The effects of the c-kit blocker glivec on the contractile response of urinary bladder.

OBJECTIVES: To investigate the effects of imatinib (Glivec) on the urinary bladder contraction and excitation induced by neurostimulation, therefore to clarify the relationships between the bladder interstitial cell of Cajal (ICC) and the neural signals. METHODS: In in vivo experiments, pelvic nerves of rats were stimulated by square-wave pulses. The contractile response was recorded before and 40 min after the administration of medications (atropine, Glivec, and ketotifen). In in vitro experiments, the bladder contractile response induced by acetylcholine with or without Glivec was evaluated. The space relationship between ICC and neural fibers were observed with double-labeled fluorescence using primary antibodies (anti-c-kit and anti-vesicular acetylcholine transferase) and secondary fluorescent antibodies (Alexa 488 and Alexa 594; Molecular Probes, Eugene, OR). RESULTS: Atropine and Glivec could significantly inhibit the bladder contractile response induced by the electrical stimulation in a dose-dependent manner, while ketotifen did not obviously affect bladder contractile response. In in vitro experiments, Glivec did not affect acetylcholine-induced bladder contractile response. The location of ICC in close proximity to cholinergic nerve fibers was confirmed by double-labeled fluorescence. CONCLUSIONS: Bladder ICC play an important role as intermediaries in the transmission of cholinergic signals from nerve to smooth muscle cells.

Operative techniques of anastomotic posterior urethroplasty for traumatic posterior urethral strictures.

OBJECTIVE: To elucidate the details of operative technique of anastomotic posterior urethroplasty for traumatic posterior urethral strictures in attempt to offer a successful result. METHODS: We reviewed the clinical data of 106 patients who had undergone anastomotic repair for posterior urethral strictures following traumatic pelvic fracture between 1979 and 2004. Patients'age ranged from 8 to 53 years (mean 27 years). Surgical repair was performed via perinea in 72 patients, modified transperineal repair in 5 and perineoabdominal repair in 29. Follow-up ranged from 1 to 23 years (mean 8 years). RESULTS: Among the 77 patients treated by perineal approaches, 69 (95.8%) were successfully repaired and 27 out of the 29 patients (93.1%) who were repaired by perineoabdominal protocols were successful. The successful results have sustained as long as 23 years in some cases. Urinary incontinence did not happen in any patients while impotence occurred as a result of the anastomotic surgery. CONCLUSIONS: Three important skills or principles will ensure a successful outcome, namely complete excision of scar tissues, a completely normal mucosa ready for anastomosis at both ends of the urethra, and a tension-free anastomosis. When the urethral stricture is below 2.5 cm long, restoration of urethral continuity can be accomplished by a perineal procedure. If the stricture is over 2.5 cm long, a modified perineal or transpubic perineoabdominal procedure should be used. In the presence of a competent bladder neck, anastomotic surgery does not result in urinary incontinence. Impotence is usually related to the original trauma and rarely (5.7%) to urethroplasty.

[Inhibition on phenotypic transformation of detrusor smooth muscle cells after bladder outlet obstruction by E2F decoy strategy].

OBJECTIVE: To investigate the inhibitory effect of E2F decoy strategy on the phenotypic transformation of detrusor smooth muscle cells (DSMCs) so as to verify the effect of the E2F decoy strategy in improvement of the bladder function after bladder outlet obstruction (BOO). METHODS: Rat DSMCs were cultured, underwent cyclic mechanical stretch so as to establish BOO model, and then were divided into 3 groups: E2F-ODN decoy group [transfected with E2F-decoy ODN tagged with carboxy-fluorescein (FAM), a at its 3' end], Mis-decoy group (transfected with mismatch E2F-decoy ODN), and control group (without transfection). Inverted fluorescence microscopy was used to detect the green fluorescence of FAM in the successfully transfected cells. The proliferation of the cells was observed by MTT method. RT-PCR was used to examine the mRNA expression of proliferating cell nuclear antigen (PCNA). Western blotting was used to detect the protein expression of PCNA and cdk2 kinase. RESULTS: FAM-labeled E2F-ODN was detected stably in the DSMCs of the E2F-ODN decoy group 24 hours after transfection. The proliferation of the DSMCs of the E2F-ODN decoy group was decreased significantly compared with the mismatch E2F-ODN decoy and control groups (both P < 0.01). The mRNA expression of PCNA, protein expression of PCNA and cdk2 kinase of the E2F-ODN decoy group were all significantly lower than those of the other 2 groups (all P < 0.01). CONCLUSION: E2F-decoy ODN can be transfected and stably expressed in DSMCs. The phenotypic transformation of DSMCs can be successfully inhibited by E2F decoy strategy, which clarifies the potential role of structural stability-based method on improvement of bladder function recovery after BOO.

