BORA regulates cell proliferation and migration in bladder cancer.

BACKGROUND: Bladder cancer is having a gradually increasing incidence in China. Except for the traditional chemotherapy drugs, there are no emerging new drugs for almost 30 years in bladder cancer. New potential therapeutic targets and biomarkers are urgently needed. METHODS: BORA is the activator of kinase Aurora A and plays an important role in cell cycle progression. To investigate the function of BORA in BCa, we established BORA knockdown and overexpression cell models for in vitro studies, xenograft and pulmonary metastasis mouse models for in vivo studies. RESULTS: Our results indicated that BORA was upregulated in human bladder cancer (BCa) compared to the normal bladder and paracancerous tissues at transcriptional and translational levels. We found that BORA was positively related to BCa cell proliferation. Furthermore, BORA knockdown induced cell cycle arrest in G2/M phase while BORA overexpression decreased the proportion of cells in G2/M, associated with PLK1-CDC25C-CDK1 alteration. Interestingly, we observed that knockdown of BORA inhibited BCa cell migration and invasion, accompanied with alterations of epithelial-mesenchymal transition (EMT) pathway related proteins. In vivo studies confirmed the inhibition effect of BORA knockdown on BCa cell growth and migration. CONCLUSIONS: Our study indicates that BORA regulates BCa cell cycle and growth, meanwhile influences cell motility by EMT, and could be a novel biomarker and potential therapeutic target in BCa.

LPS impairs steroidogenesis and ROS metabolism and induces PPAR transcriptional activity to disturb estrogen/androgen receptor expression in testicular cells.

Inflammation can deregulate the testicular functions of steroidogenesis and spermatogenesis, consequently contributing to male infertility. Animals and cells treated with lipopolysaccharide (LPS) exhibit infection- and inflammation-induced testicular dysfunction. However, the precise mechanisms affecting steroidogenesis and spermatogenesis in response to LPS-treatment remain poorly understood. We isolated distinct testicular cells including spermatocytes, round spermatids and late spermatids to analyze distribution of peroxisome proliferator-activated receptor (PPAR) family, plays central roles in the regulation of metabolism. Our results suggested Pparα/Pparγ mRNA was highly expressed in late spermatids, while Pparß mRNA was highly expressed in round spermatids. To analyze the effect of LPS on testicular cells, we established an LPS infection model using primary Sertoli cells and testicular cell lines (TM4, GC2 and MLTC1). We observed that PPARγ and SIRT1 were concentrated in the nuclear region and that the mRNA expression levels of antioxidative enzymes (Cat and Homx1) and PPARγ were upregulated in primary Sertoli cells after LPS-treatment. Moreover, luciferase reporter gene assays of the testicular cell lines revealed that the activity of the PPAR response element (PPRE) was significantly increased. Importantly, the transcriptional activity of the androgen response element was significantly reduced, whereas activity of estrogen response element was strongly induced in LPS-treated TM4 cells, consistent with the RT-PCR results. Meanwhile, the qRT-PCR results revealed that the LPS-induced upregulation of Ar mRNA in MLTC1 cells and Erß mRNA in TM4 cells were significantly recovered after treatment with the specific PPARγ-antagonist GW9662. In addition, we also found that LPS induced alterations in enzymes involved in steroidogenesis in testicular cell lines. Taken together, our results revealed that LPS may induce PPAR transcriptional activity to disturb estrogen/androgen receptor expression and impair steroidogenesis and ROS metabolism in testicular cells.

PPARγ inhibition regulates the cell cycle, proliferation and motility of bladder cancer cells.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a member of the nuclear receptor family of ligand-activated transcription factors and plays an important role in regulating cell proliferation, inflammation and lipid and glucose homeostasis. Our results revealed that PPARγ was up-regulated in human bladder cancer (BCa) tissues both at transcriptional and translational levels. Moreover, down-regulation of PPARγ mRNA or inhibition of PPARγ function (using GW9662, antagonist of PPARγ) could significantly suppress the proliferation of BCa cells. Furthermore, the cell cycle arrested in G0/G1 phase was also induced by the down-regulated PPARγ possibly through AKT-mediated up-regulation of p21/p27, whereas no significant transformation of apoptosis was observed. In addition, knockdown or inhibition of PPARγ might reduce the invasion and migration of BCa cells by affecting epithelial-mesenchymal transition-related proteins through AKT/GSK3ß signalling pathway. Additionally, in vivo studies showed that BCa cell proliferation was significantly suppressed by GW9662. In conclusion, our results indicated that PPARγ might be crucial for BCa tumorigenesis by interfering with the motility and viability of BCa cells.

