Metallization of Targeted Protein Assemblies in Cell-Derived Extracellular Matrix by Antibody-Guided Biotemplating.

Biological systems are composed of hierarchical structures made of a large number of proteins. These structures are highly sophisticated and challenging to replicate using artificial synthesis methods. To exploit these structures in materials science, biotemplating is used to achieve biocomposites that accurately mimic biological structures and impart functionality of inorganic materials, including electrical conductivity. However, the biological scaffolds used in previous studies are limited to stereotypical and simple morphologies with little synthetic diversity because of a lack of control over their morphologies. This study proposes that the specific protein assemblies within the cell-derived extracellular matrix (ECM), whose morphological features are widely tailorable, can be employed as versatile biotemplates. In a typical procedure, a fibrillar assembly of fibronectin-a constituent protein of the ECM-is metalized through an antibody-guided biotemplating approach. Specifically, the antibody-bearing nanogold is attached to the fibronectin through antibody-antigen interactions, and then metals are grown on the nanogold acting as a seed. The biomimetic structure can be adapted for hydrogen production and sensing after improving its electrical conductivity through thermal sintering or additional metal growth. This study demonstrates that cell-derived ECM can be an attractive option for addressing the diversity limitation of a conventional biotemplate.

In situ silver nanoparticle development for molecular-specific biological imaging via highly accessible microscopies.

In biological studies and diagnoses, brightfield (BF), fluorescence, and electron microscopy (EM) are used to image biomolecules inside cells. When compared, their relative advantages and disadvantages are obvious. BF microscopy is the most accessible of the three, but its resolution is limited to a few microns. EM provides a nanoscale resolution, but sample preparation is time-consuming. In this study, we present a new imaging technique, which we termed decoration microscopy (DecoM), and quantitative investigations to address the aforementioned issues in EM and BF microscopy. For molecular-specific EM imaging, DecoM labels proteins inside cells using antibodies bearing 1.4 nm gold nanoparticles (AuNPs) and grows silver layers on the AuNPs' surfaces. The cells are then dried without buffer exchange and imaged using scanning electron microscopy (SEM). Structures labeled with silver-grown AuNPs are clearly visible on SEM, even they are covered with lipid membranes. Using stochastic optical reconstruction microscopy, we show that the drying process causes negligible distortion of structures and that less structural deformation could be achieved through simple buffer exchange to hexamethyldisilazane. Using DecoM, we visualize the nanoscale alterations in microtubules by microtubule-severing proteins that cannot be observed with diffraction-limited fluorescence microscopy. We then combine DecoM with expansion microscopy to enable sub-micron resolution BF microscopy imaging. We first show that silver-grown AuNPs strongly absorb white light, and the structures labeled with them are clearly visible on BF microscopy. We then show that the application of AuNPs and silver development must follow expansion to visualize the labeled proteins clearly with sub-micron resolution.

Regenerative and anti-inflammatory effect of a novel bentonite complex on burn wounds.

BACKGROUND: Bentonite, a montmorillonite clay, has been used as a classical remedy strategy for a long time. Recently, bentonite has been used as a raw material in cosmetic products because of its antibacterial and antioxidant properties. However, the therapeutic effect of bentonite on burn injuries has not yet been identified. OBJECTIVES: The aim of this study was to explore the therapeutic effect of a novel bentonite complex, which was developed for medical use, on burn wounds and the anti-inflammatory function of bentonite clay in the complex in vitro. METHODS: A novel bentonite complex and bentonite clay were prepared for each in vivo and in vitro assay (C&L Biotech, Seoul, Korea). Burn wounds were induced on the dorsal skin of two Yucatan minipigs, and effects of tissue regeneration and anti-inflammation were assessed by histological and gene expression analysis. RESULTS: The bentonite complex improved skin regeneration in burn wounds by inducing collagen synthesis, cell proliferation and angiogenesis in vivo. Furthermore, expression of inflammatory cytokines was significantly inhibited by treatment of the bentonite complex. In vitro study for identification of anti-inflammatory effect showed that bentonite clay significantly regulated COX-2 signalling in both HacaT keratinocytes and 3D4/2 macrophage cell lines. CONCLUSIONS: This study identified the therapeutic effect of a novel bentonite complex in burn wounds by inducing skin regeneration and bentonite-mediated anti-inflammatory responses.

Multiscale Functional Metal Architectures by Antibody-Guided Metallization of Specific Protein Assemblies in Ex Vivo Multicellular Organisms.

