Oral Administration of Euglena Gracilis Z Alleviates Constipation and Cardiac Dysfunction in a Mouse Model of Isoproterenol-Induced Heart Failure.

Background: Patients with heart failure (HF) often experience gastrointestinal problems such as constipation, diarrhea, and disturbances to drug absorption. In HF, hypoperfusion and congestion cause structural and functional changes in the gut, which, in turn, lead to impaired cardiac function. Euglena gracilis Z (hereafter "Euglena"), called Midorimushi in Japanese, is a microalga that is used as a food or nutritional supplement. It is unclear whether Euglena is beneficial for bowel habitus and cardiac function in subjects with HF. Methods and Results: We injected C57BL/6 male mice subcutaneously with isoproterenol (ISO) (20 mg/kg/day) for 7 days to examine bowel movement in HF. Euglena was orally administered to mice on an ad libitum-feeding to a normal chow containing 2% dietary mixture. ISO induced a decrease in bowel movement and an increase in fecal retention in the cecum, as well as a decrease in left ventricular (LV) contraction. Euglena accelerated intestinal transit, relieved fecal retention, and prevented the alterations in gut pathology in ISO-treated mice. Euglena also suppressed ISO-induced decreases in LV contraction, although it had no significant effect on LV hypertrophy. Conclusions: The results suggested that oral administration of Euglena alleviated constipation and cardiac dysfunction in a mouse model of ISO-induced HF, and highlight the potential clinical benefit of Euglena in patients with HF in preventing constipation and contractile deterioration.

Effects of orally administered Euglena gracilis and its reserve polysaccharide, paramylon, on gastric dysplasia in A4gnt knockout mice.

Euglena gracilis is widely utilized as food or supplement to promote human and animal health, as it contains rich nutrients. In this study, we administered spray-dried powder of E. gracilis and paramylon, ß-glucan stored in E. gracilis cells, to A4gnt knockout (KO) mice. A4gnt KO mice are a mutant mouse model that spontaneously develops gastric cancer through hyperplasia-dysplasia-adenocarcinoma sequence in the antrum of the stomach, and we observed the effects of E. gracilis and paramylon on the early involvements of A4gnt KO mice. Male and female 10-week-old A4gnt KO mice and their age-matched wildtype C57BL/6J mice were orally administered with 50 mg of E. gracilis or paramylon suspended in saline or saline as a control. After 3-week administration, animals were euthanatized and the stomach was examined histopathologically and immunohistochemically. Gene expression patterns of the stomach, which have been reported to be altered with A4gnt KO, and IgA concentration in small intestine were also analyzed with real-time PCR and ELISA, respectively. Administration of Euglena significantly reduced the number of stimulated CD3-positive T-lymphocytes in pyloric mucosa of A4gnt KO mice and tend to reduce polymorphonuclear leukocytes infiltration. Euglena administration further downregulated the expression of Il11 and Cxcl1 of A4gnt KO mice. Euglena administration also affected IgA concentration in small intestinal contents of A4gnt KO mice. Paramylon administration reduced the number of CD3-positive lymphocytes in pyloric mucosa of A4gnt KO mice, and downregulated the expressions of Il11 and Ccl2 of A4gnt KO mice. Although we found no significant effects on gross and microscopic signs of gastric dysplasia and cell proliferation, the present study suggests that the administration of Euglena and paramylon may ameliorate the early involvements of A4gnt mice through the effects on inflammatory reactions in the gastric mucosa. The cancer-preventing effects should be studied with long-term experiments until actual gastric cancer formation.

The alga Euglena gracilis stimulates Faecalibacterium in the gut and contributes to increased defecation.

