Elevated NOX4 promotes tumorigenesis and acquired EGFR-TKIs resistance via enhancing IL-8/PD-L1 signaling in NSCLC.

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been widely used for human non-small-cell lung cancer (NSCLC) treatment. However, acquired resistance to EGFR-TKIs is the major barrier of treatment success, and new resistance mechanism remains to be elucidated. In this study, we found that elevated NADPH oxidase 4 (NOX4) expression was associated with acquired EGFR-TKIs resistance. Gefitinib is the first-generation FDA-approved EGFR-TKI, and osimertinib is the third-generation FDA-approved EGFR-TKI. We demonstrated that NOX4 knockdown in the EGFR-TKI resistant cells enabled the cells to become sensitive to gefitinib and osimertinib treatment, while forced expression of NOX4 in the sensitive parental cells was sufficient to induce resistance to gefitinib and osimertinib in the cells. To elucidate the mechanism of NOX4 upregulation in increasing TKIs resistance, we found that knockdown of NOX4 significantly down-regulated the expression of transcription factor YY1. YY1 bound directly to the promoter region of IL-8 to transcriptionally activate IL-8 expression. Interestingly, knockdown of NOX4 and IL-8 decreased programmed death ligand 1 (PD-L1) expression, which provide new insight on TKIs resistance and immune escape. We found that patients with higher NOX4 and IL-8 expression levels showed a shorter survival time compared to those with lower NOX4 and IL-8 expression levels in response to the anti-PD-L1 therapy. Knockdown of NOX4, YY1 or IL-8 alone inhibited angiogenesis and tumor growth. Furthermore, the combination of NOX4 inhibitor GKT137831 and gefitinib had synergistic effect to inhibit cell proliferation and tumor growth and to increase cellular apoptosis. These findings demonstrated that NOX4 and YY1 were essential for mediating the acquired EGFR-TKIs resistance. IL-8 and PD-L1 are two downstream targets of NOX4 to regulate TKIs resistance and immunotherapy. These molecules may be used as potential new biomarkers and therapeutic targets for overcoming TKIs resistance in the future.

HK2 and LDHA upregulation mediate hexavalent chromium-induced carcinogenesis, cancer development and prognosis through miR-218 inhibition.

Hexavalent chromium [Cr(VI)] is one of the most common environmental contaminants due to its tremendous industrial applications, but its effects and mechanism remain to be investigated. Our previous studies showed that Cr(VI) exposure caused malignant transformation and tumorigenesis. This study showed that glycolytic proteins HK2 and LDHA levels were statistically significant changed in blood samples of Cr(VI)-exposed workers and in Cr-T cells compared to the control subjects and parental cells. HK2 and LDHA knockdown inhibited cell proliferation and angiogenesis, and higher HK2 and LDHA expression levels are associated with advanced stages and poor prognosis of lung cancer. We found that miR-218 levels were significantly decreased and miR-218 directly targeted HK2 and LDHA for inhibiting their expression. Overexpression of miR-218 inhibited glucose consumption and lactate production in Cr-T cells. Further study found that miR-218 inhibited tumor growth and angiogenesis by decreasing HK2 and LDHA expression in vivo. MiR-218 levels were negatively correlated with HK2 and LDHA expression levels and cancer development in human lung and other cancers. These results demonstrated that miR-218/HK2/LDHA pathway is vital for regulating Cr(VI)-induced carcinogenesis and human cancer development.

Association of cigarette smoking with oral bacterial microbiota and cardiometabolic health in Chinese adults.

The interplay among cigarette smoking status, oral microbiota, and cardiometabolic health is poorly understood. We aimed to examine the association of cigarette smoking status with oral microbiota and to assess the association of the identified microbial features with cardiometabolic risk factors in a Chinese population. This study included 587 participants within the Central China Cohort, including 111 smokers and 476 non-smokers, and their oral microbiota was profiled by 16S rRNA sequencing. Both oral microbial alpha- and beta-diversity were distinct between smokers and non-smokers (p < 0.05). With adjustment for sociodemographics, alcohol and tea drinking, tooth brushing frequency, and body mass index, the relative abundance of nine genera and 26 pathways, including the genus Megasphaera and two pathways involved in inositol degradation which have potentially adverse effects on cardiometabolic health, was significantly different between two groups (FDR q < 0.20). Multiple microbial features related to cigarette smoking were found to partly mediate the associations of cigarette smoking with serum triglycerides and C-reactive protein levels (p-mediation < 0.05). In conclusion, cigarette smoking status may have impacts on the oral microbial features, which may partially mediate the associations of cigarette smoking and cardiometabolic health.

