TY - JOUR
T1 - Comparative covalent protein binding of 2,5-Hexanedione and 3-Acetyl-2,5-Hexanedione in the rat
AU - DeCaprio, Anthony P.
AU - Kinney, Elizabeth A.
AU - Lopachin, Richard M.
N1 - Funding Information:
Received 4 December 2008; accepted 12 February 2009. This work was supported by NIH ES-005172 (to APD) and ES-007912 (to RML). Address correspondence to Anthony P. DeCaprio, PhD, Department of Chemistry and Biochemistry, Florida International University, CP338B, 11200 S.W. 8th St., Miami, FL 33199, USA. E-mail: adecapr@fiu.edu
PY - 2009
Y1 - 2009
N2 - 2,5-Hexanedione (HD) is the metabolite implicated in n-hexane neurotoxicity. This γ-diketone reacts with protein lysine amines to form 2,5-dimethylpyrrole adducts. Pyrrole adduction of neurofilaments (NF) and/or other axonal proteins was proposed as a critical step in the neuropathy. While pyrrole adduction is widely accepted as necessary, subsequent pyrrole oxidation, which may result in protein cross-linking, was alternatively postulated as the critical mechanistic step. Previous studies have indicated that 3- acetyl-2,5-HD (AcHD), an analogue that forms pyrroles that do not oxidize, was not neurotoxic in rats. However, relative levels of pyrrole adduction of NF or other axonal proteins were not reported. In the present study, groups of 6 male Wistar rats were given saline, [1,6-14C]-HD (3 mmol/kg/d), or [5- 14C]-AcHD (0.1 mmol/kg/d), i.p. for 21 d. HD- and AcHD-treated rats lost 10% and gained 14% body weight, respectively, compared to a 22% gain for control rats. At termination, HD- and AcHD-treated rats exhibited mean scores of 3.5 and 1.4, respectively, for hindlimb weakness (0-5 scale). Incorporation of radiolabel from HD was 27.8 ± 3.9, 13.9 ± 2.6, and 7.8 ± 0.6 nmol/mg in plasma protein, purified globin, and axonal cytoskeletal proteins, respectively, compared to 0.6 ± 0.1, 1.6 ± 0.5, and 1.0 ± 0.1 for AcHD. Binding of HD to the NF-L, -M, and -H subunit proteins from treated animals was 4-, 24-, and 13-fold higher, respectively, that that of AcHD, indicating differing stoichiometry and patterns of NF adduction for the two diketones. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of globin and NF proteins did not demonstrate protein cross-linking for either diketone at the dose levels and time period examined. These results indicate that that the lack of neurotoxicity previously reported for AcHD may reflect differences in adduct levels at critical axonal target sites rather than an inability to form cross-linking adducts. Based on these data, further studies are required to fully assess the neurotoxic potency of AcHD and other non-cross-linking analogues as compared to HD.
AB - 2,5-Hexanedione (HD) is the metabolite implicated in n-hexane neurotoxicity. This γ-diketone reacts with protein lysine amines to form 2,5-dimethylpyrrole adducts. Pyrrole adduction of neurofilaments (NF) and/or other axonal proteins was proposed as a critical step in the neuropathy. While pyrrole adduction is widely accepted as necessary, subsequent pyrrole oxidation, which may result in protein cross-linking, was alternatively postulated as the critical mechanistic step. Previous studies have indicated that 3- acetyl-2,5-HD (AcHD), an analogue that forms pyrroles that do not oxidize, was not neurotoxic in rats. However, relative levels of pyrrole adduction of NF or other axonal proteins were not reported. In the present study, groups of 6 male Wistar rats were given saline, [1,6-14C]-HD (3 mmol/kg/d), or [5- 14C]-AcHD (0.1 mmol/kg/d), i.p. for 21 d. HD- and AcHD-treated rats lost 10% and gained 14% body weight, respectively, compared to a 22% gain for control rats. At termination, HD- and AcHD-treated rats exhibited mean scores of 3.5 and 1.4, respectively, for hindlimb weakness (0-5 scale). Incorporation of radiolabel from HD was 27.8 ± 3.9, 13.9 ± 2.6, and 7.8 ± 0.6 nmol/mg in plasma protein, purified globin, and axonal cytoskeletal proteins, respectively, compared to 0.6 ± 0.1, 1.6 ± 0.5, and 1.0 ± 0.1 for AcHD. Binding of HD to the NF-L, -M, and -H subunit proteins from treated animals was 4-, 24-, and 13-fold higher, respectively, that that of AcHD, indicating differing stoichiometry and patterns of NF adduction for the two diketones. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of globin and NF proteins did not demonstrate protein cross-linking for either diketone at the dose levels and time period examined. These results indicate that that the lack of neurotoxicity previously reported for AcHD may reflect differences in adduct levels at critical axonal target sites rather than an inability to form cross-linking adducts. Based on these data, further studies are required to fully assess the neurotoxic potency of AcHD and other non-cross-linking analogues as compared to HD.
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U2 - 10.1080/15287390902959508
DO - 10.1080/15287390902959508
M3 - Article
C2 - 19557614
AN - SCOPUS:67650287665
SN - 1528-7394
VL - 72
SP - 861
EP - 869
JO - Journal of Toxicology and Environmental Health - Part A: Current Issues
JF - Journal of Toxicology and Environmental Health - Part A: Current Issues
IS - 14
ER -