Hesperetin promotes bladder cancer cells death via the PI3K/AKT pathway by network pharmacology and molecular docking.

Patients with bladder cancer (BLCA) still show high recurrence after surgery and chemotherapy. Hesperetin (HE), as a natural compound, has attracted researchers' attention due to its low toxicity and easy access. However, the inhibitory effect of HE on BLCA remains unknown. The hub genes and enrichment pathways regulated by HE in the treatment of BLCA were predicted by network pharmacology. The molecular docking of HE and hub proteins was visualized. Colony and CCK8 assays were used to test cell proliferation, and BLCA migration was confirmed by transwell and wound healing assays. In addition, the occurrence of apoptosis and ferroptosis was demonstrated by Hoechst staining, transmission electron microscopy (TEM) and ROS (reactive oxygen species) assay. Western Blotting was performed to validate the hub proteins, target functions and pathways. SRC, PIK3R1 and MAPK1 were identified as hub targets for HE in BLCA, involving the PI3k/AKT pathway. Furthermore, HE inhibited the proliferation and migration of BLCA cells. The MMP2/MMP9 proteins were significantly inhibited by HE. The increased expression of Bax and cleaved caspase-3 indicated that HE could promote BLCA cell apoptosis. In addition, Hoechst staining revealed concentrated and illuminated apoptotic nuclei. The activation of ROS and the decline of GPX4 expression suggested that HE might induce ferroptosis as an anti-BLCA process. Shrunk mitochondria and apoptotic bodies were observed in BLCA cells treated with HE, with reduced or absent mitochondrial cristae. We propose for the first time that HE could inhibit the proliferation and migration of BLCA cells and promote apoptosis and ferroptosis. HE may act by targeting proteins such as SRC, PIK3R1 and MAPK1 and the PI3K/AKT pathway.

Reliability and validity of the electronic version of the Hopkins verbal learning test-revised in middle-aged and elderly Chinese people.

Background: The aging population is increasing, making it essential to have a standardized, convenient, and valid electronic memory test that can be accessed online for older people and caregivers. The electronic version of the Hopkins Verbal Learning Test-Revised (HVLT-R) as a test with these advantages and its reliability and validity has not yet been tested. Thus, this study examined the reliability and validity of the electronic version of the HVLT-R in middle-aged and elderly Chinese people to provide a scientific basis for its future dissemination and use. Methods: We included 1,925 healthy participants aged over 40, among whom 38 were retested after 3-6 months. In addition, 65 participants completed both the pad and paper-and-pencil versions of the HVLT-R (PAP-HVLT-R). We also recruited 42 Alzheimer's disease (AD) patients, and 45 amnestic mild cognitive impairment (aMCI) patients. All participants completed the Pad-HVLT-R, the Hong Kong Brief Cognitive Test (HKBC), the Brief Visual Memory Test-Revised (BVMT-R), and the Logical Memory Test (LM). Results: (1) Reliability: the Cronbach's α value was 0.94, the split-half reliability was 0.96. The test-retest correlation coefficients were moderate, ranging from 0.38 to 0.65 for direct variables and 0.16 to 0.52 for derived variables; (2) Concurrent validity: the Pad-HVLT-R showed a moderate correlation with the HKBC and BVMT-R, with correlation coefficients between total recall of 0.41 and 0.54, and between long-delayed recall of 0.42 and 0.59, respectively. It also showed a high correlation with the LM, with correlation coefficients of 0.72 for total recall and 0.62 for long-delayed recall; (3) Convergent validity: the Pad-HVLT-R was moderately correlated with the PAP version, with correlation coefficients ranging from 0.29 to 0.53 for direct variables and 0.15 to 0.43 for derived variables; (4) Discriminant capacity: the Pad-HVLT-R was effective in differentiating AD patients, as demonstrated by the ROC analysis with AUC values of 0.834 and 0.934 for total recall and long-delayed recall, respectively. Conclusion: (1) The electronic version of HVLT-R has good reliability and validity in middle-aged and elderly Chinese people; (2) The electronic version of HVLT-R can be used as an effective tool to distinguish AD patients from healthy people.

Role of copper ionophore-induced death in immune microenvironment and clinical prognosis of ccRCC: An integrated analysis.

Background: Clear cell renal cell carcinoma (ccRCC) is a malignancy with a high incidence rate and poor prognosis worldwide. Copper ionophore-induced death (CID) plays an important role in cancer progression. Methods: One training and three validation datasets were acquired from TCGA, GEO and ArrayExpress. K-means clustering was conducted to identify the CID subtypes. The ESTIMATE and CIBERSORT algorithms were employed to illustrate the immune microenvironment of ccRCC. LASSO Cox regression was applied to construct the CID feature-based prognostic model. The immunotherapy cohort was acquired from the literature to explore the potential risk scores for predicting immunotherapy responsiveness. Results: Two CID-related cancer subtypes of ccRCC were identified that performed different immune microenvironment characteristics and prognosis. Based on the identified subtypes, we analyzed the biological heterogeneity and constructed a gene prognostic model. The prognostic model performed well in one training dataset, three validation datasets, and different clinical pathological groups. The prognostic model has a good potential for predicting cancer immune features and immunotherapy responsiveness. Conclusion: CID plays an important role in the tumor microenvironment progression of ccRCC. The robust gene prognostic model developed can help predict cancer prognosis, immune features, and immunotherapy responsiveness.

