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BACKGROUND: Aberrant autophagy and preternatural elevated glycolysis are prevalent in bladder cancer (BLCA) and are both related to malignant progression. However, the regulatory relationship between autophagy and glycolytic metabolism remains largely unknown. We imitated starvation conditions in the tumour microenvironment and found significantly increased levels of autophagy and aerobic glycolysis, which both regulated the progression of BLCA cells. We further explored the regulatory relationships and mechanisms between them. METHODS: We used immunoblotting, immunofluorescence and transmission electron microscopy to detect autophagy levels in BLCA cells under different treatments. Lactate and glucose concentration detection demonstrated changes in glycolysis. The expression of lactate dehydrogenase A (LDHA) was detected at the transcriptional and translational levels and was also silenced by small interfering RNA, and the effects on malignant progression were further tested. The underlying mechanisms of signalling pathways were evaluated by western blot, immunofluorescence and immunoprecipitation assays. RESULTS: Starvation induced autophagy, regulated glycolysis by upregulating the expression of LDHA and caused progressive changes in BLCA cells. Mechanistically, after starvation, the ubiquitination modification of Axin1 increased, and Axin1 combined with P62 was further degraded by the autophagy-lysosome pathway. Liberated ß-catenin nuclear translocation increased, binding with LEF1/TCF4 and promoting LDHA transcriptional expression. Additionally, high expression of LDHA was observed in cancer tissues and was positively related to progression. CONCLUSION: Our study demonstrated that starvation-induced autophagy modulates glucose metabolic reprogramming by enhancing Axin1 degradation and ß-catenin nuclear translocation in BLCA, which promotes the transcriptional expression of LDHA and further malignant progression.
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BACKGROUND: Bladder cancer is one of the most prevalent malignancies worldwide. However, traditional indicators have limited predictive effects on the clinical outcomes of bladder cancer. The aim of this study was to develop and validate a glycolysis-related gene signature for predicting the prognosis of patients with bladder cancer that have limited therapeutic options. METHODS: mRNA expression profiling was obtained from patients with bladder cancer from The Cancer Genome Atlas (TCGA) database. Gene set enrichment analysis (GSEA) was conducted to identify glycolytic gene sets that were significantly different between bladder cancer tissues and paired normal tissues. A prognosis-related gene signature was constructed by univariate and multivariate Cox analysis. Kaplan-Meier curves and time-dependent receiver operating characteristic (ROC) curves were utilized to evaluate the signature. A nomogram combined with the gene signature and clinical parameters was constructed. Correlations between glycolysis-related gene signature and molecular characterization as well as cancer subtypes were analyzed. RT-qPCR was applied to analyze gene expression. Functional experiments were performed to determine the role of PKM2 in the proliferation of bladder cancer cells. RESULTS: Using a Cox proportional regression model, we established that a 4-mRNA signature (NUP205, NUPL2, PFKFB1 and PKM) was significantly associated with prognosis in bladder cancer patients. Based on the signature, patients were split into high and low risk groups, with different prognostic outcomes. The gene signature was an independent prognostic indicator for overall survival. The ability of the 4-mRNA signature to make an accurate prognosis was tested in two other validation datasets. GSEA was performed to explore the 4-mRNA related canonical pathways and biological processes, such as the cell cycle, hypoxia, p53 pathway, and PI3K/AKT/mTOR pathway. A heatmap showing the correlation between risk score and cell cycle signature was generated. RT-qPCR revealed the genes that were differentially expressed between normal and cancer tissues. Experiments showed that PKM2 plays essential roles in cell proliferation and the cell cycle. CONCLUSION: The established 4mRNA signature may act as a promising model for generating accurate prognoses for patients with bladder cancer, but the specific biological mechanism needs further verification.