Fifteen hub genes associated with progression and prognosis of clear cell renal cell carcinoma identified by coexpression analysis.

Renal cell carcinoma (RCC) is the most common type of renal tumor, and the clear cell renal cell carcinoma (ccRCC) is the most frequent subtype. In this study, our aim is to identify potential biomarkers that could effectively predict the prognosis and progression of ccRCC. First, we used The Cancer Genome Atlas (TCGA) RNA-sequencing (RNA-seq) data of ccRCC to identify 2370 differentially expressed genes (DEGs). Second, the DEGs were used to construct a coexpression network by weighted gene coexpression network analysis (WGCNA). Moreover, we identified the yellow module, which was strongly related to the histologic grade and pathological stage of ccRCC. Then, the functional annotation of the yellow module and single-samples gene-set enrichment analysis of DEGs were performed and mainly enriched in cell cycle. Subsequently, 18 candidate hub genes were screened through WGCNA and protein-protein interaction (PPI) network analysis. After verification of TCGA's ccRCC data set, Gene Expression Omnibus (GEO) data set (GSE73731) and tissue validation, we finally identified 15 hub genes that can actually predict the progression of ccRCC. In addition, by using survival analysis, we found that patients of ccRCC with high expression of each hub gene were more likely to have poor prognosis than those with low expression. The receiver operating characteristic curve showed that each hub gene could effectively distinguish between localized and advanced ccRCC. In summary, our study indicates that 15 hub genes have great predictive value for the prognosis and progression of ccRCC, and may contribute to the exploration of the pathogenesis of ccRCC.

Fatty acid oxidation inhibitor etomoxir suppresses tumor progression and induces cell cycle arrest via PPARγ-mediated pathway in bladder cancer.

Tumor cells rely on aerobic glycolysis as their main energy resource (Warburg effect). Recent research has highlighted the importance of lipid metabolism in tumor progression, and certain cancers even turn to fatty acids as the main fuel. Related studies have identified alterations of fatty acid metabolism in human bladder cancer (BCa). Our microarray analysis showed that fatty acid metabolism was activated in BCa compared with normal bladder. The free fatty acid (FFA) level was also increased in BCa compared with paracancerous tissues. Inhibition of fatty acid oxidation (FAO) with etomoxir caused lipid accumulation, decreased adenosine triphosphate (ATP) and nicotinamide adenine dinucleotide phosphate (NADPH) levels, suppressed BCa cell growth in vitro and in vivo, and reduced motility of BCa cells via affecting epithelial-mesenchymal transition (EMT)-related proteins. Furthermore, etomoxir induced BCa cell cycle arrest at G0/G1 phase through peroxisome proliferator-activated receptor (PPAR) γ-mediated pathway with alterations in fatty acid metabolism associated gene expression. The cell cycle arrest could be reversed by PPARγ antagonist GW9662. Taken together, our results suggest that inhibition of FAO with etomoxir may provide a novel avenue to investigate new therapeutic approaches to human BCa.

Longer length improvement and more covert incision: a single-center, prospective study of two innovative surgical methods "one stitch" and "four stitch" for pediatric buried penis.

BACKGROUND: To introduce the detailed procedures of two innovative surgical options for pediatric buried penis and prospectively compare their efficacy and safety. METHODS: A single-center, non-randomized, prospective study was conducted at the Zhongnan Hospital of Wuhan University, where patients were operated on using the so-called "one stitch" (OS) or "four stitch" (FS) methods. The operation time, adverse events, and satisfaction were recorded for both groups. RESULTS: Finally, 156 patients underwent the so-called OS (n = 65) or FS (n = 91) method, with a follow-up rate of 86.5% (135/156). During the perioperative period, the FS group spent much longer in surgery (P < 0.001), had more blood loss (P < 0.001), and took longer to recover from edema (P < 0.001) than the OS group. In contrast to the satisfaction after 12 months' follow up, both the objective length improvement (2.5 ± 0.6 vs 3.8 ± 0.5 cm, P < 0.001) and subjective satisfaction percent (86 vs 95%, P = 0.678) in the FS group were superior to those in the OS group. No significant differences were detected in postoperative infection, stenosis circle, scar hyperplasia, and relapse. CONCLUSIONS: In conclusion, the two surgical options for pediatric buried penis are both safe and effective. The OS method has a simple procedure, so with shorter operation time and faster postoperative recovery; though the FS method with more complex procedure, patients can acquire a satisfactory improvement of penile length almost 4 cm and more covert incision at the midline of the scrotum. We primarily recommend the FS method for patients with moderate or severe buried cases; but for mild cases, we preferred the OS method.