Feedback activation of GATA1/miR-885-5p/PLIN3 pathway decreases sunitinib sensitivity in clear cell renal cell carcinoma.

Sunitinib is the most commonly used first-line therapy for the treatment of advanced renal cell carcinoma (RCC), but intrinsic and extrinsic resistance to targeted therapies dramatically compromise the benefit of clinical outcome. Dissecting the underlying mechanisms and discovering reliable predictive biomarkers are urgently needed in clinic. Here, we discovered miR-885-5p was notably decreased after sunitinib treatment and associated with poor disease progression in clear cell renal cell carcinoma (ccRCC). In vitro and in vivo studies identified miR-885-5p inhibition contributed to sunitinib resistance. Mechanistically, sunitinib treatment reduced GATA1 expression, which in turn reduced its binding to MIR885 promoter and resulted in miR-885-5p downregulation in transcriptional level. In addition, PLIN3 was confirmed to be directly targeted by miR-885-5p and its upregulation significantly increased lipid droplets formation to decrease sunitinib sensitivity. Therefore, GATA1/miR-885-5p/ PLIN3 pathway may serve as a potential therapeutic strategy and a biomarker for sunitinib treatment in ccRCC.

Novel circRNA_0071196/miRNA­19b­3p/CIT axis is associated with proliferation and migration of bladder cancer.

Circular RNAs (circRNAs) are non­coding RNAs that are connected at the 3' and 5' ends by an exon or intron. Studies increasingly show that circRNAs play an important role in tumorigenesis by acting as a 'sponge' for microRNAs (miRNAs), which abrogates the latter's effect on their target mRNAs. To identify a possible circRNA/miRNA/mRNA network in bladder cancer (BCa), we analyzed the circRNA and mRNA expression profiles of BCa and adjacent normal bladder tissues. A total of 127 circRNAs and 1,612 mRNAs were differentially expressed in the tumor tissues, and were primarily associated with cancer­related pathways. A competing endogenous RNAs (ceRNA) network was then constructed which predicted a regulatory axis of circRNA_0071196, miRNA­19b­3p and its target gene citron Rho­interacting serine/threonine kinase (CIT). Luciferase reporter assay validated the relationship between circRNA_0071196 and miRNA­19b­3p and of the latter with CIT. Furthermore, CIT was overexpressed in the BCa tissues, and was found to be correlated with metastasis and tumor histological grade. Knockdown of CIT in the human bladder cancer cell line 5367 significantly inhibited the proliferation, migration and colony formation capacity of the cells, and also upregulated the mediators of the p53 and RhoA­ROCK signaling cascades that regulate cell cycle and migration. Taken together, our findings indicate that circRNA­0071196 upregulates CIT levels in BCa by sponging off miRNA­19b­3p, and the circRNA_0071196/miRNA­19b­3p/CIT axis is a potential therapeutic target in BCa.

Down-regulation of CIT can inhibit the growth of human bladder cancer cells.

OBJECTIVE: Our study is to examine the citron rho-interacting, serine/threonine kinase 21 (CIT) in bladder cancer. METHODS: We examined CIT level in human bladder cancer tissues by immunohistochemical staining. To explore the impact of CIT on cell proliferation and apoptosis, we down-regulated its expression in two human bladder cancer cell lines, 5367 and T24. We examined cell growth in 5367 and T24. We also performed in vivo analysis using T24 cells. We further used microarray expression profiling to investigate genes differentially expressed in T24 cells with CIT down-regulated. RESULTS: In 100 human samples, CIT was expressed by only 2 of 30 (6.7 %) controls in bladder tissues, whereas by 64 of 70 (91.4 %) cancer patients in tumor tissues (p < 0.001). in vitro analysis demonstrated that CIT knockdown represses cell proliferation by 50 % in both cells and colony formation (77 ± 5 vs. 13 ± 2, p = 0.001 for T24, 58 ± 3 vs. 1 ± 1, p < 0.001 for 5637). We also found CIT knockdown could induce cell cycle arrest, and promote apoptosis in both cells. Tumor-volume monitoring and live in vivo bladder cancer imaging in human xenograft model confirmed that CIT knockdown reduces tumor volume (668.4 ± 333.0 vs. 305.7 ± 170.4 mm3, p = 0.02) and weight (0.27 ± 0.15 vs. 0.57 ± 0.32 g, p = 0.02). Microarray analysis revealed that CIT may regulate cell cycle signalling pathway through various cell cycle regulators. CONCLUSIONS: In summary, we provided clinical and experimental evidence that CIT may promote bladder cancer through regulation of cell cycle pathway.

