Umbilical Cord Mesenchymal Stem Cells Overexpressing Heme Oxygenase-1 Promotes Symptoms Recovery in Cystitis Rats by Alleviating Neuroinflammation.

Interstitial cystitis/bladder pain syndrome (IC/BPS) seriously reduces the patient's quality of life, yet current therapies only provide partial relief. In the spinal dorsal horn (SDH), neuroinflammation plays a pivotal role in the development of IC. Injection of human umbilical cord mesenchymal stem cells (hUMSCs) to reduce inflammation is an effective strategy, and heme oxygenase-1 (HO-1) exhibits anti-nociceptive effect in neuroinflammatory pain. This study aimed to test the therapeutic effects of hUMSCs overexpressing HO-1 on cyclophosphamide-induced cystitis rat model. Cystitis rats were transplanted with altered cells and then assessed for 3 weeks. A series of behavioral measurements would be trial including suprapubic mechanical allodynia, depressive-like behaviors, micturition frequency, and short-term memory function. Additionally, western blot, immunofluorescence staining, and ELISA kit test for anti-inflammation effect. HUMSCs were capable of being transduced to overexpress HO-1. Injection of hUMSCs overexpressing HO-1 was more effective than hUMSCs alone in alleviating behavioral symptoms in rats. Furthermore, hUMSCs overexpressing HO-1 inhibited the activation of glial and TLR4/p65/NLRP3 pathway, decreased the levels of pro-inflammatory cytokines in the SDH region. Surprisingly, it markedly increased anti-inflammatory cytokine IL-10, reduced MDA content, and protected GSH concentrations in local environment. Our results suggest that injecting hUMSCs overexpressing HO-1 intrathecally can significantly promote functional outcomes in cystitis rats by reducing neuroinflammation, at least, partly through downregulating TLR4/p65/NLRP3 signaling pathway in the SDH region. This cell therapy affords a new strategy for IC/BPS treatment.

Identification of tumor antigens and immune landscapes for bladder urothelial carcinoma mRNA vaccine.

Background: Bladder urothelial carcinoma (BLCA) is associated with high mortality and recurrence. Although mRNA-based vaccines are promising treatment strategies for combating multiple solid cancers, their efficacy against BLCA remains unclear. We aimed to identify potential effective antigens of BLCA for the development of mRNA-based vaccines and screen for immune clusters to select appropriate candidates for vaccination. Methods: Gene expression microarray data and clinical information were retrieved from The Cancer Genome Atlas and GSE32894, respectively. The mRNA splicing patterns were obtained from the SpliceSeq portal. The cBioPortal for Cancer Genomics was used to visualize genetic alteration profiles. Furthermore, nonsense-mediated mRNA decay (NMD) analysis, correlation analysis, consensus clustering analysis, immune cell infiltration analysis, and weighted co-expression network analysis were conducted. Results: Six upregulated and mutated tumor antigens related to NMD, and infiltration of APCs were identified in patients with BLCA, including HP1BP3, OSBPL9, SSH3, ZCCHC8, FANCI, and EIF4A2. The patients were subdivided into two immune clusters (IC1 and IC2) with distinct clinical, cellular and molecular features. Patients in IC1 represented immunologically 'hot' phenotypes, whereas those in IC2 represented immunologically 'cold' phenotypes. Moreover, the survival rate was better in IC2 than in IC1, and the immune landscape of BLCA indicated significant inter-patient heterogeneity. Finally, CALD1, TGFB3, and ANXA6 were identified as key genes of BLCA through WGCNA analysis, and their mRNA expression levels were measured using qRT-PCR. Conclusion: HP1BP3, OSBPL9, SSH3, ZCCHC8, FANCI, and EIF4A2 were identified as potential antigens for developing mRNA-based vaccines against BLCA, and patients in IC2 might benefit more from vaccination.

PXDNL activates the motility of urothelial bladder carcinoma cells through the Wnt/ß-catenin pathway and has a prognostic value.