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Bone marrow-derived mesenchymal stem cells (BMSCs) have been recently reported to play a variety of vital roles in organ and tissue damage repair, mainly via potent paracrine activity, including secreting extracellular vesicles, such as exosomes, that serve as mediators facilitating intercellular communication and reprogramming recipient cells by delivering their contents to target cells. However, the underlying mechanisms are diverse and complex, and the influencing characteristics have rarely been studied. Accordingly, we designed this study to explore the time dependence of the effects of exosomes derived from BMSCs (BMexos) on renal ischemia-reperfusion (I/R) injury and the underlying mechanisms associated with the reperfusion time. Impressively, our study is the first to find that BMexos protected against renal I/R injury in vitro and in vivo at the very early reperfusion stages, especially 4-8 h after reperfusion in vitro and 8-16 h after reperfusion in vivo. Interestingly, we simultaneously found that endoplasmic reticulum (ER) stress was significantly suppressed following the administration of BMexos in vitro and in vivo with a similar time dependence. Additionally, we discovered that miR-199a-5p, which was abundant in the BMSCs, was transferred into renal tubular epithelial cells (NRK-52E) in a time-dependent manner and significantly inhibited I/R-induced ER stress by targeting binding immunoglobulin protein (BIP). Cocultivation with miR-199a-5p-overexpressing BMSCs amplified the suppression of ER stress and further protected against I/R injury. However, coculture with miR-199a-5p-knockdown BMSCs obviously increased ER stress and reversed the BMexos-induced protection, and silencing BIP by small interfering RNA-1098 in NRK-52E inhibited these effects. This study provides evidence that administering BMexos at the very early reperfusion stages significantly protects against renal I/R injury, and ER stress is closely linked to this protection. These results suggest a novel therapeutic strategy during the very early reperfusion stages of renal I/R injury.-Wang, C., Zhu, G., He, W., Yin, H., Lin, F., Gou, X., Li, X. BMSCs protect against renal ischemia-reperfusion injury by secreting exosomes loaded with miR-199a-5p that target BIP to inhibit endoplasmic reticulum stress at the very early reperfusion stages.
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Estresse do Retículo Endoplasmático/fisiologia , Exossomos/metabolismo , Nefropatias/metabolismo , Linfocinas/metabolismo , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Traumatismo por Reperfusão/metabolismo , Animais , Transporte Biológico/fisiologia , Linhagem Celular , Rim/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Transdução de Sinais/fisiologiaRESUMO
Renal ischemia/reperfusion (I/R) injury is the main reason for acute kidney injury (AKI) and is closely related to high morbidity and mortality. In this study, we found that exosomes from human-bone-marrow-derived mesenchymal stem cells (hBMSC-Exos) play a protective role in hypoxia/reoxygenation (H/R) injury. hBMSC-Exos were enriched in miR-199a-3p, and hBMSC-Exo treatment increased the expression level of miR-199a-3p in renal cells. We further explored the function of miR-199a-3p on H/R injury. miR-199a-3p was knocked down in hBMSCs with a miR-199a-3p inhibitor. HK-2 cells cocultured with miR-199a-3p-knockdown hBMSCs were more susceptible to H/R injury and showed more apoptosis than those cocultured with hBMSCs or miR-199a-3p-overexpressing hBMSCs. Meanwhile, we found that HK-2 cells exposed to H/R treatment incubated with hBMSC-Exos decreased semaphorin 3A (Sema3A) and activated the protein kinase B (AKT) and extracellular-signal-regulated kinase (ERK) pathways. However, HK-2 cells cocultured with miR-199a-3p-knockdown hBMSCs restored Sema3A expression and blocked the activation of the AKT and ERK pathways. Moreover, knocking down Sema3A could reactivate the AKT and ERK pathways suppressed by a miR-199a-3p inhibitor. In vivo, we injected hBMSC-Exos into mice suffering from I/R injury; this treatment induced functional recovery and histologic protection and reduced cleaved caspase-3 and Sema3A expression levels, as shown by immunohistochemistry. On the whole, this study demonstrated an antiapoptotic effect of hBMSC-Exos, which protected against I/R injury, via delivering miR-199a-3p to renal cells, downregulating Sema3A expression and thereby activating the AKT and ERK pathways. These findings reveal a novel mechanism of AKI treated with hBMSC-Exos and provide a therapeutic method for kidney diseases.
