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Keywords:

  • EP4 receptor;
  • cyclophosphamide;
  • overactive bladder;
  • cystitis

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

Study Type – Aetiology (case control)

Level of Evidence 3b

What's known on the subject? and What does the study add?

Recent evidence has suggested that up-regulation of the prostaglandin E2 (PGE2) receptor subtype 4 (EP4) receptor in the bladder is involved in bladder overactivity.

The present study found that MF191, a selective EP4 receptor antagonist, may have effects on the bladder urothelium and inflammatory cells and suppress CYP- or PGE2-induced bladder overactivity. Systemic or intravesical MF191 administration for the treatment of overactive bladder may merit clinical study.

OBJECTIVE

  • • 
    To investigate the mechanisms and urodynamic effects of a potent and selective prostaglandin E2 (PGE2) receptor subtype 4 (EP4) antagonist, MF191, on cyclophosphamide (CYP) or PGE2-induced bladder overactivity in rats.

MATERIALS AND METHODS

  • • 
    Experimental and control rats were injected with CYP (200 mg/kg i.p.) or saline on day 1. Continuous cystometrogram (CMGs) were performed on day 3.
  • • 
    In group 1, MF191 (vehicle 0.1 and 1 mg/kg) was given i.v. The bladder was then harvested for histology and immunohistochemistry. Some bladders were harvested for analysis of EP4 expression by Western blotting without a CMG study.
  • • 
    In group 2, MF191 (vehicle 10 nM, and 100 nM) was continuously infused into the bladder.
  • • 
    In group 3, bladder overactivity was induced by intravesical instillation of PGE2 (200 uM) and vehicle or MF191 (1 mg/kg) was given i.v.

RESULTS

  • • 
    CYP induced bladder inflammation, overactivity and EP4 upregulation. The CYP effects were suppressed by MF191 (1 mg/kg i.v.; intercontraction interval [ICI]: 39.4% increase, and reduced inflammatory cells infiltration, and EP4 expression).
  • • 
    Intravesical instillation of MF191 (100 nM) suppressed CYP-induced bladder overactivity (ICI: 71.8% increase).
  • • 
    PGE2-induced bladder overactivity was suppressed by MF191 (ICI: 43.2% increase).
  • • 
    MF191 had no significant effects on other CMG variables or on control rats.

CONCLUSIONS

  • • 
    MF191 might affect the bladder urothelium and inflammatory cells and suppresses CYP- or PGE2-induced bladder overactivity.
  • • 
    Systemic or intravesical MF191 administration for the treatment of overactive bladder merits clinical study.

Abbreviations
PGE2

prostaglandin E2

CYP

cyclophosphamide

CMG

cystometrogram

PT

pressure threshold

PB

pressure baseline

ICI

intercontraction interval

INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

Prostaglandin E2 (PGE2) synthesized in bladder muscle and mucosa has a complex local action in the bladder. PGE2 affects the micturition reflex under both normal and pathophysiological conditions (e.g. mucosa injury and inflammatory mediators) [1–3]. Intravesical administration of PGE2 stimulates reflex micturition through the activation of capsaicin-sensitive afferent nerves and causes bladder overactivity in rats and humans [4–6]. Furthermore, urinary PGE2 has been reported to be increased in rats with cyclophosphamide (CYP)-induced bladder inflammation and overactivity [7,8].

The effects of PGE2 are mediated by G-protein-coupled EP receptors (EP1–EP4). The differences in the functions and locations of EP receptors, and the conditions of the subjects (i.e. normal vs bladder overactivity) determine the influence of different EP receptors in micturition processing. Rahnama'i et al. [1] showed EP1 and EP2 expression in the bladder mucosa and suburothelium in the guinea pig bladder. EP1 receptor antagonists have been shown to inhibit bladder overactivity induced by spinal cord injury or BOO [5,9]. Su et al. [10] reported that intrathecal administration EP3 antagonist produced a long-lasting and robust inhibition of the visceromotor reflex response to urinary bladder distension, and suggested that centrally acting EP3 receptor antagonists may be useful in the control of detrusor overactivity. Previous studies have also shown increased expression of EP4 receptor in the bladder and spinal cord in rats with CYP-induced bladder overactivity [11], and in the bladder in patients with overactive bladder induced by BOO [12].