AIMS: Although aberrant expression of peroxidasin-like (PXDNL) has been associated with carcinogenesis, its potential role in the Urothelial Carcinoma of the Bladder (UCB) remains unknown. The present study aimed to explore the role of PXDNL in UCB carcinogenesis and its potential clinical value. MAIN METHODS: Based on The Cancer Genome Atlas (TCGA) data, bioinformatics was used to explore the potential clinical value of PXDNL. Wound healing and Transwell invasion assays were employed for the purpose of assessing the cell motility, while the Western Blotting experiments were utilized for investigating the protein expression pattern of PXDNL in UCB and investigating the Epithelial-to-Mesenchymal Transition (EMT) and Wnt/ß-catenin pathways for understanding the probable mechanisms involved. KEY FINDS: PXDNL mRNA was overexpressed in UCB tissues and indicated a poor prognosis. High PXDNL mRNA levels were also associated with advanced clinicopathological features and were regarded as independent prognostic factors for UCB. However, PXDNL showed a weak correlation with immune cell infiltration in UCB. In addition, the findings of the study verified that the existing form of the PXDNL protein was 57-kDa and it was upregulated in the UCB cell lines and tissue samples. Furthermore, silencing PXDNL inhibited, while overexpressing PXDNL promoted EMT and motility of UCB cells in vitro. Mechanistic studies showed that PXDNL activated UCB cell motility via the Wnt/ß-catenin pathway. SIGNIFICANCE: The results reveal a novel molecular target that could be further explored for developing preventive, predictive, and individualized treatment strategies for UCB.

Alternative splicing patterns reveal prognostic indicator in muscle-invasive bladder cancer.

BACKGROUND: Bladder cancer is one of the most lethal malignancy in urological system, and 20-25% of bladder cancer patients are muscle invasive with unfavorable prognosis. However, the role of alternative splicing (AS) in muscle-invasive bladder cancer (MIBC) remains to be elucidated. METHODS: Percent spliced in (PSI) data obtained from the Cancer Genome Atlas (TCGA) SpliceSeq database (n = 394) were utilized to evaluate the AS events in MIBC. Prognosis-associated AS events were screened out by univariate Cox regression. LASSO Cox regression was used to identify reliable prognostic patterns in a training set and further validated in a test set. Splicing regulatory networks were constructed by correlations between PSI of AS events and RNA expression of splicing factors. RESULTS: As a result, a total of 2589 prognosis-related AS events in MIBC were identified. Pathways of spliceosomal complex (FDR = 0.017), DNA-directed RNA polymerase II, core complex (FDR = 0.032), and base excision repair (FDR = 0.038) were observed to be significantly enriched. Additionally, we noticed that most of the prognosis-related AS events were favorable factors. According to the LASSO and multivariate Cox regression analyses, 15-AS-based signature was established with the area under curve (AUC) of 0.709, 0.823, and 0.857 at 1-, 3-, and 5- years, respectively. The MIBC patients were further divided into high- and low-risk groups based on median risk sores. Interestingly, we observed that the prevalence of FGFR3 with mutations and focal amplification was significantly higher in low-risk group. Functional and immune infiltration analysis suggested potential signaling pathways and distinct immune states between these two groups. Moreover, splicing correlation network displayed a regulatory mode of prognostic splicing factors (SF) in MIBC patients. CONCLUSIONS: This study not only provided novel insights into deciphering the possible mechanism of tumorgenesis and pathogenesis but also help refine risk stratification systems and potential treatment of decision-making for MIBC.

