Normalization of magnesium deficiency attenuated mechanical allodynia, depressive-like behaviors, and memory deficits associated with cyclophosphamide-induced cystitis by inhibiting TNF-α/NF-κB signaling in female rats.

BACKGROUND: Bladder-related pain symptoms in patients with bladder pain syndrome/interstitial cystitis (BPS/IC) are often accompanied by depression and memory deficits. Magnesium deficiency contributes to neuroinflammation and is associated with pain, depression, and memory deficits. Neuroinflammation is involved in the mechanical allodynia of cyclophosphamide (CYP)-induced cystitis. Magnesium-L-Threonate (L-TAMS) supplementation can attenuate neuroinflammation. This study aimed to determine whether and how L-TAMS influences mechanical allodynia and accompanying depressive symptoms and memory deficits in CYP-induced cystitis. METHODS: Injection of CYP (50 mg/kg, intraperitoneally, every 3 days for 3 doses) was used to establish a rat model of BPS/IC. L-TAMS was administered in drinking water (604 mg·kg-1·day-1). Mechanical allodynia in the lower abdomen was assessed with von Frey filaments using the up-down method. Forced swim test (FST) and sucrose preference test (SPT) were used to measure depressive-like behaviors. Novel object recognition test (NORT) was used to detect short-term memory function. Concentrations of Mg2+ in serum and cerebrospinal fluid (CSF) were measured by calmagite chronometry. Western blot and immunofluorescence staining measured the expression of tumor necrosis factor-α/nuclear factor-κB (TNF-α/NF-κB), interleukin-1ß (IL-1ß), and N-methyl-D-aspartate receptor type 2B subunit (NR2B) of the N-methyl-D-aspartate receptor in the L6-S1 spinal dorsal horn (SDH) and hippocampus. RESULTS: Free Mg2+ was reduced in the serum and CSF of the CYP-induced cystitis rats on days 8, 12, and 20 after the first CYP injection. Magnesium deficiency in the serum and CSF correlated with the mechanical withdrawal threshold, depressive-like behaviors, and short-term memory deficits (STMD). Oral application of L-TAMS prevented magnesium deficiency and attenuated mechanical allodynia (n = 14) and normalized depressive-like behaviors (n = 10) and STMD (n = 10). The upregulation of TNF-α/NF-κB signaling and IL-1ß in the L6-S1 SDH or hippocampus was reversed by L-TAMS. The change in NR2B expression in the SDH and hippocampus in the cystitis model was normalized by L-TAMS. CONCLUSIONS: Normalization of magnesium deficiency by L-TAMS attenuated mechanical allodynia, depressive-like behaviors, and STMD in the CYP-induced cystitis model via inhibition of TNF-α/NF-κВ signaling and normalization of NR2B expression. Our study provides evidence that L-TAMS may have therapeutic value for treating pain and comorbid depression or memory deficits in BPS/IC patients.

Neuregulin-1-ErbB signaling promotes microglia activation contributing to mechanical allodynia of cyclophosphamide-induced cystitis.

AIMS: Central sensitization playsimportant roles in cyclophosphamide (CYP)-induced cystitis. In addition, as a visceral pain, CYP-induced chronic pain shares common pathophysiological mechanisms with neuropathic pain. Previous studies demonstrated that neuregulin-1 (Nrg1)-ErbB signaling contributes to neuropathic pain, but whether and how this signaling influences mechanical allodynia in CYP-induced cystitis is unclear. This study aimed to determine whether and how Nrg1-ErbB signaling modulates mechanical allodynia in a CYP-induced cystitis rat model. METHODS: Systemic injection with CYP was used to establish a rat model of bladder pain syndrome/interstitial cystitis (BPS/IC). An irreversible ErbB family receptor inhibitor, PD168393, and exogenous Nrg1 were intrathecally injected to modulate Nrg1-ErbB signaling. Mechanical allodynia in the lower abdomen was assessed with von-Frey filaments using the up-down method. Western blot analysis and immunofluorescence staining were used to measure the expression of Nrg1-ErbB signaling, Iba-1, p-p38, and IL-1ß in the L6-S1 spinal dorsal horn (SDH). RESULTS: We observed upregulation of Nrg1-ErbB signaling as well as overexpression of the microglia activation markers Iba-1 and p-p38 and the proinflammatory factor, interleukin-1ß (IL-1ß), in the SDH of the cystitis group. Further, treatment with PD168393 attenuated mechanical allodynia in CYP-induced cystitis and inhibited microglia activation, leading to decreased production of IL-1ß. The inhibitor PD168393 reversed the algesic effect of exogenous Nrg1 on the cystitis model. CONCLUSIONS: Nrg1-ErbB signaling may promote microglia activation, contributing to mechanical allodynia of CYP-induced cystitis. Our study showed that modulation of Nrg1-ErbB signaling may have therapeutic value for treating pain symptoms in BPS/IC.

