Flagellin synergistically enhances anti-tumor effect of EGFRvIII peptide in a glioblastoma-bearing mouse brain tumor model.

BACKGROUND: Glioblastoma (GBM) is the most aggressive type of brain tumor with heterogeneity and strong invasive ability. Treatment of GBM has not improved significantly despite the progress of immunotherapy and classical therapy. Epidermal growth factor receptor variant III (EGFRvIII), one of GBM-associated mutants, is regarded as an ideal therapeutic target in EGFRvIII-expressed GBM patients because it is a tumor-specific receptor expressed only in tumors. Flagellin B (FlaB) originated from Vibrio vulnificus, is known as a strong adjuvant that enhances innate and adaptive immunity in various vaccine models. This study investigated whether FlaB synergistically could enhance the anti-tumor effect of EGFRvIII peptide (PEGFRvIII). METHODS: EGFRvIII-GL261/Fluc cells were used for glioblastoma-bearing mouse brain model. Cell-bearing mice were inoculated with PBS, FlaB alone, PEGFRvIII alone, and PEGFRvIII plus FlaB. Tumor growth based on MRI and the survival rate was investigated. T cell population was examined by flow cytometry analysis. Both cleaved caspase-3 and CD8 + lymphocytes were shown by immunohistochemistry (IHC) staining. RESULTS: The PEGFRvIII plus FlaB group showed delayed tumor growth and increased survival rate when compared to other treatment groups. As evidence of apoptosis, cleaved caspase-3 expression and DNA disruption were more increased in the PEGFRvIII plus FlaB group than in other groups. In addition, the PEGFRvIII plus FlaB group showed more increased CD8 + T cells and decreased Treg cells than other treatment groups in the brain. CONCLUSIONS: FlaB can enhance the anti-tumor effect of PEGFRvIII by increasing CD8 + T cell response in a mouse brain GBM model.

Multi-Walled Carbon Nanotube-Assisted Encapsulation Approach for Stable Perovskite Solar Cells.

Perovskite solar cells (PSCs) are regarded as the next-generation thin-film energy harvester, owing to their high performance. However, there is a lack of studies on their encapsulation technology, which is critical for resolving their shortcomings, such as their degradation by oxygen and moisture. It is determined that the moisture intrusion and the heat trapped within the encapsulating cover glass of PSCs influenced the operating stability of the devices. Therefore, we improved the moisture and oxygen barrier ability and heat releasing capability in the passivation of PSCs by adding multi-walled carbon nanotubes to the epoxy resin used for encapsulation. The 0.5 wt% of carbon nanotube-added resin-based encapsulated PSCs exhibited a more stable operation with a ca. 30% efficiency decrease compared to the ca. 63% decrease in the reference devices over one week under continuous operation. Specifically, the short-circuit current density and the fill factor, which are affected by moisture and oxygen-driven degradation, as well as the open-circuit voltage, which is affected by thermal damage, were higher for the multi-walled carbon nanotube-added encapsulated devices than the control devices, after the stability test.

Crystal structure of 5-enolpyruvylshikimate-3-phosphate synthase from a psychrophilic bacterium, Colwellia psychrerythraea 34H.

To survive at low temperatures, psychrophiles seem to produce cold-adapted enzymes with a high flexibility around active sites for high catalytic efficiency. To gain insights into the cold-adaptation of psychrophilic enzymes in atomic detail, we determined the crystal structure of 5-enolpyruvylshikimate-3-phosphate synthase (CpsEPSPS) from Colwellia psychrerythraea, a psychrophilic bacterium. EPSPS is the primary target for the broad-spectrum herbicide, glyphosate, and a promising target for the development of antimicrobial and antiparasitic agents since it is absent in animals. The crystal structure of unliganded, open CpsEPSPS was determined at 2.2 Å resolution in space group P21 with two protomers per asymmetric unit. Superposition of separate domain I and II of CpsEPSPS structure with those of Escherichia coli EPSPS (EcoEPSPS) structure showed relatively small differences of RMSD values of 0.423 Å and 0.693 Å for domains I and II, respectively, implying the residues in ligand binding and catalysis of cold-adapted CpsEPSPS showed no significant flexibility. This result is conflicting to other cases of cold-adapted proteins. We also observed that hydrogen-bond forming residues in the surface of EcoEPSPS was mutated to non- or lesser hydrogen-bond forming one in CpsEPSPS, which makes the protein surface softer and eventually makes the protein more active at low temperature. In addition, domain rotation angle between open and closed states of CpsEPSPS was smaller than those of any EPSPSs whose structures are known. The restriction of the domain closure, which reduces the entropy cost of ligand binding and catalysis, may be a novel molecular adaptations of cold-adapted enzymes.

