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The objective of this study was to evaluate whether D-allulose has teratogenic effects on rats. A prenatal developmental toxicity test was conducted in SD rats in compliance with modified OECD guidelines test number 414, prenatal developmental toxicity study. Pregnant female rats received repeated doses of 1250, 2500, or 5000 mg/kg body weight D-allulose, or a vehicle control by gavage on GD 6-15. On GD 20, one day prior to the expected day of delivery, pregnant rats were weighed and anesthetized, and laparotomized to remove the uterine and its content were weighed. Fetuses were examined macroscopically for any soft tissue and skeletal changes. The evaluation indicators included general sign observation, body weight, food consumption, animal death, corpora lutea, numbers of embryonic or fetal deaths, and viable fetuses including live birth rate, fetal resorption rate, and stillbirth rate, as well as sex, body weights, and skeletal and soft tissue alterations of fetuses. No treatment-related abnormalities were observed in prenatal developmental toxicity and fetal malformation parameters, indicating that D-allulose had no teratogenic effects on pregnant rats and fetuses. From the findings of this prenatal developmental toxicity study, the NOAEL of D-allulose was estimated to be 5000 mg/kg/day in pregnant SD rats.

The aggregation of aminoacyl transfer RNA synthetase complex-interacting multifunctional protein-2 (AIMP2) accelerates α-synuclein aggregation via direct interaction, leading to enhanced dopaminergic neurotoxicity in Parkinson's disease (PD). Thus, it would be beneficial to prevent AIMP2 aggregation to suppress α-synucleinopathy in PD. In this study, we screened small compounds that could inhibit the in vitro aggregation of AIMP2 using a 1909 small-compound library. The AIMP2 inhibitors (SAI-04, 06, and 08) with the most effective inhibition of AIMP2 aggregation bind to AIMP2, disaggregate the pre-formed AIMP2 aggregates, and prevented AIMP2/α-synuclein coaggregation and cytotoxicity in SH-SY5Y cells. Moreover, AIMP2 inhibitors prevented α-synuclein preformed fibril (PFF)-induced pathological AIMP2 aggregation in both mouse cortical and embryonic stem cell-derived human dopaminergic neurons, thereby blocking PFF-induced α-synuclein aggregation and neurotoxicity. Collectively, our results suggest that the use of brain-permeable AIMP2 aggregation inhibitors may serve as an effective therapeutic strategy for α-synucleinopathy in PD.

We developed Fenton-like reaction performing mineralized nanoparticles as novel oxidative stress amplifying anticancer agents. We fabricated Cu-doped superoxide dismutase (SOD)-loaded calcium carbonate (CaCO3)-mineralized nanoparticles (Cu/SOD-MNPs) that could simultaneously release hydrogen peroxide-generating SOD and Fenton-like reaction-inducing copper ions within cancer cells. The CaCO3 core of the Cu/SOD-MNPs effectively inhibited the release of the entrapped SOD and copper ions in physiological pH (7.4). In contrast, in acidic pH (5.0), the release of SOD and copper ions was facilitated by CaCO3 dissolution. We demonstrated that the Cu/SOD-MNPs elevated the level of highly toxic apoptosis-inducing hydroxyl radicals in the cancer cells. The Cu/SOD-MNPs showed specific toxicity for cancer cells (MCF-7 cells), whereas no or negligible toxicity was found for normal cells (NIH3T3 and HEK293 cells). This mineralized oxidative stress amplifying nanoparticle may serve as a novel cancer-specific anticancer agent.

The purpose of this study was to determine whether disease recurrence and intrinsic characteristics of ductal carcinoma in situ (DCIS) are associated with the imaging method of detection in asymptomatic women. This multicenter, retrospective study included 844 women treated for asymptomatic DCIS who had pre-operative mammography and breast ultrasonography (US) studies available. Of the 844 women, 25 (3.0%) developed recurrences. Patients in the US group had significantly lower 5- and 10-y recurrence-free survival (RFS) rates compared with patients in the mammography group (p = 0.011). US-detected DCIS showed significantly lower 5-and 10-y RFS rates compared with mammography-detected DCIS in patients50 y or with mammographically dense breasts (p = 0.002 and 0.002, respectively). US as the detection modality (hazard ratio [HR]: 4.451; 95% confidence interval [CI]: 1.530, 12.950; p = 0.006) and HER2 positivity (HR: 4.036; 95% CI: 1.438; 11.330; p = 0.008) were significantly associated with recurrence. We concluded that US as the detection modality and HER2 positivity were significantly associated with recurrence in patients treated for asymptomatic DCIS.

Highlights • RNA adjuvant was developed from the CrPV intergenic region IRES. • The RNA adjuvant functioned as an adjuvant with protein-based vaccines. • The RNA adjuvant increased vaccine efficacy and induced balanced Th1/Th2 response. • The RNA adjuvant enhanced APC chemotaxis., An ideal adjuvant should increase vaccine efficacy through balanced Th1/Th2 responses and be safe to use. Recombinant protein-based vaccines are usually formulated with aluminum (alum)-based adjuvants to ensure an adequate immune response. However, use of alum triggers a Th2-biased immune induction, and hence is not optimal. Although the adjuvanticity of RNA has been reported, a systematic and overall investigation on its efficacy is lacking. We found that single strand RNA (termed RNA adjuvant) derived from cricket paralysis virus intergenic region internal ribosome entry site induced the expression of various adjuvant-function-related genes, such as type 1 and 2 interferon (IFN) and toll-like receptor (TLR), T cell activation, and leukocyte chemotaxis in human peripheral blood mononuclear cells; furthermore, its innate and IFN transcriptome profile patterns were similar to those of a live-attenuated yellow fever vaccine. This suggests that protein-based vaccines formulated using RNA adjuvant function as live-attenuated vaccines. Application of the RNA adjuvant in mouse enhanced the efficacy of Middle East respiratory syndrome spike protein, a protein-subunit vaccine and human papillomavirus L1 protein, a virus-like particle vaccine, by activating innate immune response through TLR7 and enhancing pAPC chemotaxis, leading to a balanced Th1/Th2 responses. Moreover, the combination of alum and the RNA adjuvant synergistically induced humoral and cellular immune responses and endowed long-term immunity. Therefore, RNA adjuvants have broad applicability and can be used with all conventional vaccines to improve vaccine efficacy qualitatively and quantitively.

Based on density functional theory, we systematically study the mechanical and electronic properties of monolayer and bilayer SnS2 and SnSe2. The electronic properties of these layers can be significantly tuned by applying in-plane strains and electric fields perpendicular to the sheets. The band gaps of monolayer SnS2 and SnSe2 slightly increase with the in-plane tensile strains, and they start to decrease after critical strains (5% for monolayer SnS2 and 7% for monolayer SnSe2). The band gaps of bilayer SnS2 and SnSe2 have a similar tendency to the monolayers with smaller critical strains (1% for bilayer SnS2 and 2% for bilayer SnSe2), which enables a semiconductor-to-metal transition at 10% strain for bilayer SnSe2. We also find that an external electric field perpendicular to bilayer SnS2 and SnSe2 modulates their electronic band gaps. Semiconductor-to-metal transitions are achieved at the electric fields of 0.27 V/A for bilayer SnS2 and 0.13 V/A for bilayer SnSe2.

