In Vivo Behavior of Hydrolyzable Tannins after Oral Administration of the Trapa bispinosa Extract to Rats.

The pericarp extract of Trapa bispinosa (TBPE), which is rich in hydrolyzable tannins, has been reported to inhibit α-glucosidase and glycation reactions. We investigated the in vivo behavior of hydrolyzable tannins and related metabolites after administration of TBPE to rats. Using high pressure liquid chromatography-electrospray ionization-tandem mass spectroscopy (HPLC-ESI-MS/MS), 12 ellagitannin metabolites, such as urolithins and 6 gallotannin metabolites, produced in the collected plasma and urine were quantified. Urolithins and gallic acid metabolites reached their maximum blood concentration after 24 and 1 h of administration, respectively. Conversely, the excretion of urolithins in urine required up to 72 h and followed a sigmoidal curve, whereas gallic acid metabolites were rapidly excreted earlier after administration. The results suggest that the metabolites gallotannin and ellagitannin are responsible for the antiglycation effect of TBPE, which proceeds via different mechanisms and times. Our findings provide basic data demonstrating the functionality of hydrolyzable tannins as well as Trapa ingredients.

Differences in lower-limb biomechanics during single-leg landing considering two peripheral fatigue tasks.

Dynamic knee valgus (DKV) occurs during landing after a fatigue task involving the lower extremity. However, the manner in which different peripheral fatigue tasks affect DKV remains unknown. In this study, we investigated the DKV via electromyography during single-leg landing considering the hip-joint fatigue task (HFT) and knee-joint fatigue task (KFT) performed by healthy men. We recruited 16 healthy male participants who performed a single-leg jump-landing motion from a height of 20 cm before and after an isokinetic hip abduction/adduction task (HFT) and knee extension/flexion task (KFT). Three-dimensional motion analysis systems were attached to the left gluteus medius and quadriceps, and surface electromyography was used to analyze the lower limb kinematics, kinetics, and muscle activity. The primary effects and interactions of the task and fatigue were identified based on the two-way repeated-measures analysis of variance. The results of the average angle during landing indicated that DKV occurs in KFT, whereas HFT applies external forces that adduct and internally rotate the knee at peak vertical ground reaction force (vGRF). Furthermore, both KFT and HFT exhibited an increase in muscle activity in the quadriceps. The analysis revealed that the occurrence of DKV varies depending on the peripheral fatigue task, and the effects on average DKV during landing and DKV at peak vGRF vary depending on the peripheral fatigue task.

Quantification and characterization of biological activities of glansreginin A in black walnuts (Juglans nigra).

Glansreginin A has been reported to be an indicator of the quality of walnuts (Juglans spp.). However, bioactive properties of glansreginin A have not been adequately explored. In the present study, we quantified concentrations of glansreginin A in black walnuts (Juglans nigra) using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and performed an array of in vitro bioassays to characterize biological activities (e.g., antibacterial, antioxidant, anticancer capacities) of this compound. Results from HPLC-MS/MS analysis indicated that glansreginin A was presented in all 12 black cultivars examined and its contents were variable among black walnut cultivars, ranged from 6.8 mg/kg (Jackson) to 47.0 mg/kg (Hay). Glansreginin A possessed moderate antibacterial activities against Gram-positive pathogens (Staphylococcus aureus and Bacillus anthracis). This compound exhibited no antioxidant activities, did not induce the activity of antioxidant response element signaling pathways, and exerted no antiproliferative effects on tumorigenic alveolar epithelial cells and non-tumorigenic lung fibroblast cells.

Identification of Antioxidative Hydrolyzable Tannins in Water Chestnut.

Despite the various biological activities exhibited by water chestnut (the fruit of the Trapa genus), the phenolic compounds present in its extract require comprehensive characterization. Accordingly, we analyzed a 80% methanol extract of commercially available water chestnut and identified a new hydrolyzable tannin dimer termed trapadin A. Additionally, 22 known compounds, including 10 hydrolyzable tannin monomers and 2 dimers, were also detected in the extract. Spectroscopic and chemical methods were used to elucidate the structure of trapadin A, revealing it to be a hydrolyzable tannin dimer formed from units of tellimagrandin II and 1,2,3,6-tetra-O-galloyl-ß-d-glucose. Moreover, the 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity assay used to determine the half-maximal effective concentration values for the 23 compounds isolated from water chestnut indicated significant radical scavenging activity associated with hydrolyzable tannins. Notably, trapadin A, the new hydrolyzable tannin dimer, exhibited the highest activity value among the tested compounds.

