Activation of a c-Jun N-terminal kinase-mediated autophagy pathway attenuates the anticancer activity of gemcitabine in human bladder cancer cells.

The role of autophagy in the anticancer activity of gemcitabine (GEM) in bladder cancer is unclear. The aim of this study is to determine whether GEM activates autophagy, the role of autophagy in the anticancer activity of GEM, and the underlying mechanism by which GEM induces autophagy. Human bladder cancer cell lines T24 and BIU87 were treated with GEM in vitro. Cell viability was measured using the Cell Counting Kit-8 assay. Apoptosis was detected by annexin V assay and western blot. Autophagy was measured by western blot and transmission electron microscopy. c-Jun N-terminal kinase (JNK) activation was detected by western blot. Chemical inhibitors were used for intervention of JNK and autophagy. GEM killed bladder cancer cells, which was associated with apoptosis induction. Autophagy was effectively activated by GEM. Suppressing autophagy in GEM-treated cells significantly decreased cell viability, which was associated with increased apoptosis. GEM-induced JNK activation and suppressed B-cell lymphoma 2 expression. The JNK inhibitor SP600125 inhibited GEM-induced autophagy activation and increased GEM's cytotoxicity. GEM kills bladder cancer cells through apoptosis. Meanwhile, JNK-mediated autophagy was activated, which protects the cells against apoptosis. Therefore, inhibition of autophagy could be exploited to enhance the anticancer efficacy of GEM for treating bladder cancer.

ROS activates JNK-mediated autophagy to counteract apoptosis in mouse mesenchymal stem cells in vitro.

AIM: Transplantation of mesenchymal stem cells (MSCs) for the treatment of diabetic erectile dysfunction (ED) is hampered by apoptosis of the transplanted cells. In diabetic ED, there is increased oxidative stress and decreased NO in the corpora cavernosa, and reactive oxygen species (ROS) induce apoptosis of the transplanted cells. In this study we examined whether and how autophagy was involved in ROS-induced apoptosis of MSCs. METHODS: Mouse C3H10 MSCs were treated with H2O2 to simulate the high oxidative condition in diabetic ED. Cell viability was measured using MTT assay. Apoptosis was analyzed by flow cytometry. Apoptosis- and autophagy-related proteins were detected with Western blot assays. Intracellular autophagosome accumulation was studied using transmission electron microscopy. RESULTS: Treatment of MSCs with H2O2 (50-400 µmol/L) inhibited the cell viability in concentration- and time-dependent manners. Furthermore, H2O2 (300 µmol/L) induced apoptosis, as well as activated autophagy in MSCs. Pretreatment with lysosome inhibitor chloroquine (10 µmol/L) or PI3K inhibitor 3-methyladenine (5 mmol/L) significantly enhanced H2O2-induced cell death. Pretreatment with JNK inhibitor SP600125 (10 µmol/L) abrogated H2O2-induced accumulation of LC3-II, and attenuated H2O2-induced reduction of Bcl-2 levels in MSCs. CONCLUSION: ROS induce autophagy to counteract apoptosis in MSCs by activation of JNK. Thus, augmentation of autophagy may reduce apoptosis, prolonging MSC survival and improving MSC-based therapeutic efficacy for diabetic ED.

Panax notoginseng saponins improve erectile function through attenuation of oxidative stress, restoration of Akt activity and protection of endothelial and smooth muscle cells in diabetic rats with erectile dysfunction.