Lack of selective antagonists has been one of the major hurdles for elucidating the role of various EP receptors in the micturition reflex. In recent years, selective antagonists for various EP receptors have been developed. In the present study, we therefore used a selective EP4 antagonist, MF191, 4-{(1S)-1-[({2,5-Dimethyl-4-[4-(trifluoromethyl)benzyl]-3-thienyl}-carbonyl)-amino]ethyl}benzoic acid (28d) [13], to determine more precisely the role of EP4 receptors in the modulation of the micturition reflex. MF191 is reportedly very potent at binding to their target receptors, with Ki values in the nanomolar range, and is highly selective over other prostanoid receptors [13].

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

All experimental procedures were performed in 52 female Sprague–Dawley rats (220–280 g; 12 rats for Experiment 1, 12 rats for experiment two, 10 rats for Experiment 3, six new rats plus 12 used rats from Experiment 1 for Experiment 4 and 12 rats for Experiment 5). The study was reviewed and approved by the Institutional Animal Care and Use Committee before the study began. In Experiment 1, a cystometrogram (CMG) study, control (n= 6) or CYP-treated (n= 6) rats received i.v. MF191. In Experiment 2, a CMG study, control (n= 6) or CYP-treated (n= 6) rats received intravesical instillation of MF191. In Experiment 3, a CMG study, rats treated with intravesical instillation of PGE2 received i.v. MF191 (n= 6) or vehicle (n= 4). In Experiment 4, a histological study, all of the 12 rats from Experiment 1 and an additional six CYP-treated rats were used. In Experiment 5, Western blot analysis, control rats, CYP plus vehicle-treated rats, and CYP plus MF191-treated (1 mg/kg, i.v.) rats were used for western blotting (n= 4, for each group).

CYP INJECTION

Bladder overactivity was induced by i.p. injection of CYP, which is metabolized to acrolein, an irritant eliminated in the urine [11]. CYP (200 mg/kg; i.p.) or a corresponding volume of saline was injected on day 1.

CATHETER IMPLANTATION

On day 3, animals were anaesthetized with s.c. injection of urethane (1.2 g/kg). PE-50 tubing filled with heparinized saline (100 IU/mL) was inserted into the femoral vein in some animals for drug administration.

CMG STUDY

After i.v. catheterization, another PE-50 tubing was inserted into the bladder through the urethra and connected via a three-way stopcock to a pressure transducer and to a syringe pump for recording intravesical pressure and for infusing solutions into the bladder. Baseline CMGs were performed by filling the bladder with saline (0.08 mL/min) to elicit repetitive voiding after a 60-min equilibration period with a stable CMG pattern had been achieved. The amplitude (the peak pressure minus the basal pressure during each contraction period), pressure threshold (PT; the pressure immediately before the reflex contraction), pressure baseline (PB; the lowest bladder pressure during filling) and intercontraction interval (ICI; the time between contractions) of reflex bladder contractions were recorded. Measurements in each animal represented the mean of 3–5 bladder contractions.

MF191 ADMINISTRATION

MF191 was a gift from Merck Frost Canada Ltd. A 20 mg/mL stock solution of MF191 was made in DMSO and subsequent drug dilution was made in saline on the day of experiment. Cumulative doses of MF191 (vehicle, 0.1 and 1.0 mg/kg; Merck Frost Canada Ltd., Kirdland, QC, Canada) was injected i.v. after obtaining baseline CMGs and the effects of the drug were monitored for 60 min. Sequential doses of MF191 (vehicle, 10 nM, 100 nM) were infused intravesically for 60 min after obtaining baseline CMGs.

TRANSCARDIAC PERFUSION

After CMGs had been obtained, rats that received i.v. MF191 treatment were anaesthetized and killed via transcardiac perfusion, with Krebs buffer followed by 4% paraformaldehyde fixative. The rats were then dissected to harvest the bladder. Another six rats that received CYP and i.v. vehicle treatment without CMG were used as positive controls.

HISTOLOGY

The bladder was fixed in 10% buffered formaldehyde for 24–48 h, and then embedded in paraffin. The bladder tissue for histology was cut into 3-µm thick pieces and stained with haematoxylin and eosin.

INTRAVESICAL PGE2 AND I.V. MF191 ADMINISTRATION

Stock solutions (0.01 M) of PGE2 (Sigma®, St. Louis, MO, USA) were made in absolute ethanol and then diluted in saline (200 uM) just before use. PGE2 was infused intravesically after baseline CMGs had been obtained. MF191 (1.0 mg/kg) or vehicle was given i.v. after CMGs with 3–5 voiding contractions during intravesical PGE2 application had been obtained .

