PeFtsH5 negatively regulates the biological stress response in Phalaenopsis equestris mitochondria.

Mitochondrial homeostasis plays a crucial role in determining cell fate by direct influence on cell apoptosis and autophagy. The ATP and Zn2+-dependent protease FtsH are of paramount importance in maintaining mitochondrial homeostasis. In Phalaenopsis equestris, three mitochondrial FtsH proteases were identified, one of which was encoded by the PeFtsH5 gene. This gene encoded a distinctive mitochondrial protein featuring a unique domain within the FtsH family. Down-regulating the expression of the PeFtsH5 homolog in Nicotiana benthamiana resulted in elevated expression levels of SA synthesis-related genes, leading to enhanced disease resistance. However, this down-regulation also caused cellular damage. Similarly, in P. equestris, the down-regulation of PeFtsH5 expression promoted the expression of defense response genes, leading to accelerated apoptosis and increased ROS levels. Nonetheless, this down-regulation also positively influenced plant resistance to biotic stress. Notably, the PeFtsH5 (i-AAA) protein, as revealed by dual membrane experiments, could form homopolymers exclusively, as it did not interact with the other two mitochondrial FtsH proteases. Consequently, this mitochondrial FtsH protease functioned as a homopolymer within P. equestris cells. The findings of this study elucidated the role of PeFtsH5 in responding to biological stress and provided new insights into its potential molecular mechanism. The result presented in this study hold promise for future research endeavors examining the regulatory effects of mitochondrial proteases on mitochondrial homeostasis and the development of stress-resistant P. equestris varieties through breeding programs.

Actively tunable plasmon-induced transparency in terahertz based on Dirac semimetal metamaterials.

We numerically investigate a tunable plasmon-induced transparency based on bulk Dirac semimetal (BDS) metamaterial in the terahertz band. In the unit cell, the prominent transparent peak appears to be due to the interference between the cut wires (CWs) and split-ring resonators (SRRs). An active modulation via near-field coupling is obtained by varying the Fermi level of the BDS. Introducing photoactive silicon, it will be found that once the intensity of the pump light is adjusted, a tunable transparent peak will appear. Furthermore, by shifting the coupling distance between CWs and SRRs, the depth of the transparent peak will change accordingly. Finally, we place the structure in environments with different refractive indices, which will exhibit excellent sensitivity and facilitate the application of biochemical sensors. This simple and easy-to-fabricate metamaterial structure will have excellent potential applications in modulation, filters, and detection.

BERT-Based Approaches to Identifying Malicious URLs.

Malicious uniform resource locators (URLs) are prevalent in cyberattacks, particularly in phishing attempts aimed at stealing sensitive information or distributing malware. Therefore, it is of paramount importance to accurately detect malicious URLs. Prior research has explored the use of deep-learning models to identify malicious URLs, using the segmentation of URL strings into character-level or word-level tokens, and embedding and employing trained models to differentiate between URLs. In this study, a bidirectional encoder representation from a transformers-based (BERT) model was devised to tokenize URL strings, employing its self-attention mechanism to enhance the understanding of correlations among tokens. Subsequently, a classifier was employed to determine whether a given URL was malicious. In evaluating the proposed methods, three different types of public datasets were utilized: a dataset consisting solely of URL strings from Kaggle, a dataset containing only URL features from GitHub, and a dataset including both types of data from the University of New Brunswick, namely, ISCX 2016. The proposed system achieved accuracy rates of 98.78%, 96.71%, and 99.98% on the three datasets, respectively. Additionally, experiments were conducted on two datasets from different domains-the Internet of Things (IoT) and Domain Name System over HTTPS (DoH)-to demonstrate the versatility of the proposed model.

Hydrogen peroxide-induced oxidative damage and protective role of peroxiredoxin 6 protein via EGFR/ERK signaling pathway in RPE cells.