TM0416, a Hyperthermophilic Promiscuous Nonphosphorylated Sugar Isomerase, Catalyzes Various C5 and C6 Epimerization Reactions.

There is currently little information on nonphosphorylated sugar epimerases, which are of potential interest for producing rare sugars. We found a gene (the TM0416 gene) encoding a putative d-tagatose-3-epimerase-related protein from the hyperthermophilic bacterium Thermotoga maritima We overexpressed the TM0416 gene in Escherichia coli and purified the resulting recombinant protein for detailed characterization. Amino acid sequence alignment and a structural similarity search revealed that TM0416 is a putative nonphosphorylated sugar epimerase. The recombinant enzyme exhibited maximal C-3 epimerization of l-ribulose to l-xylulose at ∼80°C and pH 7 in the presence of 1 mM Mn2+ In addition, this enzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose, with a conversion yield of 20% after 6 h at 80°C. Remarkably, the enzyme catalyzed the isomerization of d-erythrose or d-threose to d-erythrulose significantly, with conversion yields of 71% and 54.5%, respectively, after 6 h at 80°C at pH 7. To further investigate the substrate specificity of TM0416, we determined its crystal structures in complex with divalent metal ions and l-erythrulose at resolutions of 1.5 and 1.6 Å. Detailed inspection of the structural features and biochemical data clearly demonstrated that this metalloenzyme, with a freely accessible substrate-binding site and neighboring hydrophobic residues, exhibits different and promiscuous substrate preferences, compared with its mesophilic counterparts. Therefore, this study suggests that TM0416 can be functionally classified as a novel type of l-ribulose 3-epimerase (R3E) with d-erythrose isomerase activity.IMPORTANCE Rare sugars, which occur naturally in small amounts, have attracted considerable attention in the food and drug industries. However, there is little information on nonphosphorylated sugar epimerases, which might potentially be applied for the production of rare sugars. This study describes the characterization and functional annotation of a putative nonphosphorylated sugar 3-epimerase from a hyperthermophilic bacterium. Furthermore, we determined its crystal structures in complex with divalent metal ions and l-erythrulose, highlighting its metal-dependent, bifunctional, sugar-isomerizing activity. This hyperthermophilic R3E exhibited d-erythrose/d-threose isomerase activity, with structural features near the substrate-binding site distinct from those of its mesophilic counterparts. Moreover, this metalloenzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose at 70°C. Therefore, TM0416 can be functionally classified as a novel type of promiscuous R3E with a potential for the production of rare sugars for the food and pharmaceutical industries.

Structure of the thermophilic l-Arabinose isomerase from Geobacillus kaustophilus reveals metal-mediated intersubunit interactions for activity and thermostability.

Thermophilic l-arabinose isomerase (AI), which catalyzes the interconversion of l-arabinose and l-ribulose, can be used to produce d-tagatose, a sugar substitute, from d-galactose. Unlike mesophilic AIs, thermophilic AIs are highly dependent on divalent metal ions for their catalytic activity and thermostability at elevated temperatures. However, the molecular basis underlying the substrate preferences and metal requirements of multimeric AIs remains unclear. Here we report the first crystal structure of the apo and holo forms of thermophilic Geobacillus kaustophilus AI (GKAI) in hexamer form. The structures, including those of GKAI in complex with l-arabitol, and biochemical analyses revealed not only how the substrate-binding site of GKAI is formed through displacement of residues at the intersubunit interface when it is bound to Mn(2+), but also revealed the water-mediated H-bonding networks that contribute to the structural integrity of GKAI during catalysis. These observations suggest metal-mediated isomerization reactions brought about by intersubunit interactions at elevated temperatures are responsible for the distinct active site features that promote the substrate specificity and thermostability of thermophilic AIs.