The alga Euglena gracilis (E. gracilis) has recently gained attention as a health food, but its effects on human gut microbiota remain unknown. This study aimed to determine the effect of E. gracilis on gut microbiota and defecation due to modulation of microbiota composition in vitro and in vivo. The in vitro model simulating human colonic microbiota revealed that E. gracilis addition stimulated the growth of commensal Faecalibacterium. Further, E. gracilis addition enhanced butyrate production by Faecalibacterium prausnitzii. Paramylon, an insoluble dietary fibre that accumulates in E. gracilis and is the main component of E. gracilis, did not stimulate Faecalibacterium growth in vitro. Daily ingestion of 2 g of E. gracilis for 30 days increased bowel movement frequency as well as stool volume in 28 human participants. Collectively, these findings indicate that E. gracilis components other than paramylon, stimulate the growth of Faecalibacterium to improve digestive health as well as promote defecation by increasing butyrate production.

Effects of Euglena gracilis Intake on Mood and Autonomic Activity under Mental Workload, and Subjective Sleep Quality: A Randomized, Double-Blind, Placebo-Controlled Trial.

While the human body maintains homeostasis by altering the balance in the autonomic nervous, endocrine, and immune systems, a prolonged imbalance in these systems can result in physical and mental symptoms, including a decline in sleep quality and work efficiency. Euglena gracilis (Euglena) is a single-celled microalga with the properties of both plants and animals and contains abundant nutrients, such as vitamins, minerals, amino acids, and fatty acids, which have various beneficial health effects. This study evaluated the effects of Euglena intake on the mood states and stress coping under mental workload tasks, and subjective sleep quality. We assigned men and women aged 20 to 64 years to Euglena and placebo intake groups, and measured indices related to the autonomic nervous system, psychological states, and sleep quality together with the application of workload stress before food intake, and 4, 8, and 12 weeks after commencing intake. Euglena intake regulated the autonomic nervous system under a workload and improved psychological parameters and sleep conditions. These results indicate that the consumption of Euglena may regulate the balance of the autonomic nervous system during stress and may have a favorable effect on psychological status and sleep quality.

Euglena Gracilis and ß-Glucan Paramylon Induce Ca2+ Signaling in Intestinal Tract Epithelial, Immune, and Neural Cells.

The intestinal tract contains over half of all immune cells and peripheral nerves and manages the beneficial interactions between food compounds and the host. Paramylon is a ß-1,3-glucan storage polysaccharide from Euglena gracilis (Euglena) that exerts immunostimulatory activities by affecting cytokine production. This study investigated the signaling mechanisms that regulate the beneficial interactions between food compounds and the intestinal tract using cell type-specific calcium (Ca2+) imaging in vivo and in vitro. We successfully visualized Euglena- and paramylon-mediated Ca2+ signaling in vivo in intestinal epithelial cells from mice ubiquitously expressing the Yellow Cameleon 3.60 (YC3.60) Ca2+ biosensor. Moreover, in vivo Ca2+ imaging demonstrated that the intraperitoneal injection of both Euglena and paramylon stimulated dendritic cells (DCs) in Peyer's patches, indicating that paramylon is an active component of Euglena that affects the immune system. In addition, in vitro Ca2+ imaging in dorsal root ganglia indicated that Euglena, but not paramylon, triggers Ca2+ signaling in the sensory nervous system innervating the intestine. Thus, this study is the first to successfully visualize the direct effect of ß-1,3-glucan on DCs in vivo and will help elucidate the mechanisms via which Euglena and paramylon exert various effects in the intestinal tract.

Design, synthesis, and pharmacological evaluation of 2-(4-sulfonylphenyl)-2-[(E)-pyrrolidin-1-ylimino]-N-thiazoleacetamides as glucokinase activators.

This paper presents the synthesis and glucokinase activity of novel hydrazone derivatives. The 2-(4-cyclopropylsulfonylphenyl)-2-[(E)-pyrrolidin-1-ylimino]-acetamide derivatives 5a-5h presented the in vitro glucokinase activities and in vivo blood glucose-lowering effects in mice. Particularly, 5h showed an oral hypoglycemic effect in rats at 1 mg/kg. These hydrazone derivatives are a potential new class of glucokinase activators for the treatment of type 2 diabetes.