Prediction of overall survival based upon a new ferroptosis-related gene signature in patients with clear cell renal cell carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the most common and lethal renal cell carcinoma (RCC) histological subtype. Ferroptosis is a newly discovered programmed cell death and serves an essential role in tumor occurrence and development. The purpose of this study is to analyze ferroptosis-related gene (FRG) expression profiles and to construct a multi-gene signature for predicting the prognosis of ccRCC patients. METHODS: RNA-sequencing data and clinicopathological data of ccRCC patients were downloaded from The Cancer Genome Atlas (TCGA). Differentially expressed FRGs between ccRCC and normal tissues were identified using 'limma' package in R. GO and KEGG enrichment analyses were conducted to elucidate the biological functions and pathways of differentially expressed FRGs. Consensus clustering was used to investigate the relationship between the expression of FRGs and clinical phenotypes. Univariate and the least absolute shrinkage and selection operator (LASSO) Cox regression analysis were used to screen genes related to prognosis and construct the optimal signature. Then, a nomogram was established to predict individual survival probability by combining clinical features and prognostic signature. RESULTS: A total of 19 differentially expressed FRGs were identified. Consensus clustering identified two clusters of ccRCC patients with distinguished prognostic. Functional analysis revealed that metabolism-related pathways were enriched, especially lipid metabolism. A 7-gene ferroptosis-related prognostic signature was constructed to stratify the TCGA training cohort into high- and low-risk groups where the prognosis was significantly worse in the high-risk group. The signature was identified as an independent prognostic indicator for ccRCC. These findings were validated in the testing cohort, the entire cohort, and the International Cancer Genome Consortium (ICGC) cohort. We further demonstrated that the signature-based risk score was highly associated with the ccRCC progression. Further stratified survival analysis showed that the high-risk group had a significantly lower overall survival (OS) rate than those in the low-risk group. Moreover, we constructed a nomogram that had a strong ability to forecast the OS of the ccRCC patients. CONCLUSIONS: We constructed a ferroptosis-related prognostic signature, which might provide a reliable prognosis assessment tool for the clinician to guide clinical decision-making and outcomes research.

Activation of translocator protein alleviates mechanical allodynia and bladder dysfunction in cyclophosphamide-induced cystitis through repression of BDNF-mediated neuroinflammation.

BACKGROUND: Bladder pain syndrome/interstitial cystitis (BPS/IC) is a refractory disease accompanied by bladder-related pain and hyperactivity. Studies have shown that the translocator protein (TSPO) modulates neuroinflammation and central sensitisation associated with pain. Moreover, we previously demonstrated that brain-derived neurotrophic factor (BDNF) regulates neuroinflammation and mechanical allodynia in cyclophosphamide (CYP)-induced cystitis through activation of glial cells. Here, we aimed to explore whether activation of TSPO attenuates mechanical allodynia and bladder dysfunction by regulating BDNF induced neuroinflammation in a CYP-induced cystitis model. METHODS: Injection of CYP was performed to form a rat model of BPS/IC. The expression of TSPO was regulated by intrathecal injection of the TSPO agonist Ro5-4864. The von Frey filament test was applied to evaluate suprapubic allodynia. Bladder function was assessed using filling cystometry. Western blotting was used to detect the expression of TSPO, BDNF, GFAP, Iba-1, p-p38, p-JNK, TNF-α, and IL-1ß, and double immunofluorescence was performed to localise TSPO in the L6-S1 spinal dorsal horn (SDH). RESULTS: TSPO was activated in the SDH after CYP injection and was primarily colocalised with astrocytes. Ro5-4864 reversed mechanical allodynia and bladder dysfunction induced by CYP. Moreover, the upregulation of BDNF and activation of astrocytes and microglia was suppressed by Ro5-4864, resulting in downregulation of p-p38, p-JNK, TNF-α, and IL-1ß. CONCLUSIONS: Ro5-4864 alleviated mechanical allodynia and bladder dysfunction in the CYP model, possibly by inhibiting the elevation of BDNF and consequent activation of astrocytes and microglia induced neuroinflammation. TSPO may be a potential target for the treatment of BPS/IC. SIGNIFICANCE: This study examined the mechanism underlying the ability of the translocator protein to modulate bladder pain syndrome/interstitial cystitis.