[Transurethral resection of the prostate for patients with the permanent cardiac pacemaker].

OBJECTIVE: To evaluate the transurethral resection of the prostate (TURP) for patients with the permanent cardiac pacemaker. METHODS: A retrospective study was made on TURP for 8 patients aged from 62 to 71 and equipped with the cardiac pacemaker for 2 to 7 years, because of sick sinus syndrome (5 cases), complete atrioventricular block (2 cases), and three-cord block (1 case). The pacemakers varied accordingly, Type DDD in 4, Type AAI in 3 and Type VVI in 1 of the patients. RESULTS: All the operations were successful, and all the patients experienced satisfactory recovery. CONCLUSION: Patients with the permanent cardiac pacemaker can receive TURP.

[Building immune microsphere against tumor necrosis factor-alpha (TNF-alpha)].

We have constructed the immune microsphere against tumor necrosis factor-alpha (TNF-alpha) prospectively, hoping to establish the experiment groundwork in more researches which could be used in specific elimination of the TNF-alpha by blood purification method for the future. The recombinant human tumor necrosis factor-alpha monoclonal antibody (rHTNF-alpha McAb) was wrapped on the polystyrene microsphere (PSM) carrier connecting poly-L-lysine (PLL) beforehand. They were earmarked by the fluorescein isothiocyanate (FITC) respectively. The packing conditions were examined using the inversted and fluorescence microscopes and the spectrophotometer. The results showed that the best conditions for wrapping were 20 degrees C, pH9.5 and 60 minutes. The PLL content was not changed in the washing fluid after coating, which indicated the wrapping was quite firm. At the same temperature and same coating time, the rHTNF-alpha McAb coated on the PLL was obviously substantial when the concentration of glutaraldehyde solution was 0.2%. The findings demonstrated that the built immune microsphers can be used as a novel adsorption material. This method is simple and economic, and it offers a new approach to the related studies.

[The relationship between phenotype transformation and biomechanical properties of detrusor smooth muscle cell subjected to the cyclic mechanical stretch].

OBJECTIVE: To investigate the relationship between phenotype transformation and biomechanical properties of detrusor smooth muscle cell (DSMC) subjected to the cyclic mechanical stretch. METHODS: Cultured rat DSMCS were grown on collagen-coated silicone membranes and subjected to continuous cycles of stretch-relaxation. All experiments were made on cells between passage 2 and 4. Each cycle consists of 5-second stretch and 5-second relaxation. The computer controlled vacuum induced 10% (I), 20% (II) and 30% (III) maximum elongation of the plate membrane at different designed pressures. We assessed DNA synthesis rate using tritiated thymidine incorporation assay. Using immunofluorescent assay and flow cytometer, we analysed the expression of SM-alpha-actin and proliferation of DSMC. The image analysis and micropipette aspiration systems were employed to investigate the single cell contraction and viscoelasticity. The elastic modulus K(1), K(2) and viscoelastic coefficient micro were determined using the three-element standard linear solid model, thus demonstrating the passive deformation ability of detrusor cells. RESULTS: As the basic structural changes to mechanical stretch, DSMCs underwent phenotypic modulation from their normal contractile phenotype to a "synthetic" phenotype: the DSMCs became more proliferative and the actin less organized along the cell's long axis. The cell proliferation index (CPI) of control and stretched group (10%, 20%, 30% elongation) were 0.24, 0.43, 0.58 and 0.65 respectively. After mechanical stretch, the well-spread filaments changed their orientation. Contraction and viscoelasticity of single DSMC subjected to stretch both decreased significantly compared to control. The Vmax and. DeltaLmax of group III (30% elongation) saw significant decreases compared with unstretched control (P < 0.01). K(1) and K(2) decreased with the increasing of mechanical overload, however, there was no statistic difference between groups II and group III. CONCLUSIONS: Structure determines function. Conversely, dysfunction implies the structural transformation. Functional abnormalities of BOO have the structural basis: phenotype transformation of detrusor cells. Cyclic stretch and relaxation applied to DSMCs in vitro can be used to model the increases in urodynamic load experienced by the bladder detrusor muscle under the conditions of bladder outlet obstruction. Phenotypic transformation is the structural basis of functional changes of DSMC subjected to periodic overload mechanical stretch.

