Tadalafil Effect on Metabolic Syndrome-Associated Bladder Alterations: An Experimental Study in a Rabbit Model
Linda Vignozzi, MD, PhD; Sandra Filippi, PhD; Paolo Comeglio, PhD; Ilaria Cellai, PhD; Annamaria Morelli, PhD; Elena Maneschi, PhD; Erica Sarchielli, PhD; Mauro Gacci, MD; Marco Carini, MD; Gabriella Barbara Vannelli, MD; and Mario Maggi MD
ONLINE: February 26, 2014 – The Journal of Sexual Medicine
Introduction and Aims
Historically, prostatic enlargement and lower urinary tract symptoms (LUTS) have been causally linked. More recently, this connection has been challenged, as men with an enlarged prostate do not always develop LUTS and vice versa. LUTS can also be found in women. As a result, other factors aside from an enlarged prostate, such as bladder dysfunction, should be considered.
LUTS has been associated with age-related disorders, including cardiovascular disorders and metabolic syndrome (MetS). Several MetS symptoms, such as abdominal obesity and hypertension, are related to LUTS as well. Animal studies have been helpful, but are not as applicable to humans.
The authors’ past animal research has shown that MetS can affect the entire lower urinary tract. In particular, bladders from MetS animals showed increased signaling of the Ras homolog gene family, member A (RhoA)/Rho-associated protein kinase (ROCK), which can contribute to bladder dysfunction in humans as well.
Previously, the authors tested chronic (preventative) and acute (curative) dosing of tadalafil in animals fed a high-fat diet, paying close attention to prostate changes. Both chronic and acute dosing reduced Met-S-associated prostatic abnormalities, including fibrosis and inflammation.
The aim of this study was to examine the acute (curative) effect of tadalafil dosing on Met-S associated increased signaling of RhoA/ROCK in the bladder.
Materials and Methods
MetS Rabbit Model
The study involved 94 male New Zealand White rabbits who ate a standard rabbit diet before being randomly assigned to one of two different treatment groups: the control group (43 rabbits) and the treatment group (51 rabbits). For twelve weeks, the control group ate a standard diet and the treatment group ate a high-fat diet (HFD). Fourteen rabbits in the treatment group received 2 mg/kg of tadalafil each day for one week. The tadalafil was added to their drinking water.
The rabbits were sacrificed after twelve weeks of treatment. Samples of their bladders, seminal vesicles, prostates, and testes were harvested. Biochemical and hormonal serum analyses were also completed.
The following measurements, assessments, and preparations were made:
• Glucose, total cholesterol, triglycerides, testosterone (T), and 17β estradiol
• Oral glucose tolerance tests
• In vitro contractility studies on bladder samples
• Membrane/cytosolic fractions and western blog analysis for RhoA assay
• RNA extraction and quantitative RT-PCR
• Rabbit bladder smooth muscle cell cultures
• Rabbit smooth muscle tissue migration
The authors summarized their results as follows:
• Bladder strips from HFD rabbits showed hyper-responsiveness to Y-27632, indicating RhoA/ROCK overactivity in HFD bladder compared with matched controls.
• Accordingly, the fraction of activated (translocated to the membrane) RhoA as well as ROCK expression are increased in HFD bladder.
• Tadalafil dosing normalized HFD-induced bladder hypersensitivity to Y-27632, by reducing RhoA membrane translocation and ROCK overexpression.
• Tadalafil dosing reduced mRNA expression of inflammatory, pro-fibrotic, and hypoxia markers.
• A direct inhibitory effect of tadalafil on RhoA/ROCK signaling in bladder smooth muscle cell was demonstrated by using chemotaxis assay.
• Pre-treatment with tadalafil inhibited both basal and PDGF-induced migration of bladder smooth muscle cells.
Discussion and Conclusion
The authors noted that their study confirmed that “experimental MetS is associated with severe bladder alterations, including upregulation of RhoA/ROCK signaling and abnormal expression of genes related to inflammation, hypoxia, and fibrosis. Short-term (1 week) tadalafil dosing was able to normalize all these alterations.”
The study also confirmed “the concept that RhoA/ROCK is functionally overactive in an HFD bladder, most probably mediating hypersensitivity to carbachol,” the authors wrote.
They added, “Another important observation of this study is that PDE5 expression within the bladder resulted positively associated to the expression of several genes related to inflammation and fibrosis.”
The researchers acknowledged the following limitations:
• Detrusor smooth-muscle contraction is mediated not only by the cholinergic signaling, but also by the purinergic one (mainly through purinergic receptor P2X, ligand-gated ion channel, 1 [P2X1]), the latter has not been investigated in the present study.
• In vivo urodynamic studies, to evaluate the effect of tadalafil on in vivo bladder function, were not performed. However, a previous study demonstrated that vardenafil dosing was able to ameliorate urodynamic parameters in another model of MetS-like syndrome, the SHR rat.
• Although we studied the effect of tadalafil on two downstream pathways of RhoA activation-in rbBSMC (such as migration and mRNA expression of smooth muscle genes), we did not specifically evaluate the main ROCK1 substrate, MYPT1, and its phosphorylation.
Our data demonstrate that tadalafil dosing reduced RhoA/ROCK signaling in an animal model of MetS-associated bladder alterations, by decreasing smooth muscle overactivity. Our findings add new insights into the comprehension of the mechanism of action of PDE5 inhibitors in alleviating LUTS in MetS patients.