Introduction: Damage to retinal pigment epithelium (RPE) cells caused by oxidative stress is closely related to the pathogenesis of several blinding retinal diseases, such as age-related macular degeneration (AMD), retinitis pigmentosa, and other inherited retinal degenerative conditions. However, the mechanisms of this process are poorly understood. Hence, the goal of this study was to investigate hydrogen peroxide (H2O2)-induced oxidative damage and protective role of peroxiredoxin 6 (PRDX6) protein via EGFR/ERK signaling pathway in RPE cells. Methods: Cells from a human RPE cell line (ARPE-19 cells) were treated with H2O2, and then cell viability was assessed using the methyl thiazolyl tetrazolium assay. Cell death and reactive oxygen species (ROS) were detected by flow cytometry. The levels of PRDX6, epidermal growth factor receptor (EGFR), P38 mitogen-activated protein kinase (P38MAPK), c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) were detected by Western blot assay. PRDX6 and EGFR were also detected via immunofluorescence staining. Results: Our results show that H2O2 inhibited cell viability, induced cell death, and increased ROS levels in ARPE-19 cells. It was also found that H2O2 decreased the levels of PRDX6, EGFR, and phosphorylated ERK but increased the levels of phosphorylated P38MAPK and JNK. PRDX6 overexpression was found to attenuate H2O2-induced inhibition of cell viability and increased cell death and ROS production in ARPE-19 cells. PRDX6 overexpression also increased the expression of EGFR and alleviated the H2O2-induced decrease in EGFR and phosphorylated ERK. Moreover, inhibition of epidermal growth factor-induced EGFR and ERK signaling in oxidative stress was partially blocked by PRDX6 overexpression. Discussion: Our findings indicate that PRDX6 overexpression protects RPE cells from oxidative stress damage caused by decreasing ROS production and partially blocking the inhibition of the EGFR/ERK signaling pathway induced by oxidative stress. Therefore, PRDX6 shows promise as a therapeutic target for the prevention of RPE cell damage caused by oxidative stress associated with retinal diseases.

Cross-platform comparisons for targeted bisulfite sequencing of MGISEQ-2000 and NovaSeq6000.

BACKGROUND: An accurate and reproducible next-generation sequencing platform is essential to identify malignancy-related abnormal DNA methylation changes and translate them into clinical applications including cancer detection, prognosis, and surveillance. However, high-quality DNA methylation sequencing has been challenging because poor sequence diversity of the bisulfite-converted libraries severely impairs sequencing quality and yield. In this study, we tested MGISEQ-2000 Sequencer's capability of DNA methylation sequencing with a published non-invasive pancreatic cancer detection assay, using NovaSeq6000 as the benchmark. RESULTS: We sequenced a series of synthetic cell-free DNA (cfDNA) samples with different tumor fractions and found MGISEQ-2000 yielded data with similar quality as NovaSeq6000. The methylation levels measured by MGISEQ-2000 demonstrated high consistency with NovaSeq6000. Moreover, MGISEQ-2000 showed a comparable analytic sensitivity with NovaSeq6000, suggesting its potential for clinical detection. As to evaluate the clinical performance of MGISEQ-2000, we sequenced 24 clinical samples and predicted the pathology of the samples with a clinical diagnosis model, PDACatch classifier. The clinical model performance of MGISEQ-2000's data was highly consistent with that of NovaSeq6000's data, with the area under the curve of 1. We also tested the model's robustness with MGISEQ-2000's data when reducing the sequencing depth. The results showed that MGISEQ-2000's data showed matching robustness of the PDACatch classifier with NovaSeq6000's data. CONCLUSIONS: Taken together, MGISEQ-2000 demonstrated similar data quality, consistency of the methylation levels, comparable analytic sensitivity, and matching clinical performance, supporting its application in future non-invasive early cancer detection investigations by detecting distinct methylation patterns of cfDNAs.

Cell-free DNA methylation biomarker for the diagnosis of papillary thyroid carcinoma.

BACKGROUND: Cell-free DNA (cfDNA) is being explored as biomarker for non-invasive diagnosis of cancer. We aimed to establish a cfDNA-based DNA methylation marker panel to differentially diagnose papillary thyroid carcinoma (PTC) from benign thyroid nodule (BTN). METHODS: 220 PTC- and 188 BTN patients were enrolled. Methylation markers of PTC were identified from patients' tissue and plasma by reduced representation bisulfite sequencing and methylation haplotype analyses. They were combined with PTC markers from literatures and were tested on additional PTC and BTN samples to verify PTC-detecting ability using targeted methylation sequencing. Top markers were developed into ThyMet and were tested in 113 PTC and 88 BTN cases to train and validate a PTC-plasma classifier. Integration of ThyMet and thyroid ultrasonography was explored to improve accuracy. FINDINGS: From 859 potential PTC plasma-discriminating markers that include 81 markers identified by us, the top 98 most PTC plasma-discriminating markers were selected for ThyMet. A 6-marker ThyMet classifier for PTC plasma was trained. In validation it achieved an Area Under the Curve (AUC) of 0.828, similar to thyroid ultrasonography (0.833) but at higher specificity (0.722 and 0.625 for ThyMet and ultrasonography, respectively). A combinatorial classifier by them, ThyMet-US, improved AUC to 0.923 (sensitivity = 0.957, specificity = 0.708). INTERPRETATION: The ThyMet classifier improved the specificity of differentiating PTC from BTN over ultrasonography. The combinatorial ThyMet-US classifier may be effective in preoperative diagnosis of PTC. FUNDING: This work was supported by the grants from National Natural Science Foundation of China (82072956 and 81772850).

