Finasteride

Behavioral tests were conducted by an experimenter who was unaware of treatment condition. The testing apparatus consisted of two chambers equipped with a floor made of stainless steel rods (4 mm diameter, spaced 10 mm apart). The grid floor in the dark compartment (12 cm × 20 cm × 16 cm) was connected to an electrical source so that a shock could be delivered when a rat entered the compartment. The adjoining safe compartment (30 cm × 20 cm × 16 cm) was illuminated with four 13W lights. Light intensity in the center of the illuminated chamber was 1000 lux and that in the dark chamber less than 10 lux. On the initial day of training, rats were placed in the apparatus without turning on the light and allowed to habituate for 10 min in each compartment. Finasteride (50 mg/kg suspended in Liposyn III 30%) or Liposyn III alone was administered by intraperitoneal (i.p.) injection on the same day. On the next day, each animal was placed in the illuminated compartment and we confirmed that the rat promptly entered the dark compartment. No foot shocks were given during the pre-exposure period. Another dose of finasteride or Liposyn III was injected 40 min before midazolam (MDZ) injection. MDZ (3 mg/kg, dissolved in saline with sonication) or saline was injected 20 min before trials in which a foot shock was administered when the rat entered the dark chamber. All rats were placed in the illuminated compartment and allowed to enter the dark compartment spontaneously. When rats escaped from the dark compartment, they were returned to their original home cages. On the third day, all rats were again placed in the illuminated chamber for a maximum duration of 3 min and monitored for the time elapsed before entering the dark compartment.

Whole blood was collected from experimental groups by submandibular bleed or trunk blood collection. Submandibular blood was collected 24 hours prior to restraint stress (before). THDOC (20 mg/kg) and finasteride (50 mg/kg) were dissolved in 1ml cremaphor heated to 65° and then 4 ml 0.9% injection saline was added. Mice either received vehicle (1ml cremaphor + 4ml injection saline), THDOC, or finasteride 30 min prior to a single 30 min restraint stress. The mice were allowed to recover for 30 min then decapitated and trunk blood was collected (after). Plasma was isolated by high speed centrifugation and corticosterone levels were measured by enzyme immunoassay according to manufacturer's specifications (Enzo Life Sciences). Briefly, duplicate 5 μl plasma samples were assayed and absorbance measurements at 415 nM were compared to a standard curve. Samples from different experimental groups were run in parallel.

All analyses were conducted using SUDAAN v 9.0 software (Research Triangle Park, NC) as implemented in SAS v 9.2 (Cary, NC), to account for the unequal probabilities of selection, over-sampling, and non-response that were part of the complex NHANES sampling design [18 ]. We estimated age (continuous) and multivariable-adjusted geometric mean PSA concentrations and 95% confidence intervals by statin and cholesterol-lowering drug use and for each quintile of serum total cholesterol concentration using linear regression. We repeated these analyses for HDL cholesterol and, in the fasting subsample (n=1,101), LDL cholesterol. We transformed PSA concentration using the natural logarithm because it was not normally distributed. We also estimated geometric mean PSA concentration by deciles and clinical cutpoints of total cholesterol to determine whether modeling according to quintile cutpoints accurately captured the shape of the association between total cholesterol and PSA concentration. The inferences were similar using each of the three sets of cutpoints, so we report the results by quintiles of total cholesterol.
We included in the multivariable models factors that were known or that we hypothesized would be associated with both PSA concentration and either statin use or cholesterol concentration: race/ethnicity (non-Hispanic white, non-Hispanic black, Mexican-American, other Hispanic, other), measured body mass index (cutpoints based on World Health Organization categories [19 ] with finer categorization: <22.5, 22.5–<25, 25–<27.5, 27,5–<30, 30–<32.5, 32.5–<35, and ≥35 kg/m²), and cigarette smoking (never, current, former). Information on cigarette smoking was collected by interview. Further adjustment for serum cotinine, serum C-reactive protein concentration, waist circumference, skinfold measurements, education level, poverty index ratio, dietary intake of total fat, protein, and carbohydrate, serum cholesterol concentration (in analyses of statin use), and aspirin use (2001–2002 only) did not change these results (no point estimate changed by >0.01); thus these variables were not included in the final model.
Subanalyses were conducted to limit the possible confounding influence of health status by stratifying by presence or absence of major comorbidities (cancer, myocardial infarction, stroke, angina, congestive heart failure, coronary heart disease, diabetes); information on comorbidities was self-reported during the interview. Within strata of health status we further adjusted for liver and kidney function, as defined using methods previously described [20 (link), 21 (link)], because we hypothesized that impaired function could influence PSA clearance. We also conducted analyses stratified by age (40–<50, 50–<60, 60–<70, 70–<80, ≥80 years) and BMI (<25, 25–<30, ≥30 kg/m²). Further, we conducted analyses excluding men who were taking finasteride (n=40). To be able to efficiently control for confounding and account for possible interactions among the confounders, we generated a propensity score [22 ] using the following: age, race/ethnicity, BMI, waist circumference, cigarette smoking status, alcohol consumption, hypertension, diabetes, kidney function, and liver function. We then adjusted for quintiles of the propensity score using indicator variables. Statistical interaction was assessed by including interaction terms between the potential effect modifiers as categorized above and statin use or quintiles of cholesterol concentration in the multivariable model and estimating its statistical significance using the Wald test.
All protocols for the implementation of NHANES 2001–2004 were approved by the Institutional Review Board of the National Center for Health Statistics, Centers for Disease Control and Prevention; informed consent was obtained for all participants.