Abstract
The aromatic hydrocarbon 1,2-diacetylbenzene (1,2-DAB) is a protein-reactive γ-diketone metabolite of the neurotoxic solvent 1,2-diethylbenzene (1,2-DEB). The effect of neurotoxic 1,2-DAB and its non-neurotoxic isomer 1,3-DAB has been studied on motor proteins and cytoskeletal proteins of rat spinal cord (SC). For in vitro studies, SC slices were incubated with 1, 2, 5, 10 mM of DAB isomers for 30 min at 37°C. For in vivo studies, rats received (i.p.) 20 mg/kg/day of 1,2-DAB or 1,3-DAB, or vehicle (2% acetone in saline), 5 days a week for 2 weeks. Spinal cord and sciatic nerve proteins were subjected to Western blotting using monoclonal mouse antibodies to NF-M, kinesin, dynein, and tau. Proteins were quantified and paired mean comparisons performed to assess concentration-dependent changes in native protein bands. In vitro, 1,2-DAB produced a concentration-dependent decrease of motor and cytoskeletal proteins. While dynein and tau appeared similarly affected by 1,2-DAB, kinesin was most affected by the toxicant. In vivo, 1,2-DAB affected motor and cytoskeletal proteins of sciatic nerves and spinal cord differentially. In general, sciatic nerve proteins were much more affected than spinal cord proteins. The results show that motor proteins that drive axonal transport anterogradely (kinesin) and retrogradely (dynein), cytoskeletal protein NF-M, which is slowly transported in the anterograde direction, and microtubule-associated protein, tau, which is involved in axonal transport, are differentially impacted by 1,2-DAB. By contrast, non-neurotoxic isomer 1,3-diacetylbenzene (1,3-DAB), had no adverse effect on neural proteins either in vitro or in vivo. 2D-Differential in gel electrophoresis (2D-DIGE) of sciatic nerves from neurotoxic 1,2-DAB and non-neurotoxic 1,3-DAB treated rats revealed 197 and 304 protein spots, respectively.
Similar content being viewed by others
References
Anthony DC, Boekelheide K, Anderson CW, Graham DG (1983a) The effect of 3,4-dimethyl substitution on the neurotoxicity of 2,5-hexanedione. II. Dimethyl substitution accelerates pyrrole formation and protein crosslinking. Toxicol Appl Pharmacol 71:372–382
Anthony DC, Giangaspero F, Graham DG (1983b) The spatio-temporal pattern of the axonopathy associated with the neurotoxicity of 3,4-dimethyl-2,5-hexanedione in the rat. J Neuropathol Exp Neurol 42:548–560
Cavender F (1994) Aromatic hydrocarbons. In: Clayton DE, Clayton FE (eds) Patty’s industrial hygiene and toxicology, 4th edn. Wiley, New York, vol IIB, p 1301
DeCaprio AP (2000) n-Hexane, metabolites and derivatives. In: Spencer PS, Schaumburg HH (eds) Experimental and clinical neurotoxicology. Oxford, New York, NY, pp 633–648
deSousa BN, Horrocks LA (1979) Development of rat spinal cord. I. Weight and length, with a method for rapid removal. Dev Neurosci 2:115–121
Gagnaire F, Becker MN, De Ceaurriz J (1992a) Alteration of brainstem auditory evoked potentials in diethylbenzene- and diacetylbenzene-treated rats. J Appl Toxicol 12:343–350
Gagnaire F, Becker MN, Marignac B, Bonnet P, De Ceaurriz J (1992b) Diethylbenzene inhalation-induced electrophysiological deficits in peripheral nerves and changes in brainstem auditory evoked potentials in rats. J Appl Toxicol 12:335–342
Gagnaire F, Ensminger A, Marignac B, De Ceaurriz J (1991) Possible involvement of 1,2-diacetylbenzene in diethylbenzene-induced neuropathy in rats. J Appl Toxicol 11:261–268
Gagnaire F, Marignac B, de Ceaurriz J (1990) Diethylbenzene-induced sensorimotor neuropathy in rats. J Appl Toxicol 10:105–112
Graham DG (1995) Pathogenetic studies of hexane and carbon disulfide neurotoxicity. CRC Crit Rev Toxicol 25:91–112
