Regular insulin

Human PC tissues were obtained from patients undergoing radical prostatectomy at the University of Wisconsin-Madison. All patients were consented under an Institutional Review Board (IRB) protocol #20130653. Each core was cut into ~1 mm³ pieces with a sterile scalpel and scissors. Absorbable gelatin sponges (Ethicon) were cut into pieces to fit in a 24-well tissue culture plate (Figure 4). Sponges were soaked in Ham's F-12 media (Mediatech) supplemented with 0.25 units/ml regular insulin (Sigma-Aldrich), 1 μg/mL hydrocortisone (Sigma), 5 μg/mL human transferrin (Sigma), 2.7 mg/ml dextrose (Sigma), 0.1 nM non-essential amino acids (Hyclone), 100 units/ml and 100 μg/mL Penicillin/Streptomycin respectively (Mediatech), 2 mM L-glutamine (Sigma), 25 μg/mL bovine pituitary extract (Invitrogen) 1% FBS (Gibco, Life Technologies) until fully saturated. Tissue was placed on the sponges and cultured for up to 6 days at 37C at 5% CO₂ and 0.5 ml. Media was replaced, daily.

Human prostate tissues were obtained from patients undergoing radical prostatectomy at the University of Wisconsin-Madison. All patients were consented in writing under an Institutional Review Board (IRB) protocol #20130653. Research has been performed in accordance with the Declaration of Helsinki. All the laboratory investigators were blinded to clinical information. Absorbable gelatin sponges (Ethicon, Cat# 1973) were cut into pieces to fit in a 24-well tissue culture plate. Sponges were soaked in Ham’s F-12 media (Fisher Scientific, Cat# SH3002601) supplemented with 0.25 units/ml regular insulin (Sigma-Aldrich, Cat# I9278-5ML), 1 μg/mL hydrocortisone (Sigma-Aldrich, Cat# H0888-1g), 5 μg/mL human transferrin (Sigma-Aldrich, Cat# T8158-100mg), 2.7 mg/ml dextrose, 0.1 nM non-essential amino acids (HyClone, Cat# SH30238.01), 100 units/ml and 100 μg/mL Penicillin/Streptomycin, respectively (HyClone, Cat# SV30010), 2 mM L-glutamine (Corning, Cat# 25-005-CI), 25 μg/mL bovine pituitary extract (Life Technologies, Cat# 13028014), and 1% fetal bovine serum (FBS) (Gibco, Cat# 10437028) until fully saturated. Each core was cut into ~1 mm² by 1 mm² cubes. Tissue was placed on the sponges and cultured for up to 4 days at 37 °C at 5% CO₂ and 500 μL media was replaced daily.