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Exossomos/metabolismo , Rim/lesões , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/genética , Traumatismo por Reperfusão/prevenção & controle , Animais , Apoptose/fisiologia , Caspase 3/biossíntese , Linhagem Celular , Humanos , Rim/patologia , Nefropatias/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , Semaforina-3A/genética , Semaforina-3A/metabolismoRESUMO
The biological characteristics of bladder cancer include enhanced invasion and migration, which are the main causes of death in patients. Starvation is a typical feature of the bladder cancer microenvironment and can induce autophagy. Autophagy has an important relationship with the invasion and migration of tumors. However, the role of autophagy in the invasion and migration of bladder cancer cells remains unclear. Hence, the aim of the current study was to clarify this role and underlying mechanism. In this study, we found that starvation enhanced the epithelial-mesenchymal transition (EMT)-mediated invasion and migration of T24 and 5637 cells while inducing autophagy. The inhibition of autophagy with chloroquine (CQ) or 3-methyladenine (3MA) decreased EMT-mediated invasion and migration. In addition, the expression of transforming growth factor 1 (TGF-ß1) and phosphorylated Smad3 (p-Smad3) increased after starvation. The inhibition of autophagy with CQ or 3MA also decreased the expression of TGF-ß1 and p-Smad3. The inhibitor of TGF-ß receptor sb431542 also inhibited the invasion, migration, and EMT of T24 and 5637 cells during starvation. Furthermore, recombinant TGF-ß1 induced autophagy and inhibition of the TGF-ß/Smad signaling pathway with sb431542 suppressed autophagy. In summary, our results suggested that autophagy promotes the invasion and migration of bladder cancer cells by inducing EMT through the TGF-ß1/Smad3 signaling pathway. Moreover, autophagy and TGF-ß1 can form a positive feedback loop to synergistically promote invasion and migration. Thus, our findings may provide a theoretical basis for the prevention of invasion and migration in bladder cancer.
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Autofagia , Movimento Celular , Transição Epitelial-Mesenquimal , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia , Linhagem Celular Tumoral , Humanos , Invasividade Neoplásica , Transdução de SinaisRESUMO
Rapamycin, an immunosuppressant, is widely used in patients with kidney transplant. However, the therapeutic effects of rapamycin remain controversial. Additionally, previous studies have revealed deleterious effects of rapamycin predominantly when administered for ≥24 h. Few studies, however, have focused on the short-term effects of rapamycin administered only during the initial reperfusion phase. As such, we designed this study to explore the potential effects and mechanisms of rapamycin under a specific therapeutic regimen in which rapamycin is mixed in the perfusate during the initial reperfusion phase (within 24 h). Interestingly, we found that rapamycin maintained renal function and attenuated ischemia-reperfusion (I/R)-induced apoptosis in vivo and in vitro during the initial reperfusion phase, especially at 8 h after reperfusion. Simultaneously, rapamycin activated autophagy and inhibited endoplasmic reticulum (ER) stress and 3 pathways of unfolding protein response: ATF6, PERK, and IRE1α. Interestingly, we further found that the protective effects of rapamycin were suppressed when autophagy was inhibited by chloroquine and 3-methyladenine or when ER stress was induced by thapsigargin. Moreover, in terms of the regulatory effects of rapamycin, a negative-feedback loop between autophagy and ER stress occurred, with autophagy inhibiting ER stress and increased ER stress promoting autophagy during the initial reperfusion phase of renal I/R injury. Our study provides evidence that immediate reperfusion with rapamycin during the initial reperfusion phase repairs renal function and reduces apoptosis via activating autophagy, which could further inhibit ER stress. These results suggest a novel treatment modality for application during the initial reperfusion phase of renal I/R injury caused by kidney transplantation.-Li, X., Zhu, G., Gou, X., He, W., Yin, H., Yang, X., Li, J. Negative feedback loop of autophagy and endoplasmic reticulum stress in rapamycin protection against renal ischemia-reperfusion injury during initial reperfusion phase.
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Women with hyperinsulinism and insulin resistance have reduced fertility, but the underlying mechanism is still poorly understood. Aberrant endometrial decidualization in early pregnancy was linked to pregnancy complications. In this study, we aimed to test whether elevated insulin levels compromise decidualization in early-stage pregnancy. C57BL/6J mice in high insulin-exposed group were given a subcutaneous injection of recombinant insulin at a concentration of 0.05 IU daily. During decidualization in early pregnancy, serum levels of insulin, E2, P4, LH, FSH and blood glucose were significantly altered in mice treated with high insulin levels. The number of embryo implantation sites and endometrial decidual markers BMP2, ER, PR was significantly decreased by high insulin levels in vivo. Artificial decidual induction in primary mouse endometrial stromal cells and immortal human endometrial stromal cells line were all compromised after treated with 100 nmol/L insulin levels. All these results on flow cytometry, transmission electron microscopy and western blotting of Bax, Bcl2, cleaved Caspase3, cleaved PARP proteins level showed that decidual cells apoptosis was significantly decreased. Mitochondrial transmembrane potential also significantly increased by the influence of high insulin levels. PI3K and p-Akt were much higher after insulin exposure and the compromised decidualization by high insulin treatment was rescued by PI3K/Akt inhibitor LY294002 both in vitro and in vivo. In conclusion, we demonstrated that elevated insulin levels could compromise mice decidualization in early-stage pregnancy and PI3K/p-Akt-regulated apoptosis was essential for this role. It provides a clue for future investigation on compromised reproduction in women with hyperinsulinemia.