Increased severity of inflammation correlates with elevated expression of TRPV1 nerve fibers and nerve growth factor on interstitial cystitis/bladder pain syndrome.

OBJECTIVES: Although evidence supports a role for inflammation in interstitial cystitis/bladder pain syndrome (IC/BPS), the mechanism remains unknown. We determined whether inflammation causes an elevated expression of nerve growth factor (NGF) and transient receptor potential vanilloid receptor subtype 1 (TRPV1) and correlated them with the symptoms. METHODS: Bladder biopsies were obtained from 53 IC/BPS patients and 27 controls, and hematoxylin and eosin staining, immunostaining and Western blotting were performed to detect inflammation, TRPV1-immunoreactive and PGP9.5-immunoreactive nerve fibers, and NGF, respectively. Symptoms were assessed using the Pelvic Pain/Urgency/Frequency (PUF) questionnaire and pain visual analogue scale scores. Suburothelial nerve fiber density was quantified and correlated with PUF scores. RESULTS: Increased severity of inflammation was correlated with a higher TRPV1-immunoreactive nerve fiber density (r = 0.4113, p = 0.0024) and higher NGF levels (r = 0.3775, p = 0.0052). Suburothelial TRPV1-immunoreactive nerve fiber density was significantly correlated with pain scores and urgency scores (r = 0.3320, p = 0.0145 and r = 0.3823, p = 0.0039, respectively). PGP9.5-immunoreactive nerve fibers were significantly increased in IC/BPS (p = 0.0193) and had a positive relationship with inflammation severity (r = 0.6138, p < 0.0001). CONCLUSIONS: Our study revealed increased severity of inflammation correlated with a higher expression of TRPV1-immunoreactive nerve fibers and NGF in IC/BPS and correlated with clinical symptoms.

Supine lithotomy versus prone position in minimally invasive percutaneous nephrolithotomy for upper urinary tract calculi.

OBJECTIVE: To compare operative time, safety and effectiveness of minimally invasive percutaneous nephrolithotomy (MPCNL) in the supine lithotomy versus prone position. METHODS: Between January 2008 and December 2010, a total of 109 consecutive patients with upper urinary tract calculi were enrolled and randomly divided into group A (53 patients, supine lithotomy position) and group B (56 patients, prone position). The MPCNL procedures were performed under the guidance of real-time grayscale ultrasound system. The preoperative characteristics, intraoperative and postoperative parameters were analyzed and compared. RESULTS: All patients were successfully operated. There was no significant difference between the two groups in stone-free rate (group A 90.1 vs. group B 87.5%, p = 0.45), mean blood loss, number of access tracts, calyx puncture, mean hospital stay (group A 6 ± 1.1 vs. group B 6 ± 1.5 days, p = 0.38) and complications. But the operative time was significantly shortened in supine lithotomy position (group A 56 ± 15 vs. group B 86 ± 23 min, p < 0.001). CONCLUSIONS: The effectiveness and safety of the supine lithotomy position for MPCNL were similar to the prone position. However, the supine lithotomy position has an important advantage of reducing the operative time. The supine lithotomy position could be a good choice to perform MPCNL.

Deep Genomic Sequencing of Bladder Urothelial Carcinoma in Southern Chinese Patients: A Single-Center Study.

