SOFT 2004 Meeting Abstracts - Society of Forensic Toxicologists

C17
CHANGES OF GENE EXPRESSION BEFORE AND AFTER ACETAMINOPHEN INJECTION
DETECTED BY DIFFERENTIAL DISPLAY
2
Mizuho Namiki I, Tomohisa Mori 2 , Shinobu It0 , Toshiko Sawaguche
I Dept.of Emergency Medicine,Tokyo Women's Medical University,Tokyo,Japan
2Dept.ofLegal Medicine,Tokyo Women's Medical University,Tokyo,Japan
Objectives; Acetaminophen is a familiar antipyretic analgesic. While the mechanism by which
acetaminophen causes chemical damage has been fully explained, the molecular biological dynamics that
takes place in the damaged liver has not been elucidated. The current study was conducted to advance the
molecular biological understanding in the dynamics of the gene expression during the subsequent hepatic
damage by acetaminophen.
Materials and Method: I)Preparation of mice with acetaminophen-induced hepatic damage. Following
fasting for 24 hours, 250 mglkg body weight of an acetaminophen solution was administered via a
peritoneal route to mice to serve as a model for a drug-induced hepatic disorder, while the same amount of
a physiological saline was also given via the peritoneal injection to the control animals. Both groups fasted
for an additional 24 hours thereafter. Under chloroform anesthesia, laparotomies were performed.
Following blood specimen collection from the heart, the liver was excised. 2) Serum AL T and AST
determinations were measured by using an autoanalyzer (Hitachi-7450). 3) The excised livers were frozen
with liquid nitrogen and pulverized, from which RNA was collected by using Trizol. The product was then
treated with DNAse and processed with phenol chloroform for cDNA synthesis. The DNA was amplified
by using a primer from the Fluorescence Differential Display Kit (Takara). 4) The second PCR and
acrylamide gel electrophoresis were conducted by employing the fluorescence differential display method.
After a band pattern was visualized by a molecular imager FX, the bands with eminent differences were cut
out, from which the amplified products were recovered for cloning. 5) The homology was confirmed
between each base sequence that had been obtained from the Web site NCBI BRAST and the known mouse
cDNA.
Results: The model for a drug-induced hepatic disorder displayed numerous small white blotches, which
were visible with the naked eye, around the hepatic portal system; and the results of blood chemical
analyses indicated AST and AL T levels that were significantly higher (P