Biological systems consist of hierarchical protein structures, each of which has unique 3D geometries optimized for specific functions. In the past decades, the growth of inorganic materials on specific proteins has attracted considerable attention. However, the use of specific proteins as templates has only been demonstrated in relatively simple organisms, such as viruses, limiting the range of structures that can be used as scaffolds. This study proposes a method for synthesizing metallic structures that resemble the 3D assemblies of specific proteins in mammalian cells and animal tissues. Using 1.4 nm nanogold-conjugated antibodies, specific proteins within cells and ex vivo tissues are labeled, and then the nanogold acts as nucleation sites for growth of metal particles. As proof of concept, various metal particles are grown using microtubules in cells as templates. The metal-containing cells are applied as catalysts and show catalytic stability in liquid-phase reactions due to the rigid support provided by the microtubules. Finally, this method is used to produce metal structures that replicate the specific protein assemblies of neurons in the mouse brain or the extracellular matrices in the mouse kidney and heart. This new biotemplating approach can facilitate the conversion of specific protein structures into metallic forms in ex vivo multicellular organisms.

Asymmetric Dichroic Colors in Stretchable Film with Embedded Au/Ag Alloy Nanoparticles.

We present a new type of stretchable dichroic film in which Au and Ag alloy nanoparticles (NPs) are dispersed in polydimethylsiloxane (PDMS). The alloy NPs are synthesized with different atomic compositions and sizes to modulate their plasmonic resonance frequencies and absorption and scattering cross sections. The PDMS dichroic film in which 100 nm alloy NPs with a Au/Ag ratio of 7:3 are dispersed shows exotic optical properties under tensile strain. When 40% tensile strain is applied, the film exhibits a strain-sensitive transmission and strain-insensitive reflection behavior in which the transmittance is increased up to 2.6 times, whereas the reflectance remains unchanged. Moreover, we demonstrate a stretchable anticounterfeiting film and a flexible dichroic sculpture fabricated with the PDMS composite. This work demonstrates a new type of plasmonic application that has great potential in various applications, such as special-purpose optical films, security patterns, and smart windows.

Phthalazinone Pyrazole Enhances the Hepatic Functions of Human Embryonic Stem Cell-Derived Hepatocyte-Like Cells via Suppression of the Epithelial-Mesenchymal Transition.

During liver development, nonpolarized hepatic progenitor cells differentiate into mature hepatocytes with distinct polarity. This polarity is essential for maintaining the intrinsic properties of hepatocytes. The balance between the epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) plays a decisive role in differentiation of polarized hepatocytes. In this study, we found that phthalazinone pyrazole (PP), a selective inhibitor of Aurora-A kinase (Aurora-A), suppressed the EMT during the differentiation of hepatocyte-like cells (HLCs) from human embryonic stem cells. The differentiated HLCs treated with PP at the hepatoblast stage showed enhanced hepatic morphology and functions, particularly with regard to the expression of drug metabolizing enzymes. Moreover, we found that these effects were mediated though suppression of the AKT pathway, which is involved in induction of the EMT, and upregulation of hepatocyte nuclear factor 4α expression rather than Aurora-A inhibition. In conclusion, these findings provided insights into the regulatory role of the EMT on in vitro hepatic maturation, suggesting that inhibition of the EMT may drive transformation of hepatoblast cells into mature and polarized HLCs.

A miniaturized wall-climbing segment robot inspired by caterpillar locomotion.

Caterpillars are very successful soft-bodied climbers that navigate in complex environments. This paper develops a multi-segmented robot climbing on vertical surfaces using dry adhesive pads, inspired by caterpillar locomotion. The miniaturized robot consists of four segments, and each segment uses a solenoid actuator with a permanent magnet plunger. The head and body segments adapt a novel mechanism and Scott-Russell linkages to generate a bi-directional plane motion using one solenoid actuator, resulting to reliable attaching and peeling motions of gecko pads. A tail is also attached at the back of the last segment to avoid falling or exhibiting unstable motion. Gecko-inspired adhesive pads are fabricated from polydimethylsiloxane (PDMS) with the area of 20 mm × 10 mm. We have conducted experiments on the locomotion performance of the segment robot climbing vertical surfaces for two types of locomotion, achieving the fast and stable climbing motion.

Xeno-sensing activity of the aryl hydrocarbon receptor in human pluripotent stem cell-derived hepatocyte-like cells.