[Ultrastructural observation of detrusor in BPH patients].

OBJECTIVES: To observe the detrusor ultrastructure in BPH patients and to investigate the relationship between detrusor instability and ultrastructure. METHODS: The patients were divided into groups of detrusor instability(DI) and detrusor stability(DS) according to urodynamics examination. The structure of the detrusor were observed by light microscopy and transmission electron microscopy(TEM). RESULTS: The intercellular intermediate junctions and cytoplasmic process junctions in DS were 11.34 +/- 3.23 and 4.26 +/- 1.78 respectively. The intercellular intermediate junctions decreased obviously (3.12 +/- 1.47, P < 0.01) instead of a large amount of cytoplasmic process junctions (26.37 +/- 7.14, P < 0.01) in DI. CONCLUSIONS: There is a close relevance between intercellular junctions and DI. The observation of the ultrastructure of the detrusor is helpful for the diagnosis of BPH with DI and for the clinical treatment.

[Characteristics on diagnosis and treatment of renal tuberculosis].

OBJECTIVE: To study the new characteristics on diagnosis and treatment of renal tuberculosis (RT). METHODS: Eighty-seven patients with renal tuberculosis were retrospectively reviewed; their diagnosis was established by standard microbiological and histological techniques. RESULTS: Atypical RT was diagnosed by various examinations, including urinary analysis, polymerase chain reaction of tuberculosis (PCR-TB), ultrasonography, intravenous urography (i.v.U), and computerized tomography (CT). Treatment consisted of antituberculous chemotherapy in all patients, in combination with nephrectomy (62.5%) or enterocystoplasty (4.6%). CONCLUSIONS: The differential diagnosis of RT should be emphasized, especially for atypical RT, provided a much more specific diagnosis in clinical suspicion of RT. i.v.U can not be regarded as a specific examination for RT. Computerised tomography (CT) can be used for early diagnosis of RT. Surgery for RT is still ablative.

Roles of stem cell factor on loss of interstitial cells of Cajal in bladder of diabetic rats.

OBJECTIVE: To explore the roles of stem cell factor (SCF) on the loss of interstitial cells (ICCs) in the bladder of diabetic rats, which have not been investigated. METHODS: The rats were assigned to 3 groups: normal control rats, diabetic rats, and SCF-treated diabetic rats. The diabetic rat model was created using a streptozotocin (60 mg/kg) intraperitoneal injection. The SCF and c-kit levels in bladder tissue were determined using reverse transcriptase-polymerase chain reaction and Western blot analysis. The quantity of ICCs as represented by c-kit-positive cells was examined by image analysis of immunofluorescence staining. RESULTS: Compared with the control rats, the diabetic rats exhibited a significant decrease in the SCF levels and c-kit expression and number of ICCs in the bladder tissues. All these impaired parameters were effectively restored to the control level after exogenous SCF treatment. CONCLUSION: These findings suggest that the loss of ICCs in the bladder tissue of diabetic rats can be attributed to a deficiency in endogenous SCF. The beneficial effect of exogenous SCF on diabetic depletion of ICCs could provide a theoretical rationale for the use of SCF as a potential therapeutic drug in treating patients with diabetes-related voiding dysfunction.

