Mean Bladder Weight Research Articles

Chemoprevention of Urothelial Cell Carcinoma Tumorigenesis by Dietary Flavokawain A in UPII-Mutant Ha-ras Transgenic Mice.

Non-muscle-invasive bladder cancer (NMIBC) has one of the highest recurrence rates among all solid cancers and the highest lifetime treatment cost per patient. Therefore, the development of chemoprevention strategies for reducing the occurrence and recurrence of NMIBC as well as its burdens on the healthcare system is valuable. Our aim was to determine whether flavokawain A (FKA), a kava chalcone isolated from the kava plant, can target the in vivo activated Ha-ras pathway for prevention and treatment of NMIBC. UPII-mutant Ha-ras transgenic mice that develop papillary urothelial cell carcinoma were fed orally with vehicle control or FKA-formulated food for 6 months starting at 6 weeks of age. Seventy-nine percent (15/19) of male mice fed with 6 g FKA per kilogram (kg) of food survived beyond the 6 months of treatment, while 31.6% (6/19) of control food-fed male mice survived the 6-month treatment period (p = 0.02). The mean bladder weights in FKA vs. control food-fed mice were 0.216 ± 0.033 vs. 0.342 ± 0.039 g in male mice (p = 0.0413) and 0.043 ± 0.004 vs. 0.073 ± 0.004 g in female mice (p < 0.0001); FKA reduced bladder weight by 37% and 41%, respectively. The tumor burdens, determined by the wet bladder weight, in these mice were inversely related to plasma FKA concentrations. In addition to decreased bladder weight, FKA treatment significantly reduced the incidences of hydronephrosis and hematuria. FKA-treated mice exhibited more well-differentiated tumors in the bladder and ureter. Immunohistochemical analysis of FKA-treated tumors compared to those in the control group revealed fewer Ki-67- and survivin-positive cells and an increased number of p27- and TUNEL-positive cells, indicating that FKA inhibits proliferation and induces apoptosis. Overall, the results suggest that FKA can target the in vivo activated Ha-ras pathway for the prevention and treatment of NMIBC.

Sonographic Reference Values for Bladder Wall Thickness, Detrusor Wall Thickness, and Bladder Weight in Apparently Healthy Adults in a Nigerian Population

The aim was to establish reference values of bladder wall thickness (BWT), detrusor wall thickness (DWT), and bladder weight (BW) in apparently healthy adults in a Nigerian population. Therefore, a cross-sectional study of healthy adult participants was conducted from May 2015 to April 2016. The urinary bladder was sonographically evaluated on a convenient sample of 384 adult participants. The BWT, DWT, and BW of the participants were measured and documented. The BW was estimated based on the surface area, thickness, and bladder muscle specific gravity. The mean BWT, DWT, and BW were 2.8 ± 0.3 mm, 1.3 ± 0.1 mm, and 23.3 ± 4.1 g, respectively. There was no statistically significant correlation between anthropometric variables with BWT, DWT, and BW except age, which had a weak positive correlation with BWT ( P = .05). This data set could be used for future research, in other parts of the country, for a possible nationwide nomogram.

TGF‐β1 and connexin‐43 expression in neurogenic bladder from rats with sacral spinal cord injury

Sacral spinal cord injury (SCI) could induce underactive bladder (UAB). Malfunction of connexin 43 (CX43) regulated by TGF-β1 might involve in urinary bladder dysfunction. We studied the changes of CX43 and TGF-β1/Smad3 signaling in detrusor of neurogenic bladder (NB) in sacral SCI rats. Sacral SCI was produced by hemisection (SSCH) or transection (SSCT) of spinal cord between L4 and L5 in female Wistar rats. BBB scores, residual urine volume and bladder weight as well as characteristic cystometric parameters at 6th week were used to confirm the successful establishment of NB. Western blotting and qRT-PCR were used to exam the protein and mRNA expression levels of CX43, CX45, TGF-β1, and Smad3 in detrusor. BBB scores were significantly decreased, with the lowest in SSCT rats (P < 0.01). The residual urine volume, mean bladder weight, and cystometric parameters were increased, with the highest in SSCT rats. CX43 and phospho-CX43 protein levels were significantly decreased, but those of TGF-β1, Smad3, and phospho-Smad3 were significantly increased. It was the protein and mRNA levels of CX43 but not those of CX45 which were decreased in negative accordance with those of TGF-β1 and Smad3. Those changes were more significant in SSCT than in SSCH rats. This study indicates that voiding dysfunction is related to the decreased CX43 function in detrusor from NB. TGF-β1/Smad3 signaling might be involved in the down-regulation of CX43 in SCI rats. Early regulation of CX43 might be beneficial to patients with voiding dysfunction.

Abstract 256: Delta tocopherol inhibits urothelial tumorigenesis in the UPII mutant Ha-ras transgenic mouse model and induces apoptosis via activation of the ATF4/CHOP-DR5 pathway