Although hepatocyte-like cells derived from human pluripotent stem cells (hPSC-HLCs) are considered a promising model for predicting hepatotoxicity, their application has been restricted because of the low activity of drug metabolizing enzymes (DMEs). Here we found that the low expression of xenobiotic receptors (constitutive androstane receptor, CAR; and pregnane X receptor, PXR) contributes to the low activity of DMEs in hPSC-HLCs. Most CAR- and PXR-regulated DMEs and transporters were transcriptionally down-regulated in hPSC-HLC. Transcriptional expression of CAR and PXR was highly repressed in hPSC-HLCs, whereas mRNA levels of aryl hydrocarbon receptor (AHR) were comparable to those of adult liver. Furthermore, ligand-induced transcriptional activation was observed only at AHR in hPSC-HLCs. Bisulfite sequencing analysis demonstrated that promoter hypermethylation of CAR and PXR was associated with diminished transcriptional activity in hPSC-HLCs. Treatment with AHR-selective ligands increased the transcription of AHR-dependent target genes by direct AHR-DNA binding at the xenobiotic response element. In addition, an antagonist of AHR significantly inhibited AHR-dependent target gene expression. Thus, AHR may function intrinsically as a xenosensor as well as a ligand-dependent transcription factor in hPSC-HLCs. Our results indicate that hPSC-HLCs can be used to screen toxic substances related to AHR signaling and to identify potential AHR-targeted therapeutics.

Integrative analysis of genes and miRNA alterations in human embryonic stem cells-derived neural cells after exposure to silver nanoparticles.

Given the rapid growth of engineered and customer products made of silver nanoparticles (Ag NPs), understanding their biological and toxicological effects on humans is critically important. The molecular developmental neurotoxic effects associated with exposure to Ag NPs were analyzed at the physiological and molecular levels, using an alternative cell model: human embryonic stem cell (hESC)-derived neural stem/progenitor cells (NPCs). In this study, the cytotoxic effects of Ag NPs (10-200µg/ml) were examined in these hESC-derived NPCs, which have a capacity for neurogenesis in vitro, at 6 and 24h. The results showed that Ag NPs evoked significant toxicity in hESC-derived NPCs at 24h in a dose-dependent manner. In addition, Ag NPs induced cell cycle arrest and apoptosis following a significant increase in oxidative stress in these cells. To further clarify the molecular mechanisms of the toxicological effects of Ag NPs at the transcriptional and post-transcriptional levels, the global expression profiles of genes and miRNAs were analyzed in hESC-derived NPCs after Ag NP exposure. The results showed that Ag NPs induced oxidative stress and dysfunctional neurogenesis at the molecular level in hESC-derived NPCs. Based on this hESC-derived neural cell model, these findings have increased our understanding of the molecular events underlying developmental neurotoxicity induced by Ag NPs in humans.

Differences in the Epigenetic Regulation of Cytochrome P450 Genes between Human Embryonic Stem Cell-Derived Hepatocytes and Primary Hepatocytes.

Human pluripotent stem cell-derived hepatocytes have the potential to provide in vitro model systems for drug discovery and hepatotoxicity testing. However, these cells are currently unsuitable for drug toxicity and efficacy testing because of their limited expression of genes encoding drug-metabolizing enzymes, especially cytochrome P450 (CYP) enzymes. Transcript levels of major CYP genes were much lower in human embryonic stem cell-derived hepatocytes (hESC-Hep) than in human primary hepatocytes (hPH). To verify the mechanism underlying this reduced expression of CYP genes, including CYP1A1, CYP1A2, CYP1B1, CYP2D6, and CYP2E1, we investigated their epigenetic regulation in terms of DNA methylation and histone modifications in hESC-Hep and hPH. CpG islands of CYP genes were hypermethylated in hESC-Hep, whereas they had an open chromatin structure, as represented by hypomethylation of CpG sites and permissive histone modifications, in hPH. Inhibition of DNA methyltransferases (DNMTs) during hepatic maturation induced demethylation of the CpG sites of CYP1A1 and CYP1A2, leading to the up-regulation of their transcription. Combinatorial inhibition of DNMTs and histone deacetylases (HDACs) increased the transcript levels of CYP1A1, CYP1A2, CYP1B1, and CYP2D6. Our findings suggest that limited expression of CYP genes in hESC-Hep is modulated by epigenetic regulatory factors such as DNMTs and HDACs.