Landscape analysis of alternative splicing in kidney renal clear cell carcinoma and their clinical significance.

A growing number of studies reveal that alternative splicing (AS) is associated with tumorigenesis, progression, and metastasis. Systematic analysis of alternative splicing signatures in renal cancer is lacking. In our study, we investigated the AS landscape of kidney renal clear cell carcinoma (KIRC) and identified AS predictive model to improve the prognostic prediction of KIRC. We obtained clinical data and gene expression profiles of KIRC patients from the TCGA database to evaluate AS events. The calculation results for seven types of AS events indicated that 46276 AS events from 10577 genes were identified. Next, we applied Cox regression analysis to identify 5864 prognostic-associated AS events. We used the Metascape database to verify the potential pathways of prognostic-associated AS. Moreover, we constructed KIRC prediction systems with prognostic-associated AS events by the LASSO Cox regression model. AUCs demonstrated that these prediction systems had excellent prognostic accuracy simultaneously. We identified 34 prognostic associated splicing factors (SFs) and constructed homologous regulatory networks. Furthermore, in vitro experiments were performed to validate the favorable effect of SFs FMR1 in KIRC. In conclusion, we overviewed AS events in KIRC and identified AS-based prognostic models to assist the survival prediction of KIRC patients. Our study may provide a novel predictive signature to improve the prognostic prediction of KIRC, which might facilitate KIRC patient counseling and individualized management.

Acyl-coenzyme A: cholesterol acyltransferase inhibitor avasimibe suppresses tumorigenesis and induces G1-phase cell-cycle arrest by activating PPARγ signaling pathway in bladder cancer.

Reprogramming of energy metabolism is one of the most important characteristics of tumors. Bladder cancer (BLCA) cells contain higher levels of cholesterol content compared to normal cells, and acyl-coenzyme A (CoA): cholesterol acyltransferase-1 (ACAT1) plays a crucial role in the esterification of cholesterol. Avasimibe is a drug that has been used in the treatment of atherosclerosis, and it can effectively inhibit ACAT1. We observed that ACAT1 was significantly up-regulated in BLCA and positively correlated with tumor grade. By avasimibe administration, the proliferation and migration ability of BLCA cells were reduced, while the production of ROS was strongly increased, accompanied by the up-regulated expression of ROS metabolism-related proteins SOD2 and catalase. Furthermore, BLCA cell cycle was arrested at the G1 phase, accompanied by the downregulation of cell cycle-related proteins (CCNA1/2, CCND1, CDK2 and CDK4), while the PPARγ was found to be up-regulated at both transcriptional and protein levels after avasimibe treatment. Then we found that the PPARγ antagonist GW9662 could reverse the effect of avasimibe on the cell cycle. Moreover, xenograft and pulmonary metastasis models further demonstrated that avasimibe could inhibit tumor cell growth and metastasis in vivo. Taken together, our results for the first time revealed that avasimibe can inhibit BLCA progression and metastasis, and PPARγ signaling pathway may play a key role in regulation of cell cycle distribution induced by avasimibe.

Decreased Nrf2 protein level and low sperm quality in intractable spermatocystitis.

ABSTRACT: To investigate the molecular etiology of low sperm quality in patients with intractable spermatocystitis, spermatozoa samples from patients with persistent hematospermia undergoing transurethral seminal vesiculoscopy and healthy volunteers were utilized. Spermatozoa samples were collected from the seminal vesicles through transurethral seminal vesiculoscopy or by masturbation ejaculation. Sperm quality was analyzed by a WLJY-9000 color semen analysis system. Measurement of tumor necrosis factor alpha (TNFα) and interleukin-6 (IL-6) in the seminal plasma was performed using enzyme-linked immunosorbent assay (ELISA). Measurement of H2O2 in the seminal plasma was performed with a hydrogen peroxide kit. The protein levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and phosphorylated-Nrf2 (p-Nrf2) were measured by western blot analysis and immunofluorescence assays. Low sperm quality parameters and increased levels of inflammatory cytokines (TNFα, IL-6, and H2O2) in the seminal plasma were detected among the semen samples from the patients with persistent hematospermia. Nrf2 and p-Nrf2 were strongly expressed in the nucleus and periphery of human sperm cells, according to the results of the immunofluorescence assays. The protein levels of Nrf2 and p-Nrf2 were significantly lower in the spermatozoa samples from patients with persistent hematospermia than in those from healthy volunteers with normal sperm motility. The results suggested that Nrf2 signaling might play a role in the low sperm quality of patients with intractable spermatocystitis.