Panax notoginseng saponins (PNS), which have an antioxidant property, are a widely used traditional Chinese medicine. In this study we investigated whether PNS can improve erectile function in rats with erectile dysfunction and the underlying mechanism by using a rat diabetic erectile dysfunction model. The rats were randomly divided into four groups: three PNS-treated groups (50, 100 and 150 mg/kg) and one saline-treated control group. Four weeks post treatment, electrostimulation was applied to the cavernous nerve and intracavernous pressure was measured to assess erectile function. Malondialdehyde, superoxide dismutase and glutathione were detected in the penises of all rats. Ultrastructural changes in the penises were examined by electron microscopy. Expression of Akt was detected by immunohistochemistry. The results showed that intracavernous pressure was increased in PNS-treated groups (100 and 150 mg/kg) compared to the control group. The levels of superoxide dismutase, glutathione and Akt were increased (p < 0.05) while that of malondialdehyde was decreased in the PNS groups. Ruptured endothelium, impaired smooth muscle cells and thrombus in the penises were detected by electron microscopy in the control group, but not in the PNS groups (10 and 150 mg/kg). The results suggest that PNS improves erectile function in diabetic rats. This improvement was associated with increased Akt expression, suppressed oxidative stress and restored functions of endothelial cells and smooth muscle cells in the penis.

The potential mechanism of ursolic acid in the treatment of bladder cancer based on network pharmacology and molecular docking.

OBJECTIVE: This study explored the potential molecular mechanisms of ursolic acid (UA) in bladder cancer treatment using network pharmacology and molecular docking. METHODS: The Traditional Chinese Medicine Systems Pharmacology and UniProt databases were used to screen potential targets of UA. Relevant bladder cancer target genes were extracted using the GeneCards database. All data were pooled and intercrossed to obtain common target genes of UA and bladder cancer. Gene Ontology functional annotation and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed. Molecular docking was conducted to verify the possible binding conformation between UA and bladder cancer cells. Then, in vitro experiments were performed to further validate the predicted results. RESULTS: UA exerts anti-tumor effects on bladder cancer through multiple targets and pathways. Molecular docking indicated that UA undergoes stable binding with the proteins encoded by the top six core genes (STAT3, VEGFA, CASP3, TP53, IL1B, and CCND1). The in vitro experiments verified that UA can induce bladder cancer cell apoptosis by regulating the PI3K/Akt signaling pathway. CONCLUSIONS: Our study illustrated the potential mechanism of UA in bladder cancer based on network pharmacology and molecular docking. The results will provide scientific references for follow-up studies and clinical treatment.

Deep learning system for malignancy risk prediction in cystic renal lesions: a multicenter study.

OBJECTIVES: To develop an interactive, non-invasive artificial intelligence (AI) system for malignancy risk prediction in cystic renal lesions (CRLs). METHODS: In this retrospective, multicenter diagnostic study, we evaluated 715 patients. An interactive geodesic-based 3D segmentation model was created for CRLs segmentation. A CRLs classification model was developed using spatial encoder temporal decoder (SETD) architecture. The classification model combines a 3D-ResNet50 network for extracting spatial features and a gated recurrent unit (GRU) network for decoding temporal features from multi-phase CT images. We assessed the segmentation model using sensitivity (SEN), specificity (SPE), intersection over union (IOU), and dice similarity (Dice) metrics. The classification model's performance was evaluated using the area under the receiver operator characteristic curve (AUC), accuracy score (ACC), and decision curve analysis (DCA). RESULTS: From 2012 to 2023, we included 477 CRLs (median age, 57 [IQR: 48-65]; 173 men) in the training cohort, 226 CRLs (median age, 60 [IQR: 52-69]; 77 men) in the validation cohort, and 239 CRLs (median age, 59 [IQR: 53-69]; 95 men) in the testing cohort (external validation cohort 1, cohort 2, and cohort 3). The segmentation model and SETD classifier exhibited excellent performance in both validation (AUC = 0.973, ACC = 0.916, Dice = 0.847, IOU = 0.743, SEN = 0.840, SPE = 1.000) and testing datasets (AUC = 0.998, ACC = 0.988, Dice = 0.861, IOU = 0.762, SEN = 0.876, SPE = 1.000). CONCLUSION: The AI system demonstrated excellent benign-malignant discriminatory ability across both validation and testing datasets and illustrated improved clinical decision-making utility. CRITICAL RELEVANCE STATEMENT: In this era when incidental CRLs are prevalent, this interactive, non-invasive AI system will facilitate accurate diagnosis of CRLs, reducing excessive follow-up and overtreatment. KEY POINTS: The rising prevalence of CRLs necessitates better malignancy prediction strategies. The AI system demonstrated excellent diagnostic performance in identifying malignant CRL. The AI system illustrated improved clinical decision-making utility.