Polarization-independent and ultra-sensitive biosensor with a one-dimensional topological photonic crystal.

Optical biosensor, which perceptively captures the variety of refractive index (RI) of the surrounding environment, has great potential applications in detecting property changes and types of analytes. However, the disequilibrium of light-matter interaction in different polarizations lead to the polarization-dependence and low sensitivity. Here, we propose a polarization-independent and ultrasensitive biosensor by introducing a one-dimensional topological photonic crystal (1D TPhC), where two N-period 1D photonic crystals (PhC1 and PhC2) with different topological invariants are designed for compressing the interaction region of the optical fields, and enhancing the interaction between the light and analyte. Since the strong light-matter interaction caused by the band-inversion is polarization-independent, the biosensor can obtain superior sensing performance both for TE and TM polarization modes. The sensitivity and Figure of Merit (FOM) of the designed biosensor are 1.5677×106 RIU-1 (1.3497 × 106 RIU-1) and 7.8387×1010 RIU-1deg-1 (4.4990×1010 RIU-1deg-1) for TM (TE) polarization mode, which performs two orders of magnitude enhancement compared with the reported biosensors. With the protection of the topological edge state, this biosensor has high tolerance to the thickness deviations and refractive index (RI) variations of the component materials, which can reduce the requirements on fabrication and working environment. It is anticipated that the proposed biosensor possesses excellent sensing performances, may have great potentials in environmental monitoring, medical detection, etc.

The W-Acidic Motif of Histidine Kinase WalK Is Required for Signaling and Transcriptional Regulation in Streptococcus mutans.

In Streptococcus mutans, we find that the histidine kinase WalK possesses the longest C-terminal tail (CTT) among all 14 TCSs, and this tail plays a key role in the interaction of WalK with its response regulator WalR. We demonstrate that the intrinsically disordered CTT is characterized by a conserved tryptophan residue surrounded by acidic amino acids. Mutation in the tryptophan not only disrupts the stable interaction, but also impairs the efficient phosphotransferase and phosphatase activities of WalRK. In addition, the tryptophan is important for WalK to compete with DNA containing a WalR binding motif for the WalR interaction. We further show that the tryptophan is important for in vivo transcriptional regulation and bacterial biofilm formation by S. mutans. Moreover, Staphylococcus aureus WalK also has a characteristic CTT, albeit relatively shorter, with a conserved W-acidic motif, that is required for the WalRK interaction in vitro. Together, these data reveal that the W-acidic motif of WalK is indispensable for its interaction with WalR, thereby playing a key role in the WalRK-dependent signal transduction, transcriptional regulation and biofilm formation.

Risk-standardized sepsis mortality map of the United States.

OBJECTIVE: Sepsis is the leading cause of in-hospital mortality in the United States (US). Quality improvement initiatives for improving sepsis care depend on accurate estimates of sepsis mortality. While hospital 30-day risk-standardized mortality rates have been published for patients hospitalized with acute myocardial infarction, heart failure, and pneumonia, risk-standardized mortality rates for sepsis have not been well characterized. We aimed to construct a sepsis risk-standardized mortality rate map for the United States, to illustrate disparities in sepsis care across the country. METHODS: This cross-sectional study included adults from the US Nationwide Inpatient Sample who were hospitalized with sepsis between 1 January 2010 and 30 December 2011. Hospital-level risk-standardized mortality rates were calculated using hierarchical logistic modelling, and were risk-adjusted with predicted mortality derived from (1) the Sepsis Risk Prediction Score, a logistic regression model, and (2) gradient-boosted decision trees, a supervised machine learning (ML) algorithm. RESULTS: Among 1,739,033 adults hospitalized with sepsis, 50% were female, and the median age was 71 years (interquartile range: 58-81). The national median risk-standardized mortality rate for sepsis was 18.4% (interquartile range: 17.0, 21.0) by the boosted tree model, which had better discrimination than the Sepsis Risk Prediction Score model (C-statistic 0.87 and 0.78, respectively). The highest risk-standardized mortality rates were found in Wyoming, North Dakota, and Mississippi, while the lowest were found in Arizona, Colorado, and Michigan. CONCLUSIONS: Wide variation exists in sepsis risk-standardized mortality rates across states, representing opportunities for improvement in sepsis care. This represents the first map of state-level variation of risk-standardized mortality rates in sepsis.