Abstract Epidemiological studies have reported that Vitamin E intake was inversely related to the risk of multiple cancers including human urinary bladder cancer. Tocopherols (T) are the major forms of vitamin E in the U.S. diet, exist as α-T, β-T, γ-T and δ-T. Compared to α-T, the anti-cancer effect of other Ts and their mechanisms of action remain largely unknown. We have shown that δ-T is the most effective one among the Ts in reducing the viabilities of bladder cancer cell lines with IC50s of under 10 μM, whereas there is no or minimal effect of α-T at a concentration of up to 50 μM on the viability of bladder cancer cell lines. δ-T treatment of bladder cancer cell lines, RT4 and UMUC-3, resulted in apoptosis via marked induction of death receptor-5 (DR5) expression and then activation of caspase 3, 8, and 9 leading to PARP cleavage and apoptotic morphology. siRNA knockdown of DR5 expression significantly attenuated the apoptotic effects of δ-T in bladder cancer cells. Consistent with the above results, δ-T treatment significantly upregulated the expression of ER stress sensors PERK and IRE1α, as well as downstream components GRP78, ATF4, and CHOP. These results suggested that δ-T can induce endoplasmic reticulum stress and the Unfolded Protein Response, triggering its apoptotic effect. Furthermore, homozygous male UPII mutant Ha-ras transgenic mice that mimic the development and progression of human urothelial cell carcinoma (UCC) with H-ras activation were fed with vehicle control diet (n=20) or diet supplemented with δ-T (0.2% in diet) for 150 days (n=18). The δ-T diet significantly decreased the mean bladder weights (serves as a surrogate of tumor weight) of survived Ha-ras mutant mice by 56% (control vs δ-T, 0.259 ± 0.015 gram vs. 0.114 ± 0.012 gram, P&amp;lt;0.0001) with no significant difference in overall body mass and no detectable indication of toxicity. Our results suggest that δ-T is a potent anti-cancer agent and deserves further investigation in preventing the recurrence and progression of non-muscle-invasive UCC. Citation Format: Christopher A. Blair, Maggie Wu, Tim Huynh, Hanze Hu, Arman Walia, Chung S. Yang, Xiaolin Zi. Delta tocopherol inhibits urothelial tumorigenesis in the UPII mutant Ha-ras transgenic mouse model and induces apoptosis via activation of the ATF4/CHOP-DR5 pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 256. doi:10.1158/1538-7445.AM2017-256

MP68-20 LUMBAR TO SACRAL ROOT REROUTING TO RESTORE VOIDING FUNCTION IN A FELINE SPINAL CORD INJURY MODEL: URODYNAMIC AND RETROGRADE NERVE TRACING RESULTS FROM A PILOT STUDY

You have accessJournal of UrologyUrodynamics/Lower Urinary Tract Dysfunction/Female Pelvic Medicine: Basic Research & Pathophysiology II1 Apr 2016MP68-20 LUMBAR TO SACRAL ROOT REROUTING TO RESTORE VOIDING FUNCTION IN A FELINE SPINAL CORD INJURY MODEL: URODYNAMIC AND RETROGRADE NERVE TRACING RESULTS FROM A PILOT STUDY Ornella Lam van ba, Mary F. Barbe, Romain Caremel, Shachar Aharony, Oleg Loutochin, Line Jacques, Matthew Woods, Gerald F. Tuite, Michael R. Ruggieri, Lysanne Campeau, and Jacques Corcos Ornella Lam van baOrnella Lam van ba More articles by this author , Mary F. BarbeMary F. Barbe More articles by this author , Romain CaremelRomain Caremel More articles by this author , Shachar AharonyShachar Aharony More articles by this author , Oleg LoutochinOleg Loutochin More articles by this author , Line JacquesLine Jacques More articles by this author , Matthew WoodsMatthew Woods More articles by this author , Gerald F. TuiteGerald F. Tuite More articles by this author , Michael R. RuggieriMichael R. Ruggieri More articles by this author , Lysanne CampeauLysanne Campeau More articles by this author , and Jacques CorcosJacques Corcos More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.1357AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Following complete spinal cord injury (SCI), lumbar to sacral rerouting surgery has the potential to allow voiding via a new skin-central nervous system-bladder reflex pathway. We aimed to assess whether this surgery is effective in treating neurogenic bladder in a feline SCI model METHODS Eight cats underwent SCT at T9-T10. Unilateral L7-S1 ventral root anastomosis was performed 1 month later in 6 cats. Remaining dorsal and ventral roots below L7 were transected, bilaterally. Bladders were checked three times per day and emptied with the Crede maneuver. The new neurological pathway was evaluated at 3, 5, 7 and 9 months by electrical and manual cutaneous stimulation of L7 dermatomes and urodynamics. At 9 months, cats were euthanized 2 weeks after bladder wall Fluorogold retrograde tracer injection and an assessor-blinded pathological exam of the L7-S1 root anastomosis site and lumbosacral cord ventral horns for presence of Fluorogold labeled cell bodies was performed. Outcomes included: 1) urodynamic responses to cutaneous or electrical stimulation the L7 dermatome 2) axonal growth from L7 and S1 ventral horns to the bladder; and 3) incidence of bladder hypertrophy RESULTS At 9 months, 3 of the 6 rerouted cats exhibited increased detrusor pressure provoked by unilateral cutaneous stimulation, and one cat bilaterally. Ipsilateral cutaneous stimulation produced a voiding stream in 2 cats. SCI controls did not respond. Mean bladder volume or weight was not different between rerouted animals and SCI controls. Postmortem studies determined the anastomosed ventral roots were one-sided L7 to S1 (3 right-sided rerouting and 3 left-sided). One right-rerouted cat also showed the left L7 ventral root was annealed to the right S1 root. Although some aberrant axonal regrowth was observed, all 6 rerouted animals showed regrowth of axons from the L7 ventral horn segment to the bladder using retrograde tracing, (Figure 1) including the animal with bilateral input to S1 CONCLUSIONS L7-to-S1 ventral root rerouting surgery below the level of SCT showed successful axonal regrowth to the bladder in all rerouted animals and induced urodynamic responses to cutaneous stimulation in a subset. This feasibility study paves the way for future studies for bladder re-innervation © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e897 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Ornella Lam van ba More articles by this author Mary F. Barbe More articles by this author Romain Caremel More articles by this author Shachar Aharony More articles by this author Oleg Loutochin More articles by this author Line Jacques More articles by this author Matthew Woods More articles by this author Gerald F. Tuite More articles by this author Michael R. Ruggieri More articles by this author Lysanne Campeau More articles by this author Jacques Corcos More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Antitumor Efficacy of Bacillus Calmette-Guerin Loaded Cationic Nanoparticles for Intravesical Immunotherapy of Bladder Tumor Induced Rat Model