WESTERN BLOT ANALYSIS FOR EP4 EXPRESSION

On day 3, the rats were anaesthetized and killed, 2 h after receiving vehicle or MF191 treatment, and the control rats were also anaesthetized and killed. The bladder was harvested for Western blot analysis of EP4 expression according to the standard protocol (Amersham Biosciences, Little Chalfont, UK) and our previous study [11]. The samples were homogenized in protein extraction solution (T-PER; Pierce Biotechnology, Rockford, IL, USA) before sonication and purification. The amount of total protein was measured with the Bradford protein assay method (Bio-Rad Laboratories, Hercules, CA, USA). SDS-PAGE was performed using the Laemmli buffer system. Briefly, an aliquot of the extracts equivalent to 30 µg protein was loaded onto 8% polyacrylamide gel, electrophoresed at a constant voltage of 100 V for 1 h and transferred to Hybond-P PVDF Membrane (Amersham Biosciences). The membrane was blocked with blocking agent and then immunoblotted overnight at +4 °C mouse anti-actin monoclonal antibody (Chemicon, CA, USA, 1:10 000 dilution) and anti-EP4 receptor antibody (rabbit anti-EP4 receptor polyclonal antibody, Cayman, Ann Arbor, MI, USA 1:500 dilution). After washing, the membrane was incubated with secondary antibody using 5% defatted milk powder in TBS for 2 h at room temperature using a horseradish peroxidase-linked anti-rabbit or antimouse immuglobulin G. Western blots were visualized using an enhanced chemiluminescence detection system (Amersham Biosciences). The amount of β-actin was also detected as the internal control. Quantitative analysis was done using LabWorks Image Acquisition and Analysis software.

STATISTICAL ANALYSIS

Data are expressed as sem values. We compared differences between multiple groups using anova, followed by the Tukey multiple comparisons test. Two-group analysis was done using the paired or unpaired t-test, as appropriate. Serial studies were tested using repeated measures anova with a P value <0.05 considered to indicate statistical significance.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

CMG RESPONSE TO CYP AND I.V. MF191

CYP treatment induced an increase in contraction amplitude by 108.6% (from 41.9 [4.7] to 87.4 [5.6] cmH2O) and a decrease in ICI by 38.9% (from 10.8 [1.1] to 6.6 [1.1] min) compared with the control (saline injection) group (Table 1 and Fig. 1). These CMG results reflect bladder overactivity induced by CYP injection.

Table 1. Effects of i.v. administration of MF191 on CMG variables in CYP- and saline-treated (control) rats
VariableICI, minAmplitude, cmH2OBP, cmH2OPT, cmH2O
  1. Values are mean (sem). *P < 0.01 vs baseline; **P < 0.001 vs vehicle, repeated measures anova followed by Tukey multiple comparison test for intragroup test; baseline ICI, control vs CYP-treated, P= 0.027, unpaired t-test; baseline amplitude, control vs CYP-treated, P < 0.001, unpaired t-test.

Control (n= 6)    
 Baseline10.8 (1.1)41.9 (4.7)4.1 (0.8)9.2 (0.9)
 Vehicle11.7 (1.2)39.5 (3.7)4.4 (0.7)8.1 (0.6)
 MF191 0.1 mg/kg10.3 (1.1)36.8 (4.8)4.6 (0.7)7.5 (0.8)
 MF191 1 mg/kg12.1 (0.8)34.6 (4.1)5.4 (0.7)8.0 (0.5)
CYP-treated (n= 6)    
 Baseline6.6 (1.1)87.4 (5.6)4.5 (0.4)9.4 (0.8)
 Vehicle6.0 (0.9)75.9 (8.8)4.6 (0.2)9.3 (0.6)
 MF191 0.1 mg/kg7.8 (1.5)64.6 (6.0)3.8 (0.2)8.8 (0.7)
 MF191 1 mg/kg9.2 (1.4)*,**64.7 (13.3)4.0 (0.4)9.0 (1.2)
image

Figure 1. Representative traces of continuous CMGs in urethane-anaesthetized rats. CMGs were performed in a control rat (A) with a normal voiding pattern and a rat with CYP-induced bladder overactivity (B) before and after i.v. administration of MF191 (vehicle, 0.1 mg/kg and 1.0 mg/kg).