OBJECTIVE: Bladder urothelial carcinoma (BUC) is a common urological malignancy with molecular heterogeneity. However, the genetic feature of Chinese BUC patients is still not well-identified. METHODS: We performed deep sequencing by a large panel (450 genes) on 22 BUC samples and using matched normal bladder tissue as control. Genomic alterations (GAs), pathways and Tumor Mutation Burden (TMB) were investigated. RESULTS: The frequencies of GAs (TERT, 54.5%; CREBBP, 27.3%; GATA3, 22.7%; BRAF, 18.2%; TEK, 18.2% and GLI1, 18.2%) were significantly higher in Chinese than Western BUC patients. Other GAs' frequencies were in accordance with previous study (TP53, 50.0%; KDM6A, 31.8%; KMT2D, 22.7%; etc.). Besides, we detected gene amplification in ERBB2, FRS2, FAS, etc. The gene fusion/rearrangement took place in the chromosome 11, 12, 14, 17, 19, 22, and Y. Other than cell cycle and PI3K-AKT-mTOR, mutated genes were more associated with the transcription factor, chromatin modification signaling pathways. Interestingly, the TMB value was significantly higher in the BUC patients at stages T1-T2 than T3-T4 (P = 0.025). CONCLUSION: Deep genomic sequencing of BUC can provide new clues on the unique GAs of Chinese patients and assist in therapeutic decision.

[Effects of anti-gene and antisense therapeutics on human prostate cancer xenograft in nude mice].

OBJECTIVE: To investigate the effects of triple-helix forming oligonucleotide (TFO) and antisense oligonucleotide (ASO) on androgen receptor (AR) expression and tumor growth of human prostate cancer xenografts in nude mice. METHODS: Thirty-two nude mice were inoculated with human prostate cancer cells of the line LNCaP-C4-2 were randomized into 4 equal groups: TFO treatment group, undergoing intra-tumor injection of TFO at the dose of 25 mgxkg(-1)x(2d)(-1) for 14 times, ASO treatment group, undergoing intra-tumor injection of ASO at the same dose for 14 times, SCO group, undergoing intra-rumor injection of sequence control oligonucleotide (SCO) at the same dose for 14 times, and control group. The body weight and xenograft tumor volume of the nude mice were monitored during the therapy. 28 days later venous blood samples were collected to measure the level of prostate specific antigen (PSA) by radioimmunoassay and then the mice were killed with their tumors taken out to measure the weight, and RT-PCR, immunohistochemistry, and radioligand binding assay were used to detect the AR gene mRNA and protein expression in the tumor tissues. RESULTS: By the end of experiment the volumes and weights of tumor of the ASO and ASO groups were all significantly lower than those of the control group (all P < 0.01) with the inhibition rates of 67.55% and 41.06% respectively, and the volumes and weights of tumor of the TFO group were all significantly lower than those of the ASO group (all P < 0.05). The tumor weight and AR expression levels of TFO group were significantly lower than those of ASO group (P < 0.05). The serum PSA level of TFO group was (6.6 +/- 1.0) ng/ml, significantly lower than that of the ASO group [(19.8 +/- 3.7) ng/ml, P < 0.05]. The mRNA and protein expression levels of AR of the TFO group were significantly lower than those of the other groups (all P < 0.05). There were no significant differences in all the above mentioned markers between the SCO and control groups (all P > 0.05). CONCLUSION: TFO shows significantly higher inhibitory effect on AR expression and tumor growth of human prostate cancer xenograft than ASO, and is a promising agent for prostate cancer gene therapy.

Reduced expression of alpha-tocopherol-associated protein is associated with tumor cell proliferation and the increased risk of prostate cancer recurrence.