Integrative Modeling of Multiomics Data for Predicting Tumor Mutation Burden in Patients with Lung Cancer.

Immunotherapy has been widely used in the treatment of lung cancer, and one of the most effective biomarkers for the prognosis of immunotherapy currently is tumor mutation burden (TMB). Although whole-exome sequencing (WES) could be utilized to assess TMB, several problems prevent its routine clinical application. To develop a simplified TMB prediction model, patients with lung adenocarcinoma (LUAD) in The Cancer Genome Atlas (TCGA) were randomly split into training and validation cohorts and categorized into the TMB-high (TMB-H) and TMB-low (TMB-L) groups, respectively. Based on the 610 differentially expressed genes, 50 differentially expressed miRNAs and 58 differentially methylated CpG sites between TMB-H and TMB-L patients, we constructed 4 predictive signatures and established TMB prediction model through machine learning methods that integrating the expression or methylation profiles of 7 genes, 7 miRNAs, and 6 CpG sites. The multiomics model exhibited excellent performance in predicting TMB with the area under curve (AUC) of 0.911 in the training cohort and 0.859 in the validation cohort. Besides, the significant correlation between the multiomics model score and TMB was observed. In summary, we developed a prognostic TMB prediction model by integrating multiomics data in patients with LUAD, which might facilitate the further development of quantitative real time-polymerase chain reaction- (qRT-PCR-) based TMB prediction assay.

All-Dielectric Metasurface-Based Beam Splitter with Arbitrary Splitting Ratio.

The development of optical systems is heading to multi-branch circuit design and miniaturization. A beam splitter is a common device for dividing an incident beam into two separate beams. Conventional beam splitters are constructed using coated prisms or glass plate. Their bulky size, right-angled output direction, and fixed splitting ratio greatly limit the design of optical arrangement and also hinder the system integration. Here, an all-dielectric metasurface composed of symmetric nano-rings as a beam splitter are designed by Finite-Difference Time-Domain method. By changing the inner and outer radiuses of the nano-rings, the wavefront phase of the emergence beam can be adjusted to form a phase gradient, and the incident beam of arbitrary polarization is divided into two beams according to the designed transmittance and angle. The initial phase of the emergence beam can be changed by adjusting the refractive index of the substrate or adding the silicon film to the substrate, and the splitting ratio can be adjusted from 0.5:1 to 1:1. The simulation demonstrates that the metasurface-based beam splitter is independent of polarization and the power efficiency is over 92% with a compact area of 33.6 µm × 33.6 µm. This compact metasurface-based beam splitter has promising potential for enabling new types of compact optical systems and advancing metasurface-based functional integrated photonic applications.

Circulating microRNA profiles based on direct S-Poly(T)Plus assay for detection of coronary heart disease.

Coronary heart disease (CHD) is one of the leading causes of heart-associated deaths worldwide. Conventional diagnostic techniques are ineffective and insufficient to diagnose CHD with higher accuracy. To use the circulating microRNAs (miRNAs) as non-invasive, specific and sensitive biomarkers for diagnosing of CHD, 203 patients with CHD and 144 age-matched controls (126 high-risk controls and 18 healthy volunteers) were enrolled in this study. The direct S-Poly(T)Plus method was used to identify novel miRNAs expression profile of CHD patients and to evaluate their clinical diagnostic value. This method is an RNA extraction-free and robust quantification method, which simplifies procedures, reduces variations, in particular increases the accuracy. Twelve differentially expressed miRNAs between CHD patients and high-risk controls were selected, and their performances were evaluated in validation set-1 with 96 plasma samples. Finally, six (miR-15b-5p, miR-29c-3p, miR-199a-3p, miR-320e, miR-361-5p and miR-378b) of these 12 miRNAs were verified in validation set-2 with a sensitivity of 92.8% and a specificity of 89.5%, and the AUC was 0.971 (95% confidence interval, 0.948-0.993, P < .001) in a large cohort for CHD patients diagnosis. Plasma fractionation indicated that only a small amount of miRNAs were assembled into EVs. Direct S-Poly(T)Plus method could be used for disease diagnosis and 12 unique miRNAs could be used for diagnosis of CHD.