Biochemical and structural characterization of a keratin-degrading M32 carboxypeptidase from Fervidobacterium islandicum AW-1.

Comparative genomics of the keratin-degrading extremophilic eubacterium Fervidobacterium islandicum AW-1 and the closely related Fervidobacterium nodosum with no keratinolytic activity suggested that the FIAW1_1600 gene encoding a carboxypeptidase (CP) plays an important role in keratin degradation. The presumptive 489 amino acid sequence of the gene showed a conserved HEXXH motif with low levels of sequence identity (<38%) to reported thermostable M32 CPs. To identify its functional role, the FIAW1_1600 gene was overexpressed in Escherichia coli, and the recombinant enzyme was purified and characterized in detail. F. islandicum AW-1 CP (FisCP) formed a homodimer with a molecular mass of 107 kDa, and its apoenzyme exhibited maximal activity at 80 °C and pH 7.0 in the presence of Co(2+). This metalloenzyme mainly cleaved the C-termini of peptides with a basic amino acid sequence. The crystal structure of FisCP at 2.2 Å resolution showed high levels of structural similarities (root-mean-square deviations of <1.7 Å) to those of other M32 CP homologs. Remarkably, the enzyme significantly enhanced the degradation of native chicken feathers. This study suggests that FisCP, a keratinolytic member of the thermostable M32 CP family, plays an important role in keratin degradation for cellular metabolism in F. islandicum AW-1.

Crystal structure of CagL from Helicobacter pylori K74 strain.

Helicobacter pylori (Hp) CagL is a component of the type IV secretion system (T4SS) and interacts with integrin in host cells through its flexible RGD domain to translocate CagA. Differences in CagL amino acid polymorphisms between Western and East-Asian Hps are correlated with clinical outcome. CagL of East-Asian clinical Hp isolate K74 (CagL(K74)) contains multiple residue variations upstream of RGD motif and has different integrin binding affinities compared to those of CagL from Western Hp 26695. Here, we report the crystal structure of CagL(K74). The structure displayed a six-helix bundle including two short α-helices, and the RGD motif was found in the long rigid α2 helix flanked by the conserved protease-sensitive and RGD-helper sequences, as observed in CagL(26695). However, two additional salt bridges were found between the helices compared with the CagL(26695) structure, suggesting that the putative flexible region harboring the RGD motif may be more stable in this CagL variant.

The structural basis of substrate promiscuity in UDP-hexose 4-epimerase from the hyperthermophilic Eubacterium Thermotoga maritima.

UDP-galactose 4-epimerase (GalE) catalyzes the interconversion of UDP-glucose (UDP-Glc) and UDP-galactose (UDP-Gal), which is a pivotal step in the Leloir pathway for d-galactose metabolism. Although GalE is widely distributed in prokaryotes and eukaryotes, little information is available regarding hyperthermophilic GalE. We overexpressed the TM0509 gene, encoding a putative GalE from Thermotoga maritima (TMGalE), in Escherichia coli and characterized the encoded protein. To further investigate the molecular basis of this enzyme's catalytic function, we determined the crystal structures of TMGalE and TMGalE bound to UDP-Glc at resolutions of 1.9 Å and 2.0 Å, respectively. The enzyme was determined to be a homodimer with a molecular mass of 70 kDa. The enzyme could reversibly catalyze the epimerization of UDP-GalNAc/UDP-GlcNAc as well as UDP-Gal/UDP-Glc at elevated temperatures, with an apparent optimal temperature and pH of 80 °C and 7.0, respectively. Our data showed that TM0509 is a UDP-galactosugar 4-epimerase involved in d-galactose metabolism; consequently, this study provides the first detailed characterization of a hyperthermophilic GalE. Moreover, the promiscuous substrate specificity of TMGalE, which is more similar to human GalE than E. coli GalE, supports the notion that TMGalE might exhibit the earliest form of sugar-epimerizing enzymes in the evolution of galactose metabolism.

Aerosol-Synthesized Surfactant-Free Single-Walled Carbon Nanotube-Based NO2 Sensors: Unprecedentedly High Sensitivity and Fast Recovery.

