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Volume 63, Issue 3, Pages 506-516 (March 2013) The Mechanism of Action of Phosphodiesterase Type 5 Inhibitors in the Treatment of Lower Urinary Tract Symptoms Related to Benign Prostatic Hyperplasia François Giuliano, Stefan Ückert, Mario Maggi, Lori Birder, Jay Kissel, Lars Viktrup European Urology Volume 63, Issue 3, Pages 506-516 (March 2013) DOI: 10.1016/j.eururo.2012.09.006 Copyright © 2012 European Association of Urology Terms and Conditions

Fig. 1 Phosphodiesterase type 5 (PDE5) isoenzymes in the lower urinary tract. European Urology 2013 63, 506-516DOI: (10.1016/j.eururo.2012.09.006) Copyright © 2012 European Association of Urology Terms and Conditions

Fig. 2 Phosphodiesterase type 5 (PDE5) expression and immunolocalization in human tissues. (a) An intense PDE5 immunopositivity was detected in the smooth muscle bundles and endothelial layer surrounding the vascular bed of transverse sections of human deferential artery. (b) Representative negative control image, hematoxylin counterstained, obtained by omitting the primary anti-PDE5 antibody in a transverse section of human deferential artery. (c) The prostatic gland section shows a scanty PDE5 immunostaining in fibromuscular stroma (asterisks), whereas it is mainly distributed in the endothelial and smooth muscle cells of blood vessels (arrows). (d) An intense PDE5 immunostaining was detected in both smooth muscle and endothelial component of corpora cavernosa section. Magnification ×4. Reproduced with permission from the International Society for Sexual Medicine [37]. European Urology 2013 63, 506-516DOI: (10.1016/j.eururo.2012.09.006) Copyright © 2012 European Association of Urology Terms and Conditions

Fig. 3 Immunohistochemical staining of Hypoxyprobe in rat bladder. Hypoxyprobe-positive protein adducts were revealed in hypoxic cells (PO2 <10mm Hg) of bladder transverse sections by a monoclonal antibody (magnification ×10). (a) Wistar-Kyoto rats (WKY): Only scanty positive labeling is present. (b) Untreated spontaneously hypertensive rats (SHR): Massive hypoxia is present in both the urothelium/suburothelium (arrows) and vascular endothelium (arrowheads). (c) Vardenafil-treated spontaneously hypertensive rats (SHR). Hypoxyprobe labeling is dramatically decreased; only a few endothelial (arrows) cells are positive. Reproduced with permission from the International Society for Sexual Medicine [45]. WKY=Wistar-Kyoto rats. European Urology 2013 63, 506-516DOI: (10.1016/j.eururo.2012.09.006) Copyright © 2012 European Association of Urology Terms and Conditions

Fig. 4 Immunohistochemical staining of Hypoxyprobe in rat prostate. Hypoxyprobe-positive protein adducts were revealed in hypoxic cells (PO2 <10mm Hg) of prostate transverse sections by a monoclonal antibody (magnification ×10). Images (magnification ×10; bars=50mm) are representative of each experimental group at each time point (a–c: 1 d; d–f: 7 d; g–i: 28 d). The staining was almost undetectable in WKY prostate sections (a, d, g). The prostate sections from untreated SHR were strongly immunopositive for Hypoxyprobe in the epithelial layers (arrows) surrounding prostate ducts, which were also characterized by alveolus dilation and reduction of interstitial and stromal spaces (b, e, h). Prostate sections from tadalafil-treated SHR show a net reduction of Hypoxyprobe immunopositivity after (c) 1 d, becoming absent after both (f) 7 d and (i) 28 d. Reproduced with permission from the International Society for Sexual Medicine [37]. European Urology 2013 63, 506-516DOI: (10.1016/j.eururo.2012.09.006) Copyright © 2012 European Association of Urology Terms and Conditions