Gene silence-induced downregulation of survivin inhibits bladder cancer cells.

Urinary bladder cancer accounts for approximately 3% of all cancers in humans. Treatment for urinary bladder is not satisfactory. The present study aims to elucidate the effect of gene silencing of survivin on the inhibition of bladder cancer cells. In this study, we constructed survivin shRNA-carrying lentiviral vectors. Bladder cancer cell lines, T24 cells and BJ cells, were transduced with the constructed shRNA of survivin. The frequency of apoptotic bladder cancer cells was assessed by flow cytometry. The results showed that transfection with survivin shRNA significantly inhibited cell proliferation of both T24 and BJ cells. Most T24 and BJ cells accumulated at the G2/M stage; a portion of them was at sub-G1 stage. An increase in the fraction of bladder cancer cells undergoing apoptosis was noted. Among eight apoptosis-associated proteins, the amounts of BAX and BAD were significantly increased in the survivin-deficient bladder cancer cells. The findings suggest that survivin may be a therapeutic target of bladder cancer to selectively inhibit cell proliferation of bladder cancer cells.

WITHDRAWN: Survivin knock down by short hairpin RNA inhibits proliferation and induces apoptosis in EJ bladder cancer cells.

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Bladder cancer cell­secreted exosomal miR­21 activates the PI3K/AKT pathway in macrophages to promote cancer progression.

Tumour­associated macrophages (TAMs) compose a major component of the tumour microenvironment and form in this microenvironment prior to cancer metastasis. However, the detailed mechanisms of TAM remodelling in the context of bladder cancer have not been clearly defined. The present study collected exosomes from the conditioned medium of human bladder T24 cancer cells. The effects of macrophages treated with exosomes derived from T24 cells on bladder cancer cell migration and invasion were analysed by Transwell assays. The expression levels of endogenous and exosomal microRNA­21 (miR­21) were examined by reverse transcription­quantitative PCR, while the expression level of the target protein was analysed by western blot analysis. Luciferase reporter plasmids and mutants were used to confirm direct targeting. The effects of miR­21 on bladder cancer cell migration and invasion were analysed by Transwell and Matrigel assays following miR­21 transfection. It was identified that exosomes derived from bladder cancer cells polarized THP­1 cell­derived macrophages into the M2 phenotype, and TAM­mediated pro­migratory and pro­invasive activity was determined. Moreover, it was found that miR­21 was highly expressed in exosomes derived from bladder cancer cells as well as in macrophages treated with exosomes. In addition, macrophages transfected with miR­21 exhibited M2 polarization and promoted T24 cell migratory and invasive ability. Mechanistically, exosomal miR­21 derived from bladder cancer cells inhibited phosphatase and tensin homolog activation of the PI3K/AKT signalling pathway in macrophages and enhanced STAT3 expression to promote M2 phenotypic polarization. The present results suggest that exosomal miR­21 can promote cancer progression by polarizing TAMs.

Inhibiting autophagy overcomes docetaxel resistance in castration-resistant prostate cancer cells.

BACKGROUND: This study investigates the docetaxel-resistant mechanism and explores the effect of tea polyphenols (TP) on autophagy and its related mechanism in human castration-resistant prostate cancer (CRPC) cell lines PC3 and DU145. METHODS: Immunofluorescence assay and annexin V-FITC/PI double staining flow cytometry were used to analyze the apoptosis and autophagy of PC3 and DU145 cells. The expression of autophagy-related proteins was detected by western bolt. RESULTS: Docetaxel could induce autophagy and apoptosis, together with the expression increase in p-JNK, p-Bcl-2 and Beclin1. The level of autophagy was remarkably decreased, but apoptosis was increased after combining with TP. In addition, the expression of p-mTOR was increased after combining with TP. CONCLUSION: Docetaxel induces protective autophagy in CRPC cells by JNK pathway activation and then Bcl-2 phosphorylation and Beclin1 dissociation. TP activates mTOR pathway, which ultimately inhibits docetaxel-induced autophagy and improves therapeutic efficacy of docetaxel in CRPC cells.

