Background Inactivation of the tumor suppressor p53 is critical for pathogenesis of glioma, in particular glioblastoma multiforme (GBM). MDM2, the main negative regulator of p53, binds to and forms a stable complex with p53 to regulate its activity. Hitherto, it is unclear whether the stability of the p53/MDM2 complex is affected by lncRNAs, in particular circular RNAs that are usually abundant and conserved, and frequently implicated in different oncogenic processes. Methods RIP-seq and RIP-qPCR assays were performed to determine the most enriched lncRNAs (including circular RNAs) bound by p53, followed by bioinformatic assays to estimate the relevance of their expression with p53 signaling and gliomagenesis. Subsequently, the clinical significance of CDR1as was evaluated in the largest cohort of Chinese glioma patients from CGGA (n = 325), and its expression in human glioma tissues was further evaluated by RNA FISH and RT-qPCR, respectively. Assays combining RNA FISH with protein immunofluorescence were performed to determine co-localization of CDR1as and p53, followed by CHIRP assays to confirm RNA-protein interaction. Immunoblot assays were carried out to evaluate protein expression, p53/MDM2 interaction and p53 ubiquitination in cells in which CDR1as expression was manipulated. After AGO2 or Dicer was knocked-down to inhibit miRNA biogenesis, effects of CDR1as on p53 expression, stability and activity were determined by immunoblot, RT-qPCR and luciferase reporter assays. Meanwhile, impacts of CDR1as on DNA damage were evaluated by flow cytometric assays and immunohistochemistry. Tumorigenicity assays were performed to determine the effects of CDR1as on colony formation, cell proliferation, the cell cycle and apoptosis (in vitro), and on tumor volume/weight and survival of nude mice xenografted with GBM cells (in vivo). Results CDR1as is found to bind to p53 protein. CDR1as expression decreases with increasing glioma grade and it is a reliable independent predictor of overall survival in glioma, particularly in GBM. Through a mechanism independent of acting as a miRNA sponge, CDR1as stabilizes p53 protein by preventing it from ubiquitination. CDR1as directly interacts with the p53 DBD domain that is essential for MDM2 binding, thus disrupting the p53/MDM2 complex formation. Induced upon DNA damage, CDR1as may preserve p53 function and protect cells from DNA damage. Significantly, CDR1as inhibits tumor growth in vitro and in vivo, but has little impact in cells where p53 is absent or mutated. Conclusions Rather than acting as a miRNA sponge, CDR1as functions as a tumor suppressor through binding directly to p53 at its DBD region to restrict MDM2 interaction. Thus, CDR1as binding disrupts the p53/MDM2 complex to prevent p53 from ubiquitination and degradation. CDR1as may also sense DNA damage signals and form a protective complex with p53 to preserve p53 function. Therefore, CDR1as depletion may play a potent role in promoting tumorigenesis through down-regulating p53 expression in glioma. Our results broaden further our understanding of the roles and mechanism of action of circular RNAs in general and CDR1as in particular, and can potentially open up novel therapeutic avenues for effective glioma treatment.

Diabetes Mellitus (DM)-induced bladder dysfunction is predominantly due to the long-term oxidative stress caused by hyperglycemia. Grape seed proanthocyanidin extract (GSPE) has been reported to possess a broad spectrum of pharmacological and therapeutic properties against oxidative stress. However, its protective effects against diabetic bladder dysfunction have not been clarified. This study focuses on the effects of GSPE on bladder dysfunction in diabetic rats induced by streptozotocin. After 8 weeks of GSPE administration, the bladder function of the diabetic rats was improved significantly, as indicated by both urodynamics analysis and histopathological manifestation. Moreover, the disordered activities of antioxidant enzymes (SOD and GSH-Px) and abnormal oxidative stress levels were partly reversed by treatment with GSPE. Furthermore, the level of apoptosis in the bladder caused by DM was decreased following the administration of GSPE according to the Terminal Deoxynucleotidyl Transferase (TdT)-mediated dUTP Nick-End Labeling (TUNEL) assay. Additionally, GSPE affected the expression of apoptosis-related proteins such as Bax, Bcl-2 and cleaved caspase-3. Furthermore, GSPE showed neuroprotective effects on the bladder of diabetic rats, as shown by the increased expression of nerve growth factor (NGF) and decreased expression of the precursor of nerve growth factor (proNGF). GSPE also activated nuclear erythroid2-related factor2 (Nrf2), which is a key antioxidative transcription factor, with the concomitant elevation of downstream hemeoxygenase-1 (HO-1). These findings suggested that GSPE could ameliorate diabetic bladder dysfunction and decrease the apoptosis of the bladder in diabetic rats, a finding that may be associated with its antioxidant activity and ability to activate the Nrf2 defense pathway.