Dysregulation of MiR-199a/IL8 pathway in chronic Cr (VI)-induced tumor growth and angiogenesis.

Hexavalent chromium [Cr(VI)] is a well-known environmental carcinogen. Recent studies revealed that chronic exposure of human bronchial epithelial cells (BEAS-2B, B2B) to Cr(VI) activated several signaling pathways and induced cell malignant transformation and tumor growth. However, new mechanisms of Cr(VI) in inducing carcinogenesis remains to be elucidated. This study showed that miR-199a expression levels were significantly lower in Cr(VI)-transformed Cr-T cells. By using the mouse model, the expression levels of miR-199a were significantly decreased in blood samples and lung tissues of mice intranasally exposed to Cr(VI) for 12 weeks compared to the solvent exposure control. Overexpression of miR-199a inhibited tube formation and angiogenesis. C-X-C motif chemokine ligand 8 (CXCL8, IL8) levels were significantly higher in blood samples of Cr (VI)-exposed workers compared to normal workers, and forced expression of miR-199a in the cells suppressed IL8 levels. miR-199a suppression induced expression of hypoxia-inducible factor 1α (HIF-1α) and nuclear factor kappa B (NF-κB) p65 to increase IL8 expression. With animal experiment, the results showed that miR-199a overexpression inhibited tumor growth and angiogenesis through inhibiting IL8, HIF-1α and NF-κB p65 expression in vivo. These results show that miR-199a/IL8 pathway is important in Cr(VI)-induced carcinogenesis and angiogenesis.

Cooperative In Situ Assembly of G-Quadruplex DNAzyme Nanowires for One-Step Sensing of CpG Methylation in Human Genomes.

CpG methylation is one the most predominant epigenetic modification that has been recognized as a molecular-level biomarker for various human diseases. Taking advantage of methylation-dependent cleavage and encoding flexibility in nucleic acid functions and structures, we demonstrate the cooperative in situ assembly of G-quadruplex DNAzyme nanowires for one-step sensing of CpG methylation in human genomes. This nanodevice displays good specificity and high sensitivity with a limit of detection (LOD) of 0.565 aM in vitro and 1 cell in vivo. It can distinguish 0.001% CpG methylation level from excess unmethylated DNA, quantify different CpG methylation targets from diverse human cancer cells, and even discriminate CpG methylation expressions between lung tumor and precancerous tissues. Importantly, this nanodevice can be performed isothermally in one step within 2 h in a label-free manner without any bisulfite conversion, fluorescence tagging, and PCR amplification process, providing a new platform for genomic methylation-related clinical diagnosis and biomedical research.

Development of a CRISPR-Cas-Based Biosensor for Rapid and Sensitive Detection of 8-Oxoguanine DNA Glycosylase.

8-Oxoguanine DNA glycosylase is essential for maintaining genomic integrity and stability, while its abnormal activity may lead to the disturbance in the normal DNA damage repair and the occurrence of carcinogenicity and teratogenicity. Herein, we construct a CRISPR-Cas-based biosensor for rapid and sensitive measurement of 8-oxoguanine DNA glycosylases. This biosensor involves a hairpin probe and integrates quadratic strand displacement amplification (SDA) with a CRISPR/Cas12a effector with the characteristics of rapidity (within 40 min) and isothermal assay. The presence of 8-oxoguanine DNA glycosylase can initiate the quadratic SDA to produce large amounts of activators with the assistance of polynucleotide kinase (PNK). Subsequently, the activators can bind with crRNA to activate Cas12a, cleaving signal probes and recovering Cy5 fluorescence, which can be accurately quantified by single-molecule imaging. Notably, the designed hairpin probes can effectively block the hybridization of the generated activators with free hairpin probes, endowing this biosensor with high sensitivity. In addition, the utilization of PNK instead of apurinic/apyrimidinic endonuclease (APE1) greatly simplifies the experimental procedure to only a one-step reaction. The introduction of a single-molecule detection further reduces the sample consumption and improves the sensitivity. This biosensor displays a detection limit of 4.24 × 10-9 U µL-1, and it can accurately quantify cellular human 8-oxoguanine DNA glycosylase at a single-cell level. Furthermore, this biosensor can be applied for the screening of inhibitors, the analysis of kinetic parameters, and the discrimination of cancer cells from normal cells, with potential applications in molecular diagnostic and point-of-care testing.

