Crystalline testosterone

Cyanidin-3-rutinoside, cyanidin chloride, 3,3’-diethyloxacarbocyanin iodide, resazurin, sulforhodamine B, Flutamide, and carboxymethylcellulose were purchased from Sigma-Aldrich (St. Louis, MO). Ellagic acid was obtained from Indofine Chemical (Hillsborough, NJ). Protocatechuic acid waspurchased from LKT Labs (St. Paul, MN). Urolithin A was obtained from Santa Cruz (Dallas, TX). Crystalline testosterone, testosterone propionate, and 17β-estradiol were obtained from Steraloids (Newport, RI). Noble agar was purchased from Affymetrix (Cleveland, OH). DMSO, EtOH, formaldehyde, formic acid and trichloroacetic acid were obtained from Thermo Fisher Scientific (Waltham, MA). Ketamine was obtained from Henry Schein Animal Health (Dublin, OH). Xylazine was purchased from Lloyd Laboratories (Shenandoah, IA). N-methyl-N-nitrosourea was obtained from the NCI Carcinogen Repository (MRIGlobal, Kansas City, MO).

After randomization into six experimental groups (see Study Conduct), the rats in groups to be subjected hormone treatment received two subcutaneous silastic tubing implants (internal diameter 0.078 inch; external diameter, 0.125 inch; Dow Corning, Midland, MI) tightly packed over a length of 2 cm with crystalline testosterone (Steraloids, Newport, RI) and another implant packed over a length of 1 cm with crystalline 17β-estradiol (Steraloids); control rats received empty implants [32 (link)]. The implants were sealed with medical grade silicone sealant (RTV 108, General Electric, Momentive Performance Materials, Waterford, NY) placed on top of small glass beads inserted in the tubing after filling with hormone. Immediately before implantation, the implants were rinsed in 70% ethanol and then soaked for 48 h in sterile saline at 37°C, which was replaced twice daily. The Silastic implants were implanted under the skin between the scapulae while the rats were anesthetized with fentanyl-droperidol. This treatment increases circulating estradiol levels by 5- to 10-fold in NBL rats while maintaining testosterone at physiologic levels (~3 ng/mL) [33 (link)].