BackgroundMicroRNA-222 (miR-222) has been shown to play a potential oncogenic role in bladder cancer. The aim of this study was to evaluate the expression of miR-222 in bladder cancer and its potential relevance to clinicopathological characteristics and patient survival.MethodsSurgical specimens of cancer tissue and adjacent normal tissue were obtained from 97 patients with bladder cancer. The relative expression levels of miR-222 in the cancer and the normal adjacent tissue were measured by quantitative reverse-transcriptase PCR. We analyzed their correlation with clinicopathological parameters and prognostic value.ResultsThe expression level of miR-222 was significantly higher in tumor tissues than in corresponding non-cancerous tissues (5.46 ± 1.45 versus 1.92 ± 0.65, P < 0.0001), and a high expression of miR-222 was found to be significantly associated with tumor grade (P = 0.003) and tumor stage (P = 0.005). The miR-222 expression level was classified as high or low in relation to the median value (cutoff value = 5.15). Kaplan-Meier analysis showed that patients with higher levels of miR-222 had significantly poorer survival than those with lower expression of this miRNA in patients, with a 5-year overall survival of 29.53% and 52.75%, respectively (P = 0.0034). In the multivariate Cox proportional hazards analysis, which included miR-222 level, tumor grade, tumor stage, and tumor number, high miR-222 expression was independently associated with poor survival (P < 0.001; hazard ratio 6.17; 95% CI 2.33 to 10.39).ConclusionmiR-222 overexpression is involved in the poor prognosis of bladder cancer and can be used as a biomarker for selection of cases requiring special attention.

Bufalin (BF) is a cardiotonic steroid that has recently been found to have substantial anticancer activity; however, more efforts should be directed toward clarifying the detailed molecular mechanisms underlying this activity. BF could exert its anticancer effect by inducing apoptosis in various human cancer cells and thus triggering autophagic cancer cell death. The anti-inflammatory activities of BF are potentially important for its anticancer functions. Notably, some promising synthetic BF derivatives, including poly (ethylene glycol)-based polymeric prodrug of BF and BF211, have shown potent anticancer activity. Additionally, clinical trials regarding the use of BF-related agents in patients have supported the positive effect of BF as an anticancer treatment. Currently, large-scale randomized, double-blind, placebo or positive drug parallel controlled studies are required to confirm the anticancer potential of BF in various cancer types in the clinical setting. The present review will evaluate the potential mechanisms mediated by BF in intracellular signaling events in cancer cells and various promising BF derivatives that may have greater anticancer activity, thereby clarifying BF-mediated anticancer effects. The experimental and clinical results reviewed strongly emphasize the importance of this topic in future investigations. Contents

This study in α-chloralose-anesthetized cats discovered an excitatory peroneal nerve-to-bladder reflex. A urethral catheter was used to infuse the bladder with saline and record bladder pressure changes. Electrical stimulation was applied to the superficial peroneal nerve to trigger reflex bladder activity. With the bladder distended at a volume ~90% of bladder capacity, superficial peroneal nerve stimulation (PNS) at 1-3 Hz and threshold (T) intensity for inducing muscle twitching on the posterior thigh induced large-amplitude (40-150 cmHO) bladder contractions. PNS (1-3 Hz, 1-2T) applied during cystometrograms (CMGs) when the bladder was slowly (1-3 ml/min) infused with saline significantly ( < 0.01) reduced bladder capacity to ~80% of the control capacity and significantly ( < 0.05) enhanced reflex bladder contractions. To determine the impact of PNS on tibial nerve stimulation (TNS)-induced changes in bladder function, PNS was delivered following TNS. TNS of 30-min duration produced long-lasting poststimulation inhibition and significantly ( < 0.01) increased bladder capacity to 140.5 ± 7.6% of the control capacity. During the post-TNS inhibition period, PNS (1-3 Hz, 1-4T) applied during CMGs completely restored bladder capacity to the control level and significantly ( < 0.05) increased the duration of reflex bladder contractions to ~200% of control. The excitatory peroneal nerve-to-bladder reflex could also be activated by transcutaneous PNS using skin surface electrodes attached to the dorsal surface of the foot. These results raise the possibility of developing novel neuromodulation therapies to treat underactive bladder and nonobstructive urinary retention.