This study pioneers a chemical sensor based on surfactant-free aerosol-synthesized single-walled carbon nanotube (SWCNT) films for detecting nitrogen dioxide (NO2). Unlike conventional CNTs, the SWCNTs used in this study exhibit one of the highest surface-to-volume ratios. They show minimal bundling without the need for surfactants and have the lowest number of defects among reported CNTs. Furthermore, the dry-transferrable and facile one-step lamination results in promising industrial viability. When applied to devices, the sensor shows excellent sensitivity (41.6% at 500 ppb), rapid response/recovery time (14.2/120.8 s), a remarkably low limit of detection (below ≈0.161 ppb), minimal noise, repeatability for more than 50 cycles without fluctuation, and long-term stability for longer than 6 months. This is the best performance reported for a pure CNT-based sensor. In addition, the aerosol SWCNTs demonstrate consistent gas-sensing performance even after 5000 bending cycles, indicating their suitability for wearable applications. Based on experimental and theoretical analyses, the proposed aerosol CNTs are expected to overcome the limitations associated with conventional CNT-based sensors, thereby offering a promising avenue for various sensor applications.

Redox Regulation of PTEN by Reactive Oxygen Species: Its Role in Physiological Processes.

Phosphatase and tensin homolog (PTEN) is a tumor suppressor due to its ability to regulate cell survival, growth, and proliferation by downregulating the PI3K/AKT signaling pathway. In addition, PTEN plays an essential role in other physiological events associated with cell growth demands, such as ischemia-reperfusion, nerve injury, and immune responsiveness. Therefore, recently, PTEN inhibition has emerged as a potential therapeutic intervention in these situations. Increasing evidence demonstrates that reactive oxygen species (ROS), especially hydrogen peroxide (H2O2), are produced and required for the signaling in many important cellular processes under such physiological conditions. ROS have been shown to oxidize PTEN at the cysteine residue of its active site, consequently inhibiting its function. Herein, we provide an overview of studies that highlight the role of the oxidative inhibition of PTEN in physiological processes.

Redox Regulation of Phosphatase and Tensin Homolog by Bicarbonate and Hydrogen Peroxide: Implication of Peroxymonocarbonate in Cell Signaling.

Phosphatase and tensin homolog (PTEN) is a negative regulator of the phosphoinositide 3-kinases/protein kinase B (PI3K/AKT) signaling pathway. Notably, its active site contains a cysteine residue that is susceptible to oxidation by hydrogen peroxide (H2O2). This oxidation inhibits the phosphatase function of PTEN, critically contributing to the activation of the PI3K/AKT pathway. Upon the stimulation of cell surface receptors, the activity of NADPH oxidase (NOX) generates a transient amount of H2O2, serving as a mediator in this pathway by oxidizing PTEN. The mechanism underlying this oxidation, occurring despite the presence of highly efficient and abundant cellular oxidant-protecting and reducing systems, continues to pose a perplexing conundrum. Here, we demonstrate that the presence of bicarbonate (HCO3-) promoted the rate of H2O2-mediated PTEN oxidation, probably through the formation of peroxymonocarbonate (HCO4-), and consequently potentiated the phosphorylation of AKT. Acetazolamide (ATZ), a carbonic anhydrase (CA) inhibitor, was shown to diminish the oxidation of PTEN. Thus, CA can also be considered as a modulator in this context. In essence, our findings consolidate the crucial role of HCO3- in the redox regulation of PTEN by H2O2, leading to the presumption that HCO4- is a signaling molecule during cellular physiological processes.

Crystallization and preliminary X-ray crystallographic analysis of MxaJ, a component of the methanol-oxidizing system operon from the marine bacterium Methylophaga aminisulfidivorans MPT.