Nanobacteria may be linked to testicular microlithiasis in infertility.

Testicular microlithiasis (TM) in infertility is an uncommon pathologic condition of unclear etiology that is characterized by calcium deposits within the seminiferous tubules. Nanobacteria (NB), as novel microorganisms mediating tissue calcification, have been discovered in some diseases. In this study, we hypothesized that NB may participate in the pathogenesis of TM, particularly in infertility. Seventeen infertility patients with TM detected by scrotal color Doppler ultrasonography and 17 infertility patients without TM as controls were enrolled in the study. The NB were isolated and cultured from semen samples and urine samples. After 3 to 6 weeks of culture, 10 of 17 (58.8%) semen samples and 2 urine samples from infertile patients with TM showed the growth of white granular microbes that firmly attached to the bottom of the culture flask and were visible to the naked eye. In the control group, only 1 of 17 (5.9%) semen samples from infertile patients without TM showed the growth of white granular microbes. The cultured microbes were identified by indirect immunofluorescent staining (IIFS), transmission electron microscopy (TEM), and 16s rRNA gene expression. IIFS and TEM revealed NB to be coccoid and 100 to 500 nm in diameter. The BLAST result revealed that the 16s rRNA gene sequence from the cultured microbes was 97% the same as that of the known NB. Our results showed that NB may be linked to the development of TM, which may provide a potential target for the diagnosis and treatment of infertility with TM.

Detection of nanobacteria infection in type III prostatitis.

OBJECTIVES: To investigate the relationship between nanobacterial infection and type III prostatitis. The etiology of type III prostatitis remains unclear to date, although the recently discovered nanobacteria (NB) have been implicated in this disease. METHODS: A total of 48 patients with chronic pelvic pain syndrome for whom conventional therapy had failed were selected and randomly divided into two groups, one receiving anti-NB treatment and the other receiving a placebo. The NB were isolated and cultured from expressed prostatic secretions and urine samples before and after treatment. The morphologic features were recorded and 16s rRNA gene expression was determined. The curative effect was evaluated by the NB-positive rate and symptomatic changes using the National Institutes of Health Chronic Prostatitis Symptom Index. RESULTS: After anti-NB treatment, the NB-positive rates had decreased from 62.5% to 16.7% in the expressed prostatic secretions and from 12.5% to 0% in the urine samples after prostatic massage (P <0.001). In the patients receiving a placebo, the positive rates had no obvious change in either the expressed prostatic secretions or the urine samples after prostatic massage (P >0.05). The NB were coccoid or coccobacillary and clustered in a diameter of 100 to 500 nm. The BLAST result revealed that the 16s rRNA gene sequence from the NB in the patients with chronic pelvic pain syndrome was 97%, similar to that of the known NB with identity (97%). After anti-NB treatment, the Chronic Prostatitis Symptom Index scores decreased significantly. In contrast, no change in the Chronic Prostatitis Symptom Index scores was seen after placebo treatment. CONCLUSIONS: The results of our study have shown that nanobacterial infection might be an important etiologic factor of type III prostatitis. Anti-NB treatment could be an effective therapy against refractory type III prostatitis.

Role of UPP in evaluating bladder outlet obstruction due to benign prostatic enlargement.

