Abstract
Benzene is a well-known environmental pollutant that can induce hematotoxicity, aplastic anemia, acute myelogenous leukemia, and lymphoma. However, although benzene metabolites are known to induce oxidative stress and disrupt the cell cycle, the mechanism underlying lympho/leukemogenicity is not fully understood. Caspase-4 (alias caspase-11) and -12 are inflammatory caspases implicated in inflammation and endoplasmic reticulum stress-induced apoptosis. The objectives of this study were to investigate the altered expression of caspase-4 and -12 in mouse bone marrow after benzene exposure and to determine whether their alterations are associated with benzene-induced bone marrow toxicity, especially cellular apoptosis. In addition, we evaluated whether the p53 gene is involved in regulating the mechanism, using both wild-type (WT) mice and mice lacking the p53 gene. For this study, 8-week-old C57BL/6 mice [WT and p53 knockout (KO)] were administered a benzene solution (150 mg/kg diluted in corn oil) via oral gavage once daily, 5 days/week, for 1 or 2 weeks. Blood and bone marrow cells were collected and cell counts were measured using a Coulter counter. Total mRNA and protein extracts were prepared from the harvested bone marrow cells. Then qRT-PCR and Western blotting were performed to detect changes in the caspases at the mRNA and protein level, respectively. A DNA fragmentation assay and Annexin-V staining were carried out on the bone marrow cells to detect apoptosis. Results indicated that when compared to the control, leukocyte number and bone marrow cellularity decreased significantly in WT mice. The expression of caspase-4 and -12 mRNA increased significantly after 12 days of benzene treatment in the bone marrow cells of benzene-exposed p53KO mice. However, apoptosis detection assays indicated no evidence of apoptosis in p53KO or WT mice. In addition, no changes of other apoptosis-related caspases, such as caspase-3 and -9, were found in WT or p53KO mice at the level of mRNA and proteins. These results indicated that upregulation of caspase-4 and -12 in mice lacking the p53 gene is not associated with cellular apoptosis. In conclusion, caspase-4 and -12 can be activated by benzene treatment without inducing cell apoptosis in mouse bone marrow, which are partly under the regulation of the p53 gene.
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References
Brief RS, Lynch J, Bernath T, Scala RA (1980) Benzene in the workplace. Am Ind Hyg Assoc J 41:616–623
Choi YJ, Mendoza L, Rha SJ, Sheikh-Hamad D, Baranowska-Daca E, Nguyen V, Smith CW, Nassar G, Suki WN, Truong LD (2001) Role of p53-dependent activation of caspases in chronic obstructive uropathy: evidence from p53 null mutant mice. J Am Soc Nephrol 12:983–992
Cronkite EP (1986) Benzene hematotoxicity and leukemogenesis. Blood Cells 12:129–137
Danial NN, Korsmeyer SJ (2004) Cell death: critical control points. Cell 116:205–219
Fábián Z, Csatary CM, Szeberényi J, Csatary LK (2007) p53-independent endoplasmic reticulum stress-mediated cytotoxicity of a Newcastle disease virus strain in tumor cell lines. J Virol 81:2817–2830
Faiola B, Fuller ES, Wong VA, Recio WL (2004) Gene expression profile in bone marrow and hematopoietic stem cells in mice exposed to inhaled benzene. Mut Res 549:195–212
Fan TJ, Han LH, Cong RS, Liang J (2005) Caspase family proteases and apoptosis. Acta Biochim Biophys Sin 37:719–727