The methanol-oxidizing system (mox) is essential for methylotrophic bacteria to extract energy during the oxidoreduction reaction and consists of a series of electron-transfer proteins encoded by the mox operon. One of the key enzymes is the α2ß2 methanol dehydrogenase complex (type I MDH), which converts methanol to formaldehyde during the 2e⁻ transfer through the prosthetic group pyrroloquinoline quinone. MxaJ, a product of mxaJ of the mox operon, is a component of the MDH complex and enhances the methanol-converting activity of the MDH complex. However, the exact functional mechanism of MxaJ in the complex is not clearly known. To investigate the functional role of MxaJ in MDH activity, an attempt was made to determine its crystal structure. Diffraction data were collected from a selenomethionine-substituted crystal to 1.92 Å resolution at the peak wavelength. The crystal belonged to the orthorhombic space group P212121, with unit-cell parameters a = 37.127, b = 63.761, c = 99.246 Å. The asymmetric unit contained one MxaJ molecule with a calculated Matthews coefficient of 2.11 Ų Da⁻¹ and a solvent content of 41.7%. Three-dimensional structure determination of the MxaJ protein is currently in progress by the single-wavelength anomalous dispersion technique and model building.

The Role of Oxidative Inactivation of Phosphatase PTEN and TCPTP in Fatty Liver Disease.

Alcoholic liver disease (ALD) and nonalcoholic fatty liver disease (NAFLD) are becoming increasingly prevalent worldwide. Despite the different etiologies, their spectra and histological feature are similar, from simple steatosis to more advanced stages such as steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. Studies including peroxiredoxin knockout models revealed that oxidative stress is crucial in these diseases, which present as consequences of redox imbalance. Protein tyrosine phosphatases (PTPs) are a superfamily of enzymes that are major targets of reactive oxygen species (ROS) because of an oxidation-susceptible nucleophilic cysteine in their active site. Herein, we review the oxidative inactivation of two tumor suppressor PTPs, phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and T-cell protein tyrosine phosphatase (TCPTP), and their contribution to the pathogenicity of ALD and NAFLD, respectively. This review might provide a better understanding of the pathogenic mechanisms of these diseases and help develop new therapeutic strategies to treat fatty liver disease.

Purification, crystallization and preliminary X-ray crystallographic analysis of a methanol dehydrogenase from the marine bacterium Methylophaga aminisulfidivorans MP(T).

Methylophaga aminisulfidivorans MP(T) is a marine methylotrophic bacterium that utilizes C(1) compounds such as methanol as a carbon and energy source. The released electron from oxidation flows through a methanol-oxidizing system (MOX) consisting of a series of electron-transfer proteins encoded by the mox operon. One of the key enzymes in the pathway is methanol dehydrogenase (MDH), which contains the prosthetic group pyrroloquinoline quinone (PQQ) and converts methanol to formaldehyde in the periplasm by transferring two electrons from the oxidation of one methanol molecule to the electron acceptor cytochrome c(L). In order to obtain molecular insights into the oxidation mechanism, a native heterotetrameric α(2)ß(2) MDH complex was directly purified from M. aminisulfidivorans MP(T) grown in the presence of methanol and crystallized. The crystal diffracted to 1.7 Šresolution and belonged to the monoclinic space group P2(1) (unit-cell parameters a = 63.9, b = 109.5, c = 95.6 Å, ß = 100.5°). The asymmetric unit of the crystal contained one heterotetrameric complex, with a calculated Matthews coefficient of 2.24 Å(3) Da(-1) and a solvent content of 45.0%.

Differentiating Radiation Necrosis from Brain Tumor Using Hyperpolarized Carbon-13 MR Metabolic Imaging.