Heterologous gene expression system for the production of hydrolyzable tannin intermediates in herbaceous model plants.

Aluminum toxicity is the main factor limiting the elongation of plant roots in acidic soil. The tree species Eucalyptus camaldulensis is considerably more resistant to aluminum than herbaceous model plants and crops. Hydrolyzable tannins (HTs) accumulating in E. camaldulensis roots can bind and detoxify the aluminum taken up by the roots. However, in herbaceous model plants, HTs do not accumulate and the genes involved in the HT biosynthetic pathway are largely unknown. The aim of this study was to establish a method for reconstituting the HT biosynthetic pathway in the HT non-accumulating model plant Nicotiana benthamiana. Four E. camaldulensis enzymes were transiently expressed in N. benthamiana leaves via Agrobacterium tumefaciens-mediated transformation. These enzymes included dehydroquinate dehydratase/shikimate dehydrogenases (EcDQD/SDH2 and EcDQD/SDH3), which catalyze the synthesis of gallic acid, the first intermediate of the HT biosynthetic pathway that branches off from the shikimate pathway. The others were UDP-glycosyltransferases (UGT84A25 and UGT84A26), which catalyze the conversion of gallic acid to ß-glucogallin, the second intermediate. The co-expression of the EcDQD/SDHs in transgenic N. benthamiana leaf regions promoted the synthesis of gallic acid. Moreover, the co-expression of the UGT84As in addition to the EcDQD/SDHs resulted in the biosynthesis of ß-glucogallin, the universal metabolic precursor of HTs. Thus, we successfully reconstituted a portion of the HT biosynthetic pathway in HT non-accumulating N. benthamiana plants. This heterologous gene expression system will be useful for co-expressing candidate genes involved in downstream reactions in the HT biosynthetic pathway and for clarifying their in planta functions.

Effects of Physiatrist and Physiotherapist-supervised Therapeutic Exercise on Physical Function in Frail Older Patients with Multimorbidity.

Objectives: : To ease the burden on care facilities, there is a need to reduce the number of frail older people with multiple chronic and non-communicable diseases (multimorbidity) that require long-term care. We investigated the effects of therapeutic exercise in rehabilitation treatment for older individuals with frailty and multimorbidity. Methods: : We performed 4 weeks of inpatient rehabilitation treatment for frail older patients with multimorbidity. The therapeutic exercise was performed based on whole-body evaluations by qualified physiatrists and physiotherapists. Sixty-minute sessions were conducted twice a day and six times a week in accordance with the American College of Sports Medicine guidelines. Physical functions (grip strength, isometric lower muscle strength, 10-m walking test, and 6-min walking test) were measured at admission and discharge. Results: : Of the 33 patients, 8 were unable to complete physical function evaluations, and 1 patient had a stress fracture of the right calcaneus beyond the rehabilitation time. Twenty-four patients were included in the analysis. Compared to the admission period, significant improvements were observed for all evaluated parameters post-therapy (P < 0.05). However, no significant correlation was found between the number of chronic diseases and changes to physical function (P > 0.05). Conclusions: : Physical function of frail older individuals with multimorbidity improves through rehabilitation treatments supervised by physiatrists and physiotherapists. The number of chronic illnesses and the improvements in physical function were not related. Therefore, rehabilitation treatments for older patients may target both frailty and multimorbidity. Our results will aid in the rehabilitation treatment plans for this demographic.

A Fatal Case of Disseminated Infection Caused by Community-Associated Methicillin-Resistant Staphylococcus aureus USA300 Clone.

Methicillin-resistant Staphylococcus aureus (MRSA) USA300 is a representative community-associated MRSA (CA-MRSA) clone worldwide. Herein, we report the case of a patient with USA300 clone infection who could not be salvaged. A 25-year-old man who had sex with men presented with symptoms including fever persisting for one week and skin lesions located on the buttocks. Computed tomography imaging showed multiple nodules and consolidations, especially in the peripheral lung fields, right iliac vein thrombosis, and pyogenic myositis of medial thighs bilaterally. Blood cultures revealed MRSA bacteremia. The patient's condition deteriorated rapidly, complicated by acute respiratory distress syndrome and infective endocarditis. Despite the intubation on the 6th hospital day, he died on the 9th day. Multilocus sequence typing of this patient's MRSA strain revealed sequence type 8 with a staphylococcal cassette chromosome of mec type IVa, Panton-Valentine leukocidin gene, and the arginine catabolic mobile element, indicating presence of the USA300 clone. Patients with CA-MRSA skin lesions presenting with furuncles or carbuncles on the lower body are at a higher risk of severe disease. The patient's background, appearance, and location of skin lesions are critical for the early diagnosis of severe CA-MRSA infection.

