/es061802i [15] Sabbioni, G. and Richter, E. (1999) Aromatic amines, nitroarenes, and heterocyclic aromatic amines. In: Marquardt, H., Schafer, S.G., McClellan, R.O. and Welsch, F. Eds., Toxicology, Academic Press, San Diego, 730-736. doi:10.1016/B978-012473270-4/50089-4 [16] Wershaw, R.L., et al. (2003) Mass spectrometric identification of an azobenzene derivative produced by smectitecatalyzed conversion of 3-amino-4-hydroxy-phenyl-arsonic acid to an azobenzene derivative. Talanta, 59, 1219-1226. doi:10.1016/S0039-9140(03)00032-8 [17] Bayse, G.S., Kirlin, W.G. and Kirkland, P.D. (2004) Effects of roxarsone and its metabolites on caco-2 cell proliferation. Toxicologist, 82, 1447. [18] Bayse, G.S., Jinadu, L.A., Shaw, K.A. and Wiley, K.L. (2000) The N-acetylation of arsanilic acid in vitro by mammalian enzymes. Drug Metabolism and Disposition, 28, 487-492. [19] O’Neil, M.J. (2006) The Merck Index: An Encyclopedia of Chemicals, Drugs and Biological. Merck and Company, Whitehouse Station. [20] Harris, G. and Grady, D. (2011) Pfizer suspends sales of chicken drug with arsenic. New York Times, SB, 2. [21] Ashby, J. and Tennant, R.W. (1991) Definitive relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by the US NTP. Mutation Research, 257, 229-306. doi:10.1016/0165-1110(91)90003-E [22] Parkinson, A. (2001) Biotransformation of xenobiotics. In: Klaassen, C.D. Ed., Casarett and Doull’s Toxicology, McGraw Hill, New York, 133-224. [23] Bayse, G.S., Jackson, K.M., Kirlin, W.G. and Rollins-Hairston, A. (2006) Proliferation of human caco-2 cells mediated by N-acetylation and oxidation reactions of 3-amino-4-hydroxybenzenearsonate. Toxicologist, 90, 674. [24] Bayse, G.S., et al. (2007) Permeation of benzenearsonates provides sufficient concentrations to cause Alterations in caco-2 cell proliferation. Toxicologist, 96, 1174. [25] Robinson, D.K., Hammonds-Odie, L.P., Jackson, K.M., Kirlin, W.G. and Bayse, G.S. (2010) Caco-2 cell permeation of five benzenearsonates increases likelihood of hepatic biotransformations. Toxicologist, 114, 1101. [26] Hammonds-Odie, L.P., Bayse, G.S., Jackson, K.M. and Robinson, D.K. (2006) Evaluation of benezenearsonate permeation in caco-2 cells. http://www.fasebj.org/cgi/content/meeting_abstract/20/5/LB111-b?sid=cf22ab74-c1a9-4417-b169-133f4b8aa2ce [27] Artursson, P. (1990) Epithelial transport of drugs in cell culture. I: A model for studying the passive diffusion of drugs over intestinal absorptive (caco-2) cells. Journal of Pharmaceutical Sciences, 79, 476-482. doi:10.1002/jps.2600790604 [28] Hidalgo, I.J., Raub, T.J. and Borchardt, R.T. (1989) Characterization of the human colon carcinoma cell line (ca-co-2) as a model system for intestinal epithelial permeability. Gastroenterology, 96, 736-749. [29] Prueksaritanont, T., Gorham, L.M., Hochman, J.H., Tran, L.O. and Vyas, K.P. (1996) Comparative studies of drugmetabolizing enzymes in dog, monkey, and human small intestines, and in caco-2 cells. Drug Metabolism and Disposition, 24, 634-642. [30] Hayes, J.R. and Campbell, T.C. (1986) Food additives and contaminants. In: Klaassen, C.D., Amdur, M.O. and Doull, J. Eds., Casarett and Doull’s Toxicology, Macmillan, New York, 771-800. [31] Abdo, K.M., et al. (1989) Toxic responses in F344 rats and B6CF1 mice given roxarsone in their diets for up to 13 weeks. Toxicology Letters, 45, 5-66. doi:10.1016/0378-4274(89)90159-8 [32] Ashby, J. and Paton, D. (1993) The influence of