Hydroxymethylation-Specific Ligation-Mediated Single Quantum Dot-Based Nanosensors for Sensitive Detection of 5-Hydroxymethylcytosine in Cancer Cells.

5-Hydroxymethylcytosine (5hmC) modification is a key epigenetic regulator of cellular processes in mammalian cells, and its misregulation may lead to various diseases. Herein, we develop a hydroxymethylation-specific ligation-mediated single quantum dot (QD)-based fluorescence resonance energy transfer (FRET) nanosensor for sensitive quantification of 5hmC modification in cancer cells. We design a Cy5-modified signal probe and a biotinylated capture probe for the recognition of specific 5hmC-containing genes. 5hmC in target DNA can be selectively converted by T4 ß-glucosyltransferase to produce a glycosyl-modified 5hmC, which cannot be cleaved by methylation-insensitive restriction enzyme MspI. The glycosylated 5hmC DNA may act as a template to ligate a signal probe and a capture probe, initiating hydroxymethylation-specific ligation to generate large amounts of biotin-/Cy5-modified single-stranded DNAs (ssDNAs). The assembly of biotin-/Cy5-modified ssDNAs onto a single QD through streptavidin-biotin interaction results in FRET and consequently the generation of a Cy5 signal. The nanosensor is very simple without the need for bisulfite treatment, radioactive reagents, and 5hmC-specific antibodies. Owing to excellent specificity and high amplification efficiency of hydroxymethylation-specific ligation and near-zero background of a single QD-based FRET, this nanosensor can quantify 5hmC DNA with a limit of detection of 33.61 aM and a wider linear range of 7 orders of magnitude, and it may discriminate the single-nucleotide difference among 5hmC, 5-methylcytosine, and unmodified cytosine. Moreover, this nanosensor can distinguish as low as a 0.001% 5hmC DNA in complex mixtures, and it can monitor the cellular 5hmC level and discriminate cancer cells from normal cells, holding great potential in biomedical research and clinical diagnostics.

Activatable Self-Dissociation of Watson-Crick Structures with Fluorescent Nucleotides for Sensing Multiple Human Glycosylases at Single-Cell Level.

Nucleobase oxidation and alkylation can destroy Watson-Crick base-pairing to challenge the genomic integrity. Human 8-oxoguanine glycosylase 1 (hOGG1) and alkyladenine glycosylase (hAAG) are evolved to counter these two cytotoxic lesions through base-excision repair, and their deregulations are implicated with multifactorial diseases and cancers. Herein, we demonstrate activatable self-dissociation of Watson-Crick structures with fluorescent nucleotides for sensing multiple human glycosylases at single-cell level. The presence of hOGG1 and hAAG catalyzes 8-oxoG and deoxyinosine removal in functional probe 1 to release two trigger probes (1 and 2). Then, trigger probes hybridize with functional probe 2 to activate the autocatalytic degradation of functional probes 2 (Cycle I) and 3 (Cycle II), replicating abundant trigger probes (1-4) and releasing two fluorophores (2-aminopurine (2-AP) and pyrrolo-dC (P-dC)). New trigger probes (1, 2) and (3, 4), in turn, hybridize with free functional probes 2 and 3, repeating Cycles I and II turnovers. Through multicycle self-dissociation of Watson-Crick structures, 2-AP and P-dC are exponentially accumulated for the simultaneous quantification of hOGG1 and hAAG. This nanodevice exhibits high sensitivity with a detection limit of 2.9 × 10-3 U/mL for hOOG1 and 1.5 × 10-3 U/mL for hAAG, and it can measure enzymatic kinetics, identify potential inhibitors, discriminate glycosylases between cancer and normal cell lines, and even quantify glycosylase activities in a single HeLa cell. Moreover, this assay may be rapidly and isothermally performed in one tube with only one tool enzyme in a quencher-free manner, promising a simple and powerful platform for multiple human glycosylase detection.