Age estimation based on blood DNA methylation levels in brown bears.

Age is an essential trait for understanding the ecology and management of wildlife. A conventional method of estimating age in wild animals is counting annuli formed in the cementum of teeth. This method has been used in bears despite some disadvantages, such as high invasiveness and the requirement for experienced observers. In this study, we established a novel age estimation method based on DNA methylation levels using blood collected from 49 brown bears of known ages living in both captivity and the wild. We performed bisulfite pyrosequencing and obtained methylation levels at 39 cytosine-phosphate-guanine (CpG) sites adjacent to 12 genes. The methylation levels of CpGs adjacent to four genes showed a significant correlation with age. The best model was based on DNA methylation levels at just four CpG sites adjacent to a single gene, SLC12A5, and it had high accuracy with a mean absolute error of 1.3 years and median absolute error of 1.0 year after leave-one-out cross-validation. This model represents the first epigenetic method of age estimation in brown bears, which provides benefits over tooth-based methods, including high accuracy, less invasiveness, and a simple procedure. Our model has the potential for application to other bear species, which will greatly improve ecological research, conservation, and management.

Antiallergic Activity of 3-O-Dodecyl-l-ascorbic Acid.

2-O-Alkyl-l-ascorbic acids and 3-O-alkyl-l-ascorbic acids were synthesized, and their degranulation inhibitory activities were evaluated. Among ascorbic acid derivatives with butyl, octyl, dodecyl, hexadecyl, and octadecyl groups introduced at the C-2 or C-3 positions, an AA derivative with a dodecyl group introduced at the C-3 position, 3-O-dodecyl-l-ascorbic acid (compound 8), showed the strongest inhibitory activity against antigen-stimulated degranulation. Compound 8 also inhibited calcium ionophore-stimulated degranulation. Compound 11, in which the hydroxyl group at the C-6 position of compound 8 was substituted with an amino group, and compound 12, in which the dodecyloxy group at the C-3 position of compound 8 was exchanged with a dodecylamino group, were synthesized, and these derivatives showed weaker inhibitory activity against antigen-stimulated degranulation than that of compound 8. In addition, orally administered compound 8 inhibited passive cutaneous anaphylaxis reactions in mice with a potency equal to that of oxatomide, an antiallergic agent. These results suggest that compound 8 may be a candidate for antiallergic treatment.

Neurite Outgrowth-Promoting Compounds from the Petals of Paeonia lactiflora in PC12 Cells.

Isorhamnetin-3-O-glucoside and astragalin, flavonol glucosides, were isolated from the petals of Paeonia lactiflora as neurite outgrowth-promoting compounds. Isoquercitrin, formed by demethylating the B ring of isorhamnetin-3-O-glucoside or by adding a hydroxyl group to the B ring of astragalin, was evaluated for neurite outgrowth-promoting activity and was compared with the activities of isorhamnetin-3-O-glucoside and astragalin. The activities of isorhamnetin, kaempferol, and quercetin, aglycones corresponding to isorhamnetin-3-O-glucoside, astragalin, and isoquercitrin, respectively, were also evaluated. Isorhamnetin-3-O-glucoside and astragalin showed much stronger neurite outgrowth-promoting activities than the activities of the other tested flavonoids. They exhibited relatively weak anti-oxidant activities and moderate AChE inhibitory activities compared to the activities of the other tested flavonoids. Isorhamnetin-3-O-glucoside and astragalin promoted morphological neurite outgrowth and the expression of neurofilaments induced by NGF in PC12 cells. Isorhamnetin-3-O-glucoside and astragalin might be candidate compounds as neural differentiation agents in peripheral nerves and functional food ingredients preventing cognitive decline.

Malabaricone C derived from nutmeg inhibits arachidonate 5-lipoxygenase activity and ameliorates psoriasis-like skin inflammation in mice.