Oxidative stress and the related inflammatory responses are closely associated with many diseases including cardiovascular diseases such as atherosclerosis. Especially, mitochondrial damage and inflammasome activation have been reported to be directly involved in atherogenesis. In addition, we previously reported that endothelial cells (ECs) exposed to oxidized LDL (oxLDL) release ATP, which activates P2Y2R, resulting in the expression of receptors for advanced glycation end products and adhesion molecules that are involved in the pathogenesis of atherosclerosis. Therefore, it is expected that P2Y2R activation by ATP released under inflammatory conditions may be linked to the inflammasome-mediated pathogenesis of cardiovascular diseases such as atherosclerosis. However, the exact association remains unclear. Thus, in this study, we investigated the role of P2Y2R in oxLDL-mediated inflammasome activation and the related atherosclerotic pathogenesis in ECs. ECs stimulated with oxLDL demonstrated increased intracellular production and extracellular secretion of ATP. In addition, mitochondrial reactive oxygen species (mtROS) production and mitochondrial DNA (mtDNA) expression and cytosolic release were increased in ECs stimulated with oxLDL or the P2Y2R agonists ATP and UTP. Moreover, caspase-1 activity and IL-1β production were increased in ECs stimulated with oxLDL, ATP or UTP through the modulation of mtROS production and mtDNA expression, in a P2Y2R-dependent manner. Furthermore, TLR-9 and NF-κB activation was increased in ECs in response to oxLDL, ATP or UTP, in a mtDNA-dependent manner. Taken together, our findings suggest that P2Y2R activation by ATP is involved in oxLDL-mediated inflammasome activation and subsequent IL-1β production through the modulation of mtROS-mtDNA induction and the TLR9-NF-κB signaling pathway.

Density functional theory based calculations are used to investigate the energetically and dynamically most stable structure for a crystalline unzipped graphene oxide (UGO) monolayer by considering parallel or antiparallel dipolar structures. The O atom in the UGO monolayer induces electron excessive and deficient regions, which makes Li-ions selectively trapped in the electron deficient region. Apart from this, the periodic injection of O in graphene (C4O) opens a small band gap (0.6 eV) while keeps the Dirac cone. The adsorption of Li-ions makes it metallic by donating an extra electron to the UGO monolayer. The UGO monolayer can have a high Li storage capacity of 419 mAh/g and a typical anodic open circuit voltage range of 0.94–0.19 V. The energy barrier for Li migration is almost double of the pristine graphene, which results from the localized positive charge regions in the UGO monolayer.

The goal of this study was to evaluate various clinicopathologic and imaging characteristics as independent predictors of axillary pathologic complete response (pCR) to neoadjuvant chemotherapy (NAC) and to determine the added value of a model that integrates imaging characteristics of ultrasound (US) restaging with known clinicopathologic characteristics. A total of 227 clinically node-positive breast cancer patients underwent axillary US after NAC (termed US restaging) before surgery. We constructed a clinicopathologic model with independent predictors of clinicopathologic characteristics in multivariate analyses. A combined model was created by integrating imaging characteristics with clinicopathologic characteristics. The predictive values of the models were compared using the area under the receiver operating characteristic curve. Of the 227 patients, 106 (46.7%) achieved axillary pCR. Multivariate analysis revealed that higher histologic grades (odds ratio [OR] = 4.21 and 10.11 for moderate and high grade, respectively), negative hormonal receptor status (OR = 2.88), smaller (≤1.5 cm) residual tumor size (OR = 2.83), absence of fatty hilum loss (OR = 14.06) and absence of eccentric cortical thickening of the axillary lymph node (OR = 4.42) were independently associated with the axillary pCR (all p values0.05). Integrating the imaging characteristics of the US restaging significantly increased the predictive capability of the model that applied only the clinicopathologic characteristics (c-index, 0.783 vs. 0.657; p0.001). Imaging characteristics of the US restaging were independently associated with axillary pCR after NAC and they significantly improved the predictive capability of the model that used only the clinicopathologic characteristics.

This study highlights the effects of a biomass-derived carbon coating on the properties of flexible lithium iron phosphate polymer batteries. Pure LiFePO4 (LFP) and carbon-coated LiFePO4 (C-LFP) cathode materials are synthesized by a modified mechanical activation process. LiFePO4 was uniformly coated with biomass-derived carbon by the addition of orange peel during the synthesis. C-LFP with a particle size of approximately 90 nm, having a homogeneous particle size distribution with particles smaller than that of pure LFP (140 nm), is obtained with 6.0 wt% carbon content. The electrochemical properties are evaluated at room temperature by cyclic voltammetry and charge-discharge performance analysis using porous carbon current collector with a lithium metal anode and gel polymer electrolyte that is prepared by the phase inversion method. C-LFP exhibits a high discharge capacity of 147.3 mAh g−1 (corresponding to 87% of the theoretical capacity) at 0.5 C-rate and 139.8 mAh g−1 at 1 C-rate. Good rate capability and stable cycle performance were realized for flexible lithium polymer cells employing the biomass derived-carbon coated LiFePO4.

Microbacterium foliorum is a naturally occurring bacteria in cruciferous vegetables and ripened cheese. The safety of M. foliorum SYG27B-MF has been assessed in both acute and subchronic studies and a battery of mutagenicity and clastogenicity tests. In a single dose acute study, the LD50 of M. foliorum SYG27B-MF was greater than 3 g/kg bw or 5.1 × 1016 colony forming unit (CFU)/kg bw, the highest dose tested. In a 90-day subchronic toxicity study in 80 Sprague-Dawley rats, no animals died and there were no treatment-related abnormalities at doses of 0, 500, 1000, or 2000 mg/kg bw. In a 90-day repeated toxicity test, the no-observed-adverse-effect level (NOAEL) M. foliorum SYG27B-MF was 2000 mg/kg/day or 3.4 × 1016 CFU/kg bw/day, the highest level tested. A mutagenicity study using reverse bacterial mutation tests and a genotoxicity study employing cultured hamster ovarian fibroblasts (CHO-K1) cell showed that M. foliorum SYG27B-MF was not mutagenic or clastogenic in the presence or absence metabolic activation. In an in vivo mouse micronucleus assay, M. foliorum SYG27B-MF did not induce did not induce micronuclei formation in the bone marrow cells of mice, indicating that it is non-clastogenic. The results from these studies support the safety of M. foliorum SYG27B-MF for use as a production organism for human food ingredients.

Rice can be the major contributor to exposure of inorganic arsenic (iAs). A total of 485 domestic rice (polished rice, n = 368; husked rice, n = 117) samples produced in 2014 were determined for total arsenic (tAs) and As species. The determination of As species was conducted by HPLC-ICP-MS. AsIII was the major detected species. AsV, DMA, and MMA were occurred in lower level in both types of rice. In polished rice, iAs represented 43–91% of tAs: tAs = 0.088 ± 0.021 mg/kg; iAs = 0.060 ± 0.013 mg/kg. In husked rice, iAs ranged from 55% to 91% of tAs; tAs = 0.160 ± 0.042 mg/kg, iAs = 0.114 ± 0.029 mg/kg. The iAs level of polished rice samples was below the current maximum limit of the Chinese and German standard (0.2 mg/kg for inorganic arsenic in milled rice). Also, husked rice samples did not exceed the standard for iAs. Average dietary exposure to iAs from polished rice consumption was calculated as 0.17 ± 0.21 μg/kg·bw/day which was 8.09% level of Provisional Tolerable Weekly Intake. The result obtained in the present study provide a reference data of iAs in Korean rice for the establishment of safety standards.

To enhance the electrochemical performance and identify the redox reaction of a nitroxide radical-based organic battery, poly(2,2,6,6-tetramethylpiperidinyloxy-4-vinylmethacrylate) (PTMA) as nitroxide radical polymer is synthesized and used to fill the central pore of nitrogen-doped carbon nanotubes (NCNTs). The PTMA-filled NCNTs demonstrate high rate capability, stable cycling, and high electrode stability, owing to their high charge transfer and ion diffusion; further, they prevent the dissolution of organic active material. The nitrogen doping on CNTs improves the electrochemical properties by created defects. The PTMA-filled NCNT cells have a high reversible capacity of 199.8 mAh g−1 and high energy density of 489.4 Wh kg−1 on electrode level at 0.2C with a two-step redox couple. Moreover, the cells with the two-step redox couple exhibit excellent stable cycle performance with high capacity retention and Coulombic efficiency for 3000 cycles. The nitroxide radical of PTMA can be converted to either an oxoammonium cation or an aminoxy anion; further, the redox reaction of each step exhibits different performance. The nitroxide radical–oxoammonium cation redox step in NCNTs shows a high rate capability owing to the high charge transfer rate and ion diffusion coefficient. The unusual results will help further improve the properties and commercialization of rechargeable organic batteries.