Fabrication of Two Isomorphic and Hyperstable Rare Earth-Based Metal-Organic Frameworks with Efficient Ratiometric Probe and Photocatalytic Performances.

Two isomorphic lanthanide compounds {[Ln(ddpp)(H2O)]·CH3CN}n (Ln = Eu and Gd, H4ddpp = 2,5-di(2',4'-dicarboxylphenyl)pyridine) were synthesized. Complex 1-Eu displays ultrahigh acid-base stability and thermal stability. Furthermore, luminescence measurements revealed that 1-Eu could detect quinolone antibiotics with an ultralow limit of detection in aqueous solution. The ratiometric probe properties for sensing antibiotics could be attributed to the incompletely sensitized Eu3+ ion of the ligand. Remarkably, it is interesting that 1-Gd exhibits excellent tetracycline degradation properties under visible light. Ultraviolet-visible diffuse reflectance spectroscopy and valence band X-ray photoelectron spectroscopy were carried out to investigate the photodegradation mechanisms. Moreover, a rational explanation for the fluorescent probe and photocatalysis behavior of these two complexes was also discussed with the assistance of density functional theory calculations.

miR-196a Upregulation Contributes to Gefitinib Resistance through Inhibiting GLTP Expression.

Tyrosine kinase inhibitor (TKI) therapy has greatly improved lung cancer survival in patients with epidermal growth factor receptor (EGFR) mutations. However, the development of TKI-acquired resistance is the major problem to be overcome. In this study, we found that miR-196a expression was greatly induced in gefitinib-resistant lung cancer cells. To understand the role and mechanism of miR-196a in TKI resistance, we found that miR-196a-forced expression alone increased cell resistance to gefitinib treatment in vitro and in vivo by inducing cell proliferation and inhibiting cell apoptosis. We identified the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) bound to the promoter region of miR-196a and induced miR-196a expression at the transcriptional level. NRF2-forced expression also significantly increased expression levels of miR-196a, and was an upstream inducer of miR-196a to mediate gefitinib resistance. We also found that glycolipid transfer protein (GLTP) was a functional direct target of miR-196a, and downregulation of GLTP by miR-196a was responsible for gefitinib resistance. GLTP overexpression alone was sufficient to increase the sensitivity of lung cancer cells to gefitinib treatment. Our studies identified a new role and mechanism of NRF2/miR-196a/GLTP pathway in TKI resistance and lung tumor development, which may be used as a new biomarker (s) for TKI resistance or as a new therapeutic target in the future.

Discovery of vanoxerine dihydrochloride as a CDK2/4/6 triple-inhibitor for the treatment of human hepatocellular carcinoma.

BACKGROUND: Cyclin-dependent kinases 2/4/6 (CDK2/4/6) play critical roles in cell cycle progression, and their deregulations are hallmarks of hepatocellular carcinoma (HCC). METHODS: We used the combination of computational and experimental approaches to discover a CDK2/4/6 triple-inhibitor from FDA approved small-molecule drugs for the treatment of HCC. RESULTS: We identified vanoxerine dihydrochloride as a new CDK2/4/6 inhibitor, and a strong cytotoxicdrugin human HCC QGY7703 and Huh7 cells (IC50: 3.79 µM for QGY7703and 4.04 µM for Huh7 cells). In QGY7703 and Huh7 cells, vanoxerine dihydrochloride treatment caused G1-arrest, induced apoptosis, and reduced the expressions of CDK2/4/6, cyclin D/E, retinoblastoma protein (Rb), as well as the phosphorylation of CDK2/4/6 and Rb. Drug combination study indicated that vanoxerine dihydrochloride and 5-Fu produced synergistic cytotoxicity in vitro in Huh7 cells. Finally, in vivo study in BALB/C nude mice subcutaneously xenografted with Huh7 cells, vanoxerine dihydrochloride (40 mg/kg, i.p.) injection for 21 days produced significant anti-tumor activity (p < 0.05), which was comparable to that achieved by 5-Fu (10 mg/kg, i.p.), with the combination treatment resulted in synergistic effect. Immunohistochemistry staining of the tumor tissues also revealed significantly reduced expressions of Rb and CDK2/4/6in vanoxerinedihydrochloride treatment group. CONCLUSIONS: The present study isthe first report identifying a new CDK2/4/6 triple inhibitor vanoxerine dihydrochloride, and demonstrated that this drug represents a novel therapeutic strategy for HCC treatment.