As pro-inflammatory lipid mediators, leukotrienes have pathophysiological activities in several inflammatory diseases, including psoriasis. In the biosynthesis of leukotrienes from arachidonic acid, 5-lipoxygenase catalyzes the first two steps. In the present study, we showed that nutmeg (Myristica fragrans) strongly inhibited the catalytic activity of 5-lipoxygenase. To characterize the bioactive component(s) of nutmeg, we performed 5-lipoxygenase inhibitory activity-guided fractionation of aqueous ethanol extract of nutmeg, resulting in the isolation of malabaricone C having antioxidant activity. Malabaricone C exhibited potent competitive inhibition of 5-lipoxygenase with an IC50 value of 0.2 µM. In mice with imiquimod-induced psoriasis-like skin lesions, topical application of 2 mM malabaricone C significantly ameliorated hyperplasia and inflammatory cell infiltration, and suppressed the expression of the psoriasis-associated genes S100a9, Krt1, Il17a, and Il22. Lipid metabolome analysis of these psoriasis-like skin lesions showed that malabaricone C markedly decreased the level of leukotriene B4 but did not significantly increase the other pro-inflammatory lipid mediators. These findings suggest that malabaricone C decreases LTB4 by the 5-lipoxygenase inhibition and ameliorates the symptoms of psoriasis-like skin inflammation.

Synthesis of Ascorbic Acid Derivatives with Different Types of C8 Straight Acyl Chain and Their Neurite Outgrowth-Enhancing Activities.

We previously reported that 2-O-α-D-glucopyranosyl-6-O-octanoyl-L-ascorbic acid, having a C8 straight acyl chain, at a concentration of 100 µM remarkably enhanced nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells after being metabolized to L-ascorbic acid (AA) as an active form by esterase and α-glucosidase. In this study, to examine the structure-activity relationship of 6-O-substituted AA derivatives with a C8 straight acyl chain for neurite outgrowth-promoting activity, we synthesized AA derivatives 1-4 and compared their activities for promoting NGF-induced neurite outgrowth in PC12 cells. AA derivatives 1-4 showed neurite outgrowth-enhancing activity at 100 µM, while AA derivative 2 also showed the enhancing activity at 3 µM. Furthermore, AA derivative 2 as well as AA enhanced NGF-induced neurite outgrowth after being incorporated into PC12 cells via sodium-dependent vitamin C transporter as an anion transporter. The results suggested that AA derivative 2 has neurite outgrowth-enhancing activity in its intact form at a low concentration (3 µM) and that AA derivatives 1-4 showed their activities in the form of AA, a metabolite of these derivatives, at a higher concentration (100 µM).

Comparison of the Genetic Diversity of the Captive and Wild Populations of the Tsushima Leopard Cat Using a GRAS-Di Analysis.

The Tsushima leopard cat (Prionailurus bengalensis euptilurus) (TLC) is a regional population of the Amur leopard cat (P. bengalensis euptilurus) that lives only on the Tsushima Island in Japan and is threatened with extinction. Because the TLC population is small, genetic management is important. In this study, we obtained the draft genome of the TLC and identified single-nucleotide polymorphism (SNP) markers using a genotyping by random amplicon sequencing-direct (GRAS-Di) analysis. We genotyped 31 captive individuals and 50 wild individuals, of which 48 were from a previous study. The identified SNPs were used to clarify the genetic diversity and genetic structure of the wild and captive populations of the TLC. The size of the genome was estimated to be about 2.42 Gb. The number of SNP markers developed was 139, and although PID and probability of exclusion obtained using these SNP markers were not as high as those reported in the studies of other wild species, these SNP markers could be used to identify individuals and parentage. Moreover, the genetic diversity indices of the captive population were similar to those of the wild population. These SNP markers will be useful for understanding the ecology of the TLC and planning conservation strategies.

Generation and Gene Expression Profiles of Grevy's Zebra Induced Pluripotent Stem Cells.

Induced pluripotent stem cells (iPSCs) can serve as a biological resource for functional and conservation research for various species. This realization has led to the generation of iPSCs from many species, including those identified as endangered. However, the understanding of species variation in mammalian iPSCs remains largely unknown. To gain insight into species variation in iPSCs, we generated iPSCs from a new species Grevy's zebra (Equus grevyi; gz-iPSCs), which has been listed as endangered in the IUCN (International Union for Conservation of Nature) Red List. We isolated primary fibroblast cells from an individual and successfully reprogrammed them into iPSCs. The generated gz-iPSCs continued to grow under primed-type culture condition and showed pluripotency and differentiation potential. To describe the molecular characteristics of gz-iPSCs, we performed RNA sequencing analysis. The gz-iPSC transcriptome showed robust expression of pluripotency-associated genes reported in human and mouse, suggesting evolutionary conservation among the species. This study provides insight into the iPSCs from a rare species and helps the understanding of the gene expression basis underlying mammalian pluripotent stem cells.

Designing the building space of a shopping street to use as a disaster evacuation shelter during the COVID-19 pandemic: A case study in Kobe, Japan.

This study considers the risk of a natural hazard-induced disaster occurring during a pandemic, such as the novel coronavirus (COVID-19) pandemic, and develops the idea of utilizing a shopping street with disaster-proof buildings as a temporary evacuation shelter by incorporating countermeasures against the spread of infectious diseases. Using a case study of a shopping street in Kobe, Japan, we estimate shelter capacity by considering the requirement of 6 m2 of space allotted for each person. The shelter can accommodate 1194 evacuees and provide them with food and drinks for one day, even in the worst case of lifeline disruption. This study proposes a method of designing shelter space, and demonstrates how non-homogeneous and noncontinuous spaces within shopping street buildings can be applied to prevent the spread of infection, through the classification of evacuee types and use of space and facilities designated for each type. The study further examines the liability issue of secondary infection at the shelter with reference to civic law and the roles of government in developing a distributed evacuation framework.

Characterization and Identification of Bioactive Polyphenols in the Trapabispinosa Roxb. Pericarp Extract.

In this study, we present the isolation and characterization of the structure of six gallotannins (1-6), three ellagitannins (7-9), a neolignan glucoside (10), and three related polyphenolic compounds (gallic acid, 11 and 12) from Trapa bispinosa Roxb. pericarp extract (TBE). Among the isolates, the structure of compound 10 possessing a previously unclear absolute configuration was unambiguously determined through nuclear magnetic resonance and circular dichroism analyses. The α-glucosidase activity and glycation inhibitory effects of the isolates were evaluated. Decarboxylated rugosin A (8) showed an α-glucosidase inhibitory activity, while hydrolyzable tannins revealed stronger antiglycation activity than that of the positive control. Furthermore, the identification and quantification of the TBE polyphenols were investigated by high-performance liquid chromatography coupled to ultraviolet detection and electrospray ionization mass spectrometry analysis, indicating the predominance of gallic acid, ellagic acid, and galloyl glucoses showing marked antiglycation properties. These findings suggest that there is a potential food industry application of polyphenols in TBE as a functional food with antidiabetic and antiglycation activities.

Antiallergic Activity of 6-Deoxy-2-O-methyl-6-(N-hexadecanoyl)amino-l-ascorbic Acid.

Allergy is an excessive immune response to a specific antigen. Type I allergies, such as hay fever and food allergies, have increased significantly in recent years and have become a worldwide problem. We previously reported that an ascorbic acid derivative having palmitoyl and glucosyl groups, 2-O-α-d-glucopyranosyl-6-O-hexadecanoyl-l-ascorbic acid (6-sPalm-AA-2G), showed inhibitory effects on degranulation in vitro and on the passive cutaneous anaphylaxis (PCA) reaction in mice. In this study, several palmitoyl derivatives of ascorbic acid were synthesized and a structure-activity relationship study was performed to discover more potent ascorbic acid derivatives with degranulation inhibitory activity. 6-Deoxy-2-O-methyl-6-(N-hexadecanoyl)amino-l-ascorbic acid (2-Me-6-N-Palm-AA), in which a methyl group was introduced into the hydroxyl group at the C-2 position of ascorbic acid and in which the hydroxyl group at the C-6 position was substituted with an N-palmitoyl group, exhibited much higher inhibitory activity for degranulation in vitro than did 6-sPalm-AA-2G. 2-Me-6-N-Palm-AA strongly inhibit the PCA reaction in mice at lower doses than those of 6-sPalm-AA-2G. These findings suggest that 2-Me-6-N-Palm-AA may be a promising therapeutic candidate for allergic diseases.