For bladder cancer, intravesical chemo/immunotherapy is widely used as adjuvant therapy after surgical transurethral resection. Bacillus Calmette-Guerin (BCG) is a live attenuated Mycobacterium of the same family as tuberculosis, that is capable of inducing a local inflammatory response upon instillation into the bladder. Intravesical therapy with BCG has proved to be more effective in the prophylaxis and treatment of superficial bladder tumors than most chemotherapeutic agents used for the same indication. However, compared to intravesical chemotherapy, BCG immunotherapy provokes more pronounced local and systemic reactions. In addition to the commonly induced granulomatous inflammatory changes in the bladder, which produce irritative symptoms, this therapy may cause systemic side effects varying from mild malaise and fever to, in rare instances, life-threatening or fatal sepsis. Nanoparticles with positive surface charge and mucoadhesive properties were developed to overcome these side effects. Hence, the aim of this study was to optimize and evaluate cationic chitosan (CS) nanoparticles encapsulating BCG in terms of antitumor efficacy after intravesical administration in bladder tumor, induced in rat model. It was found that nanoparticle formulations of 269-375 nm in size can be produced with 42% encapsulation efficiency. The zeta potential was positive and was suitable for intravesical administration. Antitumor efficacy was determined over the parameters of histopathological evaluation, survival rate and mean bladder weight in comparison to treatment with commercial BCG solution. Concerning survival rates, BCG-loaded chitosan nanoparticles resulted in significantly longer survival than BCG commercial product (up to 86 days of survival with no systemic side effects). When compared to healthy bladder weight averages, all groups (especially BCG commercial solution) showed higher bladder weights confirming tumor formation. Histopathological findings confirmed antitumor activity in all treatment groups and optimum findings were observed in groups treated with CS nanoparticles encapsulating BCG. At the same time, significant nanoparticle accumulation in bladder tissues was observed especially for BCG-loaded CS group. In this study, it was clearly observed that cationic CS nanoparticles provide a significantly improved perspective in intravesical immunotherapy of bladder tumors.

Functional and biochemical characteristics of urinary bladder muscarinic receptors in long-term alloxan diabetic rats.

Objective To re-examine the function of the urinary bladder in vivo as well as to determine the functional and biochemical characteristics of bladder muscarinic receptors in long-term alloxan-induced diabetes rats.Methods Two-month-old male Wistar rats were injected with alloxan and the animals showing blood glucose levels >300mg/dL together with age-paired untreated animals were kept for 11 months. Body weight, bladder weight, blood glucose, and urinary volume over a period of 24 hours were determined in both groups of animals. A voiding cystometry in conscious control and diabetic rats was performed to determine maximal micturition pressure, micturition contraction interval and duration as well as voided and post-voiding residual volume. In addition, concentration-response curves for bethanechol in isolated bladder strips, as well as [3H]-N methyl-scopolamine binding site characteristics in bladder homogenates were determined.Results Mean bladder weight was 162.5±21.2mg versus 290±37.9mg in control and treated animals, respectively (p<0.05). Micturition contraction amplitude (34.6±4.7mmHg versus 49.6±2.5mmHg), duration (14.5±1.7 seconds versus 23.33±4.6 seconds) and interval (87.5±17.02 seconds versus 281.11±20.24 seconds) were significantly greater in alloxan diabetic rats. Voided urine volume per micturition contraction was also significantly higher in diabetic animals. However the post-voiding residual volume was not statistically different. Bethanechol potency (EC50 3µM versus 5µM) and maximal effect (31.2±5.9g/g versus 36.1±6.8g/g) in isolated bladder strips as well as number (169±4fmol/mg versus 176±3fmol/mg protein) and affinity (0.69±0.1nM versus 0.57±0.1nM) of bladder muscarinic receptors were also not statistically different.Conclusion Bladder function in vivo is altered in chronic alloxan-induced diabetes rats without changes in functional and biochemical characteristics of bladder muscarinic receptors.

Changes of muscarinic receptors and connexin-43 expression as a mechanism of overactive bladder in ovariectomized rats.

After menopause, the bladder is known to become overactive. To investigate the mechanisms involved in these changes, we examined the muscarinic receptors M2, M3 and gap junction protein connexin-43 in an ovariectomized rat bladder. Twenty 10-week-old female SD rats were used. Ten rats were ovariectomized, (Ovx group) and 10 rats received a sham operation (Con group). Four weeks after the operation, urodynamic tests were performed to verify overactive bladder, and the animals were killed. The body, bladder and uterus weights were measured. The bladder specimens were prepared for immunohistochemical staining for muscarinic receptors M2, M3 and connexin-43. Western blotting was also used for the same protein measurement (M2, M3 and connexin-43). A t test with a p value of 0.05 was considered significant, and SPSS 12.0 for Windows was used to analyze the data. The mean body weight of the Ovx group (315.8 ± 18.1 g) was heavier than the Con group (270.0 ± 23.6 g) (p = 0.009). The mean uterus weight of the Ovx group (260.4 ± 186.8 g) was lighter than the Con group, (600.6 ± 175.9 g) (p = 0.028) and the mean bladder weight of the Ovx group (80.2 ± 15.9 g) was lighter than the Con group (97.4 ± 10.6 g) (p = 0.041). The mean bladder contraction of the Ovx group (5.5 ± 2.3/10 min) was more frequent than that of the Con group (3.2 ± 2.8) (p < 0.05). The expressions of M2 and M3 were not different between the Ovx and the Con group, but the expression of connexin-43 in the Ovx group was more intense than in the Con group in immunohistochemical staining. These findings were also confirmed by Western blotting results. Ovariectomized rats showed frequent bladder contraction and increased connexin-43 expression without changes in M2 and M3 receptor expression. These results imply that ovariectomy-induced overactive bladder may be due to an altered gap junction protein function rather than muscarinic receptor modification.

