BJ-1108, a 6-Amino-2,4,5-Trimethylpyridin-3-ol Analog,Inhibits Serotonin-Induced Angiogenesis and Tumor Growth through PI3K/NOX Pathway

5-Hydroxytryptamine (5-HT) induces proliferation of cancer cells and vascular cells. In addition to 5-HT production by several cancer cells including gastrointestinal and breast cancer, a significant level of 5-HT is released from activated platelets in the thrombotic environment of tumors, suggesting that inhibition of 5-HT signaling may constitute a new target for antiangiogenic anticancer drug discovery. In the current study we clearly demonstrate that 5-HT-induced angiogenesis was mediated through the 5-HT₁ receptor-linked Gβγ/Src/PI3K pathway, but not through the MAPK/ERK/p38 pathway. In addition, 5-HT induced production of NADPH oxidase (NOX)-derived reactive oxygen species (ROS). In an effort to develop new molecularly targeted anticancer agents against 5-HT action in tumor growth, we demonstrate that BJ-1108, a derivative of 6-amino-2,4,5-trimethylpyridin-3-ol, significantly inhibited 5-HT-induced angiogenesis. In addition, BJ-1108 induced a significant reduction in the size and weight of excised tumors in breast cancer cell-inoculated CAM assay, showing proportionate suppression of tumor growth along with inhibition of angiogenesis. In human umbilical vein endothelial cells (HUVECs), BJ-1108 significantly suppressed 5-HT-induced ROS generation and phosphorylation of PI3K/Akt but not of Src. Unlike NOX inhibitors, BJ-1108, which showed better antioxidant activity than vitamin C, barely suppressed superoxide anion induced by mevalonate or geranylgeranyl pyrophosphate which directly activates NOX without help from other signaling molecules in HUVECs, implying that the anti-angiogenic action of BJ-1108 was not mediated through direct action on NOX activation, or free radical scavenging activity. In conclusion, BJ-1108 inhibited 5-HT-induced angiogenesis through PI3K/NOX signaling but not through Src, ERK, or p38.     

The growth kinetics and metabolic and antioxidant activities of the functional synbiotic combination of <Emphasis Type="Italic">Lactobacillus gasseri</Emphasis> 505 and <Emphasis Type="Italic">Cudrania tricuspidata</Emphasis> leaf extract

In a previous study, the synbiotic combination of selected Lactobacillus gasseri strains and Cudrania tricuspidata leaf extract (CT) was shown to significantly improve the functionality of fermented milk, and the greatest synbiotic effect was exhibited with L. gasseri 505. The aim of the present study was to investigate the growth kinetics and fermentation metabolism of this specific synbiotic combination. Fermentation was carried out in synthetic media and milk with or without CT supplementation using L. gasseri 505. Whole genome sequencing and comparative genomics analyses were conducted to verify the novelty of strain. Titratable acidity, pH, microbial population, and organic acid production were measured during the fermentation period. The addition of CT accelerated the acidification rate, supporting the growth of L. gasseri 505, and the production of fermentation metabolites such as lactic acid and pyruvic acid also significantly increased during fermentation of both of CT-supplemented synthetic media and milk. In particular, the formic acid and propionic acid in CT were significantly utilized during fermentation of milk by L. gasseri 505. Moreover, the antioxidant capacity of CT-supplemented fermented milk increased due to the release of bioactive compounds until the exponential growth phase, after which the antioxidant activity declined due to degradation and loss of potency. Therefore, this study established that L. gasseri 505 efficiently utilized the CT-related nutrients during fermentation producing resulting metabolites with health-promoting effects, although it is necessary to control the fermentation time to obtain dairy products with optimum functionality.     

Natural triploid Lilium leichtlinii var. maximowiczii populations in Korea

To clarify the ploidy and distribution of Lilium leichtlinii var. maximowiczii populations in Korea, we inspected several mountains and northern valleys and almost all of the sea coast of South Korea. We found nine diploid populations and 19 triploid populations. Many large and small populations were distributed around northern Chungcheongbook‐do and Gyunggi‐do and all over Kangwon‐do. The diploid plants were smaller than the triploid plants in almost all of their morphological characteristics. This is the second report of natural polyploid complexes of diploids and triploids in the genus Lilium.     

The first report on the characterization of a virus isolated from Allomyrina dichotoma in the Republic of Korea

This report reveals the structure of a virus extracted from the Korean horn beetle Allomyrina dichotoma. The purified virus particle was 100% identical to Allomyrina virus lef‐8 sequence registered as KM_233709.1. The structure of this virus was confirmed to be closely related to that of the Nudiviridae family, and it was rod shaped and enveloped, and observed to be of approximately the mean length of a single viral nucleocapsid of 200–210 nm and mean diameter of 100–110 nm. These results provide an insight into the structural characteristics of the Nudiviridae family that can be used for nudiviral identification.     

