Mice were randomly divided into one control group and
two treatment groups. For deguelin testing, the groups consisted
of a control group and two groups of animals treated
with deguelin at 5.0 or 10.0 mg/kg body weight. Deguelin was
dissolved in corn oil just before administration and was
administered by gavage (intragastrically) 5 days/week for
the duration of the study. Control animals were treated with
corn oil (administered intragastrically) throughout the study.
For silibinin testing, the groups consisted of a control group
treated with AIN-76A purified pellet diet and two groups
treated with silibinin at 0.05% and 0.1% (wt/wt) doses mixed
with AIN-76A purified powder diet. Foods were prepared
once a week with a powerful KitchenAid mixer (St. Joseph,
MI) mixing for at least 1 hour. Treatments were initiated
10 days before carcinogen introduction. After 10 days of
treatment, all groups received a single intraperitoneal dose
of BP (100 mg/kg body weight) in 0.2 ml of tricaprylin. Body
weights of mice were measured every 2 weeks.
During the duration of the study, the health condition of
the mice was monitored everyday, and body weights were
measured every 2 weeks. Mice were sacrificed by CO2
asphyxiation 20 to 21 weeks after carcinogen treatment.
For the phenotyping of lung tumors in all bioassays described
above, lung tissues were fixed in Tellyesniczky’s solution
overnight, and then in 75% EtOH. Lung tumor development
was estimated by two investigators, using a Leica MZ75 dissecting
microscope, to measure the number (N ), volume (V ),
and total tumor load (NV ), as reported previously [25]. Volume
calculation was based on the formula: V (mm3
) = 4/3pr 3
.
Histopathological examinations were performed to determine
the diagnosis of lung tumors.