Effect of Inductively Coupled Plasma on the Structural and Electrical Properties of Ti-Doped ITO Films Formed by IPVD.

In this study, we investigated Ti-doped ITO films formed through ionized physical vapor deposition (IPVD) using inductively coupled plasma (ICP). Ti-doped ITO thin films showed an enhanced mobility with ICP power; owing to the improved crystallinity, and the sheet resistance of the Ti-doped ITO (30 nm) largely decreased from 295.1 to 134.5 ohm/sq, even during at room temperature. Therefore, IPVD technology offers a useful tool for transparent electrodes with a large area window-unified touch-screen panel.

Anti-inflammatory activity of SMP30 modulates NF-κB through protein tyrosine kinase/phosphatase balance.

UNLABELLED: Recent studies on senescence marker protein-30 (SMP30) have shown that it has an important functional role in the aging process, but its precise participation in cellular works has not been fully determined. We hypothesize that SMP30 plays crucial roles in signaling processes by modulating the balance of protein tyrosine kinase (PTK)/protein tyrosine phosphatase (PTP) and in activating proinflammatory NF-κB. An experimental paradigm of gain and loss of SMP30 function was established using SMP30-overexpressed YPEN-1 cells (herein referred to as "SMP30(+) cells") and SMP30 (Y/-) knockout mouse kidneys. The resulting data show that SMP30 expression suppressed oxidative stress-induced PTK/PTP dysregulation and PP1/2A inactivation in SMP30(+) cells, leading to the suppression of NF-κB activation. In the kidneys of SMP30 (Y/-) mice, SMP30 deficiency was found to induce NF-κB activation via the upstream signaling of NIK/IKK and MAPKs and to upregulate downstream NF-κB-responsive gene expression. In this study, we also demonstrate for the first time that SMP30 deficiency induced PTK activity in SMP30 (Y/-) kidneys, thereby significantly increasing the tyrosine phosphorylation of a catalytic subunit of PP2A (PP2Ac-Tyr307). Based on these findings, we propose that SMP30 involves NF-κB regulation through the PTK/PTP balance and that the age-related decrease of SMP30 causes NF-κB activation, which contributes to an exacerbation of the inflammatory process during aging. KEY MESSAGES: SMP30-deficient mice induced a shorter lifespan and redox changes. Overexpression of SMP30 prevented oxidative stress insults. The depletion of SMP30 increased redox-related PTK/PTP imbalance and PP1/PP2A inactivation. The depletion of SMP30 caused an elevation of NF-κB-responsive inflammatory markers. SMP30 may be a potent inhibitory protein against oxidative stress and chronic inflammation.

Inhalation of carbon black nanoparticles aggravates pulmonary inflammation in mice.

An increasing number of recent studies have focused on the impact of particulate matter on human health. As a model for atmospheric particulate inhalation, we investigated the effects of inhaled carbon black nanoparticles (CBNP) on mice with bleomycin-induced pulmonary fibrosis. The CNBPs were generated by a novel aerosolization process, and the mice were exposed to the aerosol for 4 hours. We found that CBNP inhalation exacerbated lung inflammation, as evidenced by histopathology analysis and by the expression levels of interleukin-6 protein, fibronectin, and interferon-γ mRNAs in lung tissues. Notably, fibronectin mRNA expression showed a statistically significant increase in expression after CBNP exposure. These data suggest that the concentration of CBNPs delivered (calculated to be 12.5 µg/m(3)) can aggravate lung inflammation in mice. Our results also suggest that the inhalation of ultrafine particles like PM 2.5 is an impactful environmental risk factor for humans, particularly in susceptible populations with predisposing lung conditions.

Polyhexamethyleneguanidine phosphate induces severe lung inflammation, fibrosis, and thymic atrophy.

Polyhexamethyleneguanidine phosphate (PHMG-P) has been widely used as a disinfectant because of its strong bactericidal activity and low toxicity. However, in 2011, the Korea Centers for Disease Control and Prevention and the Ministry of Health and Welfare reported that a suspicious outbreak of pulmonary disease might have originated from humidifier disinfectants. The purpose of this study was to assess the toxicity of PHMG-P following direct exposure to the lung. PHMG-P (0.3, 0.9, or 1.5 mg/kg) was instilled into the lungs of mice. The levels of proinflammatory markers and fibrotic markers were quantified in lung tissues and flow cytometry was used to evaluate T cell distribution in the thymus. Administration of PHMG-P induced proinflammatory cytokines elevation and infiltration of immune cells into the lungs. Histopathological analysis revealed a dose-dependent exacerbation of both inflammation and pulmonary fibrosis on day 14. PHMG-P also decreased the total cell number and the CD4(+)/CD8(+) cell ratio in the thymus, with the histopathological examination indicating severe reduction of cortex and medulla. The mRNA levels of biomarkers associated with T cell development also decreased markedly. These findings suggest that exposure of lung tissue to PHMG-P leads to pulmonary inflammation and fibrosis as well as thymic atrophy.

