This Month in Investigative Urology

Simulated Bladder Pressure StimulatesBladder Smooth Muscle Cell Proliferation

To better understand the relationship between blad-der function and growth at the cellular level, Chenet al (page 661) from Chengdu, People’s Republic ofChina used a novel method of applying cyclic hydro-dynamic pressure that simulates the bladder cycle tocell cultures. They studied the proliferation responseof human bladder smooth muscle cells (HBSMCs) todifferent pressures as well as the signal transduc-tion mechanisms for this process. The investigatorssubjected HBSMCs cultured in scaffolds to 4 differ-ent pressures (0, 100, 200 and 300 cm H2O) for 24hours, controlled by a BioDynamic® bioreactor.Flow cytometry was used to examine cell cycle dis-tribution, and polymerase chain reaction and immu-noblot were used to quantify SGK1 and AKT expres-sion and activation in each group. Additionally,SGK1 was silenced in HBSMCs using small inter-fering RNA to validate the role of SGK1 in mediat-ing pressure induced cell proliferation.

Compared with the 0 cm H2O control, HBSMCs inthe 200 and 300 cm H2O groups showed increasedcell proliferation. The expression and activity ofSGK1 were also increased, while AKT, anotherdownstream signal of PI3K, did not change signifi-cantly. In addition, SGK1 silencing abolished theincreases in cell proliferation induced by pressure.To my knowledge, this is the first report of cyclichydrodynamic pressure stimulating the prolifera-tion of HBSMCs cultured in scaffolds. The signaltransduction mechanism for this process involvesthe PI3K/SGK1 and not the PI3K/AKT signalingpathway. Better understanding of cell cycle and pro-liferative signaling pathways may provide impor-tant novel targets to treat lower urinary tract disor-ders and engineer bladder tissue.

PDGFR� Positive InterstitialCells in Human and Guinea Pig Bladders

The observation that only a subpopulation of inter-stitial cells is positive for KIT suggests that addi-tional cell types exist in the vimentin positive cellu-lar networks of the bladder wall. Monaghan et al(page 639) from Belfast, United Kingdom examinedwhether a subpopulation of interstitial cells immuno-

positive for platelet-derived growth factor receptor al-

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pha (PDGFR�) exists in human and guinea pig blad-ders. They examined guinea pig and human bladdertissues by immunohistochemistry and bright field orconfocal microscopy. Whole mount tissues and paraffinsections were labeled with antibodies to PDGFR�,vimentin, KIT and protein gene product 9.5. Proteinexpression was assessed by Western blotting.

PDGFR�� cells were present in guinea pig andhuman bladders. In the guinea pig bladder PDGFR��

cells had a branched stellate morphology and formedcellular networks in the lamina propria. In the hu-man and guinea pig detrusor PDGFR�� cells wereelongated on the boundary of smooth muscle bun-dles or were seen as groups of stellate cells in theinterbundle spaces. PDGFR�� cells were locatedclose to nerves labeled by protein gene product 9.5.Double labeling revealed that PDGFR�� cells werea subgroup of the vimentin� population. Moreover, asignificant proportion of PDGFR�� cells were alsoKIT�. Bands corresponding to PDGFR�, KIT andvimentin proteins were detected on Western blot.This study further demonstrates the complexity ofbladder interstitial cells, and that PDGFR��/KIT�

cells located in the lamina propria and detrusorlayers are a subgroup of the vimentin� population.Since PDGFR�� cells were apparently structurallyassociated with intramural nerves, these cells maybe part of bladder control mechanisms.

Magnetic Tool for Retrieval ofStone Fragments Rendered Paramagnetic

Studies have shown that peptide and polymer coatediron oxide paramagnetic microparticles that bind tocalcium stones allow the extraction of small stonefragments with magnetic tools. Tan et al (page 648)from Dallas, Texas developed a prototype magnetictool for ureteroscopic extraction of magnetized stoneparticles and compared its efficiency for retrievingmagnetized calcium oxalate monohydrate stone par-ticles with that of a conventional nitinol basket fromthe pelvi-collecting system of a bench top uretero-scopic simulator. Iron oxide microparticles were suc-cessfully bound to 1 to 1.5, 1.5 to 2 and 2 to 2.5 mmhuman calcium oxalate monohydrate stones. Sev-eral coated fragments of each size were implantedinto the collecting system of the bench top uretero-

scopic simulator and 5-minute timed stone extrac-

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tion trials were performed for each fragment sizeusing a back loaded 8Fr magnetic tool mounted on a0.038 inch guidewire or a conventional basket. Themedian number of fragments retrieved per timedtrial was compared for the magnetic tool vs thebasket using the Mann-Whitney U test.

For 1 to 1.5 mm fragments the median numberretrieved within 5 minutes was significantly higherfor the prototype magnetic tool than for the nitinolbasket (9.5 vs 3.5, p � 0.03). The magnetic tool wasmore efficient for 1.5 to 2 mm fragments but thedifference in the number of fragments retrieved wasnot statistically significant (9.5 vs 4.5, p � 0.19). For2 to 2.5 mm fragments there was no difference be-tween instruments in the number retrieved (6 pergroup, p � 1.0). The use of a prototype magnetic toolimproved the efficiency of retrieving stone particlesrendered paramagnetic measuring less than 2 mmbut showed no advantage for larger fragments. Ad-ditional refinement and investigation of this tech-nology are required before it can be applied clini-cally. However, the system has the potential toreduce the number of small retained fragments afterureteroscopic lithotripsy.

Avanafil Phosphodiesterase-5Inhibitor for Erectile Dysfunction

Kotera et al (page 668) from Saitama, Japan inves-tigated the in vitro inhibitory effects of a novel andpotent phosphodiesterase (PDE) type 5 inhibitor,avanafil, and its enhancement of penile tumescencein dogs. They performed PDE assays with the 4PDE5 inhibitors avanafil, sildenafil, vardenafil and

tadalafil using 11 PDE isozymes. In anesthetized

dogs, intracavernous pressure was measured andthe pelvic nerve was repeatedly stimulated to evoketumescence before and after administration of avanafilor sildenafil.

Avanafil specifically inhibited PDE5 activity witha 50% inhibitory concentration value of 5.2 nM butshowed higher selectivity (121-fold) against PDE6than sildenafil and vardenafil (16 to 21-fold). Thedrug showed excellent selectivity (greater than10,000-fold) against PDE1 compared with sildenafil(375-fold). Avanafil had higher selectivity againstPDE11 (greater than 19,000-fold) compared withtadalafil (25-fold) and also showed excellent selectiv-ity against all other PDEs. Following intravenousadministration in anesthetized dogs, the 200% effec-tive dose (ED200%) of avanafil and sildenafil on thepenile tumescence was 37.5 and 34.6 �g/kg, respec-tively. Following intraduodenal administration theED200% of avanafil and sildenafil on tumescence was151.7 and 79.0 �g/kg at the peak time, respectively.Time to peak response with avanafil and sildenafilwas 10 and 30 minutes, respectively, indicating amore rapid onset of avanafil.

Avanafil has a favorable PDE5 selectivity profilecompared to that of marketed PDE5 inhibitors, andthe drug shows excellent in vitro and in vivo potencyas well as rapid onset of action. Cumulative datashow that avanafil possesses a promising pharma-cological profile for the treatment of erectile dysfunc-tion. However, controlled clinical trials are requiredto prove any clinical advantages over existing PDE5inhibitors.

Karl-Erik Andersson

Section Editor