Spectral and structural analysis of a red fluorescent protein from Acropora digitifera

Fluorescent proteins (FPs) possess a wide variety of spectral properties that make them of widespread interest as optical markers. These proteins can be applied as pH indicators or metal biosensors. The discovery and characterization of new fluorescent proteins is expected to further extend their application. Here, we report the spectral and structural analysis of a red fluorescent protein from Acropora digitifera (designated AdRed). This protein shows a tetrameric state and is red emitting, with excitation and emission maxima at 567 and 612 nm, respectively. Its crystal structure shows the tetrameric interface stabilized by hydrogen bonding and salt bridges. The electron density map of the chromophore, consisting of Asp66–Tyr67–Gly68, shows the decarboxylated side chain of Asp66. Ser223, located near the chromophore, has the role of bridging His202 and Glu221, and is part of the hydrogen bond network. Mutated AdRed with Cys148Ser reveals a blue shift in fluorescence excitation and emission. Our results provide insights into understanding the molecular function of AdRed and other FPs.     

Acentriolar microtubule organization centers and Ran‐mediated microtubule formation pathways are both required in porcine oocytes

Mammalian oocytes lack centrioles but can generate bipolar spindles using several different mechanisms. For example, mouse oocytes have acentriolar microtubule organization centers (MTOCs) that contain many components of the centrosome, and which initiate microtubule polymerization. On the contrary, human oocytes lack MTOCs and the Ran‐mediated mechanisms may be responsible for spindle assembly. Complete knowledge of the different mechanisms of spindle assembly is lacking in various mammalian oocytes. In this study, we demonstrate that both MTOC‐ and Ran‐mediated microtubule nucleation are required for functional meiotic metaphase I spindle generation in porcine oocytes. Acentriolar MTOC components, including Cep192 and pericentrin, were absent in the germinal vesicle and germinal vesicle breakdown stages. However, they start to colocalize to the spindle microtubules, but are absent in the meiotic spindle poles. Knockdown of Cep192 or inhibition of Polo‐like kinase 1 activity impaired the recruitment of Cep192 and pericentrin to the spindles, impaired microtubule assembly, and decreased the polar body extrusion rate. When the Ran^GTP gradient was perturbed by the expression of dominant negative or constitutively active Ran mutants, severe defects in microtubule nucleation and cytokinesis were observed, and the localization of MTOC materials in the spindles was abolished. These results demonstrate that the stepwise involvement of MTOC‐ and Ran‐mediated microtubule assembly is crucial for the formation of meiotic spindles in porcine oocytes, indicating the diversity of spindle formation mechanisms among mammalian oocytes.     

Biological characterization of D-lactate dehydrogenase responsible for high-yield production of D-phenyllactic acid in Sporolactobacillus inulinus

PLA (3-D-phenyllactic acid) is an ideal antimicrobial and immune regulatory compound present in honey and fermented foods. Sporolactobacillus inulinus is regarded as a potent D-PLA producer that reduces phenylpyruvate (PPA) with D-lactate dehydrogenases. In this study, PLA was produced by whole-cell bioconversion of S. inulinus ATCC 15538. Three genes encoding D-lactate dehydrogenase (d-ldh1, d-ldh2, and d-ldh3) were cloned and expressed in Escherichia coli BL21 (DE3), and their biochemical and structural properties were characterized. Consequently, a high concentration of pure D-PLA (47 mM) was produced with a high conversion yield of 88%. Among the three enzymes, D-LDH1 was responsible for the efficient conversion of PPA to PLA with kinetic parameters of Km (0.36 mM), k_cat (481.10 s⁻¹), and k_cat/Km (1336.39 mM⁻¹ s⁻¹). In silico structural analysis and site-directed mutagenesis revealed that the Ile307 in D-LDH1 is a key residue for excellent PPA reduction with low steric hindrance at the substrate entrance. This study highlights that S. inulinus ATCC 15538 is an excellent PLA producer, equipped with a highly specific and efficient D-LDH1 enzyme.     

Exclusive developmental functions of gatae cis-regulatory modules in the Strongylocentrorus purpuratus embryo

The gatae gene of Strongylocentrotus purpuratus is orthologous to vertebrate gata-4,5,6 genes. This gene is expressed in the endomesoderm in the blastula and later the gut of the embryo, and is required for normal development. A gatae BAC containing a GFP reporter knocked into exon one of the gene was able to reproduce all aspects of endogenous gatae expression in the embryo. To identify putative gatae cis-regulatory modules we carried out an interspecific sequence conservation analysis with respect to a Lytechinus variegatus gatae BAC, which revealed 25 conserved non-coding sequence patches. These were individually tested in gene transfer experiments, and two modules capable of driving localized reporter expression in the embryo were identified. Module 10 produces early expression in mesoderm and endoderm cells up to the early gastrula stage, while module 24 generates late endodermal expression at gastrula and pluteus stages. Module 10 was then deleted from the gatae BAC by reciprocal recombination, resulting in total loss of reporter expression in the time frame in which it is normally active. Similar deletion of module 24 led to ubiquitous GFP expression in the gastrula and pluteus. These results show that Module 10 is uniquely necessary and sufficient to account for the early phase of gatae expression during endomesoderm specification. In addition, they imply a functional cis-regulatory module exclusion, whereby only a single module can associate with the basal promoter and drive gene expression at any given time.