High mobility group box 1 (HMGB1), a highly conserved non-histone DNA-binding protein, plays an important role in the pathogenesis of sepsis. Previously, the authors reported 13-ethylberberine (13-EBR) has anti-inflammatory and antibacterial effects. However, the effect of 13-EBR on HMGB1 release was not investigated. In the present study, it was hypothesized 13-EBR might reduce HMGB1 release by activating AMPK under septic conditions. The results obtained showed 13-EBR significantly reduced HMGB1 release from LPS-activated RAW264.7 cells, and that this reduction was reversed by silencing p38, or AMPK, or by co-treating cells with p38 MAPKinase inhibitor. 13-EBR increased the phosphorylations of p38 and AMPK, and the phosphorylation of p38 by 13-EBR was inhibited by AMPK-siRNA, indicating AMPK acted upstream of p38. In the lung tissues of LPS-treated mice, 13-EBR administration significantly increased p-AMPK but reduced inducible nitric oxide synthase (iNOS) protein levels. Hematoxylin and eosin staining revealed 13-EBR significantly reduced LPS-induced lung and liver damage. In addition, 13-EBR inhibited NF-kB in LPS-activated RAW264.7 cells, and in LPS-treated mice, 13-EBR administration significantly increased survival. Furthermore, co-administration of 13-EBR plus LPS prevented LPS-induced aortic rings hypocontractile response to phenylephrine in vitro. Taken together, these results indicate 13-EBR might offer a means of treating sepsis through AMPK activation.

d -Psicose, a rare sugar, not only has very low caloric value but possesses anti-adipogenic properties. Here, we identified target genes in adipose tissue affected by d -psicose by transcriptomic analysis and provided mechanistic explanations for the anti-adipogenic effect. C57BL/6J ob/ob mice were fed with a control or 5% d -psicose diet for 12 weeks. d -Psicose decreased final body weight, adipose tissue mass, adipocyte size, and serum total cholesterol levels. We identified 103 differentially expressed genes involved in inflammatory response, molecular transport, and lipid metabolism consequent to d -psicose administration. Genes related to inflammation and adipo/lipogenesis were significantly down-regulated, whereas those associated with β-oxidation were up-regulated by d -psicose. Our data suggest that Fos, Mmp3, Fgf21, and Abcd2 might be key target genes associated with d -psicose-induced changes in lipid metabolism and subsequent chronic inflammatory responses. d -Psicose is thus a promising sugar substitute possessing a direct gene-regulatory function related to the suppression of body fat deposition.

Objectives The aim of the present study was to identify the anticancer effects and the mechanisms of action of shikonin and its analogue isobutyrylshikonin in oral squamous carcinoma cells. Designs The cytotoxic effects of isobutyrylshikonin and shikonin in Ca9-22 and SCC-25 cells were analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry analysis of Annexin V/Propidium Iodide (PI) staining, western blot analysis and immunohistochemistry. Results Treatment with both isobutyrylshikonin and shikonin induced dose- and time-dependent apoptotic cell death in Ca9-22 cells, although the IC50 of isobutyrylshikonin was less than that of shikonin. The induction of apoptosis by both isobutyrylshikonin and shikonin was accompanied by activation of caspase-8, -9, -3, and PARP, loss of mitochondrial trans-membrane potential, and release of cytochrome c from the mitochondria. ROS mediated the apoptosis induced by isobutyrylshikonin and shikonin, indicating that ROS may play a critical role in the distinctive cytotoxic effects of isobutyrylshikonin and shikonin in Ca9-22. Isobutyrylshikonin showed a similar cytotoxic effect in SCC-25 cells at concentrations showing the effects in Ca9 cells, but not in human normal keratinocyte cells. Although there is no biological difference between isobutyrylshikonin and shikonin, isobutyrylshikonin exerts the same cytotoxic effect at a concentration 6 times lower than shikonin. Conclusions The present study suggest that isobutyrylshikonin may be a more potent chemotherapeutic agent against oral cancer cells than shikonin. In addition, our data exhibit that both isobutyrylshikonin and shikonin induce caspase-dependent apoptosis via the mitochondrial pathway through accumulation of ROS in oral squamous carcinoma cells.

A substrate-free carbon-coated FeS2-fiber-based cathode for quasi-solid-state rechargeable flexible sodium metal batteries is reported for the first time. A facile electrospinning technique followed by carbonization is employed for the synthesis of a novel cathode for a flexible sodium metal batteries. For fabrication of the solid-state flexible battery, a Na3Zr2Si2PO12-based hybrid solid electrolyte is utilized as the separator and electrolyte. A discharge capacity of about 808.26 mAh g−1 is obtained in the first charge–discharge cycle, which is greater than that of any previously reported Na/FeS2-based battery. The carbon coating on the FeS2 fiber allows the flexible battery to have an enhanced discharge capacity of 633.43 mAh g−1, even after undergoing five charge–discharge cycles at a 0.1 C rate. At a 2 C rate, a high specific capacity of 393.5 mAh g−1 is maintained, and the value increases on the return from 2 C to 0.1 C until it reaches a value similar to the initial 0.1 C rate. A superior rate capability with enhanced electrochemical stability of the quasi-solid-state sodium metal battery is also demonstrated due to the substrate-free flexible carbon-coated FeS2-fiber-based cathode. Owing to its high flexibility and electrochemical properties, the flexible solid-state sodium metal battery has considerable potential for high-performance sodium batteries.

We investigated dielectric responses of a multi-component plasma at finite temperature and applied the results to spectral analysis of dynamic structure factor of a photo-generated high density plasma in a wurtzite GaN. The behavior of effective dielectric function and dynamic structure factor in energy loss measurement are examined focusing to the effects of dynamic screening of carriers in the plasma. Features of their spectral behavior are illustrated including the roles of various elementary excitations in the three-component solid state plasma.

There is great interest in the development of cardiac stem cells (CSCs) cell-based therapeutics; thus, clinical translation requires an efficient method for attaining therapeutic quantities of these cells. Furthermore, an in vitro model to investigate the mechanisms regulating the cardiac homeostasis is crucial. We sought to develop a simple myocardial culture method for enabling both the recapitulation of myocardial homeostasis and the simultaneous isolation of CSCs. The intact myocardial fragments were encapsulated 3-dimensionally into the fibrin and cultured under dynamic conditions. The fibrin provided secure physical support and substratum to the myocardium, which mediated integrin-mediated cell signaling that allowed in situ renewal, outgrowth and cardiomyogenic differentiation of CSCs, mimicking myocardial homeostasis. Since our culture maintained the myocardial CSCs niches, it was possible to define the identity of in vitro renewed CSCs that situated in the interstitium between cardiomyocytes and microvessels. Lastly, the use of matrix-restricted fibrinolysis enabled the selective isolation of outgrown CSCs that retained the clonogenicity, long-term growth competency and cardiovascular commitment potential. Collectively, this myocardial culture might be used as an alternative tool for studying cardiac biology and developing cell-based therapeutics.

Periodic patterning with adatoms or defect is one of the methods for opening the band gap of graphene. In particular, under certain configurations controlled by the order of hydrogen and halogen atoms attached on graphene, inversion symmetry of graphene can be broken to give piezoelectricity as well as pyroelectricity. Using first-principles calculations, we examine the structural stability and electronic properties of four polar conformations of halogenated graphane (C2HX)n to understand the origin of piezoelectricity in this two-dimensional system. The formation energies and piezoelectric coefficients manifest that the four conformations of (C2HF)n are energetically stable with considerable piezoelectricity. We find that the electronic contribution of the proper piezoelectricity in (C2HF)n is mainly related to the change of the electron distribution around F atoms. By substituting flourine with chlorine, we confirm that the piezoelectricity enhances at the expense of stability degradation.