Simple Mix-and-Read Assay with Multiple Cyclic Enzymatic Repairing Amplification for Rapid and Sensitive Detection of DNA Glycosylase.

Human 8-oxoguanine DNA glycosylase (hOGG1) can initiate base excision repair of genomic 8-oxoguanine (8-oxoG), and it can locate and remove damaged 8-oxoG through extrusion and excision. Sensitive detection of hOGG1 is critical for clinical diagnosis. Herein, we develop a simple mix-and-read assay for the sensitive detection of DNA glycosylase using multiple cyclic enzymatic repairing amplification. The hOGG1 can excise the 8-oxoG base of the DNA substrate to produce an apurinic/apyrimidinic (AP) site, and then, the AP site can be cleaved by apurinic/apyrimidic endonuclease 1 (APE1), producing the substrate fragment with a free 3'-OH terminus. Subsequently, the substrate fragment can initiate cyclic enzymatic repairing amplification, generating two triggers. The resultant two triggers can function as the primers to induce three cyclic enzymatic repairing amplification, respectively, producing more and more triggers. We experimentally verify the occurrence of each cyclic enzymatic repairing amplification and uracil DNA glycosylase (UDG)-mediated exponential amplification. The amplification products can be simply detected using SYBR Green II as the fluorescent dye. This mix-and-read assay is very simple and rapid (within 40 min) without the requirement of any extra primers and modification/separation steps. This method can sensitively measure hOGG1 with a detection limit of 2.97 × 10-8 U/µL, and it can be applied for the screening of inhibitors and the monitoring of cellular hOGG1 activity at the single-cell level, providing an adaptive and flexible tool for clinical diagnosis and drug discovery.

Construction of a Quencher-Free Cascade Amplification System for Highly Specific and Sensitive Detection of Serum Circulating miRNAs.

Circulating miRNAs are a newly emerging class of noninvasive biomarkers, and the accurate quantification of their expression is essential to the biological research and early clinic diagnosis. Herein, we demonstrate the construction of a quencher-free cascade amplification system for highly sensitive detection of serum circulating miRNAs. The target miRNA can hybridize with the linear probe to induce the cyclic strand displacement amplification (SDA) (cycle I) for the production of the binding probes. The binding probe can subsequently react with the 2-aminopurine (2-AP)-hairpin probe to induce the recycling exonuclease cleavage of 2-AP-hairpin probes (cycle II), releasing the triggers and 2-AP molecules simultaneously. The released trigger can hybridize with the free linear probe to start new cycles I and II amplifications. Through multiple rounds of cascade amplifications, a large number of 2-AP molecules are released, generating an enhanced fluorescence signal. This method exhibits a large dynamic range of 8 orders of magnitude and a detection limit of 0.16 aM. It can differentiate a single-base mismatch in miR-486-5p, quantify miR-486-5p in lung cancer cells at various stages, and even discriminate the expressions of serum circulating miR-486-5p in healthy persons from that in nonsmall-cell lung carcinoma (NSCLC) patients. Moreover, this assay can be rapidly carried out in one step under isothermal condition in a label-free manner, holding promising applications in point-of-care diagnosis and prognosis of lung cancers.

A multifunctional DNA nanostructure based on multicolor FRET for nuclease activity assay.