PURPOSE: Differentiation between radiation-induced necrosis and tumor recurrence is crucial to determine proper management strategies but continues to be one of the central challenges in neuro-oncology. We hypothesized that hyperpolarized 13C MRI, a unique technique to measure real-time in vivo metabolism, would distinguish radiation necrosis from tumor on the basis of cell-intrinsic metabolic differences. The purpose of this study was to explore the feasibility of using hyperpolarized [1-13C]pyruvate for differentiating radiation necrosis from brain tumors. PROCEDURES: Radiation necrosis was initiated by employing a CT-guided 80-Gy single-dose irradiation of a half cerebrum in mice (n = 7). Intracerebral tumor was modeled with two orthotopic mouse models: GL261 glioma (n = 6) and Lewis lung carcinoma (LLC) metastasis (n = 7). 13C 3D MR spectroscopic imaging data were acquired following hyperpolarized [1-13C]pyruvate injection approximately 89 and 14 days after treatment for irradiated and tumor-bearing mice, respectively. The ratio of lactate to pyruvate (Lac/Pyr), normalized lactate, and pyruvate in contrast-enhancing lesion was compared between the radiation-induced necrosis and brain tumors. Histopathological analysis was performed from resected brains. RESULTS: Conventional MRI exhibited typical radiographic features of radiation necrosis and brain tumor with large areas of contrast enhancement and T2 hyperintensity in all animals. Normalized lactate in radiation necrosis (0.10) was significantly lower than that in glioma (0.26, P = .004) and LLC metastatic tissue (0.25, P = .00007). Similarly, Lac/Pyr in radiation necrosis (0.18) was significantly lower than that in glioma (0.55, P = .00008) and LLC metastasis (0.46, P = .000008). These results were consistent with histological findings where tumor-bearing brains were highly cellular, while irradiated brains exhibited pathological markers consistent with reparative changes from radiation necrosis. CONCLUSION: Hyperpolarized 13C MR metabolic imaging of pyruvate is a noninvasive imaging method that differentiates between radiation necrosis and brain tumors, providing a groundwork for further clinical investigation and translation for the improved management of patients with brain tumors.

Rosiglitazone reduces tau phosphorylation via JNK inhibition in the hippocampus of rats with type 2 diabetes and tau transfected SH-SY5Y cells.

Increasing evidence supports an association between Alzheimer's disease (AD) and diabetes. Rosiglitazone, a peroxisome proliferator-activated receptor-γ (PPARγ) agonist, which is an anti-diabetic agent against type 2 diabetes, is currently in Phase III clinical trials in AD patients because rosiglitazone reduces ß-amyloid (Aß) pathology and inflammation. However, few studies have investigated whether rosiglitazone affects tau phosphorylation, another critical pathological feature of AD. Thus, we investigated it using OLETF type 2 diabetic rats and streptozotocin-injected type 1 diabetic mice. Interestingly, rosiglitazone reduced tau phosphorylation only in the hippocampus of OLETF type 2 diabetes rats, and not in that of STZ-injected type 1 diabetes mice. The activity of JNK was reduced in the hippocampus of rosiglitazone-treated OLETF rats, correlating with a reduction in tau phosphorylation, however, which was not correlated with GSK3ß activity. In human tau-transfected SH-SY5Y neuronal cell line, reduction of tau phosphorylation was also associated with reduction of JNK activity, not of GSK3ß activity. Hence, rosiglitazone could be used in reducing tau phosphorylation through JNK inactivation for therapeutic effects in type 2 diabetes related Alzheimer's disease.

Non-catalytic-Region Mutations Conferring Transition of Class A ß-Lactamases Into ESBLs.

Despite class A ESBLs carrying substitutions outside catalytic regions, such as Cys69Tyr or Asn136Asp, have emerged as new clinical threats, the molecular mechanisms underlying their acquired antibiotics-hydrolytic activity remains unclear. We discovered that this non-catalytic-region (NCR) mutations induce significant dislocation of ß3-ß4 strands, conformational changes in critical residues associated with ligand binding to the lid domain, dynamic fluctuation of Ω-loop and ß3-ß4 elements. Such structural changes increase catalytic regions' flexibility, enlarge active site, and thereby accommodate third-generation cephalosporin antibiotics, ceftazidime (CAZ). Notably, the electrostatic property around the oxyanion hole of Cys69Tyr ESBL is significantly changed, resulting in possible additional stabilization of the acyl-enzyme intermediate. Interestingly, the NCR mutations are as effective for antibiotic resistance by altering the structure and dynamics in regions mediating substrate recognition and binding as single amino-acid substitutions in the catalytic region of the canonical ESBLs. We believe that our findings are crucial in developing successful therapeutic strategies against diverse class A ESBLs, including the new NCR-ESBLs.

Organic/Inorganic Hybrid Buffer in InGaZnO Transistors under Repetitive Bending Stress for High Electrical and Mechanical Stability.