Farris GM, Robinson SN, Gaido KW, Wong BA, Wona VA, Hahn WP, Shah RS (1997) Benzene-induced hematotoxicity and bone marrow compensation in B6C3F1 mice. Fundam Appl Toxicol 36:119–129
Hazel BA, Baum C, Kalf GF (1996) Hydroquinone, a bioreactive metabolite benzene, inhibits apoptosis in myeloblasts. Stem Cells 14:730–742
Hirabayashi Y, Matsuda M, Aizawa S, Kodama Y, Kanno J, Inoue T (2002) Serial transplantation of p53-deficient hemopoietic progenitor cells to assess their infinite growth potential. Exp Biol Med (Maywood) 227:474–479
Hiraku Y, Kawanishi S (1996) Oxidative DNA damage and apoptosis induced by benzene metabolites. Cancer Res 56:5172–5178
Hisahara S, Yuhan J, Momoi T, Okano H, Miura M (2001) Caspase-11 mediates oligodendrocyte cell death and pathogenesis of autoimmune-mediated demyelination. J Exp Med 193:111–122
Huff JE, Haseman JK, Demarini DM, Eustis S, Moaronpot RR, Peters AC (1989) Multiple-site carcinogenicity of benzene in Fischer 344 rats and B5C3F1 mice. Environ Health Perspect 83:125–163
Ibuki Y, Goto R (2004) Dysregulation of apoptosis by benzene metabolites and their relationship with carcinogenesis. Biochim Biophys Acta 1690:11–21
Inayat-Hussain SH, Ross D (2005) Intrinsic pathway of hydroquinone induced apoptosis occurs via both caspase-dependent and caspase-independent mechanisms. Chem Res Toxicol 18:420–427
Kang SJ, Wang S, Hara H, Peterson EP, Namura S, Amin-Hanjani S, Huang Z, Srinivasan A, Tomaselli KJ, Thornberry NA, Moskowitz MA, Yuan J (2000) Dual role of caspase-11 in mediating activation of caspase-1 and caspase-3 under pathological conditions. J Cell Biol 149:613–622
Kuo M, Shiah S, Wang C, Chuang S (1999) Suppression of apoptosis by bcl-2 to enhance benzene metabolites-induced oxidative DNA damage and mutagenesis: a possible mechanism of carcinogenesis. Mol Pharmacol 55:894–901
Lamkanfi M, Festjens N, Declercq W, Berghe TV, Vandenabeele P (2007) Caspases in cell survival, proliferation and differentiation. Cell Death Differ 14:44–55
Li J, Lee B, Lee AS (2006) Endoplasmic reticulum stress-induced apoptosis. J Biol Chem 281:7260–7270
Lynge E, Andersen A, Nilsson R, Barlow L, Pukkala E, Nordlinder R, Boffetta P, Grandjean P, Heikkila P, Horte LG, Jakobsson R, Lundberg I, Moen B, Partanen T, Riise T (1997) Risk of cancer and exposure to gasoline vapors. Am J Epidemiol 145:449–458
Maltoni C, Ciliberti A, Cotti G, Conti B, Belpoggi F (1989) Benzene, an experimental multipotential carcinogen: results of the long-term bioassays performed at the Bologna Institute of Oncology. Environ Health Perspect 82:109–124
Martinez-Velazquez M, Maldonado V, Ortega A, Melendez-zajgla J, Albores A (2006) Benzene metabolites induce apoptosis in lymphocytes. Exp Toxicol Pathol 58:65–70
Martinon F, Tschopp J (2007) Inflammatory caspases and inflammasomes: master switches of inflammation. Cell Death Differ 14:10–22
Mouse Genome Informatics (MGI) (2009) Trp53tm1Sia targeted allele detail, ID: MGI:1926340. http://www.informatics.jax.org/javawi2/servlet/WIFetch?page=alleleDetail&key=2871
NRC (National Research Counsil) (1996) Guide for the care and use of laboratory animals. National Academy Press, Washington DC
Rana SV, Verma Y (2005) Biochemical toxicity of benzene. J Environ Biol 26:157–168
Rao NR, Snyder R (1995) Oxidative modifications produced in HL-60 cells on exposure to benzene metabolites. Appl Toxicol 15:403–409