Cough Test during Tension-Free Vaginal Tape Procedure in Preventing Postoperative Urinary Retention.

Objective. To discuss the practical value of the cough test during the tension-free vaginal tape (TVT) procedure. Methods. In the first group, 41 patients of female stress incontinence received TVT operations which were performed according to the Ulmsten's method strictly, only that the stress of tape was adjusted in light of the cough test. In the second group, 44 patients of female stress incontinence received TVT operations in which the tape was put under the urethral tract without stress, not adjusted by cough test. Results. The cure rate was 38/41 (92.6%) in the cough test group and 41/44 (93.1%) in the noncough test group; detrusor pressure-uroflow study indicated that there were 11 cases in the obstruction zone in the cough test group while only 3 cases were in the obstruction zone in the noncough test group; 4 cases of urinary retention and 5 cases of voiding dysfunction were found in the cough test group, while difficulties of urination were not found in the non-cough test group. Conclusion. Adjusting the tape stress in accordance with the cough test during the TVT can increase the opportunity of urinary retention or difficulty of urination after operation. So there is no benefit of the cough test during tension-free vaginal tape procedure in preventing post-operative urinary retention.

Laparoscopic radical cystectomy for bladder cancer with prostatic and neurovascular sparing: initial experience.

PURPOSE: This study presents our initial experience with laparoscopic radical cystectomy with preservation of the neurovascular bundles and partial prostate for the treatment of bladder cancer. METHODS: Thirty-seven patients with bladder cancer were selected for the study between June 2007 and December 2009. The criteria for patient selection included prostate-specific antigen level below 4.0 ng/mL; negative involvement of the trigone and/or prostatic urethra; and no self-reported erectile dysfunction. The surgical procedure included laparoscopic prostate- and neurovascular bundles-sparing cystectomy with an ileal neobladder construction. Mean follow-up was 18 months. RESULTS: All patients underwent laparoscopic resection without requiring a traditional open procedure. The mean operation time was 215 min with a mean volume of intraoperative hemorrhage of 190 mL. After removal of the urinary catheter, all patients had a daytime urinary continence; six had a short period of nighttime urinary incontinence. Most patients reported a strong desire for sexual activity and were able to complete sexual intercourse without auxiliary measures at 3 months postoperatively. Grade-3 complications developed in 2 patients graded by the classification of Clavien system. One patient was diagnosed with pelvic recurrence 16 months postoperatively. CONCLUSION: The laparoscopic radical cystectomy with a partial prostate preservation offers the advantages of a high continence, minimal impairment of erectile function, and low recurrence rate.

Transplantation of endothelial progenitor cells transfected with VEGF165 to restore erectile function in diabetic rats.

The present study investigated the effect of transplanting endothelial progenitor cells (EPCs) transfected with the vascular endothelial growth factor gene (VEGF165) into the corpora cavernosa of rats with diabetic erectile dysfunction (ED). A rat model of diabetic ED was constructed via intraperitoneal injection of streptozotocin. After streptozotocin treatment, pre-treated EPCs from each of three groups of rats were transplanted into their corpora cavernosa. Our results, following intracavernosal pressure (ICP) monitoring, showed that ICP increased significantly among rats in the trial group when compared to the results from rats in the blank-plasmid and control groups during basal conditions and electrical stimulation (P<0.01 for both comparisons). Histological examination revealed extensive neovascularisation in the corpora cavernosa of rats in the trial group. Fluorescence microscopy indicated that many of the transplanted EPCs in the trial group survived, differentiated into endothelial cells and integrated into the sites of neovascularisation. Based on the results of this study, we conclude that transplantation of VEGF165-transfected EPCs into the corpora cavernosa of rats with diabetic ED restores erectile function.