AIM: To examine the impact and prognostic significance of alpha-tocopherol associated protein (TAP) expression in a series of prostate cancer patients. METHODS: Tissues from 87 patients underwent radical prostatectomy were examined for TAP expression by immunohistochemistry. The relationships of the staining results, the clinic pathological characteristics and the recurrence times were analyzed. RESULTS: Compared with the adjacent areas of normal and benign glands, immunoreactivity of TAP was reduced in areas of prostate cancer. A lower TAP-positive cell number per mm(2) of the largest cancer area (defined as TAP-PN) was associated with higher clinical stage (r = -0.248, P = 0.0322). Inverse associations were found among the TAP-PN and positive lymph nodes (r = -0.231, P = 0.0325), preoperative prostate-specific antigen (PSA) levels (r = -0.423, P = 0.0043), tumor size (r= -0.315, P= 0.0210) and elevated tumor cell proliferation, which was indicated by the staining of Ki-67 (r = -0.308, P = 0.0026). TAP-PN was a significant predictor of recurrence univariately (P = 0.0006), as well as multivariately, adjusted for known markers including preoperative PSA, clinical stage, Gleason score, surgical margin, extra-prostatic extension, seminal vesicle invasion and lymph node metastasis (P = 0.0012). CONCLUSION: Reduced expression of TAP was associated with the cell proliferation status of prostate cancer, adverse pathological parameters and the increased risk of recurrence.

[Treatment of erectile dysfunction by embedding deep dorsal vein of penis for 5 cases].

OBJECTIVES: To evaluate the curative effect of embedding deep dorsal vein of penis on erectile dysfunction (ED). METHODS: Three patients with venous ED and 2 patients with mixed factors of artery and vein were treated by the embedding deep dorsal vein of penis. For evaluating curative effect the 5 patients were visited after operation. RESULTS: Two months after surgery, the 3 patients of venous ED achieved satisfactory intercourse and the 2 patients with mixed factors of artery and vein had sufficient erection for intercourse after taking 50 mg Sildenafil. The 5 cases kept the above-mentioned curative effect during the follow up period between 3 to 12 months (7 months on avenage). CONCLUSIONS: The embedding deep dorsal vein surgery, having small surgical wounds and few complications, is an effective treatment for venous ED.

Presence of tumour-infiltrating FOXP3+ lymphocytes correlates with immature tumour angiogenesis in renal cell carcinomas.

BACKGROUND: FOXP3+ regulatory T cells (Tregs) inhibit effector T cell functions and are implicated in tumour progression. However, together with microvessel density (MVD) they remain controversial prognostic predictors for renal cell carcinoma (RCC), and potential associations have yet to be determined. The objective of this study was to determine the prognostic significance of Tregs and MVD and their potential relationship in RCCs. DESIGN: Paraffin-embedded tissues from 62 RCC patients were analysed using immunohistochemistry to detect FOXP3+ lymphocytes, and double immunohistochemistry to detect different microvessel types in the tumour interior, rim and normal kidney tissue, and their correlation with clinicopathological characteristics. Survival analysis was also performed. RESULTS: The presence of FOXP3+ cells in the tumour interior or the rim showed no correlation with death from RCC and other pathological characteristics. Negative correlations were noted between the immature MVD in the tumour interior or the rim and tumour size, tumour stage and overall survival; however, there was no correlation with the nuclear grade or pathological type. A positive correlation between FOXP3+ Tregs and immature MVD (r=0.363, P=0.014) and mature MVD (r=0.383, P=0.009) was confirmed in the tumour interior. However, there was no correlation between FOXP3+ Tregs and mature MVD (r=0.281, P=0.076) or immature MVD (r=0.064, P=0.692) in the tumour rim. CONCLUSIONS: In this study, a positive correlation between the presence of FOXP3+ Tregs and immature and mature MVD in RCC was confirmed, which suggests a link between suppression of immunity, tumour angiogenesis and poor prognosis.

A randomized controlled trial of postoperative tumor lysate-pulsed dendritic cells and cytokine-induced killer cells immunotherapy in patients with localized and locally advanced renal cell carcinoma.