chemical structure on the extent and sites of carcinogenesis for 522 rodent carcinogens and 55 different human carcinogen exposures. Mutation Research, 286, 3-74. doi:10.1016/0027-5107(93)90003-X [33] US Environmental Protection Agency (2011) Region 4 should strengthen oversight of Georgia’s concentrated animal feeding operation program. http://www.epa.gov/oig/reports/2011/20110623-11-P-0274.pdf [34] Thompson, C.M., Haws, L.C., Harris, M.A., Gatto, N.M. and Proctor, D.M. (2011) Application of the US EPA mode of action framework for purposes of guiding future research: A case study involving the oral carcinogenicity of hexavalent chromium. Toxicological Sciences, 119, 20-40. doi:10.1093/toxsci/kfq320 [35] Doerge, D.R., Churchwell, M.I., Marques, M.M. and Beland, F.A. (1999) Quantitative analysis of 4-aminobiphenyl-C8-deoxyguanosyl DNA adducts produced in vitro using HPLC-ES-MS. Carcinogenesis, 20, 1055-1061. doi:10.1093/carcin/20.6.1055 [36] Zhang, T., Cao, E. and Qin, J. (2002) Opposite biological effects of arsenic trioxide and arsacetin involve a different regulation of signaling in human gastric cancer MGC-803 cells. Pharmacology, 64, 160-168. doi:10.1159/000056166 [37] Park, W.H., et al. (2000) Arsenic trioxide-mediated growth inhibition in MC/CAR myeloma cells via cell cycle arrest in association with induction of cyclin-dependent kinase inhibitor, p21, and apoptosis. Cancer Research, 60, 3065-3071. [38] Vogt, B.L. and Rossman, T.G. (2001) Effects of arsenite on p53, p21 and cyclin D expression in normal human fibroblasts—A possible mechanism for arsenite’s comutagenicity. Mutation Research, 478, 159-168. doi:10.1016/S0027-5107(01)00137-3 [39] Barr, F.D., Krohmer, L.J., Hamilton, J.W. and Sheldon, L.A. (2009) Disruption of histone modification and CARM1 recruitment by arsenic represses transcription at glucocorticoid receptor-regulated promoters. PLoS ONE, 4, e6766. doi:10.1371/journal.pone.0006766 [40] Davey, J.C., et al. (2008) Arsenic as an endocrine disruptor: Arsenic disrupts retinoic acid receptor-and thyroid hormone receptor-mediated gene regulation and thyroid hormone-mediated amphibian tail metamorphosis. Environmental Health Perspectives, 116, 165-172. [41] Kozul, C.D., et al. (2009) Chronic exposure to arsenic in the drinking water alters the expression of immune response genes in mouse lung. Environmental Health Perspectives, 117, 1108-1115. doi:10.1289/ehp.0800199 [42] Meharg, A.A. and Raab, A. (2010). Getting to the bottom of arsenic standards and guidelines. Environmental Science & Technology, 44, 4395-4399. doi:10.1021/es9034304 [43] Tapio, S. and Grosche, B. (2006) Arsenic in the aetiology of cancer. Mutation Research, 612, 215-246. doi:10.1016/j.mrrev.2006.02.001 [44] Kozul-Horvath, C.D., Zandbergen, F., Jackson, B.P., Enelow, R.I. and Hamilton, J.W. (2012) Effects of lowdose drinking water arsenic on mouse fetal and postnatal growth and development. PLoS ONE, 7, e38249. doi:10.1371/journal.pone.0038249 [45] Price, L.B., et al. (2007) Elevated risk of carrying gentamicin-resistant Escherichia coli among US poultry workers. Environmental Health Perspectives, 115, 1738-1742. doi:10.1289/ehp.10191 [46] Sapkota, A.R., Price, L.B. and Silbergeld, E.K. (2006) Arsenic resistance in Campylobacter spp. isolated from retail poultry products. Applied and Environmental Microbiology, 72, 3069-3071. doi:10.1128/AEM.72.4.3069-3071.2006 [47] Marshall, B.M. and Levy, S.B. (2011) Food animals and antimicrobials: Impacts on human health. Clinical Microbiology Reviews, 2, 718-733. doi:10.1128/CMR.00002-11 [48] US Food and Drug Administration (2012) 21 CFR part 558 subpart B-specific new animal drugs for use in animal feeds. Electronic Code of Federal Regulations. http://www.gpo.gov/fdsys/pkg/CFR-2012-title21-vol6/pdf/CFR-2012-title21-vol6-sec558-530.pdf