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Apoptose/fisiologia , Endométrio/fisiologia , Insulina/sangue , Útero/patologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Cromonas/farmacologia , Decídua/efeitos dos fármacos , Decídua/patologia , Decídua/fisiologia , Decídua/fisiopatologia , Implantação do Embrião , Endométrio/citologia , Endométrio/efeitos dos fármacos , Endométrio/fisiopatologia , Feminino , Humanos , Insulina/efeitos adversos , Potencial da Membrana Mitocondrial , Camundongos Endogâmicos C57BL , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Gravidez , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células Estromais/efeitos dos fármacos , Células Estromais/patologiaRESUMO
BACKGROUND: The aim of the study is described the regulatory mechanisms and prognostic values of differentially expressed RNAs in prostate cancer and construct an mRNA signature that predicts survival. METHODS: The RNA profiles of 499 prostate cancer tissues and 52 non-prostate cancer tissues from TCGA were analyzed. The differential expression of RNAs was examined using the edgeR package. Survival was analyzed by Kaplan-Meier method. microRNA (miRNA), messenger RNA (mRNA), and long non-coding RNA (lncRNA) networks from the miRcode database were constructed, based on the differentially expressed RNAs between non-prostate and prostate cancer tissues. RESULTS: A total of 773 lncRNAs, 1417 mRNAs, and 58 miRNAs were differentially expressed between non-prostate and prostate cancer samples. The newly constructed ceRNA network comprised 63 prostate cancer-specific lncRNAs, 13 miRNAs, and 18 mRNAs. Three of 63 differentially expressed lncRNAs and 1 of 18 differentially expressed mRNAs were significantly associated with overall survival in prostate cancer (P value < 0.05). After the univariate and multivariate Cox regression analyses, 4 mRNAs (HOXB5, GPC2, PGA5, and AMBN) were screened and used to establish a predictive model for the overall survival of patients. Our ROC curve analysis revealed that the 4-mRNA signature performed well. CONCLUSION: These ceRNAs may play a critical role in the progression and metastasis of prostate cancer and are thus candidate therapeutic targets and potential prognostic biomarkers. A novel model that incorporated these candidates was established and might provide more powerful prognostic information in predicting survival in prostate cancer.
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Redes Reguladoras de Genes , Neoplasias da Próstata/genética , RNA Mensageiro/genética , RNA Neoplásico/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Humanos , Masculino , Análise Multivariada , Prognóstico , Modelos de Riscos Proporcionais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro/metabolismo , RNA Neoplásico/metabolismo , Análise de SobrevidaRESUMO
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.
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Antimetabólitos Antineoplásicos/farmacologia , Desoxicitidina/análogos & derivados , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/enzimologia , Antracenos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Linhagem Celular Tumoral , Desoxicitidina/farmacologia , Ativação Enzimática/efeitos dos fármacos , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Neoplasias da Bexiga Urinária/patologia , GencitabinaRESUMO
Background: Clear cell renal cell carcinoma (ccRCC) is one of the most common cancers worldwide, and its incidence is increasing every year. Endoplasmic reticulum stress (ERS) caused by protein misfolding has broad and profound effects on the progression and metastasis of various cancers. Accumulating evidence suggests that ERS is closely related to the occurrence and progression of ccRCC. This study aimed to identify ERS-related genes for evaluating the prognosis of ccRCC. Methods: Transcriptomic expression profiles were obtained from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA), and clinical data were downloaded from the TCGA. First, the differentially expressed genes (DEGs) were analyzed using the limma package, and the DEGs related to ERS (ERS-DEGs) were identified from the GeneCards database. Second, a function and pathway enrichment analysis and a Gene Set Enrichment Analysis (GSEA) were performed. Third, a protein-protein interaction (PPI) network was constructed to identify the hub genes, and a gene-micro RNA (miRNA) network and gene-transcription factor (TF) network were established using the hub genes. Finally, a least absolute shrinkage and selection operator (LASSO) regression analysis was conducted to establish a diagnostic model, and a Cox analysis was used to analyze the correlations between the expression of the characteristic genes and the clinical characteristics. Results: We identified 11 signature genes and established a diagnostic model. Further, the Cox analysis results revealed a correlation between the expression levels of the signature genes and the clinical characteristics. Ultimately, five signature genes (i.e., TNFSF13B, APOL1, COL5A3, and CDH5) were found to be associated with a poor prognosis. Conclusions: This study suggests that TNFSF13B, APOL1, COL5A3, and CDH5 may have potential as prognostic biomarkers in ccRCC and may provide new evidence to support targeted therapy in ccRCC.