N6-methyladenosine-related non-coding RNAs are potential prognostic and immunotherapeutic responsiveness biomarkers for bladder cancer.

BACKGROUND: Bladder cancer (BC) is a commonly occurring malignant tumor of the urinary system, demonstrating high global morbidity and mortality rates. BC currently lacks widely accepted biomarkers and its predictive, preventive, and personalized medicine (PPPM) is still unsatisfactory. N6-methyladenosine (m6A) modification and non-coding RNAs (ncRNAs) have been shown to be effective prognostic and immunotherapeutic responsiveness biomarkers and contribute to PPPM for various tumors. However, their role in BC remains unclear. METHODS: m6A-related ncRNAs (lncRNAs and miRNAs) were identified through a comprehensive analysis of TCGA, starBase, and m6A2Target databases. Using TCGA dataset (training set), univariate and least absolute shrinkage and selection operator (LASSO) regression analyses were performed to develop an m6A-related ncRNA-based prognostic risk model. Kaplan-Meier analysis of overall survival (OS) and receiver operating characteristic (ROC) curves were used to verify the prognostic evaluation power of the risk model in the GSE154261 dataset (testing set) from Gene Expression Omnibus (GEO). A nomogram containing independent prognostic factors was developed. Differences in BC clinical characteristics, m6A regulators, m6A-related ncRNAs, gene expression patterns, and differentially expressed genes (DEGs)-associated molecular networks between the high- and low-risk groups in TCGA dataset were also analyzed. Additionally, the potential applicability of the risk model in the prediction of immunotherapeutic responsiveness was evaluated based on the "IMvigor210CoreBiologies" data set. RESULTS: We identified 183 m6A-related ncRNAs, of which 14 were related to OS. LASSO regression analysis was further used to develop a prognostic risk model that included 10 m6A-related ncRNAs (BAALC-AS1, MIR324, MIR191, MIR25, AC023509.1, AL021707.1, AC026362.1, GATA2-AS1, AC012065.2, and HCP5). The risk model showed an excellent prognostic evaluation performance in both TCGA and GSE154261 datasets, with ROC curve areas under the curve (AUC) of 0.62 and 0.83, respectively. A nomogram containing 3 independent prognostic factors (risk score, age, and clinical stage) was developed and was found to demonstrate high prognostic prediction accuracy (AUC = 0.83). Moreover, the risk model could also predict BC progression. A higher risk score indicated a higher pathological grade and clinical stage. We identified 1058 DEGs between the high- and low-risk groups in TCGA dataset; these DEGs were involved in 3 molecular network systems, i.e., cellular immune response, cell adhesion, and cellular biological metabolism. Furthermore, the expression levels of 8 m6A regulators and 12 m6A-related ncRNAs were significantly different between the two groups. Finally, this risk model could be used to predict immunotherapeutic responses. CONCLUSION: Our study is the first to explore the potential application value of m6A-related ncRNAs in BC. The m6A-related ncRNA-based risk model demonstrated excellent performance in predicting prognosis and immunotherapeutic responsiveness. Based on this model, in addition to identifying high-risk patients early to provide them with focused attention and targeted prevention, we can also select beneficiaries of immunotherapy to deliver personalized medical services. Furthermore, the m6A-related ncRNAs could elucidate the molecular mechanisms of BC and lead to a new direction for the improvement of PPPM for BC. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13167-021-00259-w.

Notch1 Signaling Contributes to Mechanical Allodynia Associated with Cyclophosphamide-Induced Cystitis by Promoting Microglia Activation and Neuroinflammation.