[1] Kolpin, D.W., et al. (2002) Pharmaceuticals, hormones, and other organic wastewater contaminants in US streams: A national reconnaissance. Environmental Science & Technology, 36, 1202-1211. doi:10.1021/es020136s [2] Osterberg, D. and Wallinga, D. (2004) Addressing externalities from swine production to reduce public health and environmental impacts. American Journal of Public Health, 94, 1703-1708. doi:10.2105/AJPH.94.10.1703 [3] US Food and Drug Administration (2010) 21 CFR Part 558.530. Food and drugs: new animal drugs for use in animal feeds. Electronic code of federal regulations. http://www.gpo.gov/fdsys/pkg/CFR-2010-title21-vol6/xml/CFR-2010-title21-vol6-part558.xml#seqnum558.530 [4] Calvert, C.C. (1975) Arsenicals in animal feeds and wastes. In: Woolson, E.A. Ed., Arsenical Pesticides, American Chemical Society, Washington DC, 70-79. doi:10.1021/bk-1975-0007.ch005 [5] Garbarino, J.R., Bednar, A.J., Rutherford, D.W., Beyer, R.S. and Wershaw, R.L. (2003) Environmental fate of roxarsone in poultry litter. I. Degradation of roxarsone during composting. Environmental Science & Technology, 37, 1509-1514. doi:10.1021/es026219q [6] Jackson, B.P. and Bertsch, P.M. (2001) Determination of arsenic speciation in poultry wastes by IC-ICP-MS. Environmental Science & Technology, 35, 4868-4873. doi:10.1021/es0107172 [7] Rutherford, D.W., et al. (2003) Environmental fate of roxarsone in poultry litter. Part II. Mobility of arsenic in soils amended with poultry litter. Environmental Science & Technology, 37, 1515-1520. doi:10.1021/es026222+ [8] Hileman, B. (2007) Arsenic in chicken production. Chemical and Engineering News, 85, 34-35. doi:10.1021/cen-v085n015.p034 [9] Cody, C. (2003) Siloam springs: Farmers told to stop spreading litter. Arkansas Democrat Gazette, 1. [10] Joyner, C. (2011) Feds warn of potential water contamination. The Atlanta Journal-Constitution, 14, SA1-SA12. [11] Nachman, K.E., Graham, J.P., Price, L.B. and Silbergeld, E.K. (2005) Arsenic: A roadblock to potential animal waste management solutions. Environ Health Perspectives, 113, 1123-1124. doi:10.1289/ehp.7834 [12] Moody, J.P. and Williams, R.T. (1964) The metabolism of 4-hydroxynitrophenylarsonic acid in hens. Food and Cosmetics Toxicology, 2, 707-710. doi:10.1016/S0015-6264(64)80422-3 [13] Cortinas, I., et al. (2006) Anaerobic biotransformation of roxarsone and related N-substituted phenylarsonic acids. Environmental Science & Technology, 40, 2951-2957. doi:10.1021/es051981o [14] Stoltz, J.F., et al. (2007) Biotransformation of 3-nitro-4-hydroxybenzene arsonic acid and release of inorganic arsenic by Clostridium species. Environmental Science & Technology, 41, 818-823. doi:10.1021/es061802i [15] Sabbioni, G. and Richter, E. (1999) Aromatic amines, nitroarenes, and heterocyclic aromatic amines. In: Marquardt, H., Schafer, S.G., McClellan, R.O. and Welsch, F. Eds., Toxicology, Academic Press, San Diego, 730-736. doi:10.1016/B978-012473270-4/50089-4 [16] Wershaw, R.L., et al. (2003) Mass spectrometric identification of an azobenzene derivative produced