Nucleases play a crucial role in DNA replication, recombination and repair which are associated with cancers. Herein, we develop a four-color fluorescent probe for ratiometric detection of multiple nucleases. This four-color fluorescent probe consists of four fluorescent dyes connected by a DNA tetrahedral nanostructure with the involvement of multistep fluorescence resonance energy transfer (FRET). Based on the principle of self-assembly, the four-color fluorescent probe is constructed by integrating one acceptor with three spatially and spectrally distinct acceptors. A DNA tetrahedral nanostructure functions as a scaffold to link the acceptor dyes (i.e., diethylaminocoumarin (DEA), carboxyfluorescein (FAM), Texas Red, and Cy5). The fluorescence emissions of DEA, FAM, Texas Red and Cy5 can be observed through efficient multi-step energy transfer. This four-color fluorescent probe enables single excitation/four emissions, and it can be used for ratiometric detection of nucleases (i.e., XhoI, HindIII and KpnI) and the screening of nuclease inhibitors. Importantly, this four-color fluorescent probe can be further applied to discriminate multiple biomolecule targets by simply integrating the recognition sites of various biomolecules into the DNA tetrahedral nanostructure.

Relative Order of Acidity among Hydroxyl Groups of Oxyluciferin and Emission Light Colors in Aqueous Solution.

The magnitude of the acidity of the oxyluciferin in water in the ground and excited state is investigated, and it is found for the first time using computational approach that the enol group of the phenol-enol species is the most acidic in the ground state, but the deprotonation of the phenol of the phenol-keto form is the most favored in the excited state. The relative order of the acidity among the hydroxyl groups in the oxyluciferin is attributed to the sequence of the O-H bond lengths in the enol and phenol group of the phenol-enol form, and the phenol group of the phenol-keto species. The mechanism of determining the dominant emissive species in the excited state is proposed, and the dependence of emission light colors on the photoexcitation energy is elucidated by the high relative concentration of six chemical forms in the ground state and the absorption efficiency.

An in vitro Raman study on compositional correlations of lipids and protein with animal tissue hydration.

Raman spectroscopy is a powerful non-invasive tool for detection and classification of chemical composition of materials including biological tissues. In this work, we report an in vitro Raman study on animal skin samples with a focus on high-frequency vibrations such as symmetric CH3 stretching mode at 2934 cm-1, and the symmetric CH2 vibration mode at 2854 cm-1, OH stretching modes near 3412 cm-1, and bounded OH mode near 3284 cm-1. Raman data was acquired with a customized InGaAs based Raman spectrometer that consolidates the NIR (866 nm) light and the InGaAs detector and is particularly suitable for probing high-frequency vibrations. The Raman spectra of fat, tendon, and muscle tissues are also analyzed to determine the spectroscopic identities of CH and OH groups in skin. Our results suggest that the protein is beneficial for the maintenance of skin hydration, as it has higher water capacity and greater capability to retain water than lipids. This conclusion is consistent with the additional discovery that water exists in fat mainly as unbound type, while part of water exists as bound type in muscle.

Suppression of miR-143 contributes to overexpression of IL-6, HIF-1α and NF-κB p65 in Cr(VI)-induced human exposure and tumor growth.

Hexavalent chromium [Cr(VI)] is a known occupational and environmental contaminant and carcinogen, but new mechanisms of Cr(VI)-induced carcinogenesis remain to be elucidated. In this study, we found that expression of miR-143 is decreased, whereas that of Interleukin 6 (IL-6) is increased in blood samples of Cr(VI)-exposing workers compared with corresponding unexposed workers. In addition, IL-6 was increased in human bronchial epithelial cells (BEAS-Cr) exposed to Cr(VI) compared with unexposed BEAS-2B cells. To further investigate the mechanisms by which Cr(VI) promotes these changes, we assessed the effects of miR-143 on gene expression and found that miR-143 suppressed expression of IL-6, HIF-1α and NF-κB p65, and that inhibiting miR-143 promoted expression of IL-6, HIF-1α and NF-κB p65. Interestingly, IL-6 regulated expression of HIF-1α, and HIF-1α transcriptionally regulated expression of IL-6. Experiments in animals showed that miR-143 inhibited tumor growth and angiogenesis by regulating IL-6/HIF-1α and downstream signaling pathways in vivo. These outcomes support the hypothesis that the miR-143/IL-6/HIF-1α pathway functions to regulate Cr(VI)-induced carcinogenesis.

Lung Endothelial MicroRNA-1 Regulates Tumor Growth and Angiogenesis.