Evaluation of subchronic inhalation toxicity of methylcyclopentane in rats.

The aim of this study was to verify subchronic inhalation toxicity of methylcyclopentane (CAS No. 96-37-7) in Sprague-Dawley rats. Four groups of 10 rats of each gender were exposed to methylcyclopentane vapor by whole-body inhalation at concentrations of 0, 290, 1300, or 5870 ppm for 6h per day, 5 days/week over a 13-week period. During the study period, clinical signs, mortality, body weight, food consumption, ophthalmoscopy, urinalysis, hematology, serum biochemistry, gross pathology, organ weights, and histopathology were examined. Exposure-related clinical signs (salivation and rubbing) were observed in both genders of the 5870 ppm group. There was an increase in liver weight for both genders but the kidney weight was only higher in females than controls. However, no toxicologically significant changes were observed in body weight, food consumption, ophthalmoscopy, urinalysis, hematology, serum biochemistry, necropsy findings, or histopathology in any of the treatment groups. Under the present experimental conditions, the target organs were determined to be kidney and liver in rats. The no-observed-adverse-effect concentration was considered to be 1300 ppm/6h/day in rats.

Anti-inflammatory effects of the Zingiber officinale roscoe constituent 12-dehydrogingerdione in lipopolysaccharide-stimulated Raw 264.7 cells.

Ginger has long been used worldwide as a spice, seasoning, and wine and is also used as a traditional medicine. There have been no previous studies of the potential beneficial effects of the ginger constituent 12-dehydrogingerdione (12-DHGD). We investigated the anti-inflammatory effect of 12-DHGD on lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cytotoxicity of 12-DHGD was measured using the MTT assay, and production of prostaglandin E2 (PGE2 ) and the inflammatory cytokines interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α was measured by ELISA. Production of nitric oxide (NO) was measured using Griess reagent and expression of cyclooxygenase-2 (COX-2) and inducible NO (iNOS) enzymes was assessed by reverse transcriptase-polymerase chain reaction. Treatment of Raw 264.7 cells with 12-DHGD significantly inhibited LPS-stimulated production of NO (at 12-DHGD concentrations of 150 and 200 ng/ml), IL-6 (at 50, 100, 150, and 200 ng/ml), and PGE2 (at 200 ng/ml). Consistent with the effects on NO and PGE2 production, 12-DHGD treatment also inhibited the LPS-stimulated increase in iNOS and COX-2 mRNA levels. However, 12-DHGD did not affect production of IL-1ß or TNF-α in response to LPS. 12-DHGD, a constituent of ginger, is a potent inhibitor of proinflammatory mediator production in Raw 264.7 macrophage cells.

Nasal and pulmonary toxicity of titanium dioxide nanoparticles in rats.

In recent decades, titanium dioxide (TiO2) nanoparticles have been used in various applications, including paints, coatings, and food. However, data are lacking on the toxicological aspects associated with their use. The aim of this study was to assess the inhalation toxicity of TiO2 nanoparticles in rats by using inhalation exposure. Male Wistar rats were exposed to TiO2 nanoparticles for 2 weeks (6 hr/day, 5 days/week) at a mean mass concentration of 11.39 ± 0.31 mg/m(3). We performed time-course necropsies at 1, 7, and 15 days after exposure. Lung inflammation and injury were assessed on the basis of the total and individual cell counts in bronchoalveolar lavage fluid (BALF), and by biochemical assays, including an assay for lactate dehydrogenase (LDH). Furthermore, histopathological examination was performed to investigate the lungs and nasal cavity of rats. There were no statistically significant changes in the number of BALF cells, results of biochemical assays of BALF and serum, and results of cytokine analysis. However, we did observe histopathological changes in the nasal cavity tissue. Lesions were observed at post-exposure days 1 and 7, which resolved at post-exposure day 15. We also calculated the actual amounts of TiO2 nanoparticles inhaled by the rats. The results showed that the degree of toxicity induced by TiO2 nanoparticles correlated with the delivered quantities. In particular, exposure to small particles with a size of approximately 20 nm resulted in toxicity, even if the total particle number was relatively low.

