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Background and Objectives : Mycobacterium bovis Bacillus Calmette-Guérin (BCG) vaccine administration has been suggested to prevent glucose metabolism abnormalities and fatty liver in genetically obese ob/ob mice; however, it is not clear whether the beneficial effects of BCG are also observed in the progression of glucose intolerance induced by a high-fat diet (HFD). Therefore, the effects of BCG vaccination on changes in glucose tolerance and insulin response were investigated in HFD-fed C57BL/6 mice. Materials and Methods : We used the BCG Tokyo 172 strain to determine effects on abnormalities in glucose metabolism. For vaccination, five-week-old male mice were injected intraperitoneally with BCG and maintained on a HFD for three weeks. The mice were regularly subjected to intraperitoneal glucose tolerance and insulin tolerance tests (IGTTs and ITTs). These tests were also performed in mice transplanted with bone marrow cells from BCG-vaccinated donor mice. Results : Significant effects of BCG vaccination on blood glucose levels in the IGTTs and ITTs were observed from week 12 of the experiment. BCG vaccination significantly improved changes in fasting glucose and insulin levels, insulin resistance indexes, and glucagon-to-insulin ratios in conjunction with the HFD at the end of the experiment. Significant inhibitory effects in the IGTTs and ITTs on glucose intolerance were also observed with transplantation with bone marrow cells derived from BCG-vaccinated donor mice. Conclusions : BCG vaccination significantly delayed glucose intolerance progression, suggesting a beneficial effect of BCG on the pathogenesis of type 2 diabetes. It has also been suggested that the effects of BCG vaccination may be at least partially due to an immune memory (trained immunity) for hematopoietic stem and progenitor cells of the bone marrow.

The most widely used isoprene emission algorithm, G93 formula, estimates instantaneous leaf-level isoprene emission using the basal emission factor and light and temperature dependency parameters. The G93 parameters have been suggested to show variation depending on past weather conditions, but no study has closely examined the relationship between past meteorological data and the algorithm parameters. Here, to examine the influence of the past weather on these parameters, we monitored weather conditions, G93 parameters, isoprene synthase transcripts and protein levels, and MEP pathway metabolites in the tropical tree Ficus septica for 12 days and analyzed their relationship with cumulative temperature and light intensity. Plants were illuminated with varying (ascending and descending) light regimes, and our previously developed Ping-Pong optimization method was used to parameterize G93. The cumulative temperature of the past 5 and 7 days positively correlated with C T 2 and α, respectively, while the cumulative light intensity of the past 10 days showed the highest negative correlation with α. Concentrations of MEP pathway metabolites and IspS gene expression increased with increasing cumulative temperature. At best, the cumulative temperature of the past 2 days positively correlated with the MEP pathway metabolites and IspS gene expression, while these factors showed a biphasic positive and negative correlation with cumulative light intensity. Optimized G93 captured well the temperature and light dependency of isoprene emission at the beginning of the experiment; however, its performance significantly decreased for the latter stages of the experimental duration, especially for the descending phase. This was successfully improved through separate optimization of the ascending and descending phases, emphasizing the importance of the optimization of formula parameters and model improvement. These results have important implications for the improvement of isoprene emission algorithms, particularly under the predicted increase in future global temperatures.

In Ficus septica, the short-term control of isoprene production and, therefore, isoprene emission has been linked to the hormone balance between auxin (IAA) and jasmonic acid (JA). However, the relationship between long-term changes in isoprene emission and that of plant hormones remains unknown. This study tracked isoprene emissions from F. septica leaves, plant hormone concentrations and signalling gene expression, MEP pathway metabolite concentrations, and related enzyme gene expression for 1 year in the field to better understand the role of plant hormones and their long-term control. Seasonality of isoprenes was mainly driven by temperature- and light-dependent variations in substrate availability through the MEP route, as well as transcriptional and post-transcriptional control of isoprene synthase (IspS). Isoprene emissions are seasonally correlated with plant hormone levels. This was especially evident in the cytokinin profiles, which decreased in summer and increased in winter. Only 4-hydroxy-3-methylbut-2-butenyl-4-diphosphate (HMBDP) exhibited a positive connection with cytokinins among the MEP metabolites examined, suggesting that HMBDP and its biosynthetic enzyme, HMBDP synthase (HDS), play a role in channelling of MEP pathway metabolites to cytokinin production. Thus, it is probable that cytokinins have potential feed-forward regulation of isoprene production. Under long-term natural conditions, the hormonal balance of IAA/JA-Ile was not associated with IspS transcripts or isoprene emissions. This study builds on prior work by revealing differences between short- and long-term hormonal modulation of isoprene emissions in the tropical tree F. septica., (© 2023 Wiley-VCH GmbH. Published by John Wiley & Sons Ltd.)

