Ab2078

Expression of OT and OTR was determined in prostate tissue from our tissue bank and the tissue microarray (PR803c) was obtained from US Biomax Inc. (Rockville, MD, USA), which together comprised 7 normal prostatic tissue cores, 98 BPH tissue samples (all were treated as no-carcinoma tissue) and 101 prostatic cancer tissue cores. IHC staining was performed using antibodies against OT (ab2078, 1:200; Abcam, Shanghai, China) and OTR (ab115664, 20–40 μg/ml; Abcam). First, hematoxylin and eosin (HE) staining was used to visualize the gross tissue morphology. The secondary antibody was a goat anti-rabbit IgG (sc-2040; Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA) used at a 1:200 dilution. Hematoxylin was further counterstained for the nuclei. All the areas with the most intensity were evaluated by scanning at two different magnifications. H-Score of each core was calculated with the QuantCenter software (version 2.0, 3DHISTECH Ltd., Budapest, Hungary) which can recognize and analyze the staining intensity (0, negative; 1 weak; 2 moderate; 3 strong) and positive staining area (pixels). The H-Score was calculated as follows: H-Score=∑ (PI×I) = (percentage of cells of weak intensity ×1) + (percentage of cells of moderate intensity ×2) + (percentage of cells of strong intensity ×3).

Four hours after sevoflurane exposure, bilateral hippocampal tissue was dissected out as described above (control group, n = 6; sevo group, n = 6) and immediately homogenized with 100 mg/ml RIPA Lysis Buffer (Shenergy Biocolor Co., China) and 1 % (v/v) PMSF (Shenergy Biocolor Co., China). After 20 min of centrifugation at 13,000×g at 4 °C, the supernatant was aspirated and stored at −80 °C for further use. The extracted hippocampal proteins were electrophoresed using a 5 % stacking gel at 80 V for 20 min and 12 % separating gel at 120 V for 40 min and were then transferred onto polyvinylidene fluoride membranes (Pall Co., USA). The blots were immunoreacted with anti-OT (1:500, AB2078, Abcam, UK), anti-AVP (1:500, AB48322, Abcam, UK), and anti-GAPDH (1:1000, SC25778, Santa Cruz, USA) antibodies. The ECL-PLUS Reagents Kit (Com Win Biotech Co., Ltd, China) was utilized to detect the bands. The results were examined under a digital imaging system (ImageQuant Las4000 mini, General Electric, USA). The optical density (OD) was then analyzed with Image J software.