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Cumulative administration of MF191 (0.1 mg/kg and 1.0 mg/kg, i.v.) did not result in significant changes in the CMG variables in the control rats (Fig. 1A). However, the doses of MF191 (1.0 mg/kg) significantly increased ICI; it increased by 39.4% (from 6.6 [1.1] to 9.2 [1.4] min) compared with the pre-drug control value (P < 0.01) in the CYP-treated group (Table 1 and Fig. 1B). MF191 did not change other CMG variables. These results indicate the effect of MF191 on CYP-induced bladder overactivity.

CMG RESPONSE TO CYP AND INTRAVESICAL MF191

In a separate group of rats, CYP treatment induced an increase in contraction amplitude by 77.4% (from 28.3 [2.1] to 50.2 [3.9] cmH2O ) and a decrease in ICI by 45.0% (from 12.9 [1.1] to 7.1 [0.8] min) compared with the control (saline injection) group (Table 2 and Fig. 2). Administration of MF191 (10 nM and 100 nM intravesically) did not result in a significant change in CMG variables in control rats (Fig. 2A). However, the dose of MF191 at 100 nM significantly increased ICI by 71.8% (from 7.1 [0.8] to 12.2 [1.3] min) compared with the pre-drug control value (P < 0.05) in the CYP-treated group (Fig. 2B). MF191 did not change any other CMG variable. Changing the sequence of drug administration (MF191 from 100 nM to 10 nM, n= 2) did not alter the results and showed MF191 100 nM still had a greater effect (ICI: 73.8% increase) than 10 nM (ICI: 28.5% increase).

Table 2. Effects of intravesical administration of MF191 on CMG variables in CYP-treated rats
VariableICI, minAmplitude, cmH2OBP, cmH2OPT, cmH2O
  1. Values are mean (sem). *P < 0.01 vs baseline; **P < 0.05 vs vehicle, repeated measures anova followed by Tukey multiple comparison test for intragroup test; baseline ICI, control vs CYP-treated, P= 0.0018, unpaired t-test; baseline amplitude, control vs CYP-treated, P= 0.0006, unpaired t-test.

Control (n= 6)    
 Baseline12.9 (1.1)28.3 (2.1)5.2 (1.0)8.1 (1.2)
 Vehicle13.7 (1.2)29.2 (1.3)5.1 (1.0)6.0 (1.8)
 MF191 10 nM13.0 (1.5)27.6 (1.2)5.4 (0.9)6.2 (1.7)
 MF191 100 nM14.2 (1.6)24.4 (1.5)5.7 (0.8)6.1 (1.5)
CYP-treated (n= 6)    
 Baseline7.1 (0.8)50.2 (3.9)6.4 (1.4)9.8 (2.2)
 Vehicle7.7 (0.8)40.8 (2.6)4.7 (1.4)8.6 (2.9)
 MF191 10 nM9.9 (1.6)37.7 (4.1)4.5 (0.9)9.3 (2.4)
 MF191 100 nM12.2 (1.3)*,**39.2 (5.6)4.6 (1.1)8.3 (1.9)
image

Figure 2. Representative traces of continuous CMGs in urethane-anaesthetized rats. CMGs were performed in a control rat (A) with a normal voiding pattern and a rat with CYP-induced bladder overactivity (B) before and after intravesical administration of MF191 (vehicle, 10 nM and 100 nM).

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CMG RESPONSE TO INTRAVESICAL PGE2 AND I.V. MF191

Intravesical PGE2 induced an increase in PT by 26.0% (from 5.0 [0.9] to 6.3 [0.8] cmH2O) and a decrease in ICI by 61.4% (from 11.4 [1.5] to 4.4 [0.6] min) compared with the baseline (saline instillation [Table 3 and Fig. 3B]). MF191 partially reversed the PGE2 effect by increasing ICI by 43.2% (from 4.4 [0.6] to 6.3 [0.7] min). These results indicate that PGE2-induced bladder overactivity is mediated at least in part by activation of EP4 receptors. A further increase in PT after MF191 administration may be attributable to the incremental effect of PGE2 on PT with a longer duration of exposure to PGE2. . A previous study also showed intravesical PGE2 induced an increase in PT [5].