BACKGROUND: It remains a challenge to inhibit the local recurrence or distant metastasis of localized or locally advanced renal cell carcinoma (RCC) after surgical resection. We investigated the feasibility, safety and efficacy of immunotherapy using autologous tumor lysate (TL)-pulsed dendritic cells (DCs) and cytokine-induced killer (CIK) cells in patients with localized or locally advanced RCC. METHODS: From January 2001 to July 2009, we collected 137 patients that met the selection criteria and randomly divided them into three groups. After surgery, immunotherapy with TL-pulsed DCs-CIK cells (DC-CIK group) and interferon (IFN)-α (IFN-α group) was performed in 46 patients, respectively. The other 45 patients received no postoperative adjuvant therapy (the control group). The changes in the numbers of T lymphocyte subsets, including CD4(+)CD25(high) regulatory T cells (Treg), were determined before the operation and after immunotherapy. The overall survival was compared among the three groups. RESULTS: An increase of the CD4(+)/CD8(+) ratio and a decrease of CD4(+)CD25(high) cells were observed after TL-pulsed DC-CIK cells or IFN-a immunotherapy. All patients tolerated the TL-pulsed DC-CIK cells immunotherapy very well, and side effects in the DC-CIK group were less than in the IFN-α group. The metastasis and recurrence rates were significantly decreased after TL-pulsed DC-CIK cells or IFN-α immunotherapy compared with the control group (P < 0.01). The Log-rank test showed that the overall survival rates were significantly higher in the DC-CIK group and IFN-α group than that in the control group (P < 0.01), but there was no difference between the DC-CIK group and IFN-α group (P > 0.05). CONCLUSION: Postoperative immunotherapy with TL-pulsed DC-CIK cells may prevent recurrence/metastasis and increase the overall survival rate after surgery in localized or locally advanced RCC.

ERG rearrangement for predicting subsequent cancer diagnosis in high-grade prostatic intraepithelial neoplasia and lymph node metastasis.

PURPOSE: We aimed to analyze whether ERG rearrangement in biopsies could be used to assess subsequent cancer diagnosis in high-grade prostatic intraepithelial neoplasia (HGPIN) and the risk of lymph node metastasis in early prostate cancer. EXPERIMENTAL DESIGN: Samples from 523 patients (361 with early prostate cancer and 162 with HGPIN) were collected prospectively. On the basis of the cutoff value established previously, the 162 patients with HGPIN were stratified to two groups: one with an ERG rearrangements rate ≥1.6% (n = 59) and the other with an ERG rearrangements rate <1.6% (n = 103). For the 361 prostate cancer cases undergoing radical prostatectomy, 143 had pelvic lymph node dissection (node-positive, n = 56 and node-negative, n = 87). All ERG rearrangement FISH data were validated with ERG immunohistochemistry. RESULTS: A total of 56 (of 59, 94.9%) HGPIN cases with an ERG rearrangements rate ≥1.6% and 5 (of 103, 4.9%) HGPIN cases with an ERG rearrangements rate <1.6% were diagnosed with prostate cancer during repeat biopsy follow-ups (P < 0.001). There were significant differences in ERG rearrangement rates between lymph node-positive and -negative prostate cancer (P < 0.001). The optimal cutoff value to predict lymph node metastasis by ERG rearrangement was established, being 2.6% with a sensitivity at 80.4% [95% confidence interval (CI), 67.6-89.8] and a specificity at 85.1% (95% CI, 75.8-91.8). ERG protein expression by immunohistochemistry was highly concordant with ERG rearrangement by FISH. CONCLUSIONS: The presence of ERG rearrangement in HGPIN lesions detected on initial biopsy warrants repeat biopsies and measuring ERG rearrangement could be used for assessing the risk of lymph node metastasis in early prostate cancer.

A comparative study of clinical value of single B-mode ultrasound guidance and B-mode combined with color doppler ultrasound guidance in mini-invasive percutaneous nephrolithotomy to decrease hemorrhagic complications.

OBJECTIVES: To compare the clinical value of single B-mode ultrasonography and B-mode combined with color Doppler ultrasonography in the guidance of mini-invasive percutaneous nephrolithotomy (m-PCNL) to decrease the incidence of hemorrhagic complications. METHODS: A total of 297 patients with renal stones who had undergone m-PCNL were retrospectively categorized into 2 groups. Group 1 (187 patients) underwent m-PCNL with single B-mode ultrasound guidance and group 2 (110 patients) underwent m-PCNL with combined B-mode and color Doppler ultrasound guidance. The clinical characteristics of the patients, intraoperative and postoperative characteristics, complications, especially hemorrhagic complications, and blood transfusion rate were recorded and compared. RESULTS: No statistically significant differences in age, height, weight, stone burden, operative time, stone-free rate, or length of postoperative hospital stay were found between the 2 groups. In group 2, a statistically significant decrease in the transfusion rate was found compared with group 1 (P <.05). In group 1, 5 patients (2.6%) required a blood transfusion, 2 (1.1%) developed a renal arteriovenous fistula and required embolotherapy, 2 (1.1%) developed hemorrhage and required embolotherapy after surgery, 16 (8.6%) developed capillary hemorrhage during surgery but had no hemorrhage postoperatively. However, no serious hemorrhagic complications were found in group 2. Only 3 patients (2.7%) developed capillary hemorrhage during surgery, and no hemorrhage occurred postoperatively. CONCLUSIONS: Using combined B-mode and color Doppler ultrasound guidance during in m-PCNL resulted in the real-time detection and avoidance of the renal blood vessels during puncture and decreased the incidence of hemorrhagic complications, especially in the patients with a solitary and compensative kidney.