Anti-interleukin-10R1 monoclonal antibody in combination with bacillus Calmette–Guérin is protective against bladder cancer metastasis in a murine orthotopic tumour model and demonstrates systemic specific anti-tumour immunity

Effective treatment of bladder cancer with bacillus Calmette-Guérin (BCG) depends on the induction of a T helper type (Th) 1 immune response. Interleukin (IL)-10 down-regulates the Th1 response and is associated with BCG failure. In this study, we investigated whether blocking IL-10 signalling could enhance the BCG-induced Th1 response and anti-tumour immunity in a murine orthotopic tumour model. Treatment with BCG and anti-IL-10 receptor 1 monoclonal antibody (anti-IL-10R1 mAb) increased the interferon (IFN)-γ to IL-10 ratio in both splenocyte cultures and urine. Mice bearing luciferase-expressing MB49 (MB49-Luc) tumours were treated and followed for tumour growth by bioluminescent imaging, bladder weight and histology. Mice treated with phosphate-buffered saline (PBS) (group 1), BCG plus control immunoglobulin (Ig)G1 (group 2) or BCG plus anti-IL-10R1 mAb (group 3) showed 0, 6 and 22% tumour regression, respectively. The mean bladder weight of group 3 mice was substantially lower than those of groups 1 and 2 mice. Remarkably, 36% of group 1 and 53% of group 2 mice but no group 3 mice developed lung metastasis (P = 0·02). To investigate the mechanisms underlying the effect of combination therapy, splenocytes were stimulated with S12 peptide (serine mutation at codon 12 of the K-ras oncogene) known to be expressed in MB49-Luc cells. Induction of ras mutation-specific IFN-γ and cytotoxicity was observed in mice treated with combination therapy. These observations indicate that BCG, in combination with anti-IL-10R1 mAb, induces enhanced anti-tumour immunity that is protective against lung metastasis. Anti-IL-10R1 mAb demonstrates systemic effects and may prove useful in clinical practice for treating bladder cancer in high-risk patients.

Metformin inhibits urothelial tumorigenesis in the UPII-mutant Ha-ras transgenic mouse model

Background Bladder cancer occurs mainly in older people and is expensive to manage. As the population continues to age, bladder cancer may remain to be a major public health burden. Therefore, we hypothesized that an agent, like metformin, with anti-aging property could have preventive activity against the development and progression of human urinary bladder cancer. Materials and methods The UPII mutant Ha-ras transgenic mouse model mimics human papillary transitional urothelial cell carcinoma and exhibits enhanced mTOR activity in tumor tissues. Homozygous UPII mutant Ha-ras transgenic mice were identified through genotyping using the Southern blotting method. Genotyped Ha-ras mice were then fed orally with normal drinking water or 0.1% or 0.05% metformin in drinking water starting at 6 weeks of age and ending at 6 months of age. Death rate, body weight, tumor burden, and proliferative and apoptotic indices at the end of treatments will be evaluated by pathological and statistical analyses. Results About 62% of male, homozygous mutant Ha-ras transgenic mice which drank normal water died of urinary tract obstruction and hydronephrosis within 6 months of age, while only about 11% or 15% of mice which drank 0.1% or 0.05% metformin containing water died. Drinking metformin dramatically increased the survival of mutant Ha-ras tumor bearing mice by 51 to 47% (Ps<0.01). Metformin drinking also significantly decreased the mean bladder weights of male, homozygous mutant Ha-ras transgenic mice by up to 62%. Histological analysis of H&E stained bladder sections from metformin treated mice demonstrated more differentiated tumors compared to those in control groups. The in vivo mechanisms of metformin’s action are associated with anti-proliferation, reduction of phospho-mTOR and 4E-BP1 expression and induction of TSC2 expression in bladder tissues. Conclusions Our results demonstrated strong in vivo anti-urothelial tumorigenesis activity of metformin drinking in the UPIImutant H-ras model via inhibition of the mTOR pathway. These results suggested the potential of metformin in preventing recurrence of clinical bladder cancer and in improving quality of life for patients.

KAVA Chalcone, Flavokawain A, Inhibits Urothelial Tumorigenesis in the UPII-SV40T Transgenic Mouse Model

Flavokawain A (FKA) is the predominant chalcone identified from the kava plant. We have previously shown that FKA preferentially inhibits the growth of p53 defective bladder cancer cell lines. Here, we examined whether FKA could inhibit bladder cancer development and progression in vivo in the UPII-SV40T transgenic model that resembles human urothelial cell carcinoma (UCC) with defects in the p53 and the retinoblastoma (Rb) protein pathways. Genotyped UPII-SV40T mice were fed orally with vehicle control (AIN-93M) or FKA (6 g/kg food; 0.6%) for 318 days starting at 28 days of age. More than 64% of the male mice fed with FKA-containing food survived beyond 318 days of age, whereas only about 38% of the male mice fed with vehicle control food survived to that age (P = 0.0383). The mean bladder weights of surviving male transgenic mice with the control diet versus the FKA diet were 234.6 ± 72.5 versus 96.1 ± 69.4 mg (P = 0.0002). FKA was excreted primarily through the urinary tract and concentrated in the urine up to 8.4 μmol/L, averaging about 38 times (males) and 15 times (females) more concentrated than in the plasma (P = 0.0001). FKA treatment inhibited the occurrence of high-grade papillary UCC, a precursor to invasive urothelial cancer, by 42.1%. A decreased expression of Ki67, survivin, and X-linked inhibitor of apoptotic proteins (XIAP) and increased expression of p27 and DR5, and the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive apoptotic cells were observed in the urothelial tissue of FKA-fed mice. These results suggest a potential of FKA in preventing the recurrence and progression of non-muscle-invasive UCC.