AIMS: Notch1 signaling regulates microglia activation, which promotes neuroinflammation. Neuroinflammation plays an essential role in various kinds of pain sensation, including bladder-related pain in bladder pain syndrome/interstitial cystitis (BPS/IC). However, the impact of Notch1 signaling on mechanical allodynia in cyclophosphamide- (CYP-) induced cystitis is unclear. This study is aimed at determining whether and how Notch1 signaling modulates mechanical allodynia of CYP-induced cystitis. METHODS: CYP was peritoneally injected to establish a bladder pain syndrome/interstitial cystitis (BPS/IC) rat model. A γ-secretase inhibitor, DAPT, was intrathecally injected to modulate Notch1 signaling indirectly. Mechanical withdrawal threshold in the lower abdomen was measured with von Frey filaments using the up-down method. The expression of Notch1 signaling, Iba-1, OX-42, TNF-α, and IL-1ß in the L6-S1 spinal dorsal horn (SDH) was measured with Western blotting analysis and immunofluorescence staining. RESULTS: Notch1 and Notch intracellular domain (NICD) were both upregulated in the SDH of the cystitis group. Moreover, the expression of Notch1 and NICD was negatively correlated with the mechanical withdrawal threshold of the cystitis rats. Furthermore, treatment with DAPT attenuated mechanical allodynia in CYP-induced cystitis and inhibited microglia activation, leading to decreased production of TNF-α and IL-1ß. CONCLUSION: Notch1 signaling contributes to mechanical allodynia associated with CYP-induced cystitis by promoting microglia activation and neuroinflammation. Our study showed that inhibition of Notch1 signaling might have therapeutic value for treating pain symptoms in BPS/IC.

Case selection and implementation of tubeless percutaneous nephrolithotomy.

BACKGROUND: The tubeless percutaneous nephrolithotomy (PCNL) was proposed to eliminate the side effects of the nephrostomy tube in recent years, such as pain, channel infection, postoperative bleeding, and longer hospital stay. But there is neither clinical guidelines nor consensus about tubeless PCNL in clinical practice. The study is aimed to how to implement the tubeless PCNL step by step, including case selection preoperatively, improving the technique of the surgeon, making the correct decisions at the end of the procedure, which had not been previously examined. METHODS: From January 2017 to March 2018, 364 consecutive patients requiring PCNL were comprehensively analyzed preoperatively and patients were selected for scheduled tubeless PCNL based on four aspects. The selected patients were divided into two groups according to whether the nephrostomy tube was finally placed. The mean operative time, intraoperative blood loss, stone clearance rate, visual pain score, postoperative hospitalization days and perioperative complications were all evaluated. RESULTS: Based on the preoperative evaluation, 42 patients were selected for tubeless PCNL, among which there were finally 37 cases of completed tubeless PCNL. Compared with patients undergoing conventional PCNL, there were not statistical differences in the mean operative time (P=0.207) or intraoperative blood loss (P=0.450) in the tubeless group. Stone clearance rate was 100% in both groups. The visual pain scores in the tubeless PCNL group were lower on operation day (P=0.029), first postoperative day (P<0.001) and the day of discharge (P=0.025). The postoperative hospitalization for the tubeless PCNL group was shorter than that of the control group (P<0.001). No significant difference in grade 1 complications was seen (P=0.424), and no grade 2 or higher complications were observed in either group. CONCLUSIONS: Postoperative pain was significantly relieved and postoperative hospitalization was significantly shortened in the tubeless PCNL group. Tubeless PCNL is safe if patients are carefully selected using four criteria before operation, attention is paid to four key points and five confirmations are made during operation.

Inflammatory pseudotumor of Castleman disease and IgG4-related disease masquerading as kidney malignancy.