Inflammatory response in rat lungs with recurrent exposure to welding fumes: a transcriptomic approach.

As chronic exposure to welding fumes causes pulmonary diseases, such as pneumoconiosis, public concern has increased regarding continued exposure to these hazardous gases in the workplace. In a previous study, the inflammatory response to welding fume exposure was analysed in rat lungs in the case of recurrent exposure and recovery periods. Thus using lung samples, well-annotated by histological observation and biochemical analysis, this study examines the gene expression profiles to identify phenotype-anchored genes corresponding to lung inflammation and the repair phenomenon after recurrent welding fume exposure. Seven genes (Mmp12, Cd5l, LOC50101, LOC69183, Spp1, and Slc26a4) were found to be significantly up-regulated according to the severity of the lung injury. In addition, the transcription and translation of Trem2, which was up-regulated in response to the repair process, were validated using a real-time polymerase chain reaction, Western blotting, and immunohistochemistry. The differentially expressed genes in the exposure and recovery groups were also classified using k-means and hierarchical clustering, plus their toxicological function and canonical pathways were further analysed using Ingenuity Pathways Analysis Software. As a result, this comprehensive and integrative analysis of the transcriptional changes that occur during repeated exposure provides important information on the inflammation and repair processes after welding-fume-induced lung injury.

Silver nanoparticles induce apoptosis and G2/M arrest via PKCζ-dependent signaling in A549 lung cells.

The use of silver nanoparticles is one of the fastest growing product categories in the nanotechnology industry, with a focus on antimicrobial activity. However, thus far, toxicity data for silver nanoparticles are limited. In this study, we investigated the cytotoxic effects of silver nanoparticles (Ag NPs) and the pathway by which they affect A549 lung epithelial cells. The effects of Ag NPs on cell survival, cell cycle progression, and mRNA and protein alterations of selected cell cycle- and apoptosis-related genes were studied using formazan dye and LDH release assays, flow cytometric analysis, semi-quantitative RT-PCR, and Western blot analysis. Ag NPs reduced cell viability, increased LDH release, and modulated cell cycle distribution through the accumulation of cells at G2/M and sub-G1 phases (cell death), with a concurrent decrease in cells at G1. Ag NP treatment increased Bax and Bid mRNA levels and downregulated Bcl-2 and Bcl-w mRNAs in a dose-dependent manner. Furthermore, Ag NPs altered the mRNA levels of protein kinase C (PKC) family members. In particular, ectopic overexpression of PKCζ led to the enhancement of cellular proliferation and reduced sensitivity to Ag NPs in A549 cells. Together, these results suggest that Ag NPs induce strong toxicity and G2/M cell cycle arrest by a mechanism involving PKCζ downregulation in A549 cells.

Differential gene expression profiles of radioresistant non-small-cell lung cancer cell lines established by fractionated irradiation: tumor protein p53-inducible protein 3 confers sensitivity to ionizing radiation.

PURPOSE: Despite the widespread use of radiotherapy as a local and regional modality for the treatment of cancer, some non-small-cell lung cancers commonly develop resistance to radiation. We thus sought to clarify the molecular mechanisms underlying resistance to radiation. METHODS AND MATERIALS: We established the radioresistant cell line H460R from radiosensitive parental H460 cells. To identify the radioresistance-related genes, we performed microarray analysis and selected several candidate genes. RESULTS: Clonogenic and MTT assays showed that H460R was 10-fold more resistant to radiation than H460. Microarray analysis indicated that the expression levels of 1,463 genes were altered more than 1.5-fold in H460R compared with parental H460. To evaluate the putative functional role, we selected one interesting gene tumor protein p53-inducible protein 3 (TP53I3), because that this gene was significantly downregulated in radioresistant H460R cells and that it was predicted to link p53-dependent cell death signaling. Interestingly, messenger ribonucleic acid expression of TP53I3 differed in X-ray-irradiated H460 and H460R cells, and overexpression of TP53I3 significantly affected the cellular radiosensitivity of H460R cells. CONCLUSIONS: These results show that H460R may be useful in searching for candidate genes that are responsible for radioresistance and elucidating the molecular mechanism of radioresistance.