Direct S-Poly(T) Plus assay in quantification of microRNAs without RNA extraction and its implications in colorectal cancer biomarker studies.

BACKGROUND: Advances in microRNAs (miRNAs) biomarkers have generated disease markers with potential clinical values. However, none of these published results have been applied in clinic until today. The main reason could be the lack of simple but robust miRNA measurements. METHODS: We built up a simple but ultrasensitive RT-qPCR protocol, Direct S-Poly(T) Plus assay, for detecting miRNAs without RNA purification. In this study, the method was optimized and compared with other RNA purification-based miRNA assays, and the sensitivity was tested. Using Direct S-Poly(T) Plus method, seven potential miRNA biomarkers of colorectal cancer were validated. RESULTS: It is possible to detect approximately 100 miRNAs with minimal plasma inputs (20 µl) and time (~ 140 min) with this approach. The sensitivity of this method was 2.7-343-fold higher than that of the stem-loop method, and comparable with S-Poly(T) plus method. 7 validated miRNA biomarkers of colorectal cancer by Direct S-Poly(T) plus assay could discriminate colorectal cancer stage I from healthy individuals, and promised satisfactory discrimination with the area under receiver operating characteristic (ROC) curve ranging from 0.79 to 0.94 (p value < 0.001). CONCLUSIONS: This simple and robust protocol may have strong impact on the development of specific miRNAs as biomarkers in clinic.

Systems biology based meth-miRNA-mRNA regulatory network identifies metabolic imbalance and hyperactive cell cycle signaling involved in hepatocellular carcinoma onset and progression.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the leading cause of cancer associated deaths worldwide. Independent studies have proposed altered DNA methylation pattern and aberrant microRNA (miRNA) levels leading to abnormal expression of different genes as important regulators of disease onset and progression in HCC. Here, using systems biology approaches, we aimed to integrate methylation, miRNA profiling and gene expression data into a regulatory methylation-miRNA-mRNA (meth-miRNA-mRNA) network to better understand the onset and progression of the disease. METHODS: Patients' gene methylation, miRNA expression and gene expression data were retrieved from the NCBI GEO and TCGA databases. Differentially methylated genes, and differentially expressed miRNAs and genes were identified by comparing respective patients' data using two tailed Student's t-test. Functional annotation and pathway enrichment, miRNA-mRNA inverse pairing and gene set enrichment analyses (GSEA) were performed using DAVID, miRDIP v4.1 and GSEA tools respectively. meth-miRNA-mRNA network was constructed using Cytoscape v3.5.1. Kaplan-Meier survival analyses were performed using R script and significance was calculated by Log-rank (Mantel-Cox) test. RESULTS: We identified differentially expressed mRNAs, miRNAs, and differentially methylated genes in HCC as compared to normal adjacent tissues by analyzing gene expression, miRNA expression, and methylation profiling data of HCC patients and integrated top miRNAs along with their mRNA targets and their methylation profile into a regulatory meth-miRNA-mRNA network using systems biology approach. Pathway enrichment analyses of identified genes revealed suppressed metabolic pathways and hyperactive cell cycle signaling as key features of HCC onset and progression which we validated in 10 different HCC patients' datasets. Next, we confirmed the inverse correlation between gene methylation and its expression, and between miRNA and its targets' expression in various datasets. Furthermore, we validated the clinical significance of identified methylation, miRNA and mRNA signatures by checking their association with clinical features and survival of HCC patients. CONCLUSIONS: Overall, we suggest that simultaneous (1) reversal of hyper-methylation and/or oncogenic miRNA driven suppression of genes involved in metabolic pathways, and (2) induction of hyper-methylation and/or tumor suppressor miRNA driven suppression of genes involved in cell cycle signaling have potential of inhibiting disease aggressiveness, and predicting good survival in HCC.