BACKGROUND: With widespread clinical application of imaging techniques, renal space-occupying lesions have been identified at an increasing frequency. Here, we report two rare cases, Castleman disease (CD) and IgG4-related disease (IgG4-RD), presenting primarily with the symptoms and imaging findings of kidney malignancy. CASE PRESENTATION: In case 1, an occupying lesion located in the right renal pelvis was detected using magnetic resonance imaging in a 32-year-old female who presented with hematuria and lumbago. First misdiagnosed as carcinoma of the renal pelvis, the patient underwent right radical nephroureterectomy. However, postoperative pathological and immunohistochemistry studies finally confirmed the diagnosis of CD. In case 2, a 45-year-old male presented with the chief complaint of anuria. Nephrostomy and renal biopsy indicated lymphoma, following which, antegrade urography and computed tomography urography were performed, which revealed bilateral hydronephrosis and mass lesions around the renal pelvis. Partial resection of the masses and frozen section examination indicated the diagnosis of CD. However, the results of postoperative histopathology and immunohistochemistry combined with serum IgG4 were consistent with IgG4-RD. Both the patients recovered well after drug treatment without recurrence of the diseases. CONCLUSIONS: Inflammatory pseudotumor of CD and IgG4-RD with kidney involvement are primarily diagnosed by postoperative histopathology and can pose a preoperative diagnostic challenge because these lesions can masquerade as kidney malignancy. Therefore, we recommend core biopsy as a nonnegligible procedure to evaluate renal masses and potentially prevent unnecessary surgical treatment.

Malignant priapism as the initial presentation induced by the metastasis of renal clear cell carcinoma after radical nephrectomy.

The priapism secondary to the tumor metastasis to penis is rare. We described a case of a patient with priapism caused by the metastasis of renal cell carcinoma after radical nephrectomy. The previous reported cases found in the literature were reviewed. The pathophysiology, diagnosis, management and prognosis were discussed. Renal cell carcinoma metastasis to penis usually represents a more advanced stage of disease and carries a poor prognosis. The therapy is only palliative.

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.

Houttuynia cordata Extract Ameliorates Bladder Damage and Improves Bladder Symptoms via Anti-Inflammatory Effect in Rats with Interstitial Cystitis.

The mechanism of interstitial cystitis/bladder pain syndrome (IC/BPS) remains unclear to date, but reports showed that bladder inflammation and increasing number of activating mast cells in bladder tissues were common in patients with IC/BPS. Houttuynia cordata is widely used in Chinese traditional medicine, and its function of anti-inflammation has been proved. The purpose of this study was to investigate the efficacy and possible mechanisms of the Houttuynia cordata (HC) extract in the treatment of interstitial cystitis/bladder pain syndrome (IC/BPS). In the current study, a total of 30 adult female rats were randomly divided into three groups: sham group (n = 10), cyclophosphamide + saline (CYP + NS) group (n = 10), and cyclophosphamide + Houttuynia cordata extract (CYP + HC) group (n = 10). The animal model of IC/BPS was induced with cyclophosphamide (75 mg/kg, intraperitoneal injection, once every 3 days for 10 days) in the CYP + NS group and CYP + HC group, and sham rats received a volume-matched injection of saline. After anesthesia with urethane (0.8 g/kg, intraperitoneal injection), intravesical administration of either saline (1 ml) or Houttuynia cordata extract (1 ml, 2 g/ml) was continued once per day for a week in the CYP + NS group and CYP + HC group, respectively. Subsequently, urinary frequency, nociceptive behaviors, cystometry, bladder weight, histological changes, and cytokine (IL-6, IL-8, TNF-α) concentration were evaluated and compared among the three groups. Variables including inflammatory grade, mast cell number, proportion of activated mast cells, bladder weight, cytokine concentration of bladder homogenates, and frequency of urination significantly increased in the CYP + NS group compared with the sham group (P < 0.01) and CYP + HC group (P < 0.01). Besides, compared with the CYP + NS group, longer intercontraction interval, bigger bladder capacity, higher nociceptive threshold, fewer number of mast cells, and lower proportion of activated mast cells were found in the CYP + HC group (P < 0.01). Our study demonstrated that the Houttuynia cordata extract can effectively inhibit mast cell proliferation and activation and downregulate proinflammatory cytokine in a rat model of IC/BPS induced with cyclophosphamide and might be potentially valuable for the treatment of IC/BPS.