Chemoprevention of BBN-Induced Bladder Carcinogenesis by the Selective Estrogen Receptor Modulator Tamoxifen

Bladder cancer is the fifth most frequent tumor in men and ninth in women in the United States. Due to a high likelihood of recurrence, effective chemoprevention is a significant unmet need. Estrogen receptors (ERs), primarily ERβ, are expressed in normal urothelium and urothelial carcinoma, and blocking ER function with selective ER modulators such as tamoxifen inhibits bladder cancer cell proliferation in vitro. Herein, the chemoprotective potential of tamoxifen was evaluated in female mice exposed to the bladder-specific carcinogen, N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Carcinogen treatment resulted in a 76% tumor incidence and increased mean bladder weights in comparison to controls. In contrast, mice receiving tamoxifen concurrent (8–20 weeks) or concurrent and subsequent (8–32 weeks) to BBN administration had no change in bladder weight and only 10% to 14% incidence of tumors. Non-muscle-invasive disease was present in animals treated with tamoxifen before (5–8 weeks) or after (20–32 weeks) BBN exposure, while incidence of muscle-invasive bladder carcinoma was reduced. ERβ was present in all mice and thus is a potential mediator of the tamoxifen chemoprotective effect. Surprisingly, ERα expression, which was detected in 74% of the mice exposed to BBN alone but not in any controlmice, was correlated with tumor incidence, indicating a possible role for this receptor in carcinogen-induced urothelial tumorigenesis. Thus, these data argue that both ERα and ERβ play a role in modulating carcinogen-induced bladder tumorigenesis. Administration of tamoxifen should be tested as a chemopreventive strategy for patients at high risk for bladder cancer recurrence.

Abstract 3709: Kawain, a kavalactone, inhibits urinary bladder carcinogenesis in the UPII-mutant Ha-ras transgenic model and associated with inhibition of the mTOR pathway.

Abstract Nonmuscle invasive bladder cancer (NIBC) is the most expensive cancer to treat because of its high rate of recurrence. Therefore, there is an urgent need to develop novel agents that can prevent NIBC recurrence and progression. Kava is a traditional drink of the Pacific Islands for thousands of years and kava drink was linked to unusually low incidences of several cancers, including lung and bladder cancer, despite a high portion of smokers in these populations. Kavalactones, including kawain, are major bioactive components in the kava extract. Given genetic alterations leading to over activity of H-Ras are predominantly found in NIBC, we examined the effect of kawain on the growth of bladder tumors and the survival of the mice in the UPII-mutant Ha-ras transgenic mouse bladder model. This model mimics human NIBC and exhibits enhanced mTOR activity in the bladder tumors. We also investigated the effect of kawain on the mTOR pathway in bladder cancer cell lines. Homozygous mutant Ha-ras transgenic mice were identified through genotyping using Southern blotting analysis. The mice were fed orally with vehicle control or 6 g kawain /kg food for about 5 months starting at 6 weeks of age. More than 68% control food- fed male mice died of ureter obstruction and hydronephrosis within 6 months of age, while more than 78% of mice fed with kawain food survived beyond 6 months of age (P=0.0082, log-rank test). The mean bladder weight of male transgenic mice in kawain treatment group decreased by about 56% compared to those fed with vehicle control diet (mean bladder weights for vehicle control versus kawain treatments are 0. 32±0.13 versus 0.14±0.01, P=0.035). In addition, kawain treatment significantly reduced the incidences of hydronephrosis and hematuria. Mouse pathology analysis revealed that vehicle control treated mice exhibited poorly differentiated transitional cell carcinoma in the bladder, while that kawain treated mice presented well differentiated tumors in the bladder. The bladder tissues from kawain treated mice showed a decreased number of Ki67 positive cells compared to those in vehicle control treated mice, which suggested that kawain inhibited cell proliferation in vivo in bladder tumors. In addition, kawain inhibited the growth of bladder cancer cell lines T24 and UMUC3 with a marked suppression of 4E-BP1 expression and induction of autophagy. The growth inhibitory effect of kawain in mouse embryonic fibroblasts (MEFs) is at least in part dependent on the existence of TSC1. This result suggested that kawain inhibits the mTOR pathway. Therefore, kawain, as a naturally-occurring mTOR inhibitor, deserves further investigation for its potential in preventing the recurrence of NIBC. Citation Format: Zhongbo Liu, Xiaolin Zi. Kawain, a kavalactone, inhibits urinary bladder carcinogenesis in the UPII-mutant Ha-ras transgenic model and associated with inhibition of the mTOR pathway. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3709. doi:10.1158/1538-7445.AM2013-3709