RATIONALE: Vascular endothelial growth factor down-regulates microRNA-1 (miR-1) in the lung endothelium, and endothelial cells play a critical role in tumor progression and angiogenesis. OBJECTIVES: To examine the clinical significance of miR-1 in non-small cell lung cancer (NSCLC) and its specific role in tumor endothelium. METHODS: miR-1 levels were measured by Taqman assay. Endothelial cells were isolated by magnetic sorting. We used vascular endothelial cadherin promoter to create a vascular-specific miR-1 lentiviral vector and an inducible transgenic mouse. KRASG12D mut/Trp53-/- (KP) mice, lung-specific vascular endothelial growth factor transgenic mice, Lewis lung carcinoma xenografts, and primary endothelial cells were used to test the effects of miR-1. MEASUREMENTS AND MAIN RESULTS: In two cohorts of patients with NSCLC, miR-1 levels were lower in tumors than the cancer-free tissue. Tumor miR-1 levels correlated with the overall survival of patients with NSCLC. miR-1 levels were also lower in endothelial cells isolated from NSCLC tumors and tumor-bearing lungs of KP mouse model. We examined the significance of lower miR-1 levels by testing the effects of vascular-specific miR-1 overexpression. Vector-mediated delivery or transgenic overexpression of miR-1 in endothelial cells decreased tumor burden in KP mice, reduced the growth and vascularity of Lewis lung carcinoma xenografts, and decreased tracheal angiogenesis in vascular endothelial growth factor transgenic mice. In endothelial cells, miR-1 level was regulated through phosphoinositide 3-kinase and specifically controlled proliferation, de novo DNA synthesis, and ERK1/2 activation. Myeloproliferative leukemia oncogene was targeted by miR-1 in the lung endothelium and regulated tumor growth and angiogenesis. CONCLUSIONS: Endothelial miR-1 is down-regulated in NSCLC tumors and controls tumor progression and angiogenesis.

Advances in the mechanisms and early warning indicators of the postoperative cognitive dysfunction after the extracorporeal circulation.

The high incidence of postoperative cognitive dysfunction (POCD) after extracorporeal circulation has seriously affected the prognosis and quality of life. Its mechanism may involve the inflammatory response and oxidative stress,the excessive phosphorylation of tau protein, the decreased blood volume and oxygen in the cerebral cortex. Appropriate early warning indicators of POCD after the extracorporeal circulation should be chosen to facilitate the cross validation of the results obtained different technical approaches and thus promote the early diagnosis and treatment of POCD.

[Sodium Ferulate Attenuates Oxidative Stress Induced Inflammation via Suppressing NALP3 and NF-κB Signal Pathway].

OBJECTIVE: To study the effects of sodium ferulate on inflammation in human lung epithelial cells (A549) under oxidative stress and itsinfluence onthe expression of inflammasome NACHT-PYD-containing protein 3 (NALP3) and nuclear factor kappa B (NF-κB). METHODS: Human lung epithelial cells A549 cultured in vitro were divided into 4 groups, including control group, H2O2 (100µmol/L) stress group, NF-κB blockers group (PDTC 100 µmol/L+ H2O2 100 µmol/L), sodium ferulate (SF) intervention group (SF 400µg/mL+ H2O2 10µmol/L). The expression of NALP3,IκBα protein were evaluated by Western blot, while mRNA levels of NALP3, NF-κB (P65) were measured by qRT-PCR. The level of interleukin-1beta (ILß1) were detected by ELISA. RESULTS: H2O2 not only increased the mRNA and protein expression levels of NALP3, but also enhanced the secretion of ILß1p in human lung epithelial cells A549 (P<0. 05) when compared with control group. NF-κB blockers PDTC and sodium ferulateresisted the effects of H2O2 on A549 cells, that decreased the mRNA and protein expression of NALP3 and the mRNA expression of NF-κB (P65), reduced the degeneration of IκBα and the secretion of IL-1ß (P<0. 05) when compared to H2O2 stress group. CONCLUSION: SF may reduce the expression of NALP3 and IL-1ß by inhibiting NF-κB, so as to reduce the inflammation caused by oxidative stress.