AIMS: To study the role of UPP in assessing bladder outlet obstruction (BOO) due to benign prostatic enlargement (BPE). MATERIALS AND METHODS: Urodynamic results from 1281 elderly men with Lower urinary tract symptoms (LUTS) obtained from September 1999 through July 2005 were retrospectively analyzed. Independent sample t-test or Wilcoxon signed-rank test was used to compare the parameters of different groups according to PFS. Chi-square test was used to compare the percentage of urethral pressure profile (UPP) types between different groups. ROC curve was used to study the practical value of UPP parameters in diagnosing BOO. RESULTS: According to PFS, patients were divided into three groups: Obstructed, equivocal obstructed and unobstructed. There was significant difference between the unobstructed and obstructed group in the age of patients, bladder capacity, residual volume, voided volume, voided time, prostatic plateau area (PPA), prostatic length, functional profile length, pressure of bladder neck, and pressure of seminal colliculus, but not in maximum urethral closure pressure. All UPP parameters correlated significantly to the diagnosis of bladder outlet obstruction due to BPE. The largest area under the ROC curve was that of PPA. CONCLUSIONS: Our results indicate that PPA on UPP is useful in assessing the grade of obstruction due to BPE when PFS fails, a finding still to be evaluated by prospective studies.

Loss of ryanodine receptor calcium-release channel expression associated with overactive urinary bladder smooth muscle contractions in a detrusor instability model.

OBJECTIVE: To investigate the changes in spontaneous bladder smooth muscle contractions that occur during detrusor instability (DI), and to test the possibility that altered function or expression of ryanodine receptors (RyRs) could account for the increased bladder contractions. MATERIALS AND METHODS: After 8 weeks of partial bladder outlet obstruction, DI was confirmed in female experimental rats by filling cystometry. Muscle strips were dissected from freshly isolated bladders, and isometric tension recorded in strips from DI and normal bladders. The contractions were recorded during electrical stimulation or exposure to various agents. Western blot analysis was used to determine RyR expression in DI and normal bladder muscle. RESULTS: In DI bladder muscle, spontaneous contractile activity persisted in the presence of blockers for known neurotransmitter receptors in the bladder wall. The RyR blocker ryanodine significantly increased the spontaneous contractile frequency in normal bladder strips, but failed to affect spontaneous contractions in DI muscle. Caffeine inhibited spontaneous contractile activity in both the DI and normal strips. After administering the l-type Ca(2+) channel antagonist nimodipine, the myogenic contractile activity was abolished in normal strips; in contrast, in DI strips, the amplitude of contractions was reduced but the frequency of contractions was unchanged. Western blot analysis showed that RyR expression was lower in DI muscle than in normal bladder muscle. CONCLUSION: These results provide the first characterization of a loss of regulation of spontaneous contractile activity by RyRs in DI muscle associated with a significant decrease in RyR expression. RyRs in normal detrusor muscle act as negative-feedback regulators of spontaneous contractile activity, presumably by releasing Ca(2+) that activates Ca(2+)-dependent K(+) channels to decrease contractility. This mechanism might be weakened in DI muscle, resulting in spontaneous contractile overactivity.

Possible mechanism of referred pain in the perineum and pelvis associated with the prostate in rats.

PURPOSE: Since persistent pain in the perineum and pelvic floor associated with chronic prostatitis /chronic pelvic pain syndrome has been hypothesized to be referred pain, it might also be explained by neural mechanisms. MATERIALS AND METHODS: Dual retrograde fluorescent labeling and immunohistochemistry were identified as methods with which to investigate the neurogenic aspect of this status. The dual distribution of dorsal root ganglia (DRG) cells was determined after double retrograde fluorescent staining of the prostate and pelvic floor, and the prostate and perineum somatic nerves. Calcitonin gene-related peptide (CGRP) and substance P (SP) in dual labeled cells were determined by immunohistochemistry, giving possible insight into the cause of pelvic pain. RESULTS: Fluorescent double labeled cells were found in the lumbar and sacral DRG, while double labeled cells were distributed predominantly in L6 to S1 and L1 to L2 segment DRG in groups 1 and 2, respectively. On immunohistochemistry some of them were confirmed to contain CGRP and SP. Thus, there are crossover pathways between the prostate and pelvic floor. CONCLUSIONS: The findings that we present confirm that the peripheral process of DRG cells dichotomizes to the prostate, sphincter and somatic parties simultaneously. Some of these cells contain CGRP and SP, which indicate that referred pain in the perineum and pelvic floor may be caused by an axon reflex in the peripheral process of DRG neurons.