Immunochemical comparison of 3'-hydroxyacetanilide and acetaminophen binding in mouse liver.
The hepatotoxicity of the analgesic acetaminophen is believed to be mediated by covalent binding to critical proteins. Radiolabeled 3'-hydroxyacetanilide, a regioisomer of acetaminophen, covalently binds to proteins at levels similar to those of acetaminophen, but without toxicity. Covalent binding has recently been detected by Western blot to a 50-kDa microsomal protein that comigrated with CYP2E1 and was accompanied by a loss of the CYP2E1 activity. However, radiolabel studies previously indicated that a significant amount of the radiolabel is lost during electrophoresis. In the present study, 3'-hydroxyacetanilide covalent binding was detected immunohistochemically in liver using an anti-acetaminophen antiserum. 3'-Hydroxyacetanilide (1000 mg/kg, ip) administration to mice resulted in panlobular immunostaining in liver, with the single layer of hepatocytes surrounding the central veins having the greatest intensity of staining. Staining was most intense at 1 hr and somewhat decreased at 3 and 6 hr. In contrast, immunochemical staining indicated that covalent binding of acetaminophen (250 mg/kg, ip) was confined to the centrilobular hepatocytes, the area of the ensuing necrosis. Cobaltous chloride pretreatment decreased the total intensity of the panlobular immunostaining following 3'-hydroxyacetanilide. The CYP2E1 inhibitor diallyl sulfide decreased the intensity of immunostaining in the central vein area only. Western blot analysis indicated diallyl sulfide also eliminated binding to the microsomal 50-kDa protein. These data are consistent with centrilobular binding of 3'-hydroxyacetanilide, mediated in part by CYP2E1, and panlobular binding, mediated by other P450 enzymes.
Salminen
W F
WF
Department of Pharmacology & Therapeutics, J. Hillis Miller Health Science Center, University of Florida, Gainesville, FL 32611, USA.
Roberts
S M
SM
Pumford
N R
NR
Hinson
J A
JA
D000082:Acetaminophen / Q000378:metabolism*; D000083:Acetanilides / Q000378:metabolism* / Q000633:toxicity; D000498:Allyl Compounds; D000818:Animals; D003035:Cobalt / Q000494:pharmacology; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D007150:Immunohistochemistry; D008099:Liver / Q000166:cytology / Q000502:physiology*; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D008861:Microsomes / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D013440:Sulfides / Q000494:pharmacology
D000083:Acetanilides; D000498:Allyl Compounds; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D013440:Sulfides; D000082:Acetaminophen; D003035:Cobalt; C038491:allyl sulfide; C026137:3-hydroxyacetanilide; C018021:cobaltous chloride