Polydatin protects against calcium oxalate crystal-induced renal injury through the cytoplasmic/mitochondrial reactive oxygen species-NLRP3 inflammasome pathway.

BACKGROUND: Oxidative stress and inflammatory responses are critical factors in calcium oxalate (CaOx) crystal-induced renal injury. Reactive oxygen species (ROS) are usually produced in the cytoplasm and mitochondria and trigger the priming and activation of the NLRP3 inflammasome, thereby regulating cytokines and inflammation. Polydatin is a plant rhizome extract with anti-inflammatory, antioxidant, and antitumor effects. However, it remains not clear whether and how these pathophysiological processes exists in CaOx crystal-induced renal inflammatory injury. METHODS: Here, we measured the expression of the NLRP3 inflammasome, IL-18, IL-1ß, intracellular and mitochondrial ROS (mtROS) levels and relevant morphological changes in treated renal tubular epithelial cells (TECs) and stone-forming rats. The study further explored the action of intracellular ROS and mtROS on these inflammatory damage, and the beneficial effects and pathway of polydatin. RESULTS: We verified that CaOx crystal-induced cytoplasmic ROS and mtROS upregulation promoted the priming and activation of the NLRP3 inflammasome, thereby stimulating IL-18/1ß maturation and activation. Polydatin can relieve oxidative stress and inflammatory damage by decreasing ROS. We further demonstrated that mtROS is the main target for polydatin to exert the NLRP3 inflammasome-regulating function. The inhibition of mtROS can effectively relieve the inflammatory damage to TECs and kidney caused by CaOx crystal. CONCLUSION: These findings provide new insight into the relationship between mitochondrial damage and inflammation in nephrolithiasis and show that polydatin-mediated anti-inflammatory and antioxidative protection is a therapeutic strategy for, but not limited to, crystalline nephropathy.

Site-selected in situ polymerization for living cell surface engineering.

The construction of polymer-based mimicry on cell surface to manipulate cell behaviors and functions offers promising prospects in the field of biotechnology and cell therapy. However, precise control of polymer grafting sites is essential to successful implementation of biomimicry and functional modulation, which has been overlooked by most current research. Herein, we report a biological site-selected, in situ controlled radical polymerization platform for living cell surface engineering. The method utilizes metabolic labeling techniques to confine the growth sites of polymers and designs a Fenton-RAFT polymerization technique with cytocompatibility. Polymers grown at different sites (glycans, proteins, lipids) have different membrane retention time and exhibit differential effects on the recognition behaviors of cellular glycans. Of particular importance is the achievement of in situ copolymerization of glycomonomers on the outermost natural glycan sites of cell membrane, building a biomimetic glycocalyx with distinct recognition properties.

CTHRC1 Is a Prognostic Biomarker and Correlated With Immune Infiltrates in Kidney Renal Papillary Cell Carcinoma and Kidney Renal Clear Cell Carcinoma.

Kidney renal clear cell carcinoma (KIRC) and kidney renal papillary cell carcinoma (KIRP) are the most common RCC types. RCC has high immune infiltration levels, and immunotherapy is currently one of the most promising treatments for RCC. Collagen triple helix repeat containing 1 (CTHRC1) is an extracellular matrix protein that regulates tumor invasion and modulates the tumor microenvironment. However, the association of CTHRC1 with the prognosis and tumor-infiltrating lymphocytes of KIRP and KIRC has not been reported. We examined the CTHRC1 expression differences in multiple tumor tissues and normal tissues via exploring TIMER, Oncomine, and UALCAN databases. Then, we searched the Kaplan-Meier plotter database to evaluate the correlation of CTHRC1 mRNA level with clinical outcomes. Subsequently, the TIMER platform and TISIDB website were chosen to assess the correlation of CTHRC1 with tumor immune cell infiltration level. We further explored the causes of aberrant CTHRC1 expression in tumorigenesis. We found that CTHRC1 level was significantly elevated in KIRP and KIRC tissues relative to normal tissues. CTHRC1 expression associates with tumor stage, histology, lymph node metastasis, and poor clinical prognosis in KIRP. The CTHRC1 level correlates to tumor grade, stage, nodal metastasis, and worse survival prognosis. Additionally, CTHRC1 is positively related to different tumor-infiltrating immune cells in KIRP and KIRC. Moreover, CTHRC1 was closely correlated with the gene markers of diverse immune cells. Also, high CTHRC1 expression predicted a worse prognosis in KIRP and KIRC based on immune cells. Copy number variations (CNV) and DNA methylation might contribute to the abnormal upregulation of CTHRC1 in KIRP and KIRC. In conclusion, CTHRC1 can serve as a biomarker to predict the prognosis and immune infiltration in KIRP and KIRC.