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.

Suppression of TRPM7 Inhibited Hypoxia-Induced Migration and Invasion of Androgen-Independent Prostate Cancer Cells by Enhancing RACK1-Mediated Degradation of HIF-1α.

Transient receptor potential melastatin subfamily member 7 (TRPM7) was essential in the growth and metastatic ability of prostate cancer cells. However, the effects and the relevant molecular mechanisms of TRPM7 on metastasis of prostate cancer under hypoxic atmosphere remain unclear. This study investigated the role of TRPM7 in the metastatic ability of androgen-independent prostate cancer cells under hypoxia. First, data mining was carried out to disclose the relationship between the TRPM7 gene level and the survival of prostate cancer patients. Specific siRNAs were used to knockdown target genes. Western blotting and qPCR were employed to determine protein and gene expression, respectively. The gene transcription activity was evaluated by luciferase activity assay of promoter gene. The protein interaction was determined by coimmunoprecipitation. Wound healing and transwell assays were employed to evaluated cell migration and invasion, respectively. Open access database results showed that high expression of TRPM7 was closely related to the poor survival of prostate cancer patients. Hypoxia simultaneously increased TRPM7 expression and induced HIF-1α accumulation in androgen-independent prostate cancer cells. Knockdown of TRPM7 significantly promoted HIF-1α degradation through the proteasome and inhibited EMT changes in androgen-independent prostate cancer cells under hypoxic condition. Moreover, TRPM7 knockdown increased the phosphorylation of RACK1 and strengthened the interaction between RACK1 and HIF-1α but attenuated the binding of HSP90 to HIF-1α. Whereas knockdown of RACK1 increased the binding of HSP90 to HIF-1α. Furthermore, both TRPM7 and HIF-1α knockdown significantly suppressed hypoxia-induced Annexin A1 protein expression, and suppression of HIF-1α/Annexin A1 signaling significantly inhibited hypoxia-induced cell migration and invasion of androgen-independent prostate cancer cells. Our findings demonstrate that TRPM7 knockdown promotes HIF-1α degradation via an oxygen-independent mechanism involving increased binding of RAKC1 to HIF-1α, and TRPM7-HIF-1α-Annexin A1 signaling axis plays a crucial role in the EMT, cell migration, and invasion of androgen-independent prostate cancer cells under hypoxic conditions.

BDNF promotes activation of astrocytes and microglia contributing to neuroinflammation and mechanical allodynia in cyclophosphamide-induced cystitis.

BACKGROUND: Patients with interstitial cystitis/bladder pain syndrome (IC/BPS) often grieve over a low quality of life brought about by chronic pain. In our previous studies, we determined that neuroinflammation of the spinal dorsal horn (SDH) was associated with mechanisms of interstitial cystitis. Moreover, it has been shown that brain-derived neurotrophic factor (BDNF) participates in the regulation of neuroinflammation and pathological pain through BDNF-TrkB signaling; however, whether it plays a role in cyclophosphamide (CYP)-induced cystitis remains unclear. This study aimed to confirm whether BDNF-TrkB signaling modulates neuroinflammation and mechanical allodynia in CYP-induced cystitis and determine how it occurs. METHODS: Systemic intraperitoneal injection of CYP was performed to establish a rat cystitis model. BDNF-TrkB signaling was modulated by intraperitoneal injection of the TrkB receptor antagonist, ANA-12, or intrathecal injection of exogenous BDNF. Mechanical allodynia in the suprapubic region was assessed using the von Frey filaments test. The expression of BDNF, TrkB, p-TrkB, Iba1, GFAP, p-p38, p-JNK, IL-1ß, and TNF-α in the L6-S1 SDH was measured by Western blotting and immunofluorescence analysis. RESULTS: BDNF-TrkB signaling was upregulated significantly in the SDH after CYP was injected. Similarly, the expressions of Iba1, GFAP, p-p38, p-JNK, IL-1ß, and TNF-α in the SDH were all upregulated. Treatment with ANA-12 could attenuate mechanical allodynia, restrain activation of astrocytes and microglia and alleviate neuroinflammation. Besides, the intrathecal injection of exogenous BDNF further decreased the mechanical withdrawal threshold, promoted activation of astrocytes and microglia, and increased the release of TNF-α and IL-1ß in the SDH of our CYP-induced cystitis model. CONCLUSIONS: In our CYP-induced cystitis model, BDNF promoted the activation of astrocytes and microglia to release TNF-α and IL-1ß, aggravating neuroinflammation and leading to mechanical allodynia through BDNF-TrkB-p38/JNK signaling.