We have previously reported that phosphoenolpyruvate carboxykinase (PCK) overexpression under glycolytic conditions enables Escherichia coli to harbor a high intracellular ATP pool resulting in enhanced recombinant protein synthesis. To estimate how much PCK-mediated phosphoenolpyruvate (PEP) carboxylation is contributed to the ATP increase under engineered conditions, the kinetics of PEP carboxylation by PCK and substrate competing phosphoenolpyruvate carboxylase (PPC) were measured using recombinant enzymes. The PEP carboxylation catalytic efficiency (kcat/Km) of the recombinant PCK was 660mM(-1)min(-1), whereas that of the recombinant PPC was 1500mM(-1)min(-1). Under the presence of known allosteric effectors (fructose 1,6-bisphosphate, acetyl-CoA, ATP, malate, and aspartate) close to in vivo conditions, the catalytic efficiency of PCK-mediated PEP carboxylation (84mM(-1)min(-1)) was 28-folds lower than that of PPC (2370mM(-1)min(-1)). To verify the above results, an E. coli strain expressing native PCK and PPC under control of identical promoter was constructed by replacing PCK promoter region with that of PPC in chromosome. The native PCK activity (33nmol/mg-proteinmin) was 5-folds lower than PPC activity (160nmol/mg-proteinmin) in the cell extract from the promoter-exchanged strain. Intracellular modifications of ATP concentration by PCK activity and the consequences for biotechnology are further discussed.

The goal of this study was to investigate the differential effect of tanshinone IIA on the induction of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) by TNF-α and the possible molecular mechanisms by which it regulates ICAM-1 and VCAM-1 expression differentially. Stimulation of human umbilical vein endothelial cells (HUVEC) with TNF-α increased ICAM-1 and VCAM-1 expressions, and the pretreatment with tanshinone IIA concentration dependently inhibited VCAM-1 expression but not ICAM-1 expression. In previous study, PI3K/Akt, PKC and Jak/STAT-3 pathways were involved in the TNF-α-mediated induction of VCAM-1 but not ICAM-1. Thus, we examined the effect of tanshinone IIA on TNF-α-mediated activations of PI3K/Akt, PKC and Jak/STAT-3 pathways. Tanshinone IIA efficiently inhibited the phosphorylations of Akt, PKC and STAT-3 by TNF-α. Moreover, we determined the effect of tanshinone IIA on IRF-1 or GATAs induction and binding activity to VCAM-1 promoter since the upstream promoter region of VCAM-1 but not ICAM-1 contains IRF-1 and GATA binding motifs. Western blot analysis and ChIP assay showed that tanshinone IIA efficiently inhibited TNF-α-increased nuclear level of IRF-1 and GATA-6 and their binding affinity to VCAM-1 promoter region. Taken together, tanshinone IIA selectively inhibits TNF-α-mediated expression of VCAM-1 but not ICAM-1 through modulation of PI3/Akt, PKC and Jak/STAT-3 pathway as well as IRF-1 and GATA-6 binding activity.

Uridine diphosphogalactose-4-epimerase (UDP-galactose-4-epimerase, GalE, EC 5.1.3.2) mediates 4-epimerization of free monosaccharides in a low activity as well as original UDP-galactose substrate in a high activity (Appl Biochem Biotechnol 163:444–451). To improve GalE for monosaccharide 4-epimerization, random mutations were introduced and the effects of single mutations on the conversion of tagatose were evaluated. Among 3060 random mutated GalE variants, two clones showing the improved conversion activity of tagatose 4-epimerization were selected and their mutation points were sequenced. To investigate the effect of each mutations, the 5 mutation points (D58E, N100S, P193S, I196N, and T317S) found in the two selected clones were further introduced into the wild-type GalE and the variant enzymes were purified for kinetic study. The enzyme carrying D58E single mutation (GalE D58E), which is located near the cofactor binding pocket in a 3D structure docking model, showed the kinetic parameters of K M = 388 mM and k cat = 1.77 min −1 for tagatose substrate and it was 3.3-fold greater k cat / K M than the wild-type enzyme. When the N179S mutation, which is located near the substrate-binding pocket according to our previous study, was further combined with GalE D58E, the kinetic parameters of the double mutant enzyme was found to have 3.7-fold higher k cat / K M than the wild-type. The effect of the mutations on the activity improvement based on the 3D structure docking model and further biotechnological potential is discussed.

Screening patients at high risk of recurrence of cancer would allow for more accurate and personalized treatment. In this study, we tried to identify the prognosis-related protein profile by applying two different quantitative proteomic techniques, difference in-gel electrophoresis and cleavable isotope-coded affinity tag method. Six tumor tissues were obtained from stage IV colorectal cancer (CRC) patients, of which three have survived more than five years (good prognostic group, GPG) and the other three died within 25 months (poor prognostic group, PPG) after palliative surgery and subsequent chemotherapy treatment. From the two independent quantitative analyses, we identified 175 proteins with abundance ratios greater than 2-fold. Gene ontology analysis revealed that proteins related to cellular assembly/organization and movement were generally increased in the PPG. Twenty-two proteins, including caveolin-1, were chosen for confirmatory studies by Western blot and immunohistochemistry. The Western blot data for each individual protein were analyzed with Mann–Whitney tests, and a multi-marker panel was generated by logistic regression analysis. Five proteins, fatty acid binding protein 1, intelectin 1, transitional endoplasmic reticulum ATPase, transgelin and tropomyosin 2, were significantly different between the two prognostic groups within 95% confidence. No single protein could completely distinguish the two groups from each other. However, a combination of the five proteins effectively distinguished PPG from GPG patients (AUC = 1). Our study suggests a multi-marker panel composed of proteome signatures that provide accurate predictive information and potentially assist personalized therapy. This article is part of a Special Issue entitled: Proteomics: The clinical link.

Cell adhesion molecules (CAMs) are involved in a variety of pathologies including cancer, inflammation, pathogenic infections and autoimmune disease. In particular, VCAM-1, rather than ICAM-1, plays a major role in the initiation of atherosclerosis and tumor progression. Therefore, we attempted to elucidate differential mechanisms that regulate VCAM-1 and ICAM-1 expressions. Down-regulation of JNK by a specific inhibitor (SP600125) or dominant negative (DN) JNK1 plasmid enhanced TNF-α-induced VCAM-1 but not ICAM-1 expression. Moreover, transfection with a JNK1-overexpressing vector resulted in the inhibition of VCAM-1 expression stimulated by TNF-α in HUVECs, suggesting that JNK negatively regulates TNF-α-induced VCAM-1 expression in endothelial cells (ECs). Next, we investigated whether JNK signaling affects IRF-1 and/or GATA6, which are transcription factors that mediate TNF-α induction of VCAM-1 but not ICAM-1. The DN-JNK1 plasmid-transfected cells enhanced TNF-α up-regulation of IRF-1 whereas JNK1-overexpressing cells displayed down-regulation; however, neither DN-JNK1 transfection nor JNK1 overexpression affected GATA6 protein levels in the nuclear fraction. Chromatin immunoprecipitation (ChIP) assay confirmed that the inhibition of JNK by DN-JNK1 transfection increases the binding of IRF-1 to the VCAM-1 promoter whereas the overexpression of JNK1 inhibits IRF-1 binding to the VCAM-1 promoter. However, neither DN-JNK1 nor JNK1 overexpression altered GATA6 affinity for the VCAM-1 promoter region. We also examined whether MKP-7 affects ICAM-1 or VCAM-1 by regulating JNK. TNF-α-induced phosphor-JNK levels increased after 5min, peaked at 10 min, and decreased after 30 min. Interestingly, MKP-7 protein levels increased after 30 min, when phosphor-JNK induction by TNF-α was decreased. In addition, silencing MKP-7 with specific siRNA resulted in an increase in phosphor-JNK and inhibited the expression of VCAM-1 but not ICAM-1. Moreover, silencing MKP-7 caused the down-regulation of IRF-1 protein levels and binding to the VCAM-1 promoter. Thus, we suggest that MKP-7, a negative regulator of JNK, regulates VCAM-1 expression in activated endothelial cells through IRF-1 but not GATA6.

Mounting an efficient immune response to pathogens while avoiding damage to host tissues is the central task of the immune system. Emerging evidence has highlighted the contribution of the CD8(+) lineage of regulatory T cells to the maintenance of self-tolerance. Specific recognition of the MHC class Ib molecule Qa-1 complexed to peptides expressed by activated CD4(+) T cells by regulatory CD8(+) T cells triggers an inhibitory interaction that prevents autoimmune responses. Conversely, defective Qa-1-restricted CD8(+) regulatory activity can result in development of systemic autoimmune disease. Here, we review recent research into the cellular and molecular basis of these regulatory T cells, their mechanism of suppressive activity and the potential application of these insights into new treatments for autoimmune disease and cancer.

Ethnopharmacological relevance The rhizomes of Cyperus rotundus have been used as traditional folk medicine for the treatment of inflammatory diseases. However, the mechanism by which extract of rhizomes of Cyperus rotundus (ECR) elicits anti-inflammation has not been extensively investigated so far. The aim of the present study was to test whether heme oxygenase (HO)-1 induction is involved in the anti-inflammatory action of ECR. Materials and methods Induction of HO-1 and inhibition of inducible nitric oxide synthase (iNOS)/NO production by ECR and its 12 constituents (3 monoterpenes, 5 sesquiterpenes, and 4 aromatic compounds) were investigated using RAW264.7 cells in vitro. In addition, anti-inflammatory action of ECR and its two active ingredients (nookkatone, valencene) were confirmed in sepsis animal model in vivo. Results ECR increased HO-1 expression in a concentration-dependent manner, which was correlated with significant inhibition of iNOS/NO production in LPS-activated RAW264.7 cells. Among 12 compounds isolated from ECR, mostly sesquiterpenes induced stronger HO-1 expression than monoterpenes in macrophage cells. Nootkatone and valencene (sesquiterpenes) significantly inhibited iNOS expression and NO production in LPS-simulated RAW264.7 cells. Inhibition of iNOS expression by nootkatone, valencene, and ECR were significantly reduced in siHO-1 RNA transfected cells. Furthermore, all three showed marked inhibition of high mobility group box-1 (HMGB1) in LPS-activated macrophages and increased survival rates in cecal ligation and puncture (CLP)-induced sepsis in mice. Conclusions Taken together, we concluded that possible anti-inflammatory mechanism of ECR is, at least, due to HO-1 induction, in which sesquiterpenes such as nootkatone and valencene play a crucial role.

High mobility group box (HMGB)-1 plays an important role in sepsis-associated death in experimental studies. Heme oxygenase-1 (HO-1) inducers were reported to reduce HMGB1 release in experimental sepsis. Previously, we reported on the importance of the β₁-adrenergic receptor and protein kinase A pathway in the regulation of HO-1 expression by isoproterenol (ISO) in RAW 264.7 cells. We investigated whether ISO reduces HMGB1 release in LPS-activated RAW 264.7 cells and improves survival rate in septic mice due to HO-1 induction. ISO concentration-dependently increased HO-1 via Nrf-2 translocation and inhibited release of HMGB1 through the β₁-adrenergic receptor (β₁-AR) in LPS-activated RAW 264.7 cells. This conclusion was supported by the finding that dobutamine but not salbutamol increased HO-1 expression in both RAW 264.7 cells. ISO failed to inhibit HMGB1 release when HO-1 expression was suppressed by ZnPPIX, an HO-1 inhibitor in RAW 264.7 cells. ISO significantly inhibited phosphorylation of IκB-α and NF-κB-driven luciferase activity in LPS-activated RAW 264.7 cells. In addition, LY294002, a PI3K inhibitor, and SB203580, a p38 MAPK inhibitor, significantly inhibited not only HO-1 induction but also HMGB1 release by ISO. Importantly, ISO increased HO-1 protein expression in heart and lung tissues, reduced HMGB1 in plasma and increased survival rate in CLP-treated septic mice, which was significantly reversed by co-treatment with ZnPPIX. Taken together, we conclude that inhibition of HMGB1 release during sepsis via β₁-AR-mediated HO-1 induction is a novel mechanism for the beneficial effects of ISO in the treatment of sepsis.

Although aldose reductase (AR) has been implicated in the cellular response to oxidative stress, the role of AR in ultraviolet-B (UVB)-induced cellular injury has not been investigated. Here, we show that an increased expression of AR in human keratinocytes modulates UVB-induced apoptotic cell death and senescence. Overexpression of AR in HaCaT cells significantly attenuated UVB-induced cellular damage and apoptosis, with a decreased generation of reactive oxygen species (ROS) and aldehydes. Ablation of AR with small interfering RNA or inhibition of AR activity abolished these effects. We also show that increased AR activity suppressed UVB-induced activation of the p38 and c-Jun N-terminal kinases, but did not affect the extracellular signal-regulated kinase and phosphatidylinositol 3-kinase pathways. Similarly, UVB-induced translocation of Bax and Bcl-2 to mitochondria and cytosol, respectively, was markedly attenuated in cells overexpressing AR. Knockdown or inhibition of AR activity in primary cultured keratinocytes enhanced UVB-induced cellular senescence and increased the level of a cell-cycle regulatory protein, p53. Finally, cellular apoptosis induced by UVB radiation was significantly reduced in the epidermis of transgenic mice overexpressing human AR. These findings suggest that AR plays an important role in the cellular response to oxidative stress by sequestering ROS and reactive aldehydes generated in keratinocytes.

Ethnopharmacological relevance The methanol extract of Cimicifugae Rhizome has been traditionally used in various disorders including inflammation. Aim of the study The aim of the study is to explore whether anti-inflammatory action of 3 active compounds, two triterpenoid glycosides (cimiside E, 23-O-actylshengmanol-3-xyloside) and one furanocoumarin (isoimperatorin), isolated from Cimicifugae Rhizome is related with peroxisome proliferator-activated receptor-γ (PPAR-γ) expression in human umbilical endothelial cell line, EA.hy926 cells. Materials and methods Cell viability and production of reactive oxygen species were performed. In addition, adhesion of monocyte into endothelial cells and western blot for expression of adhesion molecules and signal proteins were investigated in tumor necrosis factor-α (TNF-α)-activated cells. Results Pretreatment of test compounds significantly reduced reactive oxygen species (ROS) production and expression of vascular cell adhesion molecule-1 (VCAM-1), but not intercellular cell adhesion molecule-1 (ICAM-1). Three compounds all dose-dependently increased not only PPAR-γ expression in EA.hy926 cells but inhibited TNF-α-induced phosphorylation of Akt, extracelullar-signal-regulated kinase (ERK) and protein kinase C (PKC) with different specificity. Finally, they prevented TNF-α-induced adhesion of U937 monocytic cells to EA.hy926 cells. Conclusions The present results show that cimiside E, 23-O-actylshengmanol-3-xyloside, isoimperatorin isolated from Cimicifugae Rhizome selectively inhibits TNF-α-induced expression of VCAM-1 at least by upregulation of PPAR-γ, and signals for ERK1/2, PI3K, and PKC are involved in this effect.

Activators of PPAR have been demonstrated to inhibit the induction of VCAM-1 but not ICAM-1 in human endothelial cells (EC). During the screening of anti-inflammatory activity of traditional herbs, we found 7,8-didehydrocimigenol (7,8-DHC), one of active triterpenoids of Cimicifugae rhizoma (C. rhizoma) increases PPAR-γ expression in EC in a time- and dose-dependent manner. Therefore, we asked whether 7,8-DHC selectively inhibits the expression of VCAM-1 but not ICAM-1 in TNF-α-activated EC via upregulation of PPAR-γ. Treatment with 7,8-DHC or PPAR-γ agonists (GW1929, troglitazone) inhibited the expression of VCAM-1 but not ICAM-1. Furthermore, the selective inhibition of VCAM-1 expression was inhibited by PPAR-γ antagonist, GW9662, or siPPAR-γ-transfected cells. 7,8-DHC significantly inhibited NF-kB activity via inhibition of phosphorylation of IkB and it also inhibited phosphorylation of ERK1/2 and Akt but not PKC. Finally, attachment of monocytes (U937) to EC by TNF-α was significantly reduced by 7,8-DHC. These results indicate that upregualtion of PPAR-γ by 7,8-DHC in EC inhibits NF-kB activity of TNF-α-activated EC which leads to selective inhibition of VCAM-1 expression. In addition, ERK1/2 and Akt signal pathways are involved in differential regulation by 7,8-DHC. We concluded that 7,8-DHC can be used for the treatment of cardiovascular disorders such as atherosclerosis.

Chronic exposure to manganese (Mn), which can be an occupational hazard or can result from liver failure, is associated with adverse motor and cognitive outcomes. Evidence from previous neuroimaging and magnetic resonance spectroscopy studies suggested alteration of function in Mn-exposed brains. However, the effect of chronic exposure of the human brain to Mn on white matter (WM) structure has not yet been determined. In the present study, we used diffusion tensor imaging (DTI) to investigate whether welders exposed to Mn demonstrate differences in WM integrity, compared with control subjects. Fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (AD), and radial diffusivity (RD) were measured on a voxel-wise basis in 30 male welders with exposure to Mn and in 19 age- and gender-matched control subjects. Direct comparison between welders and controls using investigator-independent Statistical Parametric Mapping (SPM) voxel-wise analysis of DTI metrics revealed a reduction of FA in the corpus callosum (CC) and frontal WM in Mn-exposed welders. Further, marked increases in RD and negligible changes in AD suggested that the microstructural changes in the CC and frontal WM result from compromised radial directionality of fibers in these areas, caused primarily by demyelination. Correlation analysis with neurobehavioral performance also suggested that the microstructural abnormalities were associated with subtle motor and cognitive differences in welders.

SUMMARY PARP inhibitors have shown promising clinical activities for patients with BRCA mutations and are changing the landscape of ovarian cancer treatment. However, the therapeutic mechanisms of action for PARP inhibition in the interaction of tumors with the tumor microenvironment and the host immune system remain unclear. We find that PARP inhibition by olaparib triggers robust local and systemic antitumor immunity involving both adaptive and innate immune responses through a STING-dependent antitumor immune response in mice bearing Brca1-deficient ovarian tumors. This effect is further augmented when olaparib is combined with PD-1 blockade. Our findings thus provide a molecular mechanism underlying antitumor activity by PARP inhibition and lay a foundation to improve therapeutic outcome for cancer patients., In Brief Ding et al. show that PARP inhibition in Brca1-deficient tumors elicits strong antitumor immunity involving activation of both innate and adaptive immune responses, a process that is dependent on STING pathway activation. In addition, they show that addition of PD-1 blockade augments the therapeutic efficacy of PARP inhibitor treatment., Graphical Abstract

Phosphotase and tensin homolog deleted on chromosome 10 (PTEN) is a potent negative regulator of PI3K/Akt pathway. Here, we tried to elucidate the role of PTEN in the regulation of endothelial adhesion molecules, vascular cell adhesion molecule (VCAM)-1 and intracellular adhesion molecule (ICAM)-1, induced by TNF-α in human endothelial cells (ECs). Transfection with PTEN overexpressing vector resulted in the significant decrease in phosphorylation of Akt in TNF-α-treated ECs. PTEN strongly inhibited VCAM-1 but not ICAM-1, however this inhibitory effect was reversed by co-transfection with constitutively active-Akt (CA-Akt-HA) in TNF-α-stimulated ECs. Additionally, silencing of PTEN with specific siRNA showed significant increase of phosphor-Akt compared with TNF-α alone treated ECs. siPTEN significantly upregulated VCAM-1 but was indifferent to ICAM-1 in TNF-α-treated cells. Further, chromatin immunoprecipitation (ChIP) assay showed that PTEN targets GATA-6 but not IRF-1 binding to VCAM-1 promoter. In addition, GATA-6 is associated with glycogen synthesis kinase-3beta (GSK-3β) which is in turn regulated by PTEN-dependent Akt activity. Finally, PTEN significantly prevented monocyte adhesion to TNF-α-induced ECs probably through VCAM-1 regulation. It is concluded that PTEN selectively inhibits expression of VCAM-1 but not ICAM-1 through modulation of PI3K/Akt/GSK-3β/GATA-6 signaling cascade in TNF-α-treated ECs.

This paper examines the strategic use of intermediaries by companies to better meet their technology needs from the resource-based theory and network theory perspectives. The results of this study are based on a sample of Korean IT companies and indicate the particular importance of cooperation with government support organizations and public R&D institutions in building relational assets. Intermediaries played a crucial role in enabling and facilitating joint development projects and promoting technology transfer among the companies in general. The results of this study suggest that intermediaries, as they are linked to other organizations within networks and are centrally situated within those networks where information and resource flow, can coordinate and control the exchange of information and resources within networks and enjoy timely and privileged access to such information, giving them the means to expand their strategic social capital.

Statins have recently been shown to produce anti-cardiac hypertrophic effects via the regulation of small GTPases. However, the effects of statins on G protein-mediated cardiac hypertrophy, which is the main pathway of cardiac hypertrophy, have not yet been studied. We sought to evaluate whether statin treatment directly suppresses cardiac hypertrophy through a large G protein-coupled pathway regardless of the regulation of small GTPases. Using neonatal rat cardiomyocytes, we evaluated norepinephrine-induced cardiac hypertrophy for suppressibility of rosuvastatin and the pathways involved by analyzing total protein/DNA content, cell surface area, immunoblotting and RT-PCR for the signal transduction molecule. In a concentration-dependent manner, rosuvastatin inhibited total protein synthesis and downregulated basal and norepinephrine-induced expressions of myosin light chain2 and the c-fos proto-oncogene in cardiomyocytes. Treatment with norepinephrine induced cardiac hypertrophy accompanied by G(h) expression and membrane translocation. Rosuvastatin inhibited G(h) protein activity in cardiomyocytes by inhibiting basal and norepinephrine-stimulated mRNA transcription, protein expression and membrane translocation; however, norepinephrine-stimulated G(q) protein expression was not inhibited. In addition, the norepinephrine-stimulated protein kinase C (PKC)-mitogen-activated protein kinase (MEK 1,2)-extracellular signal-regulated kinases (ERKs) signaling cascade was inhibited by pretreatment with rosuvastatin. Rosuvastatin treatment also helped maintain expression levels of SERCA2a and intracellular calcium concentration. G(h) protein is a novel target of statins in myocardial hypertrophy, and statin treatment may directly suppress cardiac hypertrophy through a large G(h) protein-coupled pathway regardless of the regulation of small GTPases.