The isoprene emission rate from plants is simulated by a function of light intensity and leaf temperature, and the G-93 formula is the most extensively applied algorithm for this purpose. Isoprene is biosynthesized by the enzyme isoprene synthase (IspS), and instantly emitted from the leaf. Enzyme kinetics of IspS and substrate availability are important factors involved in the short-term leaf-level control of isoprene emissions. It is thus assumed that the parameters of G-93 may correlate with the kinetics of IspSs, however, at present there is no data available on the relationship between these two parameters. In this investigation, six IspS genes from tropical trees were cloned, their properties characterized, and the relationship between the enzyme kinetics of IspSs and the parameters of G-93 examined. There was a negative correlation between the enzyme kinetics of IspS K m and parameter C T1 of G93, which is used to define the temperature dependency of isoprene emissions. However, performance constant of IspS (k cat /K m ) only showed slight positive correlation with C T1 .suggesting that the enzyme kinetics of IspS has limited significance in controlling the temperature response of isoprene emissions. The molecular structure of IspS was further elucidated using a molecular dynamics simulation with a focus on the active site in the 6 α-helices bundle. The simulation of the enzyme-substrate complex of IspS from B. variegata predicted a new metal binding domain in helix F (E383) and catalytic motif FXRDRLXE in the A-C loop that could involve the deprotonation of dimethylallyl diphosphate (DMADP) to form a carbocation. Notably, after the binding of a metal ion and DMADP, the active-site closure mechanism was found to involve conformational alterations in the helix H-α1 and transition from a loose to tight enclosure of the 6 α-helices bundles to tune the active pocket size. The characteristics identified for the IspSs from tropical trees could help to explain regional isoprene emissions in tropical areas., (© 2022. The Author(s) under exclusive licence to The Botanical Society of Japan.)

Leucaena leucocephala growing in the tropics and subtropics serves as potential forage for livestock because its foliage is rich in protein, fiber, and minerals. However, its use for livestock feed has been hindered by toxic nonprotein amino acid mimosine. Therefore, it is necessary to develop a method to reduce or eliminate mimosine from foliage. A previous study found that the fermentation of L. leucocephala foliage reduced the mimosine content and prompted the authors to isolate potent mimosine degrading microorganisms and characterize the mimosinase for the complete elimination of mimosine in the L. leucocephala foliage. The soil screening of the L. leucocephala tree surroundings led to the isolation of Arthrobacter sp. Ryudai-S1, which can degrade and assimilate mimosine as a nitrogen and carbon source. Mimosinase in this strain was found to be thermostable and showed strong activity. Docking model's inspection and the interaction energy calculation between mimosine-pyridoxal-5'-phosphate (PLP) complex and the active site of this enzyme identified 11 important amino acid residues that stabilized the binding. Of these amino acid residues, mutation experiment suggested that Tyr-263' and Phe-34 stabilizes the substrate binding and play a critical role in guiding the substrate to proper positions to accomplish high catalytic efficacy and selectivity. These observations suggest that Arthrobacter sp. Ryudai-S1 could be potentially useful for the development of L. leucocephala feed with reduced mimosine content., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Plant hormone signalling and the circadian clock have been implicated in the transcriptional control of isoprene biosynthesis. To gain more insight into the hormonal control of isoprene biosynthesis, the present study measured plant hormone concentrations in jasmonic acid (JA)-treated leaves of our previous model study, examined their relationship with gene expression of isoprene synthase (IspS) and hormone signalling transcription factors. Of the plant hormones, IAA and JA-Ile and their related transcription factors (MYC2 and SAUR21) were significantly correlated with IspS gene expression. Concentrations of cytokinins, isopentenyladenine (iP), trans-zeatin riboside (tZR) and cis-zeatin riboside (cZR), were similarly significantly correlated with IspS expression. However, there was no significant correlation between their related transcription factor (ARR-B) and IspS expression. The circadian clock-related gene PRR7, but not the transcription factor LHY, was highly correlated with IspS expression. These results suggest that the hormonal balance between JA-Ile and IAA plays a central role in transcriptional regulation of IspS through the transcription factors MYC2 and SAUR21, the early auxin responsive genes. The putative cis-acting elements for SAUR on the IspS promoter (TGTCNN and CATATG), in addition to the G-box for MYC2, support the above proposal. These results provide insightful information on the core components of plant hormone-related regulation of IspS under coordination with the circadian clock genes., (© 2022 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.)

