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. Author manuscript; available in PMC: 2016 Apr 7.
Published in final edited form as: J Sex Med. 2009 Mar;6(Suppl 3):269–278. doi: 10.1111/j.1743-6109.2008.01184.x

Figure 4.

Figure 4

Protein and mRNA expression levels of RhoA ROCK I, and ROCK II in lean and obese-diabetic corpus cavernosal tissues following rosuvastatin or saline treatment. Representative protein blots and mean expression levels of RhoA (A), ROCK I (C), and ROCK II (E) in lean and obese-diabetic penile tissue as detected by western blot. Frozen corpus cavernosal tissue was homogenized in lysis buffer supplemented with protease and phosphatase inhibitor cocktails. Proteins were resolved by polyacrylamide gel electrophoresis and transferred onto nitrocellulose membranes. The membranes were blocked with Tris-buffered saline containing 0.1% Tween 20 and 4% BSA. Incubation with specific primary antibodies (ROCK I and II from BD Transduction Labs [Franklin Lakes, NJ, USA], and RhoA from Cell Signaling Technology [Beverly, MA, USA]) and then incubated with horseradish peroxidase-conjugated anti-mouse immunoglobulin G. Immunoreactive bands were visualized using ECL Plus and image acquisition was performed on a Typhoon 9410 workstation (Amersham Bioscience, Piscataway, NJ, USA) and analyzed using UN-SCAN-IT™ gel version 5.1. Reported are means ± standard error of the mean (SEM) for three animals in each group. Mean mRNA expression of RhoA (B), Rho-kinase I (D), and II (F) as detected by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) in lean and obese-diabetic corpus cavernosal tissues following rosuvastatin treatment. Total RNA was extracted from harvested cavernosal tissue using TriReagent (Sigma-Aldrich) and cDNA was generated using the High Capacity cDNA kit from Applied Biosystems (Foster City, CA, USA). Real-time RT-PCR was performed using TaqMan primers and probes mixes using glyceraldehyde-3-phosphate dehydrogenase as the reference gene on an iCycler iQ (Bio-Rad Laboratories, Hercules, CA, USA). Reported are means ± SEM for four to six animals in each group. Statistical difference from control within group *P < 0.05 or ***P < 0.001. L C = lean control; L S = lean statin; O C = obese control; O S = obese statin.