Table 3. Effects of intravesical administration of PGE2 on CMG variables in control (vehicle-treated) and MF191-treated (1 mg/kg, i.v.) rats
VariableICI, minAmplitude, cmH2OBP, cmH2OPT, cmH2O
  1. Values are mean (sem). P values in the control group, paired t-test. Baseline vs PGE2: ICI 0.0058, Amplitude 0.2765, BP 0.2985, PT 0.0243. PGE2 vs vehicle: ICI 0.6376, Amplitude 0.4327, BP 0.4639, PT 0.3272. P values in the MF191 group, paired t-test. Baseline vs PGE2: ICI 0.0043, Amplitude 0.4456, BP 0.1307, PT 0.0193. PGE2 vs MF191: ICI 0.0019, Amplitude 0.2625, BP 0.2739, PT < 0.0001.

Control (n= 4)    
 Baseline13.7 (1.5)35.9 (10.8)2.1 (0.4)5.0 (0.5)
 PGE24.9 (1.1)40.0 (13.0)2.8 (0.2)6.1 (0.7)
 ehicle5.0 (1.0)37.6 (10.4)3.0 (0.3)6.8 (0.5)
MF191 (n= 6)    
 Baseline11.4 (1.5)44.0 (10.6)2.0 (0.2)5.0 (0.9)
 PGE24.4 (0.6)48.9 (10.6)2.5 (0.2)6.3 (0.8)
 MF191 1 mg/kg6.3 (0.7)46.0 (10.9)2.7 (0.4)8.5 (0.9)
image

Figure 3. Representative traces of continuous CMGs in urethane-anaesthetized rats. CMGs were performed in rats with intravesical PGE2-induced bladder overactivity and received i.v. administration of vehicle (A) or MF191 (1.0 mg/kg) (B).

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HISTOLOGICAL RESPONSE TO CYP AND MF191 TREATMENT

Photomicrographs of bladder sections showed mucosal oedematous change (marked thickening of the suburothelial layer), and accumulation of inflammatory cells in the CYP-treated rat (Fig. 4B) compared with the control rat (Fig. 4A). I.v. MF191 acutely halted the infiltration of inflammatory cells, but was not able to block the oedematous changes (Fig. 4C).

image

Figure 4. Photomicrographs of bladder sections. (A) Control rats; (B) CYP-treated rats; (C) CYP-treated rats with Intravenous MF191 administration. Control rats showed intact urothelium (A), no evidence of inflammation. CYP-treated rats had inflammatory cells accumulation (B, arrow; inlet), and oedematous change. In CYP-treated rats with i.v. MF191 administration, MF191 did not alter the oedematous change, but decreased the inflammatory cells infiltration (C). Magnification X 100.

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EP4 EXPRESSION ON CYP AND MF191 TREATMENT

As shown in Fig. 5, Western blotting showed that EP4 protein level was increased 4.68-fold after CYP treatment (mean EP4 protein level, control rats: 0.19 [0.02], CYP-treated rats: 0.89 [0.03], P < 0.001), and the CYP effect was suppressed by MF191 (mean EP4 protein level: 0.09 [0.02], P < 0.001 compared with the CYP-treated group).

image

Figure 5. Western blotting for detecting EP4 expression in the bladder. On day 3, the protein amount of EP4 was significantly increased in the CYP-treated rat (central) than the control rat (left), and these changes were significantly reduced in the MF191-treated rat (right).

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

In the present study, we showed that treatment with i.v. or intravesical MF191 significantly increased ICI compared with the pre-drug control value in CYP-treated rats, but had no significant effects on the other CMG variables. MF191 also increased ICI, which was decreased by intravesical PGE2 administration. These results indicate the effect of systemic or local administration of MF191 on CYP or PGE2-induced bladder overactivity. Furthermore, MF191 suppressed CYP-induced inflammatory cell infiltration and decreased EP4 expression. These results indicated the effect of MF191 on CYP-induced bladder inflammation.

It has been shown that activation of EP4 receptor activates adenylyl cyclase and increases the level of inctracellular cAMP, which might induce excitable effects on the peripheral nerves and a change in ion channel permeability, resulting in afferent hyperexcitability [14]. Lin et al. [15] reported that AH23848, an EP4 antagonist, attenuates PGE2-mediated sensitization of capsaicin-induced currents in cultured DRG neurons. Chuang et al. [16] showed that acute CYP treatment increased EP4 expression in the bladder mucosa region and submucosal inflammatory cells and induced bladder overactivity, which was partially suppressed by i.v. injection of AH23848. Ma et al. [17] showed that a selective EP4 antagonist, L-161982, was able to suppress interleukin-6 release from invading macrophage with EP4 up-regulation in a partial sciatic nerve ligation model of rats. Up-regulation of EP4 receptors in the nervous system, bladder urothelium and immune cells may therefore be involved in the hyperactive neuroimmune interaction, resulting in neurogenic inflammation and afferent hypersensitivity, which can be suppressed by EP4 antagonists.

