Regulation of c-Myc Expression by Ahnak Promotes Induced Pluripotent Stem Cell Generation

We have previously reported that Ahnak-mediated TGFβ signaling leads to down-regulation of c-Myc expression. Here, we show that inhibition of Ahnak can promote generation of induced pluripotent stem cells (iPSC) via up-regulation of endogenous c-Myc. Consistent with the c-Myc inhibitory role of Ahnak, mouse embryonic fibroblasts from Ahnak-deficient mouse (Ahnak^−/− MEF) show an increased level of c-Myc expression compared with wild type MEF. Generation of iPSC with just three of the four Yamanaka factors, Oct4, Sox2, and Klf4 (hereafter 3F), was significantly enhanced in Ahnak^−/− MEF. Similar results were obtained when Ahnak-specific shRNA was applied to wild type MEF. Of note, expressionof Ahnak was significantly induced during the formation of embryoid bodies from embryonic stem cells, suggesting that Ahnak-mediated c-Myc inhibition is involved in embryoid body formation and the initial differentiation of pluripotent stem cells. The iPSC from 3F-infected Ahnak^−/− MEF cells (Ahnak^−/−-iPSC-3F) showed expression of all stem cell markers examined and the capability to form three primary germ layers. Moreover, injection of Ahnak^−/−-iPSC-3F into athymic nude mice led to development of teratoma containing tissues from all three primary germ layers, indicating that iPSC from Ahnak^−/− MEF are bona fide pluripotent stem cells. Taken together, these data provide evidence for a new role for Ahnak in cell fate determination during development and suggest that manipulation of Ahnak and the associated signaling pathway may provide a means to regulate iPSC generation.     

Synthesis of L-Sangivamycin and Toyocamycin Analogues and Their Inhibitory Activities of SER/THR Protein Kinases

Abstract

Novel _l-sangivamycin and toyocamycin analogues were synthesized and evaluated for Cdc2 protein kinase activity. Among the compounds tested, _l-xylose derivative and _l-arabinose derivative exhibited potent inhibitory activity against Cdc2 protein kinase with IC₅₀ values of 3.7 and 1.6 μM, respectively.     

Administration of vitamin E attenuates airway inflammation through restoration of Nrf2 in a mouse model of asthma

Accumulating evidence reveals that ROS is one of the key mediators that contribute to the development of asthma. Studies on antioxidants have shown to have beneficial effects on asthma management. However, we still do not know the precise mechanism, and the effects depend on age. This study was conducted to assess the levels of ROS and the effect of antioxidants in younger and older mice using an eosinophilic asthma model. We analyzed airway hyperresponsiveness (AHR), cytokines in bronchoalveolar lavage fluid (BALF), inflammatory cell counts, and the expression levels of NFκB, Nrf2, EPx, and EDN in the lung tissue, as well as the level of ROS in the lung tissue and BALF. The degree of eosinophilia and the levels of IL-5, ROS, and NFκB were significantly increased, whereas the endogenous levels of vitamin E and Nrf2 were decreased in the lung and BALF in the older mice compared to younger mice. The administration of vitamin E attenuated AHR, airway inflammation, and the level of IL-13 and ROS and enhanced the Nrf2 level in the older mice compared to the younger mice. Taken together, vitamin E treatment may have the therapeutic potential through restoration of the Nrf2 level, especially in elderly asthma.     

Characterization of tyrosinase inhibitory constituents from the aerial parts of Humulus japonicus using LC-MS/MS coupled online assay

In the screening of natural products for the development as cosmetic ingredients, the EtOAc-soluble fraction of Humulus japonicus showed tyrosinase inhibitory activity. HPLC-MS/MS coupled online tyrosinase assay of EtOAc-soluble fraction of H. japonicus characterized the twenty-eight constituents including two unknown ones and their tyrosinase inhibitory activity. Fractionation of H. japonicus using various chromatographic techniques yielded thirty-eight compounds. The chemical structures of isolated compounds were identified by spectroscopic analysis. As characterized by HPLC-MS/MS analysis, we isolated twenty-four predicted compounds and further identified two unknown ones, named humulusides A (1) and B (2). Additional ten compounds were also identified by purification. Tyrosinase inhibitory activity of isolated compounds were evaluated, which was closely correlated with the results from HPLC-MS/MS coupled online tyrosinase assay. Consistent with predicted data, two major compounds, trans-N-coumaroyltyramine (14) and cis-N-coumaroyltyramine (15) showed tyrosinase inhibition with IC₅₀ values of 40.6 and 36.4?μM. Taken together, H. japonicus is suggested as whitening ingredient in cosmetic products. In addition, HPLC-MS/MS coupled tyrosinase assay is powerful tool for predicting active compounds with short time and limited amounts, although identification of new compounds and verification of predicted data are also needs to be demonstrated by further experiment.     

Enhancement of Growth Characteristics and Biological Activities in <Emphasis Type="Italic">Astragalus membranaceus</Emphasis> Using Artificial Light Sources

We investigated the effects of light emitting plasma (LEP) and red, blue, green, and white light emitting diode (LED) artificial light sources on plant growth characteristics and antioxidant activities in Astragalus membranaceus B. The highest germination rate (91.33 ± 2.08%) was produced by growth under red LED light. Root growth under LEP light was superior to that under the other light treatments. We measured antioxidant activity using a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, and determined the total phenol and flavonoid contents and tyrosinase inhibitory activity in response to the various light treatments. Antioxidant activities were 53.6% as the highest level of activity under LEP light. Growth under LEP light also produced the highest total phenolic and flavonoid contents of 36.05 and 5.94 mg/mL, respectively. Extracts from plants grown under LEP light caused the highest inhibition of tyrosinase activity with inhibition of 35.37, 61.87 and 65.49%, respectively, for extract concentrations of 100, 500, and 1000 μg/mL compared with other artificial light treatments. These results suggest that LEP light may be an important artificial light source for the enhancement of plant growth and biological activity and indicate the importance of further study of the use of LEP light sources for agricultural production.     

The effect of storage temperature on the biological activity of extracellular vesicles for the complement system

Extracellular vesicles (EVs) are mediators of intercellular communication by transporting cargo containing proteins, lipids, mRNA, and miRNA. There is increasing evidence that EVs have various roles in regulating migration, invasion, stemness, survival, and immune functions. Previously, we have found that EVs from Kaposi’s sarcoma-associated herpesvirus (KSHV)-infected human endothelial cells have the potential to activate the complement system. Although many studies have shown that the physical properties of EVs can be changed by their storage condition, there have been few studies for the stability of biological activity of EVs in various storage conditions. In this study, we investigated various conditions to identify the best conditions to store EVs with functional stability for 25 d. Furthermore, the correlation between the function and other characteristics of EVs, including the expression of EV markers, size distribution, and particle number, were also analyzed. Our results demonstrated that storage temperature is an important factor to maintain the activity of EVs and would be useful information for basic research and clinical application using EVs.     

High-fat diet-induced lipidome perturbations in the cortex,hippocampus, hypothalamus,and olfactory bulb of mice

Given their important role in neuronal function, there has been an increasing focus on altered lipid levels in brain disorders. The effect of a high-fat (HF) diet on the lipid profiles of the cortex, hippocampus, hypothalamus, and olfactory bulb of the mouse brain was investigated using nanoflow ultrahigh pressure liquid chromatography-electrospray ionization-tandem mass spectrometry in the current study. For 8?weeks, two groups of 5-week-old mice were fed either an HF or normal diet (6 mice from each group analyzed as the F and N groups, respectively). The remaining mice in both groups then received a 4-week normal diet. Each group was then subdivided into two groups for another 4-week HF or normal diet. Quantitative analysis of 270 of the 359 lipids identified from brain tissue revealed that an HF diet significantly affected the brain lipidome in all brain regions that were analyzed. The HF diet significantly increased diacylglycerols, which play a role in insulin resistance in all regions that were analyzed. Although the HF diet increased most lipid species, the majority of phosphatidylserine species were decreased, while lysophosphatidylserine species, with the same acyl chain, were substantially increased. This result can be attributed to increased oxidative stress due to the HF diet. Further, weight-cycling (yo-yo effect) was found more critical for the perturbation of brain lipid profiles than weight gain without a preliminary experience of an HF diet. The present study reveals systematic alterations in brain lipid levels upon HF diet analyzed either by lipid class and molecular levels.     

Expression and characterization of glutamate decarboxylase from <Emphasis Type="Italic">Lactobacillus brevis</Emphasis> HYE1 isolated from <Emphasis Type="Italic">kimchi</Emphasis>

A putative gene (gadlbhye1) encoding glutamate decarboxylase (GAD) was cloned from Lactobacillus brevis HYE1 isolated from kimchi, a traditional Korean fermented vegetable. The amino acid sequences of GADLbHYE1 showed 48% homology with the GadA family and 99% identity with the GadB family from L. brevis. The cloned GADLbHYE1 was functionally expressed in Escherichia coli using inducible expression vectors. The expressed recombinant GADLbHYE1 was successfully purified by Ni–NTA affinity chromatography, and had a molecular mass of 54 kDa with optimal hydrolysis activity at 55 °C and pH 4.0. Its thermal stability was determined to be higher than that of other GADs from L. brevis, based on its melting temperature (75.18 °C). Kinetic parameters including K_m and V_max values for GADLbHYE1 were 4.99 mmol/L and 0.224 mmol/L/min, respectively. In addition, the production of gamma-aminobutyric acid in E. coli BL21 harboring gadlbhye1/pET28a was increased by adding pyridoxine as a cheaper coenzyme.     

Red and Blue Light Emitting Diodes (LEDs) Participate in Mitigation of Hyperhydricity in In Vitro-Grown Carnation Genotypes (<Emphasis Type="Italic">Dianthus Caryophyllus</Emphasis>)

The present study was to determine the factors that can reduce hyperhydricity in in vitro-propagated carnation genotypes. The carnation genotypes (Green Beauty, Purple Beauty, and Inca Magic) were grown in vitro under normal and hyperhydric conditions in white fluorescent light (FL) in which half of the hyperhydric plants were grown in red and blue LEDs (light emitting diodes). It was observed that hyperhydricity leads to oxidative stress in terms of TBARS (thiobarbituric acid reactive substances) content, whereas stress was alleviated by R (red) and B (blue) LEDs. The multiprotein complex proteins such as ATPase (RCI?+?LHC1) PSII-core dimer, PSII-monomer/ATPs synthase, and PSII-monomer/cyt b₆f had decreased levels in hyperhydric conditions grown in white FL; however, the expression level of these photosynthetic proteins was retained in hyperhydric plants grown in R and B LEDs. Moreover, the immunoblots of two photosynthetic proteins (PsaA and PsbA) and stress-responsive proteins such as superoxide dismutase, ascorbate peroxidase, and catalase showed recovery of hyperhydricity in carnation genotypes grown in R and B LEDs. Our present study signifies that red (R) and blue light (B) LEDs reduced the hyperhydricity to control levels by maintaining the composition of thylakoid proteins and antioxidative defense mechanisms in carnation genotypes.     

Semi-rational engineering of CYP153A35 to enhance ω-hydroxylation activity toward palmitic acid

Applied Microbiology and Biotechnology - CYP153A35 from Gordonia alkanivorans was recently characterized as fatty acid ω-hydroxylase. To enhance the catalytic activity of CYP153A35 toward...