by smectitecatalyzed conversion of 3-amino-4-hydroxy-phenyl-arsonic acid to an azobenzene derivative. Talanta, 59, 1219-1226. doi:10.1016/S0039-9140(03)00032-8 [17] Bayse, G.S., Kirlin, W.G. and Kirkland, P.D. (2004) Effects of roxarsone and its metabolites on caco-2 cell proliferation. Toxicologist, 82, 1447. [18] Bayse, G.S., Jinadu, L.A., Shaw, K.A. and Wiley, K.L. (2000) The N-acetylation of arsanilic acid in vitro by mammalian enzymes. Drug Metabolism and Disposition, 28, 487-492. [19] O’Neil, M.J. (2006) The Merck Index: An Encyclopedia of Chemicals, Drugs and Biological. Merck and Company, Whitehouse Station. [20] Harris, G. and Grady, D. (2011) Pfizer suspends sales of chicken drug with arsenic. New York Times, SB, 2. [21] Ashby, J. and Tennant, R.W. (1991) Definitive relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by the US NTP. Mutation Research, 257, 229-306. doi:10.1016/0165-1110(91)90003-E [22] Parkinson, A. (2001) Biotransformation of xenobiotics. In: Klaassen, C.D. Ed., Casarett and Doull’s Toxicology, McGraw Hill, New York, 133-224. [23] Bayse, G.S., Jackson, K.M., Kirlin, W.G. and Rollins-Hairston, A. (2006) Proliferation of human caco-2 cells mediated by N-acetylation and oxidation reactions of 3-amino-4-hydroxybenzenearsonate. Toxicologist, 90, 674. [24] Bayse, G.S., et al. (2007) Permeation of benzenearsonates provides sufficient concentrations to cause Alterations in caco-2 cell proliferation. Toxicologist, 96, 1174. [25] Robinson, D.K., Hammonds-Odie, L.P., Jackson, K.M., Kirlin, W.G. and Bayse, G.S. (2010) Caco-2 cell permeation of five benzenearsonates increases likelihood of hepatic biotransformations. Toxicologist, 114, 1101. [26] Hammonds-Odie, L.P., Bayse, G.S., Jackson, K.M. and Robinson, D.K. (2006) Evaluation of benezenearsonate permeation in caco-2 cells. http://www.fasebj.org/cgi/content/meeting_abstract/20/5/LB111-b?sid=cf22ab74-c1a9-4417-b169-133f4b8aa2ce [27] Artursson, P. (1990) Epithelial transport of drugs in cell culture. I: A model for studying the passive diffusion of drugs over intestinal absorptive (caco-2) cells. Journal of Pharmaceutical Sciences, 79, 476-482. doi:10.1002/jps.2600790604 [28] Hidalgo, I.J., Raub, T.J. and Borchardt, R.T. (1989) Characterization of the human colon carcinoma cell line (ca-co-2) as a model system for intestinal epithelial permeability. Gastroenterology, 96, 736-749. [29] Prueksaritanont, T., Gorham, L.M., Hochman, J.H., Tran, L.O. and Vyas, K.P. (1996) Comparative studies of drugmetabolizing enzymes in dog, monkey, and human small intestines, and in caco-2 cells. Drug Metabolism and Disposition, 24, 634-642. [30] Hayes, J.R. and Campbell, T.C. (1986) Food additives and contaminants. In: Klaassen, C.D., Amdur, M.O. and Doull, J. Eds., Casarett and Doull’s Toxicology, Macmillan, New York, 771-800. [31] Abdo, K.M., et al. (1989) Toxic responses in F344 rats and B6CF1 mice given roxarsone in their diets for up to 13 weeks. Toxicology Letters, 45, 5-66. doi:10.1016/0378-4274(89)90159-8 [32] Ashby, J. and Paton, D. (1993) The influence of chemical structure on the extent and sites of carcinogenesis for 522 rodent carcinogens and 55 different human carcinogen exposures. Mutation Research, 286, 3-74. doi:10.1016/0027-5107(93)90003-X [33] US Environmental Protection Agency (2011) Region 4 should strengthen oversight of Georgia’s concentrated animal feeding operation program. http://www.epa.gov/oig/reports/2011/20110623-11-P-0274.pdf [34] Thompson, C.M., Haws, L.C., Harris, M.A., Gatto, N.M. and Proctor, D.M. (2011) Application of the US EPA mode of action framework for purposes of guiding future research: A case study involving the oral carcinogenicity of hexavalent chromium. Toxicological Sciences, 119, 20-40. doi:10.1093/toxsci/kfq320 [35] Doerge, D.R., Churchwell, M.I., Marques, M.M. and Beland, F.A. (1999) Quantitative analysis of 4-aminobiphenyl-C8-deoxyguanosyl DNA adducts produced in vitro using HPLC-ES-MS. Carcinogenesis, 20, 1055-1061. doi:10.1093/carcin/20.6.1055 [36] Zhang, T., Cao, E. and Qin, J. (2002) Opposite biological effects of arsenic trioxide and arsacetin involve a different regulation of signaling in human gastric cancer MGC-803 cells. Pharmacology, 64, 160-168. doi:10.1159/000056166 [37] Park, W.H., et al. (2000) Arsenic trioxide-mediated growth inhibition in MC/CAR myeloma cells via cell cycle arrest in association with induction of cyclin-dependent kinase inhibitor, p21, and apoptosis. Cancer Research, 60, 3065-3071. [38] Vogt, B.L. and Rossman, T.G. (2001) Effects of arsenite on p53, p21 and cyclin D expression in normal human fibroblasts—A possible mechanism for arsenite’s comutagenicity. Mutation Research, 478, 159-168. doi:10.1016/S0027-5107(01)00137-3 [39] Barr, F.D., Krohmer, L.J., Hamilton, J.W. and Sheldon, L.A. (2009) Disruption of histone modification and CARM1 recruitment by arsenic represses transcription at glucocorticoid receptor-regulated promoters. PLoS ONE, 4, e6766. doi:10.1371/journal.pone.0006766 [40] Davey, J.C., et al. (2008) Arsenic as an endocrine disruptor: Arsenic disrupts retinoic acid receptor-and thyroid hormone receptor-mediated gene regulation and thyroid hormone-mediated amphibian tail metamorphosis. Environmental Health Perspectives, 116, 165-172. [41] Kozul, C.D., et al. (2009) Chronic exposure to arsenic in the drinking water alters the expression of immune response genes in mouse lung. Environmental Health Perspectives, 117, 1108-1115. doi:10.1289/ehp.0800199 [42] Meharg, A.A. and Raab, A. (2010). Getting to the bottom of arsenic standards and guidelines. Environmental Science & Technology, 44, 4395-4399. doi:10.1021/es9034304 [43] Tapio, S. and Grosche, B. (2006) Arsenic in the aetiology of cancer. Mutation Research, 612, 215-246. doi:10.1016/j.mrrev.2006.02.001 [44] Kozul-Horvath, C.D., Zandbergen, F., Jackson, B.P., Enelow, R.I. and Hamilton, J.W. (2012) Effects of lowdose drinking water arsenic on mouse fetal and postnatal growth and development. PLoS ONE, 7, e38249. doi:10.1371/journal.pone.0038249 [45] Price, L.B., et al. (2007) Elevated risk of carrying gentamicin-resistant Escherichia coli among US poultry workers. Environmental Health Perspectives, 115, 1738-1742. doi:10.1289/ehp.10191 [46] Sapkota, A.R., Price, L.B. and Silbergeld, E.K. (2006) Arsenic resistance in Campylobacter spp. isolated from retail poultry products. Applied and Environmental Microbiology, 72, 3069-3071. doi:10.1128/AEM.72.4.3069-3071.2006 [47] Marshall, B.M. and Levy, S.B. (2011) Food animals and antimicrobials: Impacts on human health. Clinical Microbiology Reviews, 2, 718-733. doi:10.1128/CMR.00002-11 [48] US Food and Drug Administration (2012) 21 CFR part 558 subpart B-specific new animal drugs for use in animal feeds. Electronic Code of Federal Regulations. http://www.gpo.gov/fdsys/pkg/CFR-2012-title21-vol6/pdf/CFR-2012-title21-vol6-sec558-530.pdf