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Background: Removable partial denture (RPD) design is crucial to long-term success in dental treatment, but shortcomings in RPD design training and competency acquisition among dental students have persisted for decades. Digital production is increasing in prevalence in stomatology, and a digital RPD (D-RPD) module, under the framework of the certified Objective Manipulative Skill Examination of Dental Technicians (OMEDT) system reported in our previous work, may improve on existing RPD training models for students. Objective: We aimed to determine the efficacy of a virtual 3D simulation-based progressive digital training module for RPD design compared to traditional training. Methods: We developed a prospective cohort study including dental technology students at the Stomatology College of Chongqing Medical University. Cohort 1 received traditional RPD design training (7 wk). Cohort 2 received D-RPD module training based on text and 2D sketches (7 wk). Cohort 3 received D-RPD module pilot training based on text and 2D sketches (4 wk) and continued to receive training based on 3D virtual casts of real patients (3 wk). RPD design tests based on virtual casts were conducted at 1 month and 1 year after training. We collected RPD design scores and the time spent to perform each assessment. Results: We collected the RPD design scores and the time spent to perform each assessment at 1 month and 1 year after training. The study recruited 109 students, including 58 (53.2%) female and 51 male (56.8%) students. Cohort 1 scored the lowest and cohort 3 scored the highest in both tests (cohorts 1-3 at 1 mo: mean score 65.8, SD 21.5; mean score 81.9, SD 6.88; and mean score 85.3, SD 8.55, respectively; P<.001; cohorts 1-3 at 1 y: mean score 60.3, SD 16.7; mean score 75.5, SD 3.90; and mean score 90.9, SD 4.3, respectively; P<.001). The difference between cohorts in the time spent was not statistically significant at 1 month (cohorts 1-3: mean 2407.8, SD 1370.3 s; mean 1835.0, SD 1329.2 s; and mean 1790.3, SD 1195.5 s, respectively; P=.06) but was statistically significant at 1 year (cohorts 1-3: mean 2049.16, SD 1099.0 s; mean 1857.33, SD 587.39 s; and mean 2524.3, SD 566.37 s, respectively; P<.001). Intracohort comparisons indicated that the differences in scores at 1 month and 1 year were not statistically significant for cohort 1 (95% CI -2.1 to 13.0; P=.16), while cohort 3 obtained significantly higher scores 1 year later (95% CI 2.5-8.7; P=.001), and cohort 2 obtained significantly lower scores 1 year later (95% CI -8.8 to -3.9; P<.001). Conclusions: Cohort 3 obtained the highest score at both time points with retention of competency at 1 year, indicating that progressive D-RPD training including virtual 3D simulation facilitated improved competency in RPD design. The adoption of D-RPD training may benefit learning outcomes.
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Dental anatomy education for dental technology students should be developed in alignment with digital dental laboratory practices. We hypothesized that a virtually assisted sketching-based dental anatomy teaching module could improve students' acquisition of skills essential for digital restoration design. The second-year dental technology curriculum included a novel virtual technology-assisted sketching-based module for dental anatomy education. Pre- and post-course assessments evaluated students' skill sets and knowledge bases. Computer-aided design (CAD) scores were analyzed after one year to assess how the skills students developed through this module impacted their subsequent CAD performance. Participants who undertook the dental sketching-based teaching module demonstrated significantly improved theoretical knowledge of dental anatomy, dental aesthetic perception, and spatial reasoning skills. A partial least squares structural equation model indicated that the positive effects of this module on subsequent CAD performance were indirectly mediated by dental aesthetic perception, spatial reasoning, and practice time. A virtually assisted sketching-based dental anatomy teaching module significantly improved students' acquisition of skills and knowledge and positively mediated dental technology students' CAD performance.