It has been suggested that isoprene synthesis by isoprene synthase (IspS) proceeds via a substrate-assisted mechanism. The authors observed a non-enzymatic isoprene formation by Mn 2+ , which represents the basis of IspS enzyme reaction. Because IspS and many other terpene synthases require Mn 2+ metal ions as cofactor, this study characterized the formation reaction for the first time. Metal ions including Mn 2+ non-enzymatically produced both isoprene and 2-methyl-3-buten-2-ol (2-MBO) from dimethylallyl pyrophosphate (DMADP). Isoprene formation was most enhanced by Fe 2+ and, to a lesser extent, by Mn 2+ or Cu 2+ . Ni 2+ , Co 2+ , Mg 2+ , and Ba 2+ exhibited a low activity to generate both isoprene and 2-MBO. The proportion of isoprene and 2-MBO varied with the Mn 2+ concentration: isoprene predominated over 2-MBO at a higher Mn 2+ concentration. Similarly, isoprene formation by Mn 2+ increased exponentially as temperature increased with predominance of isoprene over 2-MBO at higher temperature. Both isoprene and 2-MBO formation was enhanced by acidic and neutral pH compared to alkaline conditions. Molecular dynamic simulation of DMADP suggested that the formation reaction is initiated by deprotonation of hydrogen on allyl terminal carbon by phosphate oxygen and generates carbocation and allyl anion intermediates. This is followed by quenching to produce isoprene or by hydroxyl addition to form 2-MBO. Thus, this study provided an insight into reaction mechanism of isoprene and 2-MBO biosynthesis and highlighted some parts of isoprene emission from terrestrial plants, which could be formed by non-enzymatic mechanism., (© 2022. The Author(s).)

Despite its major role in global isoprene emission, information on the environmental control of isoprene emission from tropical trees has remained scarce. Thus, in this study, we examined the relationship between parameters of G-93 isoprene emission formula (C T1 , C T2 , and α), growth temperature and light intensity, photosynthesis (ɸ, P max ), isoprene synthase (IspS) level, and 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway metabolites using sunlit and shaded leaves of four tropical trees. The results showed that the temperature dependence of isoprene emission from shaded leaves did not differ significantly from sunlit leaves. In contrast, there was a lower saturation irradiance in shaded leaves than in sunlit leaves, the same as temperate plants. The photosynthesis rate of shaded leaves showed lower saturation irradiance, similar to the light dependence of isoprene emission. In most cases, the concentration of MEP pathway metabolites was of lower tendency in shaded leaves versus in sunlit leaves, whereas no significant difference was noted in IspS level between sunlit and shaded leaves. Correlation analysis between these parameters found that C T1 of the G-93 parameter was positively correlated with the concentration of DXP and DMADP, whereas C T2 correlated with the concentration of MEP and the average air temperature for the past 48 h. Similarly, α closely associated with the initial slope (ɸ) of photosynthesis rate, and the basal emission factor is also linked to the photon flux of past days. These results suggest that growth conditions may control the temperature dependence of isoprene emission from tropical trees via the changes in the profiles of MEP pathway metabolites, causing alteration in the parameters of the isoprene emission formula., (© 2021. The Botanical Society of Japan.)

The dynamic ability of adipocytes in adipose tissue to store lipid in response to changes in the nutritional input and inflammatory elicitors has a major impact on human health. Previously, we established laminarin-coated beads or LCB as an inflammatory elicitor for adipocytes. However, it was not clear whether LCB inhibits lipid accumulation in adipocytes. Here, we show that LCB acts in the early stage of adipogenesis through both interleukin-1 receptor-associated kinases (IRAK) and spleen tyrosine kinase (SYK) pathways, resulting in the activation of the AMP-activated protein kinase (AMPK) and nuclear factor-κB (NF-κB) complexes, which subsequently cause cell cycle arrest, downregulation of the key transcription factors and enzymes responsible for adipogenesis, inhibition of adipogenesis, and stimulation of an inflammatory response. While LCB could effectively block lipid accumulation during the early stage of adipogenesis, it could stimulate an inflammatory response at any stage of differentiation. Additionally, our results raise a possibility that toll-like receptor 2 (TLR2) and C-type lectin domain family 7 member A (CLEC7A/Dectin-1) might be potential β-glucan receptors on the fat cells. Together, we present the mechanism of LCB, as fungal-like particles, that elicits an inflammatory response and inhibits adipogenesis at the early stage of differentiation., (© 2021. The Author(s).)

Facultative halophyte Kandelia obovata plants were exposed to mild (1.5% NaCl) and severe (3% NaCl) salt stress with or without sodium nitroprusside (SNP; 100 µM; a NO donor), hemoglobin (Hb, 100 µM; a NO scavenger), or Nω-nitro-L-arginine methyl ester (L-NAME, 100 µM; a NO synthase inhibitor). The plants were significantly affected by severe salt stress. They showed decreases in seedling growth, stomatal conductance, intercellular CO 2 concentration, SPAD value, photosynthetic rate, transpiration rate, water use efficiency, and disrupted antioxidant defense systems, overproduction of reactive oxygen species, and visible oxidative damage. Salt stress also induced ion toxicity and disrupted nutrient homeostasis, as indicated by elevated leaf and root Na + contents, decreased K + contents, lower K + /Na + ratios, and decreased Ca contents while increasing osmolyte (proline) levels. Treatment of salt-stressed plants with SNP increased endogenous NO levels, reduced ion toxicity, and improved nutrient homeostasis while further increasing Pro levels to maintain osmotic balance. SNP treatment also improved gas exchange parameters and enhanced antioxidant enzymes' activities (catalase, ascorbate peroxidase, monodehydroascorbate reductase, and dehydroascorbate reductase). Treatment with Hb and l-NAME reversed these beneficial SNP effects and exacerbated salt damage, confirming that SNP promoted stress recovery and improved plant growth under salt stress.