Graham DG (1999) Neurotoxicants and the cytoskeleton. Curr Opin Neurol 12:733–737
Graham DG, Szakal-Quin G, Priest JF, Anthony DC (1984) In vitro evidence that covalent crosslinking of neurofilaments occurs in gamma-diketone neuropathy. Proc Natl Acad Sci USA 81:4979–4982
Hirokawa N, Takemura R (2003) Biochemical and molecular characterization of diseases linked to motor proteins. Trends Biochem Sci 28:558–565
Hoffman PN, Lasek RJ (1975) The slow component of axonal transport: identification of major structural polypeptides of the axon and their generality among mammalian neurons. J Cell Biol 66:351–366
Holm S (1979) A simple sequentially rejective multiple test procedure. Scand J Stat 6:65–70
Kim MS, Hashemi SB, Spencer PS, Sabri MI (2002) Amino acid and protein targets of 1,2-diacetylbenzene, a potent aromatic γ-diketone that induces proximal neurofilamentous axonopathy. Toxicol Appl Pharmacol 183:55–65
Kim MS, Kayton RJ, Muniz J, Austin DR, Spencer PS, Sabri MI (1999) 1,2-diacetylbenzene neurotoxicity: a model to study the role of Schwann cells in maintenance of axonal integrity in toxic states. Microsc Microanal 5:1218–1219
Kim MS, Sabri MI, Miller VH, Kayton RJ, Dixon DA, Spencer PS (2001) 1,2-diacetylbenzene, the neurotoxic metabolite of a chromogenic aromatic organic solvent, induces proximal axonopathy. Toxicol Appl Pharmacol 177:121–131
Krapfenbauer K, Fountoulakis M, Lubec G (2003) A rat brain protein expression map including cytosolic and enriched mitochondrial and microsomal fractions. Electrophoresis 24:1847–1870
Lehning EJ, Dyer KS, Jortner BS, LoPachin RM (1995) Axonal atrophy is a specific component of 2,5-hexanedione peripheral neuropathy. Toxicol Appl Pharmacol 135:58–66
Lehning EJ, Jortner BS, Fox JH, Arezzo JC, Kitano T, LoPachin RM (2000) γ-diketone peripheral neuropathy. I. Quantitative morphometric analyses of axonal atrophy and swelling. Toxicol Appl Pharmacol 165:127–140
LoPachin RM (2000) Redefining toxic distal axonopathies. Toxicol Lett 112–113: 23–33
LoPachin RM, Lehning EJ (1977) The relevance of axonal swelling and atrophy to γ-diketone neurotoxicity. Neurotoxicology 18:7–28
LoPachin RM, Jortner BS, Reid ML, Das S (2003) γ-diketone central neuropathy: quantitative morphometric analysis of axons in rat spinal cord white matter regions and nerve roots. Toxicol Appl Pharmacol 193:29–46
Miller RG (1986) Beyond ANOVA: basics of applied statistics. Chapman & Hall, CRC, New York
Monaco S, Wongmongkolrit T, Shearson CM, Patton A, Schaetzle B, Autilio-Gambett L, Gambetti P, Sayre LM (1990) Giant axonopathy characterized by intermediate location of axonal enlargements and acceleration of neurofilament transport. Brain Res 519:73–81
Payan SJ, Beydon D, Cossec B, Ensminger A, Fabry JP, Ferrari E (1999) Toxicokinetics of 1,2-diethylbenzene in male Sprague-Dawley rats-part 1: excretion and metabolism of [(14)C]1,2-diethylbenzene. Drug Metab Dispos 27:1470–1478
Politis MJ, Pellegrino RG, Spencer PS (1980) Ultrastructural studies of the dying-back process. V. Axonal neurofilaments accumulate at sites of 2,5-hexanedione application: evidence for nerve fibre dysfunction in experimental hexacarbon neuropathy. J Neurocytol 9:505–516
R Development Core Team (2005) R: a language and environment for statistical computing. Vienna, Austria, R Foundation for Statistical Computing, http//www.R-project.org
Sabri MI (1992) Effect of 2,5-hexanedione and 3.4-dimethyl-2,5-hexanedione on retrograde axonal transport in sciatic nerve. Neurochem Res 17:835–839
Sabri MI, Hashemi S, Chohan S, Khalil S, Cranson AB, Tshala-Katumbay DD, Palmer VS, Pounds JG, Spencer PS (2005) A role for stathmin in nerve and testes damage? Soc Toxicol Ann Meet 44:168–168
