4‐Hydroxy‐2‐nonenal‐ Modified Proteins in Bone Marrow of Phenol/Hydroquinone Treated Rats: Implications for Benzene‐Mediated Hematotoxicity

Abstract

Benzene exposure causes myelotoxicity and leukemia. Phenol and hydroquinone (PHE/HQ; 1.1/0.9 mmol/kg, ip, 19h), metabolites of benzene, significantly decrease lymphocyte counts (58–89%) in rats. HQ readily oxidizes to 1,4‐BQ and conjugates with GSH, and multi‐substituted HQ‐GSHs were detected in bone marrow after PHE/HQ. HQ‐GSHs are more efficient generators of superoxide anion than HQ/1,4‐BQ, and thus retain the ability to arylate proteins and to redox cycle. However, the precise molecular mechanism(s) by which benzene/HQ‐GSH conjugates induce hematotoxicity is unknown. Using anti‐4‐hydroxy‐2‐nonenal (4HNE) specific antibodies on 2D western, and MS/MS peptide sequencing analysis of immunopositive proteins isolated from bone marrow lysate of PHE/HQ treated rats, we revealed the presence of 9 4HNE‐adducted proteins. In particular, protein disulfide isomerase (PDI) and peroxiredoxin 2 (PRDX2) were immunopositive for 4HNE adduction. In models of alcoholic liver disease, both PDI and PRDX6 have reduced activity when adducted by 4HNE. Moreover, a 4HNE adduction site was identified on calreticulin, on a critical cysteine residue involved in its binding to misfolded proteins. Analysis of the structural and functional significance of these protein modifications may provide new insight into the progression of benzene‐induced leukemia. (P30ES006694, RO1GM70890, Sci. Found. of AZ, T32ES007091)