Cirsium brevicaule A. GRAY is a wild perennial herb, and its roots (CbR) have traditionally been used as both food and medicine on the Japanese islands of Okinawa and Amami. The present study evaluated the antiadipogenic effect of CbR using mouse embryonic fibroblast cell line 3T3-L1 from JCRB cell bank. Dried CbR powder was serially extracted with solvents of various polarities, and these crude extracts were tested for antiadipogenic activity. Treatment with the methanol extract of CbR showed a significant suppression of lipid accumulation in 3T3-L1 cells. Methanol extract of CbR was then fractionated and subjected to further activity analyses. The phenylpropanoid glycosidic molecule syringin was identified as an active compound. Syringin dose dependently suppressed lipid accumulation of 3T3-L1 cells without cytotoxicity, and significantly reduced the expressions of peroxisome proliferator-activated receptor gamma, the master regulator of adipogenesis, and other differentiation markers. It was demonstrated that syringin effectively enhanced the phosphorylation of the AMP-activated protein kinase and acetyl-CoA carboxylase. These results indicate that syringin attenuates adipocyte differentiation, adipogenesis, and promotes lipid metabolism; thus, syringin may potentially serve as a therapeutic candidate for treatment of obesity.

Infrared spectroscopy in the gas phase was used to study the formation reaction of the CH⋯O hydrogen bonding complex involving the CH group of trifluoromethane, as a hydrogen donor, and the carbonyl group of methyl acetate, as a hydrogen acceptor, under different (T, p) conditions. The hydrogen-bonded carbonyl stretch of the molecular pair was monitored in dilute mixtures of methyl acetate in trifluoromethane at near-critical temperatures, from gas- to liquid-like densities. In the gas region, it was possible to discriminate the carbonyl signal of the hydrogen-bonded complex from that of the free ester and have access to their relative concentration. The equilibrium constant of the hydrogen bonding reaction and the standard enthalpy and entropy changes in the process were determined using the spectroscopic data. CH⋯O bonding was favored by lowering temperature or pressurizing F 3 CH in the mixture, remaining essentially no free carbonyl groups about the critical density. The carbonyl band of the hydrogen-bonded pair appeared as a single symmetric peak up to liquid-like densities, suggesting that the 1:1 methyl acetate-trifluoromethane complex has the most abundant stoichiometry. Spectral features as frequency shift and bandwidth of the hydrogen-bonded carbonyl were studied as a function of temperature and solvent-density. A bathochromic (red) vibrational shift was registered for the bound carbonyl band against density, with a sudden change in behavior in the near-critical region, while the width of this band remains mostly unresponsive.

Cysteine biosynthesis is directed by the successive commitments of serine acetyltransferase, and O-acetylserine (thiol) lyase (OASTL) compounds, which subsequently frame the decameric cysteine synthase complex. The isoforms of OASTL are found in three compartments of the cell: the cytosol, plastid, and mitochondria. In this investigation, we first isolated putative chloroplastic OASTL (Ch-OASTL) from Leucaena leucocephala, and the Ch-OASTL was then expressed in BL21-competent Escherichia coli. The putative Ch-OASTL cDNA clone had 1,543 base pairs with 391 amino acids in its open reading frame and a molecular weight of 41.54 kDa. The purified protein product exhibited cysteine synthesis ability, but not mimosine synthesis activity. However, they both make the common α-aminoacrylate intermediate in their first half reaction scheme with the conventional substrate O-acetyl serine (OAS). Hence, we considered putative Ch-OASTL a cysteine-specific enzyme. Kinetic studies demonstrated that the optimum pH for cysteine synthesis was 7.0, and the optimum temperature was 40 °C. In the cysteine synthesis assay, the K m and k cat values were 838 ± 26 µM and 72.83 s -1 for OAS, respectively, and 60 ± 2 µM and 2.43 s -1 for Na 2 S, respectively. We can infer that putative Ch-OASTL regulatory role is considered a sensor for sulfur constraint conditions, and it acts as a forerunner of various metabolic compound molecules.