The role and function of PPARγ in bladder cancer.

Peroxisome proliferator-activated receptor gamma (PPARγ), a member of the nuclear receptor superfamily, participates in multiple physiological and pathological processes. Extensive studies have revealed the relationship between PPARγ and various tumors. However, the expression and function of PPARγ in bladder cancer seem to be controversial. It has been demonstrated that PPARγ affects the occurrence and progression of bladder cancer by regulating proliferation, apoptosis, metastasis, and reactive oxygen species (ROS) and lipid metabolism, probably through PPARγ-SIRT1 feedback loops, the PI3K-Akt signaling pathway, and the WNT/ß-catenin signaling pathway. Considering the frequent relapses after chemotherapy, some researchers have focused on the relationship between PPARγ and chemotherapy sensitivity in bladder cancer. Moreover, the feasibility of PPARγ ligands as potential therapeutic targets for bladder cancer has been uncovered. Taken together, this review summarizes the relevant literature and our findings to explore the complicated role and function of PPARγ in bladder cancer.

Conditional reprogramming: Modeling urological cancer and translation to clinics.

Patient-derived models, including cell models (organoids and conditionally reprogrammed cells [CRCs]) and patient-derived xenografts, are urgently needed for both basic and translational cancer research. Conditional reprogramming (CR) technique refers to a co-culture system of primary human normal or tumor cells with irradiated murine fibroblasts in the presence of a Rho-associated kinase inhibitor to allow the primary cells to acquire stem cell properties and the ability to proliferate indefinitely in vitro without any exogenous gene or viral transfection. Considering its robust features, the CR technique may facilitate cancer research in many aspects. Under in vitro culturing, malignant CRCs can share certain genetic aberrations and tumor phenotypes with their parental specimens. Thus, tumor CRCs can promisingly be utilized for the study of cancer biology, the discovery of novel therapies, and the promotion of precision medicine. For normal CRCs, the characteristics of normal karyotype maintenance and lineage commitment suggest their potential in toxicity testing and regenerative medicine. In this review, we discuss the applications, limitations, and future potential of CRCs in modeling urological cancer and translation to clinics.

TRPM8 Inhibition Regulates the Proliferation, Migration and ROS Metabolism of Bladder Cancer Cells.

INTRODUCTION: Based on accumulating evidence, transient receptor potential (TRP) ion channels may play important roles in the occurrence and the progression of cancer. TRP melastatin 8 (TRPM8), a member of the TRP family, functions as a Ca2+-permeable channel and regulates various physiological and pathological processes. However, the effects of TRPM8 on bladder cancer (BCa) and its underlying mechanisms have not been elucidated. METHODS: BCa tissues and matched noncancerous tissues were collected to examine the expression of the TRPM8 mRNA and protein using qRT-PCR, Western blotting and immunofluorescence staining. Meanwhile, the effect of knockdown or inhibition of the activity of the TRPM8 protein on the proliferation, migration and ROS metabolism of bladder cancer cells was detected using the MTT assay, clonogenic survival assay, Transwell chamber migration assay, and reactive oxygen species (ROS) detection, respectively. Furthermore, a mouse model transplanted with BCa cells was established to assess tumor growth after TRPM8 expression was inhibited in vivo. RESULTS: Compared with the noncancerous tissues, the levels of TRPM8 in BCa tissues were significantly increased. Knockdown or inhibition of the activity of the TRPM8 protein in BCa cells reduced cell proliferation and migration. Moreover, the production of ROS was increased in cells treated with siTRPM8, which was accompanied by increased levels of Catalase, HO-1 and SOD2. Furthermore, a mouse model transplanted with the stable TRPM8-deficient T24 cell line was established, demonstrating that knockdown of TRPM8 delayed tumor growth in vivo. DISCUSSION: TRPM8 might play an essential for BCa tumor progression and metastasis by interfering with BCa cell proliferation, motility, ROS metabolism and migration.