Circulating Plasma miRNAs as Potential Biomarkers of Non-Small Cell Lung Cancer Obtained by High-Throughput Real-Time PCR Profiling.

BACKGROUND: Because of limited stability and sensitivity, circulating miRNAs as noninvasive biomarkers have not so far been used for early diagnosis and prognosis of non-small cell lung cancer (NSCLC) in clinic. Therefore, it is imperative to find more reliable biomarker(s). METHODS: We performed one of most sensitive qRT-PCR assays, S-Poly(T) Plus, to select differently expressed miRNAs from genome-wide miRNA profiling. miRNA candidates were validated through a three-phase selection and two validation processes with 437 NSCLC cases and 415 controls. RESULTS: A unique set of 7 and 9 miRNAs differed significantly in adenocarcinoma (ADC) and squamous cell carcinoma (SCC) samples compared with those in controls, of which, there were 5 universal biomarkers for NSCLC (ADC or SCC). Ten of 11 miRNAs could discriminate early stage (stage I) of NSCLC from healthy individuals. Risk score was obtained from the validation set-1 and was tested using the ROC curves with a high area under ROC curve of 0.89 in ADC and 0.96 in SCC. Ultimately, potential biomarkers and the risk score were verified by the validation set-2 with a sensitivity of 94% and a specificity of 91.6% in ADC, and a sensitivity of 98.5% and a specificity of 51.5% in SCC, respectively. CONCLUSIONS: Taken together, 7 miRNAs and 9 miRNAs may provide noninvasive biomarkers for diagnosis and prognosis in ADC and SCC, respectively. IMPACT: On the basis of our sensitive and accurate method, we hope that these candidate miRNAs may have strong impact on the early lung cancer diagnosis.

Conformational dynamics of the essential sensor histidine kinase WalK.

Two-component systems (TCSs) are key elements in bacterial signal transduction in response to environmental stresses. TCSs generally consist of sensor histidine kinases (SKs) and their cognate response regulators (RRs). Many SKs exhibit autokinase, phosphoryltransferase and phosphatase activities, which regulate RR activity through a phosphorylation and dephosphorylation cycle. However, how SKs perform different enzymatic activities is poorly understood. Here, several crystal structures of the minimal catalytic region of WalK, an essential SK from Lactobacillus plantarum that shares 60% sequence identity with its homologue VicK from Streptococcus mutans, are presented. WalK adopts an asymmetrical closed structure in the presence of ATP or ADP, in which one of the CA domains is positioned close to the DHp domain, thus leading both the ß- and γ-phosphates of ATP/ADP to form hydrogen bonds to the ℇ- but not the δ-nitrogen of the phosphorylatable histidine in the DHp domain. In addition, the DHp domain in the ATP/ADP-bound state has a 25.7° asymmetrical helical bending coordinated with the repositioning of the CA domain; these processes are mutually exclusive and alternate in response to helicity changes that are possibly regulated by upstream signals. In the absence of ATP or ADP, however, WalK adopts a completely symmetric open structure with its DHp domain centred between two outward-reaching CA domains. In summary, these structures of WalK reveal the intrinsic dynamic properties of an SK structure as a molecular basis for multifunctionality.

Oxidative stress affects retinal pigment epithelial cell survival through epidermal growth factor receptor/AKT signaling pathway.

AIM: To investigate the cross-talk between oxidative stress and the epidermal growth factor receptor (EGFR)/AKT signaling pathway in retinal pigment epithelial (RPE) cells. METHODS: Human RPE cell lines (ARPE-19 cell) were treated with different doses of epidermal growth factor (EGF) and hydrogen peroxide (H2O2). Cell viability was determined by a methyl thiazolyl tetrazolium assay. Cell proliferation was examined by a bromodeoxyuridine (BrdU) incorporation assay. EGFR/AKT signaling was detected by Western blot. EGFR localization was also detected by immunofluorescence. In addition, EGFR/AKT signaling was intervened upon by EGFR inhibitor (erlotinib), PI3K inhibitor (A66) and AKT inhibitor (MK-2206), respectively. H2O2-induced oxidative stress was blocked by antioxidant N-acetylcysteine (NAC). RESULTS: EGF treatment increased ARPE-19 cell viability and proliferation through inducing phosphorylation of EGFR and AKT. H2O2 inhibited ARPE-19 cell viability and proliferation and also suppressed EGF-stimulated increase of RPE cell viability and proliferation by affecting the EGFR/AKT signaling pathway. EGFR inhibitor erlotinib blocked EGF-induced phosphorylation of EGFR and AKT, while A66 and MK-2206 only blocked EGF-induced phosphorylation of AKT. EGF-induced phosphorylation and endocytosis of EGFR were also affected by H2O2 treatment. In addition, antioxidant NAC attenuated H2O2-induced inhibition of ARPE-19 cell viability through alleviating reduction of EGFR, and phosphorylated and total AKT proteins. CONCLUSION: Oxidative stress affects RPE cell viability and proliferation through interfering with the EGFR/AKT signaling pathway. The EGFR/AKT signaling pathway may be an important target in oxidative stress-induced RPE cell dysfunction.