Dihydropyranocoumarins (DPCs) were isolated from Peucedanum japonicum Thunb as anti-obesity compounds in 3T3-L1 adipocytes assay; however, it is uncertain whether DPC exerts anti-obesity activity in vivo. Therefore, this study evaluated the oral intake of pure DPCs in mice fed a high-fat diet, and also attempted to enhance its activity by nanoparticulation. Increases in body weight gain and fat accumulation in white adipose tissues were significantly suppressed by the dietary intake of DPCs (1.943 mg/mouse/day). DPCs intake also significantly decreased the mean size of adipocytes and upregulated mRNA levels of thermogenesis-related genes. Nanoparticulation of DPCs with polylactic-co-glycolic acid (PLGA) dramatically increased its activity almost 100-fold over that of a non-nanoparticulated form. Thus, our findings clearly demonstrated the anti-obesity activity of DPCs in vivo and suggested that PLGA nanoparticle encapsulation was useful to enhance the anti-obesity activity of DPCs with the aim to develop natural and safe anti-obesity agents., Competing Interests: The authors declare no conflict of interest.

Mimosinase degrades the non-protein amino acid mimosine and is thought to have evolved from cystathionine β-lyase (CBL) via gene duplication. However, no study has, to date, compared the molecular characteristics of mimosinase and CBL. We therefore cloned mimosinase and CBL from the Mimosoideae subfamily member Mimosa pudica (Mp) and explored the molecular relationship between mimosinase and CBL for the first time. The recombinant Mp mimosinase degraded both mimosine and cystathionine with a much higher turnover number (k cat ) for mimosine compared with cystathionine, and Mp CBL utilized only cystathionine as a substrate. The critical residues implicated in the substrate binding of Arabidopsis thaliana CBL (Tyr-127, Arg-129, Tyr-181, and Arg-440) were highly conserved in both Mp mimosinase and CBL. However, homology modeling and molecular simulation of these enzymes predicted variations in the residues that interact with substrates. A mutation experiment on Mp mimosinase revealed that the disruption of a disulfide bond in the vicinity of the pyridoxal-5'-phosphate domain increased the enzyme's preference toward cystathionine. Treatment of Mp mimosinase with a disulfide-cleavage agent also decreased mimosinase activity. Furthermore, mutation near the conserved binding residue altered the substrate preference between mimosine and cystathionine. Molecular dynamics simulations of Mp mimosinase suggested a closer coordination of the residues that interact with mimosine at the active site compared with cystathionine, indicating a more compact pocket size for mimosine degradation. This study thus may provide new insights into the molecular diversification of CBL, a C-S lyase, into the C-N lyase mimosinase in the Mimosoideae subfamily.

Plant hormones and the circadian rhythm have been implicated in coordinated control of isoprene emission in plants. To gain insights into the signalling networks, foliar application of plant hormones was conducted in a native emitter, Ficus septica. Spraying of 50 μM jasmonic acid (JA) gradually decreased isoprene emission by 88% compared with initial levels within 5 days, and emission increased after relief from JA application. We further explored the molecular regulatory mechanism of isoprene emission by analysing photosynthetic rate, gene expression of 2-C-methyl-D-erythrytol 4-phosphate (MEP) pathway, hormone signalling and circadian rhythm processes, and metabolite pool sizes of MEP pathway. Results show that isoprene emission strongly correlated with isoprene synthase (IspS) gene expression and IspS protein levels over the period of JA treatment, indicating transcriptional and possible translational modulation of IspS by JA. Application of JA coordinately modulated genes in the auxin, cytokinin (CK), and circadian rhythm signal transduction pathways. Among the transcriptional factors analysed, MYC2 (JA) and LHY (circadian clock) negatively correlated with isoprene emission. Putative cis-elements predicted on IspS promoter (G-box for MYC2 and circadian for LHY) supports our proposal that isoprene emission is regulated by coordinated transcriptional modulation of IspS gene by phytohormone and circadian rhythm signalling., (© 2019 John Wiley & Sons Ltd.)