875 TREATMENT WITH A CB2 AGONIST DECREASES SEVERITY OF ESTABLISHED CYSTITIS

You have accessJournal of UrologyInfections/Inflammation of the Genitourinary Tract: Interstitial Cystitis1 Apr 2013875 TREATMENT WITH A CB2 AGONIST DECREASES SEVERITY OF ESTABLISHED CYSTITIS Zunyi Wang, Peiqing Wang, and Dale Bjorling Zunyi WangZunyi Wang Madison, WI More articles by this author , Peiqing WangPeiqing Wang Madison, WI More articles by this author , and Dale BjorlingDale Bjorling Madison, WI More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.444AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Painful bladder syndrome is a chronic painful disorder characterized by frequency, urgency, and pain that is estimated to affect 2.7% to 6.5% of American women. The etiology and pathogenesis of painful bladder syndrome remain unknown, and effective treatments are lacking. Cannabinoids exert potent analgesic and anti-inflammatory effects. Cannabinoid receptor 2 (CB2) is expressed in the bladder, particularly in urothelial cells, and bladder inflammation increases CB2 expression. We investigated whether treatment with the selective CB2 agonist GP1a ameliorates severity of cystitis and referred hyperalgesia after induction of cystitis by acrolein. METHODS Cystitis was induced by intravesical instillation of acrolein (0.5 mM, 200 μl) in female C57 mice. Mice were treated with GP1a (Tocris, 10 mg/kg, ip) or vehicle at 3, 24, and 30 hours after instillation of acrolein. Mice were tested for mechanical sensitivity of hind paws at 3, 24, and 48 hours, respectively. Mice were also placed on filter paper for measurement of urine spots at 24 hours. Mice were sacrificed at 48 hours, and bladders were collected, weighed, and processed for histological analysis. RESULTS Forty-eight hours after acrolein, bladders of all animals demonstrated histological evidence of inflammation, primarily characterized by submucosal edema. Mean wet bladder weight (mg/g body weight) was greater in vehicle-treated mice than that in CB2 agonist-treated animals, (2.82 ± 0.17 vs 1.92 ± 0.25, n=5, p < 0.05), and histology revealed that edema was more severe in the vehicle-treated mice. Basal mechanical sensitivity was virtually the same in vehicle- and GP1a-treated mice (2.9 ± 0.01 g). After acrolein, mechanical sensitivity threshold was 0.46 ± 0.13, 0.28 ± 0.03 and 0.54 ± 0.16 g in vehicle-treated group, and was 0.55 ± 0.1, 0.78 ± 0.09 and 1.17 ± 0.41 g in CB2 agonist-treated group at 3, 24 and 48 hours, respectively. The mechanical sensitivity was significantly less (p<0.05, n=5, at 24 and 48 hours) in vehicle-treated than CB2 agonist-treated mice. The number of small diameter urine spots was also significantly reduced; 24.8 ± 6.9 (vehicle-treated) vs 4 ± 3.3 (GP1a-treated), n=5, p < 0.05. CONCLUSIONS Treatment with a selective CB2 agonist after induction of cystitis reduced severity of acrolein-induced cystitis and inhibited bladder inflammation-induced increased peripheral sensitivity to mechanical stimuli. Our data indicate that the CB2 receptor is a potential therapeutic target for treatment of bladder inflammation and subsequent changes in pain perception. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e361 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Zunyi Wang Madison, WI More articles by this author Peiqing Wang Madison, WI More articles by this author Dale Bjorling Madison, WI More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Effects of Mirodenafil, a Phosphodiesterase-5 Inhibitor, on Female Rat Bladder in a Partial Bladder Outlet Obstruction Model: Physiological and Immunohistochemical Aspects

PurposeWe investigated the effects of mirodenafil, a phosphodiesterase-5 inhibitor developed in South Korea, on the female rat bladder in a partial bladder outlet obstruction (BOO) model.Materials and MethodsThirty-six female Sprague-Dawley rats were divided into four groups: the control group, BOO without medication group, BOO with mirodenafil 1 mg/kg group, and BOO with mirodenafil 4 mg/kg group. Mirodenafil was administered orally for 2 weeks after the induction of BOO. Two weeks after BOO, the rats in each group underwent cystometry under urethane anesthesia. After cystometry, the bladder was excised to perform immunohistochemical staining for connexin 43.ResultsThe three BOO groups showed significant increases in mean bladder weight compared with the control group. Baseline pressure, threshold pressure, and maximum contraction pressure were not significantly different between the four groups. Although the contraction interval was decreased in all BOO groups compared with the control group, it was prolonged in the two groups treated with mirodenafil compared with the untreated BOO group. In the immunohistochemical examination, connexin 43 staining intensity in the lamina propria increased in the three BOO groups compared with the control group. The two groups treated with mirodenafil, however, showed decreased connexin 43 staining compared with the untreated BOO group.ConclusionsMirodenafil may increase the contraction intervals of female rat bladders in a partial BOO model. Decreasing bladder overactivity by mirodenafil may be related to intracellular communication mechanisms involving connexin 43.

Mesenchymal Stem Cells Overexpressing Hepatocyte Growth Factor (HGF) Inhibit Collagen Deposit and Improve Bladder Function in Rat Model of Bladder Outlet Obstruction