Introduction Hyperglycemia is a risk factor for cardiovascular complications in diabetic state. Hyperglycemia-induced oxidative stress up-regulates intracellular adhesion molecule-1 (ICAM-1) which aggravates endothelial dysfunction, although the underlying mechanisms remain unclear. We hypothesized that carbon monoxide (CO) attenuates ICAM-1 expression induced by high glucose in endothelial cells through activation of AMP-activated protein kinase (AMPK)/peroxisome proliferator-activated receptor gamma (PPAR-γ) pathway. Methods Human umbilical vein endothelial cells (HUVEC) were pre-treated with CO releasing molecule-2 (CORM-2) alone or in combination with troglitazone or GW1929, PPAR-γ agonists or GW9662, PPAR-γ antagonist and then cells were co-treated with high glucose (25 mM) for 48 h for detection of ICAM-1 expression by Western blot or luciferase assay. The involvement of AMPK on PPAR-γ and ICAM-1 expressions was tested using pharmacological inducer or inhibitor, as well as transient transfection with AMPK-DN vector. Results CO derived from CORM-2 down-regulated ICAM-1 expression induced by high glucose. CORM-2 induced the activity of PPAR-γ at 24 h, and AMPK from 5 min to 3 h. PPAR-γ agonists significantly suppressed ICAM-1 expression, whereas in the presence of antagonist (GW9662) CORM-2 failed to inhibit ICAM-1. Thus inhibition of ICAM-1 was dependent on activation of PPAR-γ. Transfection with AMPK-DN or AMPK inhibitor resulted in attenuation of inducible effect of CORM-2 on PPAR-γ and subsequently suppressive effect on ICAM-1 expression. Conclusion Our results indicate that PPAR-γ and AMPK pathways activated by CO are required for attenuation of ICAM-1 expression induced by high glucose. Thus, this study highlights a new property for CO derived from CORM-2 as anti-atherogenic drug for diabetic patients.

The aim of the present study was to evaluate the protective effect of palmatine, one of active ingredients of Coptidis rhizoma, against myocardial ischemia-reperfusion (I/R) injury is due to its antioxidant and anti-inflammatory action. Adult male rats were subjected to 30 min of ischemia and 6 or 24h of reperfusion. Rats were randomized to receive vehicle or palmatine 1h before reperfusion. Infarct size, myocardial function, and the antioxidant enzyme activity, such as malonaldehyde (MDA), lactate dehydrogenase (LDH), creatine phosphokinase (CK), superoxide dismutase (SOD) and catalase (CAT) were measured. Palmatine significantly improved I/R-induced myocardial dysfunction by increasing the values of the first derivative (+/-dp/dt) of left ventricular pressure and decreased infarct size by 50% (P

Maximal activity of the immobilized d -psicose 3-epimerase from Agrobacterium tumefaciens on Duolite A568 beads was achieved at pH 9.0 and 55 °C with borate, and at pH 8.5 and 50 °C without borate. The half-lives of the immobilized enzyme at 50 °C with and without borate were increased 4.2- and 128-fold compared to that of the free enzyme without borate, respectively. The immobilized enzyme with borate produced 441 g l−1 psicose from 700 g l−1 fructose at pH 9.0 and 55 °C, whereas 193 g l−1 psicose was produced without borate at pH 8.5 and 50 °C after 120 min in a batch reaction. The immobilized enzyme in a packed-bed bioreactor without borate was produced continuously 325 g l−1 psicose from 500 g l−1 fructose at a dilution rate of 1.62 h−1 over a 236 h period with productivity of 527 g l−1 h−1 while that without borate produced 146 g l−1 psicose at 4.15 h−1 over a 384-h period with productivity of 606 g l−1 h−1. The operational half-lives of the enzyme with and without borate in the bioreactor were 601 and 645 h, respectively. In the present study, psicose was produced stably with high productivity using the immobilized d -psicose 3-epimerase in the presence of borate.

We investigated pharmacological effects of rutin isolated form Lonicera japonica on H2O2-induced cell death in H9c2 cells in vitro and rat myocardial ischemia-reperfusion injury model in vivo. Western blot analysis showed that H2O2 increased expression of cleaved form of caspase-3 and proapoptotic Bax protein, but decreased antiapoptotic Bcl-2 protein in H9c2 cell. However, treatment with rutin decreased expression of both cleaved from of caspase-3 and increased Bcl-2/Bax ratio in H9c2 cells. The protective effect of rutin was inhibited not by JNK inhibitor or p38 MAPK inhibitor but by PI3K inhibitor or ERK inhibitor. Rutin increased phosphorylation of ERK and Akt in H9c2 cells. These anti-apoptotic effects of rutin were confirmed both by annexin-V and TUNEL assay. Furthermore, rutin improved I/R-induced myocardial contractile function and reduced infarct size. Rutin administration also inhibited apoptosis in myocardial tissues in I/R rats by increasing Bcl-2/bax ratio and decreasing active caspase-3 expression. These results suggest that rutin reduced oxidative stress-mediated myocardial damage in vitro model and in vivo model, which might be useful in treatment of myocardial infarction.

We investigated the molecular mechanism by which ascorbic acid (AA) induces apoptosis in human gastric cancer cells, AGS cells. High concentration (more than 5mM) of AA increased cellular iron uptake by increasing transferrin receptor (TfR) expression and induced AGS cell apoptosis which was inhibited by catalase. Interestingly, p38 mitogen-activated protein kinase (MAPK) inhibitor inhibited the upregulation of TfR and increased cell survival by AA. TfR-siRNA-transfected cells reduced apoptosis by AA. H(2)O(2) increased TfR expression in AGS cells. Taken together, we concluded that high concentration of AA, through H(2)O(2), induces apoptosis of AGS cells by p38-MAPK-dependent upregulation of TfR.

Yeast-based functional screening for inhibitors of Bcl-2-associated X protein (Bax)-induced cell death in yeast identified ADP-ribosylation factor 4 (ARF4) as a novel anti-apoptotic gene in human glioblastoma-derived U373MG cells. Yeast or U373MG cells that overexpressed ARF4 exhibited reduced reactive oxygen species (ROS) generation in response to Bax or N-(4-hydroxyphenyl)retinamide (4-HPR), respectively, which suggests that ROS play a role in the inhibition of cell death by ARF4. The 4-HPR-mediated phosphorylation of c-JUN N-terminal kinase, p38, and extracellular signal-regulated kinase was markedly suppressed in U373MG cells that stably expressed ARF4. Stable ARF4 transfectants were also refractory to 4-HPR-induced mitochondrial translocation of Bax, release of mitochondrial cytochrome c, and activation of caspase-3. Our results suggest that ARF4 participates in the regulation of glioblastoma apoptosis through the inhibition of stress-mediated apoptotic signals.

We tested the hypothesis that daidzein may reduce myocardial damage by both inhibiting the release of cytokines and limiting the nuclear translocation of NF-kappaB.Male Sprague-Dawley rats were anesthetized, and the left anterior descending coronary artery (LAD) was ligated for 25 min. Twenty-four hours after reperfusion was established, the hemodynamics and infarct size were examined.Treatment with daidzein (10 mg/kg, i.p.) 1 h prior to the ischemia/reperfusion procedure (I/R) reduced the infarct size by 52.8% (P0.05). Daidzein also significantly improved I/R-induced myocardial contractile dysfunction by improving the left ventricular diastolic pressure and the positive and negative maximal values of the first derivative of the left ventricular pressure. In addition, daidzein reduced the plasma levels of TNF-alpha and IL-6 in I/R rats and decreased malondialdehyde levels, myeloperoxidase activity, catalase activity and neutrophil infiltration in I/R rat myocardium. Interestingly, daidzein inhibited I/R-induced myocardial apoptosis by decreasing DNA strand breaks and cleaved caspase-3 activity. Furthermore, daidzein inhibited both the nuclear translocation of NF-kappaB in I/R rat hearts and the H(2)O(2)-induced activation of NF-kappaB-luciferase activity in human umbilical vein endothelial cells.This study reveals that the administration of daidzein in vivo attenuates I/R-induced myocardial damage via inhibition of NF-kappaB activation, which in turn may suppress inflammatory cytokine expression.