Rinsky RA, Young RK, Smith AB (1981) Leukemia in benzene workers. Am J Ind Med 2:217–245
Ross D (2000) The role of metabolism and specific metabolites in benzene-induced toxicity: evidence and issues. J Toxicol Environ Health A 61:357–372
Sabourin PJ, Chen BT, Lucier G, Birnbaum LS, Fisher E, Henderson RF (1987) Effect of dose on the absorption and excretion of [14C] benzene administration orally or by inhalation in rats and mice. Toxicol Appl Pharmacol 87:325–336
Scott AM, Saleh M (2007) The inflammatory caspases: guardians against infections and sepsis. Cell Death Differ 14:23–31
Snyder CA, Goldstein BD, Sellakumar AR, Bromberg E, Laskin S, Albert RE (1980) The inhibition toxicology of benzene: incidence of hematopoietic neoplasms and hematotoxicity in AKR/J and C57BL/6J mice. Toxicol Appl Pharmacol 54:323–331
Snyder R, Hedli CC (1996) An overview of benzene metabolism. Environ Health Perspect 104(suppl 6):1165–1171
Snyder CA, Sellakumar AR, James DJ, Albert RE (1988) The Carcinogenicity of discontinuous inhaled benzene exposures in CD-1 and C57BL/6 mice. Arch Toxicol 62:331–335
Suk K, Kim SY, Kim H (2002) Essential role of caspase-11 in activation-induced cell death of rat astrocytes. J Neurochem 80:230–238
Tsukada T, Tomooka Y, Takai S, Ueda Y, Nishikawa S, Yagi T, Tokunaga T, Takeda N, Suda Y, Abe S, Matsuso I, Ikawa Y, Aizawa S (1993) Enhanced proliferative potential in culture of cells from p53-defecient mice. Oncogene 8:3313–3322
Wallace L (1996) Environmental exposure to benzene. Environ Health Perspect 104:1129–1136
Yagi T, Tokunaga T, Furuta Y, Nada S, Yoshida M, Tsukada T, Saga Y, Takeda N, Ikawa Y, Aizawa S (1993) A novel ES cell line, TT2, with high germline-differentiating potency. Anal Biochem 214:70–76
Yi JY, Yoon BI (2008) Benzene-induced hematotoxicity and myelotoxicity by short-term repeated oral administration in mice. Lab Anim Res 24:99–103
Yoon BI, Hirabayashi Y, Kawasaki Y, Kodama Y, Kaneko T, Kim DY, Inoue T (2001) Mechanism of action of benzene toxicity: cell cycle suppression in hemopoietic progenitor cells (CFU-GM). Exp Hematol 29:278–285
Yoon BI, Li GX, Kitada K, Kawasaki Y, Igarashi K, Kodama Y, Inoue T, Kobayashi K, Kanno J, Kim DY, Inoue T, Hirabayashi Y (2003) Mechanism of benzene-induced hematotoxicity and leukemogenicity: cDNA microarry analyses using mouse bone marrow tissue. Environ Health Perspect 111:1411–1420
Yoshida K, Aizawa S, Watanabe K, Hirabayashi Y, Inoue T (2002) Stem-cell leukemia: p53 deficiency mediated suppression of leukemic differentiation in C3H/He myeloid leukemia. Leuk Res 26:1085–1092
Yu K, Yang KY, Ren XZ, Chen Y, Liu XH (2007) Amifostine protects bone marrow from benzene-induced hematotoxicity in mice. Int J Toxicol 26:315–323
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This work was supported by the Korea Research Foundation Grant funded by the Korean Government (MOEHRD) (KRF-2005-042-E00154).
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J.-Y. Yi and Y. Hirabayashi contributed equally for this study.
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Yi, JY., Hirabayashi, Y., Choi, YK. et al. Benzene activates caspase-4 and -12 at the transcription level, without an association with apoptosis, in mouse bone marrow cells lacking the p53 gene. Arch Toxicol 83, 795–803 (2009). https://doi.org/10.1007/s00204-009-0420-4
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DOI: https://doi.org/10.1007/s00204-009-0420-4