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Anatomia , Desenho Assistido por Computador , Instrução por Computador , Currículo , Educação em Odontologia , Avaliação Educacional , Humanos , Educação em Odontologia/métodos , Feminino , Anatomia/educação , Masculino , Avaliação Educacional/estatística & dados numéricos , Instrução por Computador/métodos , Estudantes de Odontologia , Aprendizagem , Adulto Jovem , Prótese Dentária , Planejamento de Prótese Dentária , Restauração Dentária Permanente/métodos , AdultoRESUMO
Autophagy-dependent cisplatin resistance poses a challenge in bladder cancer treatment. SIRT1, a protein deacetylase, is involved in autophagy regulation. However, the precise mechanism through which SIRT1 mediates cisplatin resistance in bladder cancer via autophagy remains unclear. In this study, we developed a cisplatin-resistant T24/DDP cell line to investigate this mechanism. The apoptosis rate and cell viability were assessed using flow cytometry and the CCK8 method. The expression levels of the relevant RNA and protein were determined using RT-qPCR and a Western blot analysis, respectively. Immunoprecipitation was utilized to validate the interaction between SIRT1 and Beclin1, as well as to determine the acetylation level of Beclin1. The findings indicated the successful construction of the T24/DDP cell line, which exhibited autophagy-dependent cisplatin resistance. Inhibiting autophagy significantly reduced the drug resistance index of these cells. The T24/DDP cell line showed a high SIRT1 expression level. The overexpression of SIRT1 activated autophagy, thereby further promoting cisplatin resistance in the T24/DDP cell line. Conversely, inhibiting autophagy counteracted the cisplatin-resistance-promoting effects of SIRT1. Silencing SIRT1 led to increased acetylation of Beclin1, the inhibition of autophagy, and a reduction in the cisplatin resistance of the T24/DDP cell line. Introducing a double mutation (lysine 430 and 437 to arginine, 2KR) in Beclin-1 inhibited acetylation and activated autophagy, effectively reversing the decreased cisplatin resistance resulting from SIRT1 silencing. In summary, our study elucidated that SIRT1 promotes cisplatin resistance in human bladder cancer T24 cells through Beclin1-deacetylation-mediated autophagy activation. These findings suggest a potential new strategy for reversing cisplatin resistance in bladder cancer.
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In recent years, antibody-drug conjugate (ADC) therapy targeting human epidermal growth factor receptor 2 (HER2) has been proven to be beneficial in patients with advanced urothelial carcinoma of the bladder (UCB); however, the role of HER2 in UCB remains obscure. Thus, the present retrospective single-center study was performed to evaluate the expression of HER2 in UCB and its prognostic significance. The HER2 status of 108 patients with UCB who underwent radical cystectomy was assessed using immunohistochemistry, and its association with the recurrence and survival rates of patients was analyzed. HER2 overexpression was observed in 57.4% of the patients; this was significantly associated with higher tumor grades (P=0.006) and stages (P<0.001). Kaplan-Meier analysis suggested that patients with HER2 overexpression had a shorter 5-year overall survival rate (P=0.005) and recurrence-free survival rate (P=0.003). Multivariate Cox regression analysis indicated that HER2 overexpression was a high-risk independent predictor of UCB recurrence (hazard ratio, 3.61; 95% confidence interval, 1.07-12.18; P=0.039). On the whole, these findings demonstrate that evaluating the HER2 status may improve the prediction of cancer recurrence and may thus guide the selection of patients that will benefit the most from HER2-ADC therapies.
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Bladder cancer is a highly heterogeneous and aggressive malignancy with a poor prognosis. EGF/EGFR activation causes the detachment of SHC-binding protein 1 (SHCBP1) from SHC adapter protein 1 (SHC1), which subsequently translocates into the nucleus and promotes cancer development via multiple signaling pathways. However, the role of the EGF-SHCBP1 axis in bladder cancer progression remains unexplored. Herein, we report that SHCBP1 is upregulated in bladder cancer tissues and cells, with cytoplasmic or nuclear localization. Released SHCBP1 responds to EGF stimulation by translocating into the nucleus following Ser273 phosphorylation. Depletion of SHCBP1 reduces EGF-induced cell migration and invasiveness of bladder cancer cells. Mechanistically, SHCBP1 binds to RACGAP1 via its N-terminal domain of amino acids 1 ~ 428, and this interaction is enhanced following EGF treatment. Furthermore, SHCBP1 facilitates cell migration by inhibiting RACGAP-mediated GTP-RAC1 inactivation, whose activity is indispensable for cell movement. Collectively, we demonstrate that the EGF-SHCBP1-RACGAP1-RAC1 axis acts as a novel regulatory mechanism of bladder cancer progression, which offers a new clinical therapeutic strategy to combat bladder cancer.