Accelerated Wound Healing on the Skin Using a Film Dressing with ß-Glucan Paramylon.

BACKGROUND/AIM: ß-Glucan has been shown to modulate immune system and potentially aid wound healing. A naturally-available ß-glucan, paramylon, is available in the form of a film, which would be an ideal form to use in wound care. The aim of this study was to examine the therapeutic efficacy of paramylon film as a wound dressing. MATERIALS AND METHODS: An acute wound was created on the skin of the posterior aspect of mice and wound healing was observed for 5 days. Mice were treated with either paramylon film or conventional cellulose film. RESULTS: The time course of changes in wound size revealed that paramylon film dressing application leads to significantly faster wound contraction than conventional cellulose film. The dressing suppressed elevation of the inflammatory cytokines interferon gamma, interleukin-6, and vascular endothelial growth factor. CONCLUSION: ß-Glucan paramylon film can facilitate wound healing by inhibiting inflammatory aggression and has potential application as a novel wound dressing.

Supernova: A Versatile Vector System for Single-Cell Labeling and Gene Function Studies in vivo.

Here we describe "Supernova" series of vector systems that enable single-cell labeling and labeled cell-specific gene manipulation, when introduced by in utero electroporation (IUE) or adeno-associated virus (AAV)-mediated gene delivery. In Supernova, sparse labeling relies on low TRE leakage. In a small population of cells with over-threshold leakage, initial tTA-independent weak expression is enhanced by tTA/TRE-positive feedback along with a site-specific recombination system (e.g., Cre/loxP, Flpe/FRT). Sparse and bright labeling by Supernova with little background enables the visualization of the morphological details of individual neurons in densely packed brain areas such as the cortex and hippocampus, both during development and in adulthood. Sparseness levels are adjustable. Labeled cell-specific gene knockout was accomplished by introducing Cre/loxP-based Supernova vectors into floxed mice. Furthermore, by combining with RNAi, TALEN, and CRISPR/Cas9 technologies, IUE-based Supernova achieved labeled cell-specific gene knockdown and editing/knockout without requiring genetically altered mice. Thus, Supernova system is highly extensible and widely applicable for single-cell analyses in complex organs, such as the mammalian brain.

Cells of a common developmental origin regulate REM/non-REM sleep and wakefulness in mice.

Mammalian sleep comprises rapid eye movement (REM) sleep and non-REM (NREM) sleep. To functionally isolate from the complex mixture of neurons populating the brainstem pons those involved in switching between REM and NREM sleep, we chemogenetically manipulated neurons of a specific embryonic cell lineage in mice. We identified excitatory glutamatergic neurons that inhibit REM sleep and promote NREM sleep. These neurons shared a common developmental origin with neurons promoting wakefulness; both derived from a pool of proneural hindbrain cells expressing Atoh1 at embryonic day 10.5. We also identified inhibitory γ-aminobutyric acid-releasing neurons that act downstream to inhibit REM sleep. Artificial reduction or prolongation of REM sleep in turn affected slow-wave activity during subsequent NREM sleep, implicating REM sleep in the regulation of NREM sleep.

Developmental RacGAP α2-Chimaerin Signaling Is a Determinant of the Morphological Features of Dendritic Spines in Adulthood.