Correction: Auranofin Inhibits Retinal Pigment Epithelium Cell Survival through Reactive Oxygen Species-Dependent Epidermal Growth Factor Receptor/ Mitogen-Activated Protein Kinase Signaling Pathway.

[This corrects the article DOI: 10.1371/journal.pone.0166386.].

Auranofin Inhibits Retinal Pigment Epithelium Cell Survival through Reactive Oxygen Species-Dependent Epidermal Growth Factor Receptor/ Mitogen-Activated Protein Kinase Signaling Pathway.

Abnormal survival of retinal pigment epithelium (RPE) cells contributes to the pathogenesis of proliferative vitreoretinopathy (PVR), a sight-threatening disease. In this study, we explored the effect of the anti-rheumatic agent auranofin (AF) on RPE cell survival and studied the underlying signaling mechanisms in vitro. Our results showed that AF inhibited ARPE-19 cell survival in a dose and time-dependent manner. Application of AF induced several effects: a significant decrease in total epidermal growth factor receptor (EGFR) and an increase in phosphorylated EGFR and mitogen-activated protein kinase (MAPK), including extracellular signal-regulated kinase (ERK), P38 mitogen-activated protein kinase (P38MAPK), c-Jun N-terminal kinase (JNK), c-Jun, mitogen activated protein kinase activated protein kinase 2(MAPKAPK2), and heat shock protein 27 (HSP27). AF also inhibited epidermal growth factor (EGF)-dependent cell proliferation and migration through affecting EGFR/MAPK signaling. The antioxidant N-acetylcysteine (NAC) blocked the AF-induced increase of reactive oxygen species (ROS) production, the reduction of total EGFR, and the phosphorylation of multiple nodes in EGFR/MAPK signaling pathway. P38MAPK inhibitor SB203580, but not inhibitors of EGFR (erlotinib), ERK (FR180204) and JNK (SP600125), suppressed AF-induced phosphorylation of EGFR/p38MAPK/MAPKAPK2/Hsp27. In conclusion, the ROS-dependent phosphorylation of EGFR/MAPK is an important signaling pathway for AF-induced inhibition of RPE cell survival, and AF may have the potential for treatment of abnormal survival of RPE cells in PVR.

Discussion on robot-assisted laparoscopic cystectomy and Ileal neobladder surgery preoperative care.

To investigate the clinical applications of robot-assisted radical cystectomy with orthotopic ileal neobladder (RARC-INB) and make a preliminary summary. Retrospective analysis the clinical data of 12 patients underwent robotic bladder cancer (da vinci surgical system) assisted laparoscopic cystectomy with ileal orthotopic neobladder from March 2015 to April 2015. 12 cases were successful, with no surgical intervention, and organ damage occurred. The operation time was 330~470 min, which average (390.0±61.5) min; blood loss was 90~870 ml, which average (185.0±88.3) ml. One case of intraoperative blood transfusion was 400 ml. The enjoin eating time of postoperative intestinal ventilation was 3~6 d, and the average time was (4.0±1.5) d. Removal of ureteral stents time was 14~28 d and the average time was (21±7) d. Removal of the catheter time was 18~28 d and the average time was (23±5) d. Postoperative hospital stay 19~29 d and the average time was (24±5) dRARC-INB make the surgical tends to simplify, which was conducive to surgeon intraoperative control and assurance. RARC-INB make the surgical tends to use less trauma, less bleeding, complete lymphadenectomy, quick recovery, etc. It is a safe, effective and reliablethe method in the treatment of invasive bladder cancer. So the method should be widely applied.