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Núcleo Celular/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Proteínas Adaptadoras da Sinalização Shc/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células , Progressão da Doença , Fator de Crescimento Epidérmico/farmacologia , Humanos , Hidrólise , Ligação Proteica , Transdução de Sinais , Proteína 1 de Transformação que Contém Domínio 2 de Homologia de Src/metabolismo , Neoplasias da Bexiga Urinária/patologiaRESUMO
Bladder cancer is a common malignant tumour worldwide. Epithelial-mesenchymal transition (EMT)-related biomarkers can be used for early diagnosis and prognosis of cancer patients. To explore, accurate prediction models are essential to the diagnosis and treatment for bladder cancer. In the present study, an EMT-related long noncoding RNA (lncRNA) model was developed to predict the prognosis of patients with bladder cancer. Firstly, the EMT-related lncRNAs were identified by Pearson correlation analysis, and a prognostic EMT-related lncRNA signature was constructed through univariate and multivariate Cox regression analyses. Then, the diagnostic efficacy and the clinically predictive capacity of the signature were assessed. Finally, Gene set enrichment analysis (GSEA) and functional enrichment analysis were carried out with bioinformatics. An EMT-related lncRNA signature consisting of TTC28-AS1, LINC02446, AL662844.4, AC105942.1, AL049840.3, SNHG26, USP30-AS1, PSMB8-AS1, AL031775.1, AC073534.1, U62317.2, C5orf56, AJ271736.1, and AL139385.1 was constructed. The diagnostic efficacy of the signature was evaluated by the time-dependent receiver-operating characteristic (ROC) curves, in which all the values of the area under the ROC (AUC) were more than 0.73. A nomogram established by integrating clinical variables and the risk score confirmed that the signature had a good clinically predict capacity. GSEA analysis revealed that some cancer-related and EMT-related pathways were enriched in high-risk groups, while immune-related pathways were enriched in low-risk groups. Functional enrichment analysis showed that EMT was associated with abundant GO terms or signaling pathways. In short, our research showed that the 14 EMT-related lncRNA signature may predict the prognosis and progression of patients with bladder cancer.
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Transição Epitelial-Mesenquimal/genética , RNA Longo não Codificante/genética , Neoplasias da Bexiga Urinária/patologia , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Prognóstico , Neoplasias da Bexiga Urinária/genéticaRESUMO
Cisplatin-based chemotherapy is the standard regimen for patients with bladder cancer, but its effectiveness is limited by high toxicity and the development of drug resistance. ß-elemene (ß-ELE), a compound extracted from Rhizoma zedoariae, has antitumor activity in various malignancies and exhibits low toxicity. However, the effects and specific mechanism of ß-ELE in bladder cancer remain unclear. The present study aimed to investigate the antitumor activity and possible mechanisms of ß-ELE alone and in combination with cisplatin in bladder cancer cells. Cell viability was determined using Cell Counting Kit-8. Cell cycle and reactive oxygen species (ROS) analyses were performed by flow cytometry. Apoptosis was detected by Hoechst 33258 and Annexin-V/propidium iodide staining. Mitochondrial membrane potential was determined by staining with a JC-1 probe, flow cytometry and fluorescence microscopy. Protein expression was detected by western blotting. The results revealed that ß-ELE significantly inhibited the proliferation of various bladder cancer cell lines and induced cell cycle arrest at G0/G1-phase in T24 and 5637 cells. Compared with cisplatin alone, co-treatment with ß-ELE increased cisplatin-mediated cytotoxicity against T24 cells, which resulted in the loss of mitochondrial membrane potential and release of cytochrome c into the cytoplasm. Co-treatment with ß-ELE and cisplatin enhanced ROS accumulation and activation of 5'AMP-activated protein kinase (AMPK), which induced apoptosis. The results of the present study suggested that ß-ELE inhibited the proliferation of bladder cancer cells in vitro and enhanced cisplatin-induced mitochondria-dependent apoptosis via the ROS-AMPK signaling pathway. Combination therapy with ß-ELE requires further investigation as a potential treatment of bladder cancer.