We investigated the influence of the multilayered hybrid buffer consisting of Al2O3/PA (polyacrylic) organic layer/Al2O3 on the electrical and mechanical properties of amorphous InGaZnO (a-IGZO) thin-film transistors (TFTs). The multilayered organic/inorganic hybrid buffer has multiple beneficial effects on the flexible TFTs under repetitive bending stress. First, compared to the PA or Al2O3 single-layered buffer, the multilayered hybrid buffer showed an improved WVTR value of 1.1 × 10-4 g/m2 day. Even after 40,000 bending cycles, the WVTR value of the hybrid buffer increased only by 17%, while the WVTR value of the Al2O3 layer doubled after cyclical bending stress. We also confirmed that the hybrid buffer has advantages in mechanical durability of the TFT layers because of the change in the position of the neutral plane and the strain reduction effect by the PA organic layer. When we fabricate a top-gate a-IGZO TFT with the hybrid buffer layer (HB TFT), the device shows Vth = 0.74 V, µFE = 14.4 cm2/V·s, a subthreshold slope of 0.27 V/dec, and hysteresis of 0.21 V, which are superior to that of TFTs fabricated on an Al2O3 single-layer buffer (IB TFT). From the X-ray photoelectron spectroscopy and elastic recoil detection analysis, the difference in the electrical performance of TFTs could be explained by hydrogen-related molecules. After annealing at 270 °C, the amounts of hydrogen found in the a-IGZO layer for the IB, HB, and OB TFTs were 3.57 × 1021, 5.77 × 1021, and 7.34 × 1021 atoms/cm3, respectively. A top-gate bottom-contact structured a-IGZO TFT fabricated on the PA layer (OB TFT) showed a gate dielectric breakdown because of excessively high hydrogen content and high nonbonding oxygen content. On the other hand, HB TFTs showed better positive bias stability because of the higher hydrogen concentration, as hydrogen (when not excessive) is beneficial in passivating electron traps. Finally, we conducted 60,000 repetitive bending cycles on IB TFTs and HB TFTs with various bending radii down to 1.5 mm. The HB TFT shows improved mechanical durability and exhibits less electrical degradation during and after repetitive bending stress, compared to the IB TFT.

Runx3 inhibits IL-4 production in T cells via physical interaction with NFAT.

Interleukin (IL)-4 plays a key role in T helper 2 (Th2) cell differentiation favoring humoral immune response. Regulation of IL-4 gene expression, therefore, is critically important for Th2 dependent responses and Th2 dominant disorders. In T cells, IL-4 gene expression is regulated positively or negatively by a combination of several transcription factors. Recently, enhanced IL-4 production was reported in Runx3 knockout mice; this implies negative regulation of IL-4 by Runx3. Runx proteins are transcription factors that have a Runt domain and have essential functions in development. In this study, the molecular mechanism that downregulates IL-4 expression was investigated. Runx3 inhibited IL-4 production in EL-4 T cells stimulated with PMA/ionomycin. Runx3-mediated IL-4 inhibition was NFAT-dependent, and Runx3 was physically associated with NFAT. Therefore, our results suggest that the interaction between NFAT and Runx3 is a mechanism that causes the negative regulation of IL-4, along with previously reported repression by T-bet.

Increased IL-2 production in T cells by xanthohumol through enhanced NF-AT and AP-1 activity.

Xanthohumol (XN) is a major chalcone found in hop, which is used to add bitterness and flavor to beer. In this study, we investigated the effects of XN on the production of interlukin-2 (IL-2), a potent T cell growth factor. Treatment with XN significantly increased IL-2 production in mouse EL-4 T cells activated with phorbol 12-myristate 13-acetate (PMA) plus ionomycin (Io) in a dose-dependent manner. To further characterize its regulatory mechanism of XN on increased IL-2 production, the effects of XN on IL-2 promoter activity and the activity of several transcription factors modulating IL-2 expression were analyzed. XN enhanced activity of the IL-2 promoter, which contains distal and proximal regulatory elements in PMA/Io-activated EL-4 T cells. Furthermore, the activity of NF-AT and AP-1 was enhanced but NF-kappaB activity was not influenced by XN in PMA/Io-activated EL-4 T cells. These results suggest that XN increased IL-2 production at the transcriptional levels via the up-regulation of NF-AT and AP-1 in PMA/Io-activated EL-4 T cells.