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.     

A Novel Model of Intravital Platelet Imaging Using CD41-ZsGreen1 Transgenic Rats

Platelets play pivotal roles in both hemostasis and thrombosis. Although models of intravital platelet imaging are available for thrombosis studies in mice, few are available for rat studies. The present effort aimed to generate fluorescent platelets in rats and assess their dynamics in a rat model of arterial injury. We generated CD41-ZsGreen1 transgenic rats, in which green fluorescence protein ZsGreen1 was expressed specifically in megakaryocytes and thus platelets. The transgenic rats exhibited normal hematological and biochemical values with the exception of body weight and erythroid parameters, which were slightly lower than those of wild-type rats. Platelet aggregation, induced by 20 μM ADP and 10 μg/ml collagen, and blood clotting times were not significantly different between transgenic and wild-type rats. Saphenous arteries of transgenic rats were injured with 10% FeCl₃, and the formation of fluorescent thrombi was evaluated using confocal microscopy. FeCl₃ caused time-dependent increases in the mean fluorescence intensity of injured arteries of vehicle-treated rats. Prasugrel (3 mg/kg, p.o.), administered 2 h before FeCl₃, significantly inhibited fluorescence compared with vehicle-treated rats (4.5 ± 0.4 vs. 14.9 ± 2.4 arbitrary fluorescence units at 30 min, respectively, n = 8, P = 0.0037). These data indicate that CD41-ZsGreen1 transgenic rats represent a useful model for intravital imaging of platelet-mediated thrombus formation and the evaluation of antithrombotic agents.     

A new NO ledge in Chlamydomonas: when the old nitrate reductase meets amidoxime reducing component to produce nitric oxide

This article comments on: A dual system formed by the ARC and NR molybdoenzymes mediates nitrite‐dependent NO production in Chlamydomonas     

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.     

Crystal structure of a hypothetical protein,TTHA0829 from <Emphasis Type="Italic">Thermus thermophilus</Emphasis> HB8, composed of cystathionine-β-synthase (CBS) and aspartate-kinase chorismate-mutase tyrA (ACT) domains

TTHA0829 from Thermus thermophilus HB8 has a molecular mass of 22,754 Da and is composed of 210 amino acid residues. The expression of TTHA0829 is remarkably elevated in the latter half of logarithmic growth phase. TTHA0829 can form either a tetrameric or dimeric structure, and main-chain folding provides an N-terminal cystathionine-β-synthase (CBS) domain and a C-terminal aspartate-kinase chorismate-mutase tyrA (ACT) domain. Both CBS and ACT are regulatory domains to which a small ligand molecule can bind. The CBS domain is found in proteins from organisms belonging to all kingdoms and is observed frequently as two or four tandem copies. This domain is considered as a small intracellular module with a regulatory function and is typically found adjacent to the active (or functional) site of several enzymes and integral membrane proteins. The ACT domain comprises four β-strands and two α-helices in a βαββαβ motif typical of intracellular small molecule binding domains that help control metabolism, solute transport and signal transduction. We discuss the possible role of TTHA0829 based on its structure and expression pattern. The results imply that TTHA0829 acts as a cell-stress sensor or a metabolite acceptor.     

LC–MS/MS-based metabolome analysis detected changes in the metabolic profiles of small and large intestinal adenomatous polyps in <Emphasis Type="Italic">Apc</Emphasis><Superscript><Emphasis Type="Italic">Min</Emphasis>/+</Superscript> mice

Introduction

The adenomatous polyposis coli (APC) gene is a tumor suppressor gene that is inactivated in the initiation of colorectal neoplasia. Apc ^Min/+ mice, which possess a heterozygous APC mutation, develop numerous adenomatous polyps, which are similar to those observed in familial adenomatous polyposis (FAP) in humans. However, unlike FAP patients, Apc ^Min/+ mice predominantly develop adenomatous polyps in the small intestine. The metabolic changes associated with the development of polyps in the small and large intestine remain to be investigated.

Objectives

The objective of this study was to elucidate the metabolic changes associated with intestinal polyp formation.

Methods

We compared the metabolite levels of pairs of polyp and non-polyp tissues obtained from the small intestines (n = 12) or large intestines (n = 7) of Apc ^Min/+ mice. To do this, we analyzed the tissue samples using two methods, liquid chromatography-tandem mass spectrometry (1) with a pentafluorophenylpropyl column for cation analysis, and (2) with a C18 reversed phase column coupled to an ion-pair reagent for anion analysis.

Results

Pathway mapping of the metabolites whose levels were significantly altered revealed that the polyp tissue of the small intestine contained significantly higher levels of intermediates involved in glycolysis, the pentose phosphate pathway, nucleotide metabolism, or glutathione biosynthesis than in the equivalent non-polyp tissue. In addition, significantly higher levels of methionine cycle intermediates were detected in the polyp tissues of both the large and small intestines. Organ-dependent (small vs. large intestine) differences were also detected in the levels of most amino acids and urea cycle intermediates.

Conclusion

Our results indicate that various metabolic changes are associated with polyp development, and understanding these alterations could make it possible to evaluate the treatment response of colorectal cancer earlier.

     

GENE SILENCING BY PARENTAL RNA INTERFERENCE IN THE GREEN RICE LEAFHOPPER,Nephotettix cincticeps (HEMIPTERA: CICADELLIDAE)

RNA interference (RNAi) has been widely used for investigating gene function in many nonmodel insect species. Parental RNAi causes gene knockdown in the next generation through the administration of double‐strand RNA (dsRNA) to the mother generation. In this study, we demonstrate that parental RNAi mediated gene silencing is effective in determining the gene function of the cuticle and the salivary glands in green rice leafhopper (GRH), Nephotettix cincticeps (Uhler). Injection of dsRNA of NcLac2 (9 ng/female) to female parents caused a strong knockdown of laccase‐2 gene of first instar nymphs, which eventually led to high mortality rates and depigmentation of side lines on the body. The effects of parental RNAi on the mortality of the nymphs were maintained through 12–14 days after the injections. We also confirmed the effectiveness of parental RNAi induced silencing on the gene expressed in the salivary gland, the gene product of which is passed from instar to instar. The parental RNAi method can be used to examine gene function by phenotyping many offspring nymphs with injection of dsRNA into a small number of parent females, and may be applicable to high‐efficiency determination of gene functions in this species.