Modified apical dissection of the prostate improves early continence in laparoscopic radical prostatectomy: technique and initial results.

OBJECTIVES: Incontinence after radical retropubic prostatectomy (RRP) is one of the greatest worries for all patients. One of the possible reasons for this urinary incontinence is a postoperative deficiency of the external striated urethral sphincter (EUS) complex and continence nerves. This study evaluated the application of a modified simple technique to dissection of the apical prostate in laparoscopic radical prostatectomy (LRP) and assessed the rate of urinary continence. METHODS: A total of 104 patients were randomly selected using envelopes and enrolled in this study. A standard LRP was performed in 52 patients (standard LRP group) and a modified technique for simple dissection of the apical prostate in LRP was performed in another 52 patients (modified LRP group). The urethra was dissected and transected at the apex of the prostate, proximal to the nerve-distributing rhabdosphincter using sharp scissors to avoid damage to the EUS complex and continence nerves. In all patients, a pad test was performed on 3, 30 and 90 days postoperatively and correlated with urinary continence. Continence was defined as zero pads or a liner used for security reasons only. RESULTS: After catheter removal, the continence rates were regained in 66, 85 and 96% of patients in the modified LRP group compared with 28, 55 and 91% of the patients in the standard LRP group at 3, 30 and 90 days, respectively. A statistically significant difference was present at 3 and 30 days (p < 0.01, respectively). At 90 days, the difference, although still present, was not statistically significant (p > 0.05). CONCLUSIONS: In this preliminary study, the technique of simplified apical dissection of the prostate in LRP appears to be an easy and feasible technique in early recovery of urinary continence. Further long-term and larger sample studies are necessary to elucidate the modified technique in LRP on early restoration of urinary continence.

Detection of TMPRSS2-ETS fusions by a multiprobe fluorescence in situ hybridization assay for the early diagnosis of prostate cancer: a pilot study.

Fusion of the prostate-specific and androgen-regulated transmembrane-serine protease gene (TMPRSS2) with the erythroblast transformation-specific (ETS) family members is the most common genetic alteration in prostate cancer. However, the biological and clinical role of TMPRSS2-ETS fusions in prostate cancer, especially in problematic prostate needle core biopsies, has not been rigorously evaluated. We randomly collected 85 specimens including 50 archival prostate cancer tissue blocks, 15 normal prostate specimens, and 20 benign prostatic hyperplasia specimens for TMPRSS2-ETS fusion analyses. Moreover, the fusion status in an additional 20 patients with initial negative biopsies who progressed to biopsy-positive prostate cancer at subsequent follow-ups was also characterized. Fluorescently labeled probes specific for ERG-related rearrangements involving the TMPRSS2-ERG fusion as well as TMPRSS2-ETV1 and TMPRSS2-ETV4 were used to assess samples for gene rearrangements indicative of malignancy under a design of sequential trial. Rearrangements involving TMPRSS2-ETS fusions were detected in 90.0% of the 50 postoperative prostate cancer samples. The positive rate for the rearrangements in the initial prostate cancer-negative biopsies of 20 patients who eventually progressed to prostate cancer was 60.0% (12/20). Our preliminary study demonstrates that the clinical utility of TMPRSS2-ETS fusion detection as a biomarker and ancillary diagnostic tool for the early diagnosis of prostate cancer is promising, given this approach shows significant high sensitivity and specificity in detection.