It has been suggested that PGE2 increases TRPV1 activity via EP1 receptor activation in a protein kinase C-dependent manner as well as through protein kinase A via EP4 [18]. Lee et al. showed that EP1 receptor antagonist suppressed intravesical PGE2-induced detrusor overactivity [4]. The present study showed the similar effects of an EP4 receptor antagonist, MF191, on intravesical PGE2-induced bladder overactivity. These results suggest that both EP1 and EP4 receptors contribute to the sensitization of bladder afferent nerves and PGE2-induced bladder overactivity. Our previous study showed that CYP treatment increased EP4 expression in the bladder mucosa [11,16], which was in line with the present finding that local treatment with intravesical MF191 can suppress CYP-induced bladder overactivity. Thus, the site of drug action could be at the urothelium because i.v. or intravesical treatments had similar effects. The present study also showed that MF191 can induce down-regulation of EP4 receptor in association with suppression of inflammatory cell infiltration. Previous studies have shown up-regulation of EP4 rceptor in the inflammatory cells and macrophages [16,17]; therefore, MF191 effects on suppression of EP4 expression could be attributable to suppression of inflammatory cell infiltration.

Furthermore, i.v. MF191 administration did not significantly change the CYP-induced bladder oedematous reaction, but significantly decreased the inflammatory cell infiltration and increased ICI. Thus, we suggest that MF191 acute injection may modulate neuroimmuno-interaction, and partially inhibit bladder inflammation and afferent nerves. These discrepancies after EP4 antagonist treatment suggest that the full recovery of bladder inflammatory reaction may need repeated doses to require a certain level of MF191 to show its effect on later stages of inflammation. However, we can't exclude the contribution of other receptors or other combinations of EP receptor subtypes in mediating the CYP effects.

In addition, MF191 had no significant effect on the micturition reflex in the control group. These characteristics of MF191, which suppressed bladder overactivity without affecting normal physiological function, imply that MF191 may be useful in the control of overactive bladder without compromising normal micturition.

Prostanoids, PGE2 in particular, have been implicated as an endogenous mediator of bladder function in the normal physiological state and under pathophysiological conditions through activation of four EP receptor subtypes at peripheral and/or central levels. In different animal models and different bladder conditions, the only role of EP receptors may be in affecting the micturition reflex, alone or in combination. It is plausible that a selective EP4 receptor antagonist, like MF191, which is a high-affinity, full antagonist against the EP4 receptor with a Ki of 1.4 nM and an IC50 of 4.5 nM [13], may ameliorate the symptoms of overactive bladder without the potential cardiovascular side effects observed with NSAID and COX2 inhibitors.

The present study has some limitations. First, the model of CYP cystitis is not perfect, given the known inflammatory effect of this compound and resultant bladder overactivity. The CYP model of bladder overactivity may be a good method of studying chemical inflammatory mechanisms [19], but is probably not a good translational model because chemical cystitis is uncommon. Second, the study's main deficiency is that it lacks a photomicrograph showing actual EP4 staining and a reduction in staining after MF191 treatment. Further study with more persuasive immunohistochemical figures to allow identification of specific bladder wall regions, which show changes in EP4 expression following CYP or PGE2 and treatment with MF191, might provide more informative data on the mechanism of MF191. Third, the study lacks measurement of non-voiding contractions. Further study in another bladder overactivity model with more chronic change, such as a BOO model, and measurement of non-voiding contractions, might provide more insight into the role of EP4 receptor or MF191 in the bladder overactivity.

In conclusion, EP4 receptor antagonist MF191 may have effects on the bladder urothelium, inhibition of inflammatory cell infiltration and suppressed CYP or PGE2-induced bladder overactivity. Systemic or intravesical MF191 administration merits clinical study for treatment of overactive bladder in humans; however, caution should be exerted when predicting what will be effective clinically.

ACKNOWLEDGEMENTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

Funding support: National Science Council Taiwan 97-2314-B-182A-076-MY3, 100-2314-B-182A-021-MY3; Kaohsiung Chang Gung Memorial Hospital CMRPG 891461.

MF191 was a gift from Merck Frost Canada Ltd.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES