Micrometer-thick and porous nanocomposite coating for electrochemical sensors with exceptional antifouling and electroconducting properties.

Development of coating technologies for electrochemical sensors that consistently exhibit antifouling activities in diverse and complex biological environments over extended time is vital for effective medical devices and diagnostics. Here, we describe a micrometer-thick, porous nanocomposite coating with both antifouling and electroconducting properties that enhances the sensitivity of electrochemical sensors. Nozzle printing of oil-in-water emulsion is used to create a 1 micrometer thick coating composed of cross-linked albumin with interconnected pores and gold nanowires. The layer resists biofouling and maintains rapid electron transfer kinetics for over one month when exposed directly to complex biological fluids, including serum and nasopharyngeal secretions. Compared to a thinner (nanometer thick) antifouling coating made with drop casting or a spin coating of the same thickness, the thick porous nanocomposite sensor exhibits sensitivities that are enhanced by 3.75- to 17-fold when three different target biomolecules are tested. As a result, emulsion-coated, multiplexed electrochemical sensors can carry out simultaneous detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acid, antigen, and host antibody in clinical specimens with high sensitivity and specificity. This thick porous emulsion coating technology holds promise in addressing hurdles currently restricting the application of electrochemical sensors for point-of-care diagnostics, implantable devices, and other healthcare monitoring systems.

Self-Mixed Biphasic Liquid Metal Composite with Ultra-High Stretchability and Strain-Insensitivity for Neuromorphic Circuits.

Neuromorphic circuits that can function under extreme deformations are important for various data-driven wearable and robotic applications. Herein, biphasic liquid metal particle (BMP) with unprecedented stretchability and strain-insensitivity (ΔR/R0 = 1.4@ 1200% strain) is developed to realize a stretchable neuromorphic circuit that mimics a spike-based biologic sensory system. The BMP consists of liquid metal particles (LMPs) and rigid liquid metal particles (RLMPs), which are homogeneously mixed via spontaneous solutal-Marangoni mixing flow during coating. This permits facile single step patterning directly on various substrates at room temperature. BMP is highly conductive (2.3 × 106 S/m) without any post activation steps. BMP interconnects are utilized for a sensory system, which is capable of distinguishing variations of biaxial strains with a spiking neural network, thus demonstrating their potential for various sensing and signal processing applications.

Ultrathin Mixed Ionic-Electronic Conducting Interlayer via the Solution Shearing Technique for High-Performance Lithium-Sulfur Batteries.

The commercialization of lithium-sulfur (Li-S) batteries has been hampered by diverse challenges, including the shuttle phenomenon and low electrical/ionic conductivity of lithium sulfide and sulfur. To address these issues, extensive research has been devoted to developing multifunctional interlayers. However, interlayers capable of simultaneously suppressing the polysulfide (PS) shuttle and ensuring stable electrical and ionic conductivity are relatively uncommon. Moreover, the use of thick and heavy interlayers results in an unavoidable decline in the energy density of Li-S batteries. We developed an ultrathin (750 nm), lightweight (0.182 mg cm-2) interlayer that facilitates mixed ionic-electronic conduction using the solution shearing technique. The interlayer, composed of carbon nanotube (CNT)/Nafion/poly-3,4-ethylenedioxythiophene:tetracyanoborate (PEDOT:TCB), effectively suppresses the shuttle phenomenon through the synergistic segregation and adsorption effects on PSs by Nafion and CNT/PEDOT, respectively. Furthermore, the electrical/ionic conductivity of the interlayer can be improved via counterion exchange and homogeneous Li+ ion flux/good wettability from SO3- functional group of Nafion, respectively. Enhanced sulfur utilization and reaction kinetics through polysulfide shuttle inhibition and facilitated electron/ion transfer by interlayer enable a high discharge capacity of 1029 mA h g-1 in the Li-S pouch cell under a high sulfur loading of 5.3 mg cm-2 and low electrolyte/sulfur ratio of 5 µL mg-1.

Identifying Key Residues in Lysine Decarboxylase for Soluble Expression Using Consensus Design Soluble Mutant Screening (ConsenSing).

Although recent advances in deep learning approaches for protein engineering have enabled quick prediction of hot spot residues improving protein solubility, the predictions do not always correspond to an actual increase in solubility under experimental conditions. Therefore, developing methods that rapidly confirm the linkage between computational predictions and empirical results is essential to the success of improving protein solubility of target proteins. Here, we present a simple hybrid approach to computationally predict hot spots possibly improving protein solubility by sequence-based analysis and empirically explore valuable mutants using split GFP as a reporter system. Our approach, Consensus design Soluble Mutant Screening (ConsenSing), utilizes consensus sequence prediction to find hot spots for improvement of protein solubility and constructs a mutant library using Darwin assembly to cover all possible mutations in one pot but still keeps the library as compact as possible. This approach allowed us to identify multiple mutants of Escherichia coli lysine decarboxylase, LdcC, with substantial increases in soluble expression. Further investigation led us to pinpoint a single critical residue for the soluble expression of LdcC and unveiled its mechanism for such improvement. Our approach demonstrated that following a protein's natural evolutionary path provides insights to improve protein solubility and/or increase protein expression by a single residue mutation, which can significantly change the profile of protein solubility.

Deep-Learning Algorithm and Concomitant Biomarker Identification for NSCLC Prediction Using Multi-Omics Data Integration.

Early diagnosis of lung cancer to increase the survival rate, which is currently at a low range of mid-30%, remains a critical need. Despite this, multi-omics data have rarely been applied to non-small-cell lung cancer (NSCLC) diagnosis. We developed a multi-omics data-affinitive artificial intelligence algorithm based on the graph convolutional network that integrates mRNA expression, DNA methylation, and DNA sequencing data. This NSCLC prediction model achieved a 93.7% macro F1-score, indicating that values for false positives and negatives were substantially low, which is desirable for accurate classification. Gene ontology enrichment and pathway analysis of features revealed that two major subtypes of NSCLC, lung adenocarcinoma and lung squamous cell carcinoma, have both specific and common GO biological processes. Numerous biomarkers (i.e., microRNA, long non-coding RNA, differentially methylated regions) were newly identified, whereas some biomarkers were consistent with previous findings in NSCLC (e.g., SPRR1B). Thus, using multi-omics data integration, we developed a promising cancer prediction algorithm.

Constructing multi-enzymatic cascade reactions for selective production of 6-bromoindirubin from tryptophan in Escherichia coli.

6-Bromoindirubin (6BrIR), found in Murex sea snails, is a precursor of indirubin-derivatives anticancer drugs. However, its synthesis remains limited due to uncharacterized biosynthetic pathways and difficulties in site-specific bromination and oxidation at the indole ring. Here, we present an efficient 6BrIR production strategy in Escherichia coli by using four enzymes, that is, tryptophan 6-halogenase fused with flavin reductase Fre (Fre-L3-SttH), tryptophanase (TnaA), toluene 4-monooxygenase (PmT4MO), and flavin-containing monooxygenase (MaFMO). Although most indole oxygenases preferentially oxygenate the electronically active C3 position of indole, PmT4MO was newly characterized to perform C2 oxygenation of 6-bromoindole with 45% yield to produce 6-bromo-2-oxindole. In addition, 6BrIR was selectively generated without indigo and indirubin byproducts by controlling the reducing power of cysteine and oxygen supply during the MaFMO reaction. These approaches led to 34.1 mg/L 6BrIR productions, making it possible to produce the critical precursor of the anticancer drugs only from natural ingredients such as tryptophan, NaBr, and oxygen.

Synthesis of soluble melanin nanoparticles under acidic conditions using Burkholderia cepacia tyrosinase and their characterization.

Melanin nanoparticles (MNPs) used for biomedical applications are often synthesized via the chemical auto-oxidation of catecholic monomers such as dopamine and 3,4-dihydroxyphenylalanine (DOPA) under alkaline conditions. However, the synthetic method for the chemical synthesis of MNP (cMNP) is relatively straightforward and more robust to control their homogenous particle size and morphology than the corresponding enzymatic synthetic methods. In this study, we demonstrated that the simple enzymatic synthesis of MNPs (eMNPs) with homogenous and soluble (<20 nm diameter) properties is possible using dopamine and Burkholderia cepacia tyrosinase (BcTy) under acidic conditions (i.e., pH 3.0). BcTy was highly reactive under pH 5.0, where the natural and chemical oxidation of catechol is complex, and thus melanin was synthesized via the hydroxylation of phenolic substrates. The detailed chemical analysis and characterization of the physical properties of the eMNPs confirmed the higher preservation of the catechol and primary amine moieties in the monomer substrate such as dopamine under acidic conditions. The eMNPs showed enhanced antioxidant activity and conferred stickiness to the formed hydrogel compared to the chemical auto-oxidation method owing to the large number of hydroxyl groups remaining such as catechol and quinone moieties. Because of these advantages and characteristics, the synthesis of MNPs using BcTy under acidic conditions can open a new path for their biomedical applications.

An Integrative Multiomics Approach to Characterize Prebiotic Inulin Effects on Faecalibacterium prausnitzii.

Faecalibacterium prausnitzii, a major commensal bacterium in the human gut, is well known for its anti-inflammatory effects, which improve host intestinal health. Although several studies have reported that inulin, a well-known prebiotic, increases the abundance of F. prausnitzii in the intestine, the mechanism underlying this effect remains unclear. In this study, we applied liquid chromatography tandem mass spectrometry (LC-MS/MS)-based multiomics approaches to identify biological and enzymatic mechanisms of F. prausnitzii involved in the selective digestion of inulin. First, to determine the preference for dietary carbohydrates, we compared the growth of F. prausnitzii in several carbon sources and observed selective growth in inulin. In addition, an LC-MS/MS-based intracellular proteomic and metabolic profiling was performed to determine the quantitative changes in specific proteins and metabolites of F. prausnitzii when grown on inulin. Interestingly, proteomic analysis revealed that the putative proteins involved in inulin-type fructan utilization by F. prausnitzii, particularly ß-fructosidase and amylosucrase were upregulated in the presence of inulin. To investigate the function of these proteins, we overexpressed bfrA and ams, genes encoding ß-fructosidase and amylosucrase, respectively, in Escherichia coli, and observed their ability to degrade fructan. In addition, the enzyme activity assay demonstrated that intracellular fructan hydrolases degrade the inulin-type fructans taken up by fructan ATP-binding cassette transporters. Furthermore, we showed that the fructose uptake activity of F. prausnitzii was enhanced by the fructose phosphotransferase system transporter when inulin was used as a carbon source. Intracellular metabolomic analysis indicated that F. prausnitzii could use fructose, the product of inulin-type fructan degradation, as an energy source for inulin utilization. Taken together, this study provided molecular insights regarding the metabolism of F. prauznitzii for inulin, which stimulates the growth and activity of the beneficial bacterium in the intestine.

Microfluidic Screening-Assisted Machine Learning to Investigate Vertical Phase Separation of Small Molecule:Polymer Blend.

Solution-based thin-film processing is a widely utilized technique for the fabrication of various devices. In particular, the tunability of the ink composition and coating condition allows precise control of thin-film properties and device performance. Despite the advantage of having such tunability, the sheer number of possible combinations of experimental parameters render it infeasible to efficiently optimize device performance and analyze the correlation between experimental parameters and device performance. In this work, a microfluidic screening-embedded thin-film processing technique is developed, through which thin-films of varying ratios of small molecule semiconductor:polymer blend are simultaneously generated and screened in a time- and resource-efficient manner. Moreover, utilizing the thin-films of varying combinations of experimental parameters, machine learning models are trained to predict the transistor performance. Gaussian Process Regression (GPR) algorithms tuned by Bayesian optimization shows the best predictive accuracy amongst the trained models, which enables narrowing down of the combinations of experimental parameters and investigation of the degree of vertical phase separation under the predicted parameter space. The technique can serve as a guideline for elucidating the underlying complex parameter-property-performance correlations in solution-based thin-film processing, thereby accelerating the optimization of various thin-film devices in the future.

Computational Method-Based Optimization of Carbon Nanotube Thin-Film Immunosensor for Rapid Detection of SARS-CoV-2 Virus.

The recent global spread of COVID-19 stresses the importance of developing diagnostic testing that is rapid and does not require specialized laboratories. In this regard, nanomaterial thin-film-based immunosensors fabricated via solution processing are promising, potentially due to their mass manufacturability, on-site detection, and high sensitivity that enable direct detection of virus without the need for molecular amplification. However, thus far, thin-film-based biosensors have been fabricated without properly analyzing how the thin-film properties are correlated with the biosensor performance, limiting the understanding of property-performance relationships and the optimization process. Herein, the correlations between various thin-film properties and the sensitivity of carbon nanotube thin-film-based immunosensors are systematically analyzed, through which optimal sensitivity is attained. Sensitivities toward SARS-CoV-2 nucleocapsid protein in buffer solution and in the lysed virus are 0.024 [fg/mL]-1 and 0.048 [copies/mL]-1, respectively, which are sufficient for diagnosing patients in the early stages of COVID-19. The technique, therefore, can potentially elucidate complex relationships between properties and performance of biosensors, thereby enabling systematic optimization to further advance the applicability of biosensors for accurate and rapid point-of-care (POC) diagnosis.

Investigation of the Effect of 3D Meniscus Geometry on Fluid Dynamics and Crystallization via In Situ Optical Microscopy-Assisted Mathematical Modeling.

Solution-based thin-film solidification is a complex process involving various transport phenomena that are intricately dependent on multiple experimental parameters. The difficulty of analyzing this process experimentally or conducting exact numerical simulation make it challenging to understand, predict, and control the solidification process. In this work, a simple and effective technique to analyze the thin-film solidification process during solution shearing, based on 3D geometrical model of the meniscus, is proposed. The 3D meniscus geometry, which changes depending on the experimental parameters, is attained using high-speed side-view and top-view in situ microscopy. Thereafter, mass and momentum transport mathematical models are applied to obtain numerical solutions of transport phenomena within the meniscus. Utilizing these results, the underlying mechanism of dendritic growth of small molecule organic semiconductor is elucidated, which has previously been unknown. The 3D meniscus modeling is particularly important for this analysis, as dendrite formation is strongly dependent on the meniscus geometry near the contact line and mass transport variation perpendicular to the coating direction. This technique enables the study of complex relationship between experimental parameters and solidification process, which is widely applicable to various materials and coating systems; whereby, better understanding of thin-film growth and device performance optimization is possible.

Alu cell-free DNA concentration, Alu index, and LINE-1 hypomethylation as a cancer predictor.

INTRODUCTION: The liquid biopsy approach, a less-invasive diagnostic tool, enables the detection of disease-specific genetic and epigenetic aberrations. Approximately 66-69% of the human genome may be composed of transposable repetitive elements, including Alu and LINE-1. This study aimed to investigate whether Alu-derived cell-free DNA (cfDNA) concentrations, Alu index, and LINE-1 methylation could be used to distinguish patients with cancers from healthy individuals. METHODS: Two sets of primers, shorter and longer Alu fragments, were used to amplify Alu elements, followed by the quantitation of Alu DNA concentration and its integrity index. LINE-1 methylation status was then analyzed with quantitative PCR using methylation- and unmethylation-specific TaqMan probes. RESULTS: Both Alu index and LINE-1 methylation level were significantly different in comparison between patients with lung or breast cancer and the healthy controls. The area under the ROC curve of the Alu index and LINE-1 hypomethylation was 0.742 and 0.848 for lung cancer, respectively, and 0.724 and 0.890 for breast cancer, respectively. However, Alu longer fragment DNA concentration was significantly correlated with Alu index in comparison to LINE-1 hypomethylation. Regression analysis suggested that the LINE-1 methylation level, rather than the Alu index, was a good discriminator for lung and breast cancers. CONCLUSIONS: This study investigated the genome-wide Alu index and LINE-1 methylation status; their associations with cancers suggested that these combinatory panels could be implemented as a triage test to discriminate cancer patients from healthy individuals.

Production of Tyrian purple indigoid dye from tryptophan in Escherichia coli.

Tyrian purple, mainly composed of 6,6'-dibromoindigo (6BrIG), is an ancient dye extracted from sea snails and was recently demonstrated as a biocompatible semiconductor material. However, its synthesis remains limited due to uncharacterized biosynthetic pathways and the difficulty of regiospecific bromination. Here, we introduce an effective 6BrIG production strategy in Escherichia coli using tryptophan 6-halogenase SttH, tryptophanase TnaA and flavin-containing monooxygenase MaFMO. Since tryptophan halogenases are expressed in highly insoluble forms in E. coli, a flavin reductase (Fre) that regenerates FADH2 for the halogenase reaction was used as an N-terminal soluble tag of SttH. A consecutive two-cell reaction system was designed to overproduce regiospecifically brominated precursors of 6BrIG by spatiotemporal separation of bromination and bromotryptophan degradation. These approaches led to 315.0 mg l-1 6BrIG production from tryptophan and successful synthesis of regiospecifically dihalogenated indigos. Furthermore, it was demonstrated that 6BrIG overproducing cells can be directly used as a bacterial dye.

Numerical Simulations and In Situ Optical Microscopy Connecting Flow Pattern, Crystallization, and Thin-Film Properties for Organic Transistors with Superior Device-to-Device Uniformity.

Currently, due to the lack of precise control of flow behavior and the understanding of how it influences thin-film crystallization, strict tuning of thin-film properties during solution-based coating is difficult. In this work, a continuous-flow microfluidic-channel-based meniscus-guided coating (CoMiC) is introduced, which is a system that enables manipulation of flow patterns and analysis connecting flow pattern, crystallization, and thin-film properties. Continuous supply of a solution of an organic semiconductor with various flow patterns is generated using microfluidic channels. 3D numerical simulations and in situ microscopy allow the tracking of the flow pattern along its entire path (from within the microfluidic channel to near the liquid-solid boundary), and enable direct observation of thin-film crystallization process. In particular, the generation of chaotic flow results in unprecedented device-to-device uniformity, with coefficient of variation (CV) of 7.3% and average mobility of 2.04 cm2 V-1 s-1 in doped TIPS-pentacene. Furthermore, CV and average mobility of 9.6% and 11.4 cm2 V-1 s-1 are achieved, respectively, in a small molecule:polymer blend system. CoMiC can serve as a guideline for elucidating the relation between flow behavior, liquid-to-solid phase transition, and device performance, which has thus far been unknown.

Effects of fermented rice bran on DEN-induced oxidative stress in mice: GSTP1, LINE-1 methylation, and telomere length ratio.

N-diethylnitrosamine (DEN), a well-known carcinogen, not only induces excessive reactive oxygen species but also suppresses DNA methylation. This study investigated the effect of fermented rice bran (FRB) treatment on DEN-induced oxidative stress through DNA methylation and telomere length analysis. To evaluate the potential protective role of FRB in oxidative stress, two different doses of FRB, DEN, and their combination were administered to mice that were preadapted or not to FRB. Glutathione-S-transferase P1 (GSTP1) methylation levels significantly decreased at 2 and 24 hr after FRB and DEN co-administration in mice with and without pre-adaptation. Moreover, GSTP1 mRNA was upregulated under DEN-induced oxidative stress. Furthermore, changes in long interspersed nuclear element-1 methylation were observed from the viewpoint of genomic instability. In addition, FRB preadapted mice displayed a lower telomere length ratio than the non-adapted mice, suggesting that FRB adaptation offers advantages over the non-adapted conditions in terms of inflammation suppression. PRACTICAL APPLICATIONS: DEN induces excessive ROS, which is associated with oxidative stress on DNA and other cellular components, resulting in inflammation. This study shows that FRB may alleviate DEN-triggered oxidative stress, based on changes in GSTP1, LINE-1 methylation, and telomere length ratios, thereby, revealing the potential of dietary intervention during inflammation. Furthermore, this study furthers the current understanding of DNA methylation mechanisms underlying the antioxidant and anti-inflammatory effects of functional food components. These results indicate that dietary inclusion of FRB may help decrease oxidative DNA damage and its associated inflammation at early stages of a disease.

Characterization of a Tryptophan 6-Halogenase from Streptomyces albus and Its Regioselectivity Determinants.

Tryptophan halogenases are found in diverse organisms and catalyze regiospecific halogenation. They play an important role in the biosynthesis of halogenated indole alkaloids, which are biologically active and of therapeutic importance. Here, a tryptophan 6-halogenase (SatH) from Streptomyces albus was characterized by using a whole-cell reaction system in Escherichia coli. SatH showed substrate specificity for chloride and bromide ions, leading to regiospecific halogenation at the C6-position of l-tryptophan. In addition, SatH exhibited higher performance in bromination than that of previously reported tryptophan halogenases in the whole-cell reaction system. Through structure-based protein mutagenesis, it has been revealed that two consecutive residues, A78/V79 in SatH and G77/I78 in PyrH, are key determinants in the regioselectivity difference between tryptophan 6- and 5-halogenases. Substituting the AV with GI residues switched the regioselectivity of SatH by moving the orientation of tryptophan. These data contribute to an understanding of the key residues that determine the regioselectivity of tryptophan halogenases.

Versatile Processing of Metal-Organic Framework-Fluoropolymer Composite Inks with Chemical Resistance and Sensor Applications.

We report a new class of metal-organic framework (MOF) inks with a water-repellent, photocurable fluoropolymer (PFPE) having up to 90 wt % MOF loading. These MOF inks are enabled to process various MOFs through spray coating, pen writing, stencil printing, and molding at room temperature. Upon UV curing, the hydrophobic PFPE matrix efficiently blocks water permeation but allows accessibility of chemicals into the MOF pores, thereby freeing the MOF to perform its unique function. Moreover, by introducing functional MOFs we successfully demonstrated a water-tolerant chemosensor for a class of aromatic pollutants in water and a chemical-resistant thermosensor for visualizing temperature image. This approach would open up innumerable opportunities for those MOFs that are otherwise dormant.

NLRP1 and NTN1, Deregulated Blood Differentially Methylated Regions in Mild Cognitive Impairment Patients.

Epigenetic dysregulation has been known to be involved in neurodegenerative diseases, including amnestic mild cognitive impairment (MCI). The aim of this study was to investigate the genome-wide DNA methylation analysis, in order to identify epigenetic dysregulation in blood from patients with MCI. Here, we investigated whether epigenetic dysregulation in MCI and whether such an aberration could be detected in blood circulation. Genome-wide bisulfite sequencing targeted 84 million bases covering 3.7 million CpG sites was comparatively analyzed in MCI and control groups. And correlation between DNA methylation and transcriptomic changes was sought. Significant differentially methylated regions (DMRs) distinguishing the MCI and control groups were identified and functionally annotated. Most DMRs specific to MCI were enriched between - 2 kb and + 2 kb of the CpG island start sites located within or near gene promoters. Representative hypo- and hypermethylated DMRs in MCI were confirmed to be correlated to mRNA expression changes with the comparative delta Ct method. DNA methylation aberrations involving metal ion homeostasis, axon growth, inflammasome, and others in this study may be less-invasive, easily measurable blood biomarker candidates for MCI.

Inorganic Polymer Micropillar-Based Solution Shearing of Large-Area Organic Semiconductor Thin Films with Pillar-Size-Dependent Crystal Size.

It is demonstrated that the crystal size of small-molecule organic semiconductors can be controlled during solution shearing by tuning the shape and dimensions of the micropillars on the blade. Increasing the size and spacing of the rectangular pillars increases the crystal size, resulting in higher thin-film mobility. This phenomenon is attributed as the microstructure changing the degree and density of the meniscus line curvature, thereby controlling the nucleation process. The use of allylhybridpolycarbosilane (AHPCS), an inorganic polymer, is also demonstrated as the microstructured blade for solution shearing, which has high resistance to organic solvents, can easily be microstructured via molding, and is flexible and durable. Finally, it is shown that solution shearing can be performed on a curved surface using a curved blade. These demonstrations bring solution shearing closer to industrial applications and expand its applicability to various printed flexible electronics.

Effect of ionizing radiation at low dose on transgenerational carcinogenesis by epigenetic regulation.

The objective of this study was to determine the effect of ionizing radiation (IR) exposure of parents on carcinogenesis of the next generation focusing on the epigenetic perspective to clarify the relationship between radiation dose and carcinogenesis in F1 generation SD rats. F1 generations from pregnant rats (F0) who were exposed to gamma rays were divided into three groups according to the dose of radiation: 10 rad, 30 rad, and untreated. They were intraperitoneally injected with 50 mg/kg of diethylnitrosamine (DEN). Carcinogenesis was analyzed by examining expression levels of tumor suppressor genes (TSG) and other related genes by methylation-specific polymerase chain reaction (MSP). DNA methylation in liver tissues was evaluated to discern epigenetic regulation of transgenerational carcinogenesis vulnerability following IR exposure. Numerous studies have proved that transcriptional inactivation due to hypermethylation of TSG preceded carcinogenesis. Results of this study revealed hypermethylation of tumor suppressor gene SOCS1 in group treated with 30 rad. In addition, genes related to DNA damage response pathway (GSTP1, ATM, DGKA, PARP1, and SIRT6) were epigenetically inactivated in all DEN treated groups. In the case of proto-oncogene c-Myc, DNA hypermethylation was identified in the group with low dose of IR (10 rad). Results of this study indicated that each TSG had different radiation threshold level (dose-independent way) and DEN treatment could affect DNA methylation profile irrelevant of ionizing radiation dose.