Evaluation of the improved tubeless cutaneous ureterostomy technique following radical cystectomy in cases of invasive bladder cancer complicated by peritoneal metastasis.

Radical cystectomy, as the most common surgical treatment for patients with invasive bladder cancer (IBC) complicated by peritoneal metastasis, is usually accompanied by a urinary diversion procedure. In this study, we evaluated the improved tubeless cutaneous ureterostomy technique by comparing the resulting clinical effects with either a traditional ureterostomy and an ileal conduit urinary diversion. Clinical data from 85 patients who underwent 1 of the 3 procedures between April 2012 and April 2015 were analyzed retrospectively. In total, 30 patients underwent improved tubeless cutaneous ureterostomy, 28 patients underwent a traditional cutaneous ureterostomy and 27 underwent an ileal conduit urinary diversion following radical cystectomy. The incidence of complications, including stoma infection, nipple atrophy, terminal necrosis, urine leakage, external orifice stenosis, uronephrosis and ureterectasia in the group of patients treated with the improved tubeless ureterostomy technique was significantly lower than that of the patients in the other 2 groups, and the difference was statistically significant (P<0.05). In addition, the duration of the surgery, intra-operative bleeding, the duration of the hospitalization period and the time to extubation in the patients treated with the improved tubeless ureterostomy technique were significantly decreased (P<0.05) compared with the patients in the other 2 groups. Finally, the health-related quality of life of the patients treated with the improved tubeless ureterostomy technique was significantly higher (P<0.05) than that of the patients in the other 2 groups. The findings of our study demonstrated that the use of the improved tubeless cutaneous ureterostomy technique following radical cystectomy in patients with IBC complicated by peritoneal metastasis resulted in improved clinical effects. Thus, improved tubeless cutaneous ureterostomy may be a promising alternative for enhancing the quality of life of patients with IBC.

Downregulation of long non-coding RNA TRIM52-AS1 functions as a tumor suppressor in renal cell carcinoma.

Long non-coding RNAs (lncRNAs) are important regulators of gene expression, interacting with the major pathways of cell growth, proliferation, differentiation and survival. Alterations in the function of lncRNAs promote tumor formation, progression and metastasis. The purpose of the present study was to identify novel tumor suppressor lncRNAs, and elucidate their physiological function and mechanism in renal cell carcinoma (RCC). The results of the present study revealed that the expression of the lncRNA, TRIM52­AS1, was downregulated in RCC, which was demonstrated using reverse transcription­quantitative polymerase chain reaction analysis. Furthermore, the effects of TRIM52­AS1 on proliferation, cell migration and apoptosis were analyzed using a wound scratch assay, a 3­(4,5­dimethylthiazol­2yl)­2,5­diphenyl tetrazolium bromide assay and flow cytometric analysis, respectively. The overexpression of TRIM52­AS1 using a synthesized vector significantly suppressed cell migration and proliferation, and induced apoptosis of the RCC cells in vitro, and interference of its expression led to the opposite effects. The present study was the first, to the best of our knowledge, to demonstrate that TRIM52­AS1 functions as a tumor suppressor in RCC. Further investigation is required to elucidate the molecular mechanisms underlying the effects of TRIM52-AS1 in the development of RCC.

MicroRNA-137 upregulation increases bladder cancer cell proliferation and invasion by targeting PAQR3.

There is increasing evidence suggesting that dysregulation of some microRNAs (miRNAs) may contribute to tumor progression and metastasis and have been proposed to be key regulators of diverse biological processes such as transcriptional regulation, cell growth and tumorigenesis. Previous studies have shown that miR-137 is dysregulated in some malignancies, but its role in bladder cancer is still unknown. In our study, we find that miR-137 is up-regulated in human bladder cancer tissues and cell lines. Moreover, the higher level of miR-137 was associated with pM or pTNM stage in clinical bladder cancer patients. Enforced expression of miR-137 in bladder cancer cells significantly enhanced their proliferation, migration and invasion. Bioinformatics analysis identified the tumor suppressor gene PAQR3 as a potential miR-137 target gene. Further studies indicated that miR-137 suppressed the expression of PAQR3 by binding to its 3'-untranslated region. Silencing of PAQR3 by small interfering RNAs phenocopied the effects of miR-137 overexpression, whereas restoration of PAQR3 in bladder cancer cells bladder cancer cells overexpressing miR-137, partially reversed the suppressive effects of miR-137. These findings indicate that miR-137 could be a potential oncogene in bladder cancer.