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There are no reliable criteria to assess risk of progression of nonmuscle invasive bladder cancer to muscle invasive bladder cancer. The aim of the present study was to identify potential markers based on gene expression profiling to improve predictive power of disease progression and prognosis in patients with bladder cancer. In the present study, we screened seventythree differentially expressed genes by analyzing bladder cancer samples with or without progression. Fortyseven prognosisrelated genes were screened, 13 of which were identified to build a progressionassociated gene signature using the LASSO regression method. Based on this 13mRNA signature, patients were divided into high and lowrisk groups, with different prognostic outcomes. The gene signature was an independent prognostic factor for overall survival. Receiver operating characteristic analysis suggested that the signature performed well in the validation cohort and its predictive power outperformed other several published signatures. CTHRC1, MMP11, AEBP1, SNCAIP, COL1A1 and S100A8 were identified as hub genes and their expression levels were detected using reverse transcriptasequantitative polymerase chain reaction. The expression of CTHRC1 was elevated in aggressive bladder cancer compared with noninvasive type, which suggests CTHRC1 may be a valuable biomarker for prediction of prognosis and progression of bladder cancer. Collectively, this 13mRNA signature may be useful in predicting disease progression and prognosis, thereby contributing to individualized management of patients with bladder cancer.
Assuntos
Biomarcadores Tumorais/genética , Perfilação da Expressão Gênica/métodos , Redes Reguladoras de Genes , Neoplasias da Bexiga Urinária/mortalidade , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Prognóstico , Curva ROC , Análise de Sobrevida , Neoplasias da Bexiga Urinária/genéticaRESUMO
The spread of coronavirus disease 2019 (COVID-19) around the world has put a heavy burden on human society and is also a great challenge facing medical staff. This study aimed to assess the difficulties faced by health care personnel (HCP) in using personal protective equipment (PPE) in clinical practice during the COVID-19 outbreak in Wuhan, China. One hundred twenty medical staff from the First Affiliated Hospital of Chongqing Medical University presented to the Wuhan First Hospital to provide medical assistance, from whom 20 HCP volunteered to participate in a focus group discussion attended by infection control nurse leaders. Participants' responses and discussions were recorded, and the content was analyzed for themes. Observed difficulties included inappropriate PPE sizes, the design of the PPE and its complexity of use, doubts related to the quality and effectiveness of PPE, potential risks during doffing, space layout between clean and contaminated area, and poor comfort with PPE use. Other factors, such as the support environment, management, processes, preparedness, HCP, and equipment can also have a positive or negative impact on the use of PPE. Future efforts to optimize PPE use should focus on strengthening training for HCP using real items for increasing compliance with standardized protocols, improving PPE design, and performing further research on the risks, benefits, and best practices of PPE use.
Assuntos
COVID-19/epidemiologia , Controle de Infecções/métodos , Enfermeiras e Enfermeiros/psicologia , Enfermeiras e Enfermeiros/estatística & dados numéricos , Equipamento de Proteção Individual/estatística & dados numéricos , Adulto , China/epidemiologia , Meio Ambiente , Feminino , Grupos Focais , Humanos , Liderança , Masculino , Pessoa de Meia-Idade , Pandemias , SARS-CoV-2RESUMO
BACKGROUND: Bladder cancer is one of the most common urinary malignancies. This study is aimed at providing some promising molecular biomarkers for bladder cancer (BC) by investigating the correlation between C1QTNF6 expression and clinical characteristics as well as prognosis in patients with bladder cancer. METHODS: Sequencing profiles of C1QTNF6 mRNA in BC patients were collected to evaluate the distinctive gene expression, between normal bladder mucosa and BC, according to the TCGA and GEO databases. The association between C1QTNF6 expression and the clinical features as well as the disease prognosis was evaluated using two independent cohorts. The expression of C1QTNF6 in normal bladder and BC cells was examined by western blotting and PCR, so the underlying molecular mechanism could be further investigated. RESULTS: C1QTNF6 mRNA levels were found to be differentially expressed in two independent public cohorts, including the TCGA database and GSE13507 dataset from GEO. The protein and RNA levels of C1QTNF6 in BC cells were both elevated when compared to normal bladder cell lines. High C1QTNF6 expression was detected in advanced T/M stages, pathological grade, and AJCC stage when compared to the low C1QTNF6 expression group. The underlying mechanism related to this differential expression could be explained by cell migration and invasion assays, where bladder cancer cells 5637 and T24 had a significant reduction on migration and invasion ability upon knockdown of C1QTNF6 expression. The low C1QTNF6 expression group presented a more prominent OS advantage over the high-expression group in both TCGA and GSE13507 cohorts. Moreover, the protein content in tissues was further validated using the HPA database and TMA. Survival analyses also indicated that the high C1QTNF6 expression group had an unfavorable OS when compared to the low-expression group. CONCLUSIONS: High C1QTNF6 expression may serve as a predictor of poor prognosis in bladder cancer patients, and the underlying mechanism is possibly associated with changes on cancer cell migration and invasion ability.