Bladder outlet obstruction (BOO) caused by collagen deposit is one of the most common problems in elderly male. This study was performed to examine the capability of human mesenchymal stem cells (MSCs) overexpressing hepatocyte growth factor (HGF) to inhibit collagen deposition in rat model of bladder outlet obstruction (BOO). HGF is known for its antifibrotic effect and the most promising agent for treating bladder fibrosis. BM3.B10 stable immortalized human MSC line (B10) was transduced to encode human HGF with a retroviral vector was prepared (B10.HGF). Two weeks after the onset of BOO, B10, and B10.HGF cells were injected into the rat's bladder wall. After 4 weeks, bladder tissues were harvested and Masson's trichrome staining was performed. Transgene expression in HGF-expressing B10 cells was demonstrated by reverse transcriptase polymerase chain reaction and immunohistochemical staining, and the high levels of HGF secreted by B10.HGF cells was confirmed by ELISA. The mean bladder weight in BOO rats was 5.8 times of the normal controls, while in animals grafted with B10.HGF cells, the weight was down to four times of the control [90.2 ± 1.6 (control), 89.9 ± 2.8 (sham), 527.9 ± 150.9 (BOO), 447.7 ± 41.0 (BOO + B10), and 362.7 ± 113.2 (BOO + B10.HGF)]. The mean percentage of collagen area increased in BOO rats, while in the animals transplanted with B10.HGF cells, the collagen area decreased to the normal control level [12.2 ± 1.3, (control), 12.8 ± 1.1 (sham), 26.6 ± 2.7 (BOO), 19.9 ± 6.0 (BOO + B10), and 13.3 ± 2.1 (BOO + B10.HGF)]. The expression of collagen and TGF-b protein increased after BOO, while the expression of HGF and c-met protein increased in the group with B10.HGF transplantation after BOO. Intercontraction interval decreased after BOO, but it recovered after B10.HGF transplantation. Maximal voiding pressure (MVP) increased after BOO, and it recovered to levels of the normal control after transplantation of B10.HGF cells. Residual urine volume (RU) increased after BOO, but the RU increase was not reversed by transplantation of B10.HGF cells. Human MSCs overexpressing HGF inhibited collagen deposition and improved cystometric parameters in bladder outlet obstruction of rats. The present study indicates that transplantation of MSCs modified to overexpress HGF could serve as a novel therapeutic strategy against bladder fibrosis in patients with bladder outlet obstruction.

Abstract 618: Rhodiola rosea L. extract, SHR-5, and metformin exhibit potent activity against bladder carcinogenesis in the UPII-mutant Ha-ras transgenic model.

Abstract The incidence of human urinary bladder cancer increases markedly with age, suggesting a mechanistic connection between aging and bladder carcinogenesis and a potential use of anti-aging agents in bladder cancer chemoprevention. Rhodiola rosea, growing in high altitude or cold regions of the world, has been reported to have anti-aging effects in Drosophila. Metformin, a popular anti-diabetics drug, has also shown anti-aging effect in mice. In addition, both agents have been shown to inhibit the mTOR pathway. Therefore, we examined the effect of SHR-5 and Metformin on bladder carcinogenesis in the UPII-mutant Ha-ras transgenic mouse bladder model. This model mimics human papillary transitional cell carcinoma and exhibits enhanced mTOR activity in the bladder tumors. The homozygous mutant Ha-ras transgenic mice were identified through genotyping using Southern blotting analysis. 0.625% SHR-5 or 0.1% Metformin in drinking water was given to the mice starting at 6 weeks of age and ending at 6 months of age. About 42% of homozygous mutant Ha-ras male transgenic mice which drank normal water died of urinary tract obstruction and hydronephrosis within 6 months of age, while more than 93% and 89% of mice which drank SHR-5 or Metformin containing water survived. The mean bladder weights in male mice drinking SHR-5 or Metformin decreased by 69% and 62%, respectively, compared to control group (Bladder weights for mice drinking normal water versus SHR-5 or Metformin are 0.3382± 0.061versus 0.1278± 0.02 or 0.1053±0.015 gram, P=0.0197 or 0.0082). The mean bladder weights in female mice which drank SHR-5 or Metformin containing water also decreased by about 40% or 39.8 %, respectively, compared to those which drank normal water (Bladder weights for normal water versus SHR-5 or Metformin: 0.07149 ±0.0036 versus 0.04301 ±0.0032 or 0.04295±0.0115 gram, P&amp;lt;0.0001 or =0.0007). Chemstrip tests showed that mice which drank SHR-5 or metformin water developed less hematurine than those which drank normal water. Histological analysis of H&amp;E staining bladder sections from SHR-5 and Metformin treated mice demonstrated more differentiated tumors compared to those from the transgenic mice which drank normal water. Immunohistochemistry staining revealed that SHR-5 and Metformin treatments decreased cell proliferation in the urothelium. Taken together, our results suggest a potential new method for bladder cancer prevention by drinking SHR-5 or Metformin. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 618. doi:1538-7445.AM2012-618

Abstract 619: Flavokawain A inhibits urinary bladder carcinogenesis in the UPII-SV40T transgenic mouse bladder cancer model.

Abstract The UPII-SV40T transgenic model of the mouse bladder reproduces a precursor to recurrent and invasive Transitional Cell Carcinoma (TCC): carcinoma in situ (CIS). This model therefore is useful for testing the potential of agents for preventing bladder cancer recurrence. Flavokawain A is the predominant chalcone identified from the kava plant. We have recently shown that flavokawain A exhibited strong growth inhibitory effects on human bladder cancer cell lines both in vitro in cell cultures and in vivo in a xenograft mouse model. In this study, we examine whether flavokawain A can also inhibit bladder cancer development in the UPII-SV40T transgenic model. The transgenic mice were identified through genotyping using Southern blotting analysis. Both heterozygous and homozygous UPII-SV40T mice were fed orally with vehicle control or 6 g flavokawain A /kg food for 9 months starting at 6 weeks of age. More than 54% control food- fed male mice died of ureter obstruction and nephrohydrosis within 10.5 months of age, while more than 67% of mice fed with 6g flavokawain A /kg food survived beyond 10.5 months of age. The mean bladder weight of male transgenic mice that were fed with 6g flavokawain /kg food for 9 months decreased by about 46% compared to those fed with vehicle control diet (mean bladder weights for vehicle control versus flavokawain A treatments are 0.2031±0.016 versus 0.1108±0.02565, P=0.0037). The mean bladder weights of female transgenic mice that were fed with 6g flavokawain /kg food for 9 months decreased by 31% compared to those fed with vehicle control diet (mean bladder weights for vehicle control versus flavokawain A treatments are 0.0466±0.0052 versus 0.03225±0.004689, p=0.0556). Mouse pathology analysis revealed that vehicle control treated mice exhibited poorly differentiated transitional cell carcinoma in the bladder, while that flavokawain A treated mice presented well differentiated tumors in the bladder. The bladder tissues from flavokawain A treated mice showed a decreased number of Ki67 positive cells compared to those in vehicle control treated mice, which suggested that flavokawain A inhibited cell proliferation in vivo in bladder tumors. These results suggested a potential of flavokawain A in preventing the recurrence of non-muscle invasive urinary bladder cancer by treating CIS. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 619. doi:1538-7445.AM2012-619