Isoprene is emitted by many plants and is thought to function as an antioxidant under stressful conditions. However, the detailed regulatory mechanism of isoprene emission in relation to the antioxidant system remains unclear. Therefore, in this study, we explored the molecular regulatory mechanism of isoprene emission under short-term drought stress in the tropical tree Ficus septica Burm.f. We found that the soil moisture content gradually decreased from 55% on Day 1 (D1) to 23% (wilting point) on D5 after withholding water for 4 days and then returning to the initial level following re-watering on D6. On D5, drought-stressed plants had more than twofold higher isoprene emission and 90.6% lower photosynthesis rates, 99.5% lower stomatal conductance and 82.3% lower transpiration rates than well-watered control plants. It was also estimated that the isoprene concentration inside the leaf greatly increased on D5 due to the increased isoprene emission rate and reduced stomatal conductance. Among the traits related to the 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway, which is responsible for isoprene biosynthesis, the isoprene synthase (IspS) protein level was positively correlated with the isoprene emission rate in stressed plants. The transcripts of the antioxidant genes peroxidase 2 (POD2), POD4, copper-zinc superoxide dismutase 2 (Cu-ZnSOD2) and manganese superoxide dismutase 1 (Mn-SOD1) also increased during the drying period, while those of ascorbate peroxidase 1 (APX1) decreased. However, there was only a weak correlation between isoprene emission and antioxidant enzyme gene expression, indicating that the regulation of isoprene biosynthesis is not directly linked to the antioxidant defense network in drought-stressed F. septica. These findings suggest that the post-transcriptional regulation of IspS led to the observed change in isoprene emission rate, which enhanced the quenching of reactive oxygen species (ROS) and, in combination with the increased antioxidant enzyme activity, conferred tolerance to drought stress in this species., (© The Author(s) 2018. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Nitric oxide (NO) is an important plant signaling molecule that has a vital role in abiotic stress tolerance. In the present study, we assessed drought-induced (15 and 30% PEG, polyethylene glycol) damage in wheat ( Triticum aestivum L. cv. Prodip) seedlings and mitigation by the synergistic effect of exogenous Arg (0.5 mM l-Arginine) and an NO donor (0.5 mM sodium nitroprusside, SNP). Drought stress sharply decreased the leaf relative water content (RWC) but markedly increased the proline (Pro) content in wheat seedlings. Drought stress caused overproduction of reactive oxygen species (ROS) and methylglyoxal (MG) due to the inefficiency of antioxidant enzymes, the glyoxalase system, and the ascorbate-glutathione pool. However, supplementation with the NO donor and Arg enhanced the antioxidant defense system (both non-enzymatic and enzymatic components) in drought-stressed seedlings. Application of the NO donor and Arg also enhanced the glyoxalase system and reduced the MG content by increasing the activities of the glyoxalase system enzymes (Gly I and Gly II), which restored the leaf RWC and further increased the Pro content under drought stress conditions. Exogenous NO donor and Arg application enhanced the endogenous NO content, which positively regulated the antioxidant system and reduced ROS production. Thus, the present study reveals the crucial roles of Arg and NO in enhancing drought stress tolerance in wheat seedlings by upgrading their water status and reducing oxidative stress and MG toxicity., Competing Interests: Compliance with ethical standardsThe authors declare no conflict of interest.

This study examined the effect of triterpenoid on the salt tolerance of lanosterol synthase deficient yeast mutant GIL77. The expression of the triterpenoid synthase gene under GAL1 promoter in GIL77 increased the triterpenoid concentration of both whole cell and plasma membrane fractions. Without the induction of the genes, the growth curve of BgbAS or RsM1 transformant depicted patterns similar to control cells in both the presence and absence of salt with growth inhibition at 500 mM NaCl. The induction of BgbAS and RsM1 gene expression slightly repressed growth compared with control cells in the absence of NaCl. The growth of GIL77 was significantly suppressed by the expression of BgbAS or RsM1 under salinity conditions. Of the triterpenoid synthase genes, BgbAS rather than RsM1 was found to strongly inhibit the growth of GIL77 cells under salt stressed conditions. The expression of the triterpenoid synthase gene in GIL77 also influenced their tolerance to other abiotic stresses. In contrast to the endogenous synthesis, the exogenous supply of triterpenoid in the culture medium appeared to occur in the plasma membrane fraction and enhanced the salt tolerance of GIL77. This study thus discussed the physiological significance of triterpenoid in relation to its possible role in modulating salt tolerance.

Being sessile organisms, plants are frequently exposed to various environmental stresses that cause several physiological disorders and even death. Oxidative stress is one of the common consequences of abiotic stress in plants, which is caused by excess generation of reactive oxygen species (ROS). Sometimes ROS production exceeds the capacity of antioxidant defense systems, which leads to oxidative stress. In line with ROS, plants also produce a high amount of methylglyoxal (MG), which is an α-oxoaldehyde compound, highly reactive, cytotoxic, and produced via different enzymatic and non-enzymatic reactions. This MG can impair cells or cell components and can even destroy DNA or cause mutation. Under stress conditions, MG concentration in plants can be increased 2- to 6-fold compared with normal conditions depending on the plant species. However, plants have a system developed to detoxify this MG consisting of two major enzymes: glyoxalase I (Gly I) and glyoxalase II (Gly II), and hence known as the glyoxalase system. Recently, a novel glyoxalase enzyme, named glyoxalase III (Gly III), has been detected in plants, providing a shorter pathway for MG detoxification, which is also a signpost in the research of abiotic stress tolerance. Glutathione (GSH) acts as a co-factor for this system. Therefore, this system not only detoxifies MG but also plays a role in maintaining GSH homeostasis and subsequent ROS detoxification. Upregulation of both Gly I and Gly II as well as their overexpression in plant species showed enhanced tolerance to various abiotic stresses including salinity, drought, metal toxicity, and extreme temperature. In the past few decades, a considerable amount of reports have indicated that both antioxidant defense and glyoxalase systems have strong interactions in conferring abiotic stress tolerance in plants through the detoxification of ROS and MG. In this review, we will focus on the mechanisms of these interactions and the coordinated action of these systems towards stress tolerance., Competing Interests: The authors declare no conflict of interest.