Morphological characteristics of dendritic spines form the basis of cognitive ability. However, molecular mechanisms involved in fine-tuning of spine morphology during development are not fully understood. Moreover, it is unclear whether, and to what extent, these developmental mechanisms determine the normal adult spine morphological features. Here, we provide evidence that α2-isoform of Rac-specific GTPase-activating protein α-chimaerin (α2-chimaerin) is involved in spine morphological refinement during late postnatal period, and furthermore show that this developmental α2-chimaerin function affects adult spine morphologies. We used a series of mice with global and conditional knock-out of α-chimaerin isoforms (α1-chimaerin and α2-chimaerin). α2-Chimaerin disruption, but not α1-chimaerin disruption, in the mouse results in an increased size (and density) of spines in the hippocampus. In contrast, overexpression of α2-chimaerin in developing hippocampal neurons induces a decrease of spine size. Disruption of α2-chimaerin suppressed EphA-mediated spine morphogenesis in cultured developing hippocampal neurons. α2-Chimaerin disruption that begins during the juvenile stage results in an increased size of spines in the hippocampus. Meanwhile, spine morphologies are unaltered when α2-chimaerin is deleted only in adulthood. Consistent with these spine morphological results, disruption of α2-chimaerin beginning in the juvenile stage led to an increase in contextual fear learning in adulthood; whereas contextual learning was recently shown to be unaffected when α2-chimaerin was deleted only in adulthood. Together, these results suggest that α2-chimaerin signaling in developmental stages contributes to determination of the morphological features of adult spines and establishment of normal cognitive ability. SIGNIFICANCE STATEMENT: Recent studies of neurodevelopmental disorders in humans and their animal models have led to an attractive hypothesis that spine morphogenesis during development forms the basis of adult cognition. In particular, the roles of Rac and its regulators, such as Rac-specific GTPase-activating proteins (RacGAPs) and Rac guanine nucleotide exchange factors, are a topic of focus in spine morphogenesis and cognitive ability. Using a series of mice with global and conditional knock-out (KO) of RacGAP α-chimaerin isoforms (α1-chimaerin and α2-chimaerin), we provide compelling evidence demonstrating that α2-chimaerin is involved in spine morphological refinement during late postnatal development and that this developmental α2-chimaerin function affects adult spine morphologies. Furthermore, our results clearly showed that α2-chimaerin signaling during late postnatal development contributes to normal cognitive ability in adult mice.

SAR, pharmacokinetics, safety, and efficacy of glucokinase activating 2-(4-sulfonylphenyl)-N-thiazol-2-ylacetamides: discovery of PSN-GK1.

Allosteric activators of the glucose-sensing enzyme glucokinase (GK) are currently attracting much interest as potential antidiabetic therapies because they can achieve powerful blood glucose lowering through actions in multiple organs. Here, the optimization of a weakly active high-throughput screening hit to (2 R)-2-(4-cyclopropanesulfonylphenyl)- N-(5-fluorothiazol-2-yl)-3-(tetrahydropyran-4-yl)propionamide (PSN-GK1), a potent GK activator with an improved pharmacokinetic and safety profile, is described. Following oral administration, this compound elicited robust glucose lowering in rats.

Glucokinase-activating ureas.

The synthesis, SAR and biological evaluation of a series of ureas that activate glucokinase, a target for diabetes therapy as a result of its critical role in the regulation of whole-body glucose homeostasis, are described. Some of the urea-containing glucokinase activators lowered blood glucose levels in vivo following oral dosing to C57BL/6J mice.

Modifications and structure-activity relationships at the 2-position of 4-sulfonamidopyrimidine derivatives as potent endothelin antagonists.

To improve water solubility and to study structure-activity relationships, we modified the structure of the pyrimidine nucleus of each of a series of potent ET(A) antagonists, 3a and 4a, at the 2-position. In a previous study, each of these antagonists showed an extremely high affinity for the ET(A) receptor in porcine aortic membrane (IC(50) 3a; < 0.001 nM, 4a; 0.0039 nM). Two modification methods, one being the addition of organolithium followed by DDQ oxidation and the other being the nucleophilic substitution of 2-(methylsulfonyl)pyrimidine, were applied individually to synthesize 2-substituted-4-sulfonamidopyrimidine derivatives. The introduction of aryl, heteroaryl, alkyl, amino, alkoxy, or alkylthio groups into the 2-position varied the affinity. Derivatives with hydrophilic groups at the 2-position showed higher water solubility but tended to reduce the affinity for the ET(A) receptor.