Immunohistochemical Detection of Polyunsaturated Fatty Acid Oxidation Markers in Acetaminophen-Induced Liver Injury in Rats

Abstract

ABSTRACT. To clarfity whether polyunsaturated fatty acid (PUFA) oxidation is involved in the mechanism of acetaminophen (APAP)-induced apoptotic cell death, the production and localization of PUFA oxidation markers N  -propanoyl-modified lysine, N  -hexanoylmodified lysine, 4-hydroxyhexenal-modified histidine and crotonaldehyde-modified lysine were evaluated in the development of APAPinduced liver injury. The immunoexpression of these markers in the liver was examined up to 24 hr post-APAP intraperitoneal injection in rats (1 g/kg body weight). The histopathological changes in the liver appeared 3 hr after APAP injection and became exacerbated with time. Proapoptotic protein Bax immunoreactivity was first detected in the degenerative hepatocytes 3 hr after the injection and areas positively immunostained for Bax reached a peak level at 6 hr, and then decreased at 12 and 24 hr. There was a significant increase in the TUNEL-positive rate at 12 and 24 hr. Immunohistological expression of all these oxidation markers was first detected in the degenerative hepatocytes 3 hr after the injection, and earlier than the occurrence of hepatocyte apoptosis. Immunohistochemical expression of these markers were observed in almost all degenerative hepatocytes 3-24 hr after APAP injection. Areas positively immunostained for these markers reached a peak level at 6 hr, and then decreased at 12 and 24 hr. The results thus suggest that the generation of PUFA oxidation markers may be the signature of early events preceding the induction of liver cell apoptosis and thus useful for early detection of oxidative stress-related liver cell injury.

Response to the Comments on Caspase-Mediated Anti-Apoptotic Effect of Ginsenoside Rg5, a Main Rare Ginsenoside, on Acetaminophen-Induced Hepatotoxicity in Mice

“…In fact, secondary antibodies incubation under the same conditions without first antibodies and for the same incubation period in negative controls did not show any staining of liver tissues in our experiment. The results from two recent papers by Sun et al and Zhang et al are in line with us that APAP causes pro-apoptotic mitochondrial Bax translocation. , Similar to our findings, there is no protein expression of Bax in controls evidenced by IHC analysis.…”

Lipid Hydroperoxide-Derived Modification of Biomolecules

“…Another alkylamide-type adduct, PRL, has also been detected in rabbit atherosclerotic plaques ), the liver of an aminoacetophenone-injected rat (Sun et al 2012), and the hippocampus region of the aged rat brain (Hisaka et al, manuscript in preparation) by immunological methods. In particular, HEL is described as one biomarker for oxidative stress (see Sect.…”

Immunochemical Detection of Lipid Hydroperoxide- and Aldehyde-Modified Proteins in Diseases

“…In the previous study (Sun et al 2012), the immunoexpression of PUFA oxidation markers PRL, HEL, 4-HHE-histidine and CRA-lysine was examined up to 24 h post-APAP intraperitoneal injection in rats (1 g/kg body weight). Three hr after the postintraperitoneal injection of APAP in rats, vacuolated hepatocytes were observed in the centrilobular region of the hepatic lobule (Fig.…”