Sabri MI, Hashemi SB, Spencer PS (2003) Neurotoxicity and reactivity of 1,2-diacetylbenzene (1,2-DAB) with motor and cytoskeletal proteins in rat spinal cord and sciatic nerves. Toxicol Lett 144:s130–s130
Sabri M, Spencer P, Testino A, Sickles D (2006) Selective disruption of kinesin-based microtubule motility by neurotoxic 1,2-diacetylbenzene. Soc. Toxicol, 45th Annual Meeting, Abs.# 2426
Sickles DW (1989) Toxic neurofilamentous axonopathies and anterograde axonal transport. II. The effects of single doses of neurotoxic and non-neurotoxic diketones and β-β′-iminodipropionitrile iminodipropionitrile (IDPN) on the rate and capacity of transport. Neurotoxicology 10:103–111
Spencer PS (2000) Diethylbenzene. In: Spencer PS, Schaumburg HH (eds) Experimental and clinical neurotoxicology, 2nd edn. Oxford, New York, NY, pp 490–492
Spencer PS, Schaumburg HH (1977) Ultrastructural studies of the dying-back process. III. The evolution of experimental peripheral giant axonal degeneration. J Neuropathol Exp Neurol 36:276–299
Thrall KD, Gies RA, Cartmell AM, Wu H, Woodstock AD, Klein JA (2007) The distribution kinetics of 1,2-diethylbenzene and its metabolite, 1,2-diacetylbenzene in the F344 male rat. J Toxicol Environ Health A 70:67–72
Trimpin S, Hsu VL, Spencer PS, Deinzer ML (2006) Time-dependent 2,5-hexanedione-adduction of lysine under physiological conditions: Structure determination, chemical mechanisms and biological implications of isoindolium salt. Org Biomol Chem (under revision)
Tshala-Katumbay DD, Palmer VS, Kayton RJ, Sabri MI, Spencer PS (2005) A new murine model of giant proximal axonopathy. Acta Neuropathol (Berl) 109:405–410
Zagoren JC, Politis MJ, Spencer PS (1983) Rapid reorganization of the axonal cytoskeleton induced by a gamma diketone. Brain Res 270:162–164
Zhan CG, Dixon DA, Sabri MI, Kim MS, Spencer PS (2002) Theoretical determination of chromophores in the chromogenic effects of aromatic neurotoxicants. J Am Chem Soc 124:2744–2752
Zhan CG, Spencer PS, Dixon DA (2003) Computational insights into the chemical structures and mechanisms of the chromomeric and neurotoxin effects of aromatic γ-diketones. J Phys Chem 107:2853–2861
Zhu M, Spink DC, Bank S, Chen X, Decaprio AP (1993) Analysis of alkylpyrrole autoxidation products by high-performance liquid chromatography with thermospray mass spectrometry and UV photodiode-array detection. J Chromatogr 628:37–47
Zhu M, Spink DC, Yan B, Bank S, Decaprio AP (1995) Inhibition of 2,5-hexanedione-induced protein crosslinking by biological thiols: chemical mechanisms and toxicological implications. Chem Res Toxicol 8:764–771
Acknowledgments
We thank Dan Austin for assistance with graphics and Juan Muniz for GC-MS analysis of DAB isomers. This publication was made possible by Grants ES10338 and ES 11384 from the National Institute for Environmental Health Sciences (NIEHS), National Institute of Health (NIH). The contents of this paper are solely the responsibility of the authors and do not necessarily represent the official view of the NIEHS, NIH.
Author information
Authors and Affiliations
Corresponding author
Additional information
This paper is dedicated to my long-time friend Naren L. Banik, Ph.D.
Rights and permissions
About this article
Cite this article
Sabri, M.I., Hashemi, S.B., Lasarev, M.R. et al. Axonopathy-Inducing 1,2-Diacetylbenzene Forms Adducts with Motor and Cytoskeletal Proteins Required for Axonal Transport. Neurochem Res 32, 2152–2159 (2007). https://doi.org/10.1007/s11064-007-9392-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11064-007-9392-z