RNA m6A Methyltransferase METTL3 Promotes The Growth Of Prostate Cancer By Regulating Hedgehog Pathway.

PURPOSE: N6-methyladenosine (m6A) is the most abundant internal modification on eukaryotic mRNA and gained increasing attention recently. More and more evidence suggest that m6A methylation plays crucial role in tumor genesis and development. However, its role in prostate cancer remains largely unknown. METHODS: METTL3 expression status in prostate cancer was analyzed by using TCGA database and Western blotting. m6A content was analyzed by using RNA Methylation Quantification Kit. The role of METTL3 in prostate cancer cells was determined by proliferation, survival, colony formation, and invasion assays. The m6A level of GLI1 RNA was detected by methylated RNA immunoprecipitation (MeRIP) assay. In vivo role of METTL3 was studied on xenograft models. RESULTS: We found that m6A methyltransferase METTL3 was overexpressed in prostate cancer cell lines, together with increased m6A content. Functionally, silencing of METTL3 by shRNA in prostate cancer cell lines resulted in decreased m6A content, cell proliferation, survival, colony formation, and invasion. Interestingly, overexpression of wild-type METTL3 abrogated the repression effect of METTL3 depletion on m6A content, cell proliferation, survival, colony formation, and invasion, while the overexpression of m6A catalytic site mutant METTL3 was unable to rescue the inhibitory effect caused by METTL3 depletion. Further mechanism analysis demonstrated that METTL3 silence decreased the m6A modification and expression of GLI1, an important component of hedgehog pathway, which led to cell apoptosis. Moreover, depletion of METTL3 inhibited tumor growth in vivo. CONCLUSION: Our results suggested that the m6A methyltransferase METTL3 promotes the growth and motility of prostate cancer cells by regulating hedgehog pathway.

The Self-Retaining Barbed Suture for Parenchymal Repair in Laparoscopic Partial Nephrectomy: A Systematic Review and Meta-Analysis.

Objectives. The warm ischemia time (WIT) is key to successful laparoscopic partial nephrectomy (LPN). The aim of this study was to perform a meta-analysis comparing the self-retaining barbed suture (SRBS) with a non-SRBS for parenchymal repair during LPN. Methods. A systematic search of PubMed, Scopus, and the Cochrane Library was performed up to March 2018. Inclusion criteria for this study were randomized controlled trials (RCTs) and observational comparative studies assessing the SRBS and non-SRBS for parenchymal repair during LPN. Outcomes of interest included WIT, complications, overall operative time, estimated blood loss, length of hospital stay, and change of renal function. Results. One RCT and 7 retrospective studies were identified, which included a total of 461 cases. Compared with the non-SRBS, use of the SRBS for parenchymal repair during LPN was associated with shorter WIT (P < .00001), shorter overall operative time (P < .00001), lower estimated blood loss (P = .02), and better renal function preservation (P = .001). There was no significant difference between the SRBS and non-SRBS with regard to complications (P = .08) and length of hospital stay (P = .25). Conclusions. The SRBS for parenchymal repair during LPN can significantly shorten the WIT and overall operative time, decrease blood loss, and preserve renal function.

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.