Recently we proposed that COX-2 induction precedes expression of HO-1 in ischemic preconditioned rat brain. In the current study, we investigated the molecular mechanism by which prostaglandin E 2 , one of COX-2 metabolites, induces HO-1 in rat C6 brain cells. We demonstrated that concentration of PGE 2 increased HO-1 expression in C6 cells in vitro . The effects of PGE 2 were mimicked by PGE 2 receptor EP 2 agonists, 11-deoxy PGE 2 , and cAMP analog, dibutyl-cAMP. HO-1 expression by PGE 2 was inhibited by LY294002, PI3K inhibitor and H89, PKA inhibitor. The EP 2 -specific antagonist, AH8006 also inhibited PGE 2 -mediated HO-1 expression in a concentration-dependent manner. Finally, PGE 2 inhibited GOX-induced apoptosis as assayed by FACS analysis or DNA strand breaks assay, and this cell death was reversed by ZnPPIX, HO-1 inhibitor. In addition to HO-1 induction, PGE 2 also increased phosphorylation of Bad by PKA- and PI3K-depednent manner. Taken together, we conclude that PGE 2 induces HO-1 protein expression through PKA and PI3K signaling pathways via EP 2 receptor in C6 cells. The induction of HO-1 along with increase of p-Bad by PGE 2 is responsible for anti-apoptosis against oxidant stress.

Propofol, a rapidly acting, short duration, intravenous hypnotic anesthetic induction agent, is often used in clinical situations where myocardial ischemia/ reperfusion (I/R) injury is a threat. The aim of the present study was to evaluate the protective effect of propofol on myocardial I/R injury in rat due to apoptosis. Myocardial I/R injury were induced by occluding the left anterior descending (LAD) coronary artery for 25 min followed by either 2 h or 6 h reperfusion. Apoptosis was evaluated by Western blot analysis (Bcl-2, Bax expression), DNA strand breaks, TUNEL analysis and measuring myocardial caspase-3 activity. Propofol significantly reduced infarct size and improved I/R-induced myocardial contractile dysfunction by improving left ventricular diastolic pressure and positive and negative maximal values of the first derivative (+dp/dt) of left ventricular pressure. Propofol increased Bcl-2/Bax expression ratio and decreased caspase-3 activity in I/R rat hearts, which resulted in reduction of myocardial apoptosis as evidenced by TUNEL analysis and DNA laddering experiments. In an in vitro study, propofol increased H9c2 cell viability against oxidative stress induced by glucose oxidase (GOX) in a dose-dependent manner. These data suggest propofol limits I/R injury with an associated reduction in apoptotic cell death in vivo.

Percutaneous transluminal coronary angioplasty (PTCA) is a common procedure for treating atherosclerosis, but its efficacy is limited because of the occurrence of restenosis within 3-6 months after angioplasty. Restenosis is induced by the remodeling of the vessel wall and/or the accumulation of cells and extracellular matrix (ECM) in the intimal layer. Therefore, the matrix metalloproteinase (MMP) system may be a potential therapeutic target for the treatment of restenosis or atherosclerosis. Cordycepin is reported to possess many pharmacological activities including immunological stimulating, anti-cancer, antioxidant, and anti-inflammatory activities. The effect of cordycepin on restenosis has not yet been clearly elucidated. Therefore, in the present study, we tested the role of cordycepin on the MMP system in vascular smooth muscle cells. In the carotid artery of a balloon-injured Sprague-Dawley (SD) rat, neointimal formation was reduced by treatment with cordycepin (20 microM/day, i.p), which inhibited the proliferation of rat aortic smooth muscle cells (RaoSMCs). To investigate the mechanism by which cordycepin inhibits the remodeling of the vessel wall and/or the accumulation of cells and ECM, we examined the activation of MMP systems in collagen type I-activated RaoSMCs. Cordycepin markedly inhibited the activation of MMP-2 and -9 as well as the expression of extracellular matrix metalloproteinase inducer (EMMPRIN) in a dose-dependent manner in collagen type I-activated RaoSMCs. Moreover, cordycepin suppressed cycloxygenase-2 (COX-2) expression related to hyperplasia of RAoSMCs. Taken together, these data suggest that cordycepin may induce antiproliferation in RAoSMCs via the modulation of vessel wall remodeling. Therefore, cordycepin may be a potential therapeutic approach to treat restenosis.

Bisacurone, one of the active compounds of the traditionally used indigenous herb Curcuma longa Linne (Zingiberaceae), has anti-oxidant, anti-inflammatory, and anti-metastatic activities. We studied how the level of vascular cell adhesion molecule-1 (VCAM-1), one of the key molecules in the development of atherosclerosis as well as carcinogenesis and metastasis, might be affected by bisacurone in tumor necrosis factor-alpha (TNF-alpha)-activated human umbilical vein endothelial cells (HUVECs). Bisacurone dose-dependently inhibited TNF-alpha-mediated expression of VCAM-1. It showed significant suppressive effect on ROS generation in response to TNF-alpha stimulation and it blocked nuclear factor-kappa B (NF-kappaB) p65 translocation into the nucleus and phosphorylation of inhibitory factor kappaBalpha (IkappaBalpha). It also inhibited phosphorylation of Akt and PKC, which are upstream in the regulation of VCAM-1 by TNF-alpha. Furthermore, bisacurone decreased U937 monocyte and human oral cancer cell (Hep-2, QLL-I, SCC-15) adhesion to HUVECs stimulated by TNF-alpha, suggesting that it may inhibit the binding of these cells by regulating the expression of critical adhesion molecules by TNF-alpha. Thus, bisacurone may be beneficial in the treatment of inflammatory diseases, such as atherosclerosis, where inflammatory monocytes are involved in their pathology, and, moreover, in the development of tumors.

The osmotic response element (ORE) differs from the nuclear factor-kappaB (NF-kappaB) binding sequence by a single base pair; therefore, we investigated the involvement of NF-kappaB in the induction of aldose reductase (AR) by curcumin. Curcumin, an herb-derived polyphenolic compound, elicited an increase in the expression and promoter activity of the AR gene in a nuclear factor-erythroid 2-related factor 2 (Nrf2)-dependent manner. Small interfering RNA (siRNA) against p65 or BAY11-7082, an inhibitor of NF-kappaB, significantly suppressed the curcumin and/or Nrf2-induced increase in expression levels and promoter activity of the AR gene. BAY11-7082 or siRNA against p65 also attenuated the curcumin-induced increase in the promoter activity of the wild type AR-ORE(wt) gene, but not that of the mutated AR-ORE(mt), indicating that the ORE is essential for the response to NF-kappaB. The expression of p65, the promoter activity and DNA binding activity of NF-kappaB were enhanced in the presence of curcumin in cells that were transfected with Nrf2 compared to those treated with curcumin alone. Cells that had been preincubated with curcumin demonstrated resistance to reactive oxygen species-induced cell damage through the suppressive effects in the generation of reactive aldehydes. These effects were significantly attenuated in the presence of BAY11-7082, indicating the involvement of NF-kappaB in the cellular response of AR to oxidative stress and toxic aldehydes.

Overexpression of the gene for heme oxygenase (HO)-1 leads to a reduction in pressor responsiveness to angiotensin II (Ang II) in experimental animals. Using rat vascular smooth muscle cells (VSMCs), we tested whether YS 49 [1-(α-naphtylmethyl)-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline] inhibits Ang II-stimulated proliferation of VSMCs via induction of HO-1. YS 49 induced HO-1 protein production in a dose-and time-dependent manner in VSMCs. Treatment with YS 49 significantly and dose-dependently inhibited Ang II-induced VSMC proliferation, ROS production, and phosphorylation of JNK, but not P38 MAP kinase or ERK1/2. The antiproliferation effect of YS 49 was reversed by pretreatment with the HO-1 inhibitor zinc protoporphyrin IX (ZnPPIX), or with hemoglobin, a carbon monoxide (CO) scavenger. Similarly, VSMC proliferation, ROS production and phosphorylation of JNK by Ang II were significantly inhibited in VSMCs transfected with the HO-1 gene. Thus, HO-1 and the HO-1 product CO play, at least in part, a crucial role in Ang II-stimulated VSMC proliferation through the regulation of ROS production and JNK phosphorylation. Therefore, YS 49 has potential as a therapeutic strategy for the pathogenesis of Ang II-related vascular diseases such as hypertension and atherosclerosis, via the induction of HO-1 gene activity.