We identified a nitidine- (NTD-) accumulating organelle and evaluated the net cytotoxicity of accumulated NTD. To evaluate tumor cell selectivity of the drug, we evaluated its selective cytotoxicity against 39 human cancer cell lines (JFCR39 panel), and the profile was compared with those of known anticancer drugs. Organelle specificity of NTD was visualized using organelle-targeted fluorescent proteins. Real-time analysis of cell growth, proliferation, and cytotoxicity was performed using the xCELLigence system. Selectivity of NTD in the JFCR39 panel was evaluated. Mitochondria-specific accumulation of NTD was observed. Real-time cytotoxicity analysis suggested that the mechanism of NTD-induced cell death is independent of the cell cycle. Short-term treatment indicated that this cytotoxicity only resulted from the accumulation of NTD into the mitochondria. The results from the JFCR39 panel indicated that NTD-mediated cytotoxicity resulted from unique mechanisms compared with those of other known anticancer drugs. These results suggested that the cytotoxicity of NTD is only induced by its accumulation in mitochondria. The drug triggered mitochondrial dysfunction in less than 2 h. Similarity analysis of the selectivity of NTD in 39 tumor cell lines strongly supported the unique tumor cell specificity of NTD. Thus, these features indicate that NTD may be a promising antitumor drug for new combination chemotherapies.

Isoprene emission from plants is very sensitive to environmental temperature both at short-term and long-term scales. Our previous study demonstrated suppression of isoprene emission by cold temperatures in a high emitting tropical tree Ficus septica and revealed a strong correlation of emission to isoprene synthase (IspS) protein levels. When challenged with decreasing daily temperatures from 30 to 12 °C, F. septica completely stopped isoprene emission at 12 °C, only to recover on the second day after re-exposure to 30 °C. Here, we explored this regulation of isoprene emission in response to environmental temperature by a comprehensive analysis of transcriptome data, gene expressions and metabolite pools of the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway. MEP pathway genes and metabolites dynamics did not support substrate-level limitations as major control over observed basal emission, but transcriptome data, network inferences and putative regulatory elements on IspS promoter suggested transcriptional regulation of IspS gene through circadian rhythm and phytohormone signalling processes. Expression levels of 29 genes involved in these pathways were examined by quantitative real-time PCR. We propose that temperature controls over basal isoprene emission at a time-scale of hours to few days are regulated by phytohormone-mediated transcriptional modulation of IspS gene under synchronization by the circadian clock., (© 2016 John Wiley & Sons Ltd.)

Isoprene is the most abundant type of nonmethane, biogenic volatile organic compound in the atmosphere, and it is produced mainly by terrestrial plants. The tropical tree species Ficus septica Burm. F. (Rosales: Moraceae) has been shown to cease isoprene emissions when exposed to temperatures of 12 °C or lower and to re-induce isoprene synthesis upon subsequent exposure to temperatures of 30 °C or higher for 24 h. To elucidate the regulation of genes underlying the disabling and then induction of isoprene emission during acclimatization to ambient temperature, we conducted gene expression analyses of F. septica plants under changing temperature using quantitative real-time polymerase chain reaction and western blotting. Transcription levels were analyzed for 17 genes that are involved in metabolic pathways potentially associated with isoprene biosynthesis, including isoprene synthase (ispS). The protein levels of ispS were also measured. Changes in transcription and protein levels of the ispS gene, but not in the other assessed genes, showed identical temporal patterns to isoprene emission capacity under the changing temperature regime. The ispS protein levels strongly and positively correlated with isoprene emission capacity (R(2) = 0.92). These results suggest that transcriptional regulation of ispS gave rise to the temporal variation in isoprene emission capacity in response to changing temperature., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: The leaves of Cirsium brevicaule A. GRAY (CL) significantly decreased hepatic lipid accumulation and the expression of fatty acid synthase gene (FASN) in mice., Purpose: We aimed to purify and identify the active compound(s) from CL and determine the inhibitory mechanism of expression of FASN., Methods: We purified monogalactosyldiacylglycerol (MGDG) from extracts of CL (CL-MGDG) and showed that it was the active CL component through analyses of its effects on the expression of genes of human breast cancer cell line, SKBR-3., Results: The content and fatty acid composition of CL-MGDG are distinctly different from those of other vegetable-derived MGDGs. Treatment of SKBR-3 cells with MGDG decreased the level of FASN mRNA as well as the levels of mRNA encoding other protein involved in lipogenesis. Further, MGDG treatments significantly inhibited luciferase activities of constructs containing liver X receptor response element in FASN promoter region without altering the levels of mRNA encoding transcription factors. MGDG and the FASN inhibitor C75 decreased the viabilities of SKBR-3 cells in a concentration-dependent manner. CL-MGDG more potently inhibited cell viability than a commercial MGDG preparation., Conclusions: CL represents a good source of glycoglycerolipids with potential as functional ingredients of food., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Three isoprene synthase (IspS) cDNA clones have been isolated from tropical trees (Ficus septica, F. virgata, and Casuarina equisetifolia), and their enzyme properties have been compared with those of Populus alba IspS. Phylogenetic analysis of the deduced amino acid sequences with known monoterpene synthase resolved IspS from F. septica and F. virgata and other IspSs in a clade together with TPS-b clade I, whereas IspS from C. equisetifolia was within another clade, sister to TPS-b clade II. The optimum reaction temperature was 40 °C for the IspSs isolated from the tropical trees, and 45 °C for P. alba IspS. The optimum pH of the IspSs from the tropical trees peaked between pH 8 and pH10 contrasting with the rather broad optimum pH (7.5-10.5) of P. alba IspS. IspSs from F. septica and F. virgata were activated solely by Mg(2+), whereas IspS from C. equisetifolia was dependent more on Mn(2+) than on Mg(2+). Michaelis constant (Km) values of IspSs from tropical trees were lower than that of P. alba IspS. Analysis of inter fragment interaction energy of IspS-substrate complex model and crystal structure of bornyl diphosphate synthase (1N20) found that the coordination geometry of amino acids with higher attraction force is similar at the active site of C. equisetifolia IspS and bornyl diphosphate synthase. These observations suggest the occurrence of another group of IspSs in TPS-b subfamily and extend the knowledge on biochemical regulatory mechanism of isoprene emission from tropical trees.