Angiotensin II type 1 (AT‐1) receptor inhibition partially prevents the urodynamic and detrusor changes associated with bladder outlet obstruction: a mouse model

Study Type - Therapy (case control) Level of Evidence 3b What's known on the subject? and What does the study add? Angiotensin II is the main effector peptide in the bladder local renin-angiotensin system. This experiment demonstrates the role of this local renin-angiotensin system with respect to bladder outlet obstruction. • To determine if treatment with an angiotensin II type 1 (AT-1) receptor antagonist, losartan, can prevent the structural and functional changes that occur in a mouse model of bladder outlet obstruction (BOO). • Twenty-four Balb/CAN mice underwent partial urethral obstruction for 6 weeks. • Twelve mice were given oral losartan (10 mg/kg/day), and 12 were not. Six mice served as unobstructed controls, and six unobstructed mice were given oral losartan (10 mg/kg/day) to determine the effect of angiotensin II inhibition on the normal bladder. • Bladder capacity (C), detrusor pressure during voiding (Pdet) and volume at first non-voiding contraction (NVC1) as a percentage of C were recorded after 6 weeks. • Bladders were stained with haematoxylin and eosin for measurement of detrusor muscular thickness, and graded as 1 = atrophy (<100 µm thick), 2 = normal (100-200 µm thick), 3 = hypertrophy (>200 µm thick) compared with controls. • Compared with controls, BOO mice had greater C (153.5 ± 20.9 vs 57.5 ± 7.4 µl, P < 0.01), higher Pdet (28.8 ± 2.1 vs 12.1 ± 2.1 mm Hg), lower NVC1 (median = 24% vs 54% P= 0.03). BOO mice manifested greater bladder weight (93.2 ± 11.7 mg vs 26.8 ± 2.40 mg, P < 0.01) and greater detrusor muscle thickness (median 3 vs 2, P= 0.02). • Compared with untreated BOO mice, mice treated with losartan had greater mean C (248.8 ± 28.6 vs 153.5 ± 20.9 µL, P= 0.01), no significant change in mean Pdet (24.7 ± 1.6 vs 28.8 ± 2.1 mm Hg, P= 0.2) and a higher mean NVC1 (47% vs 24%, P= 0.02). • Treatment with losartan mediated an insignificant reduction in mean bladder weight (68.1 ± 9.1 mg vs 93.2 ± 11.7 mg, P= 0.10), but a significant reduction in detrusor muscle thickness (median 2 vs 3, P= 0.02). Losartan did not mediate any significant structural or functional changes in the unobstructed mouse bladder. • In a mouse model of BOO, treatment with an AT-1 antagonist partially prevented the urodynamic and structural changes that otherwise occur with BOO.

Abstract 855: Flavokawain A inhibits urinary bladder cancer in UPII-mutant Ha-ras transgenic mouse bladder cancer model

Abstract Flavokawain A is the predominant chalcone identified from kava extract. We have recently shown that flavokawain A exhibited strong growth inhibitory effects on human bladder cancer cells both in vitro in cell cultures and in vivo in a xenograft mouse model. To examine whether flavokawain A affects specific-stages or course of bladder cancer development, the UPII-mutant Ha-ras transgenic mouse bladder model that mimics human papillary transitional cell carcinoma was used. The homozygous mutant Ha-ras transgenic mice were identified through genotyping using Southern blotting analysis and fed orally with 1g flavokawain A/kg food or 6 g flavokawain A /kg food for 3 or 6 months starting at 6 weeks of age. After treatments, organs are collected and weighted. H&amp;E staining and immunohistochemstry were used to evaluate bladder cancer pathology and expression of biomarkers. All control food- fed male mice died of ureter obstruction and nephrohydrosis within 6 months of treatment, while more than 50% of mice fed with 6g flavokawain A /kg food for 6 months survived beyond the 6 months of treatment. The mean bladder weight in 6g flavokawain /kg food fed male mice also decreased by 66% compared to control treated mice for 3 months. Mouse pathology analysis showed that control mice exhibited poorly differentiated transitional cell carcinoma, while that flavokawain A treated mice presented well differentiated tumors. Compared to the dose of 6g/kg, a 1g/kg dose of flavokawain A is less effective in inhibition of bladder cancer development in this transgenic model. In addition, immunohistochemistry staining of Skp2 revealed that Skp2 oncoprotein expression in urothelium was also significantly decreased in flavorkawain A treatment group compared to control group. Taken together, our results suggest the potential usefulness of flavokawain A in treatment of superficial urinary bladder cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 855. doi:10.1158/1538-7445.AM2011-855