Inflammation and immune response profoundly influence metabolic syndrome and fatty acid metabolism. To analyze influence of systemic inflammatory response to metabolic syndrome, we inoculated an attenuated vaccine strain of Mycobacterium bovis Bacillus Calmette-Guérin (BCG) into leptin-deficient ob/ob mice. BCG administration significantly decreased epididymal white adipose tissue weight, serum insulin levels, and a homeostasis model assessment of insulin resistance. Serum high molecular weight (HMW) adiponectin level and HMW/total adiponectin ratio of the BCG treated mice were significantly higher than those of control mice. Hepatic triglyceride accumulation and macrovesicular steatosis were markedly alleviated, and the enzymatic activities and mRNA levels of lipogenic-related genes in liver were significantly decreased in the BCG injected mice. We also exposed human hepatocellular carcinoma HepG2 cells to high levels of palmitate, which enhanced endoplasmic reticulum stress-related gene expression and impaired insulin-stimulated Akt phosphorylation (Ser473). BCG treatment ameliorated both of these detrimental events. The present study therefore suggested that BCG administration suppressed development of nonalcoholic fatty liver disease, at least partly, by alleviating fatty acid-induced insulin resistance in the liver.

Objectives: Partially purified hexane phase (HP) of Peucedanum japonicum Thunb (PJT) was identified as an antiobesity candidate. However, the compound responsible for the antiobesity activity remained unknown. Thus, in this study we isolated the active compound, to determine the mechanisms related to antiobesity activity in vitro., Methods: The HP was fractionated, and the effect on the triacylglycerol (TG) content was evaluated in 3T3-L1 preadipocytes and HepG2 hepatocytes. On the basis of comprehensive spectroscopic analyses, the structure of the active compound was identified as pteryxin, a known compound in PJT. However, to our knowledge, its biological activities against obesity have not been reported previously. The dose-dependent effect on the TG content, and the gene expressions related to adipogenesis, fatty acid catabolism, energy expenditure, lipolysis, and lipogenesis due to pteryxin (10, 15, and 20 μg/mL) were examined in vitro., Results: Pteryxin dose dependently suppressed TG content in both 3T3-L1 adipocytes (by 52.7%, 53.8%, and 57.4%, respectively; P < 0.05) and HepG2 hepatocytes (by 25.2%, 34.1%, and 27.4%, respectively; P < 0.05). Sterol regulatory element-binding protein-1 (SREBP-1c), fatty acid synthase (FASN), and acetyl-coenzyme A carboxylase-1 (ACC1) were down-regulated in pteryxin-treated 3T3-L1 adipocytes (by 18%, 36.1%, and 38.2%, P < 0.05) and HepG2 hepatocytes (by 72.3%, 62.9%, and 38.8%, respectively; P < 0.05). The adipocyte size marker gene, paternally expressed gene1/mesoderm specific transcript (MEST) was down-regulated (by 42.8%; P < 0.05), and hormone-sensitive lipase, a lipid catabolizing gene was up-regulated (by 15.1%; P < 0.05) in pteryxin-treated adipocytes. The uncoupling protein 2 (by 77.5%; P < 0.05) and adiponectin (by 76.3%; P > 0.05) were up-regulated due to pteryxin., Conclusion: Our study demonstrated that pteryxin in PJT plays the key role in regulating the lipid metabolism-related gene network and improving energy production in vitro. Thus, the results suggest pteryxin as a new natural compound to be used as an antiobesity drug in the pharmaceutical industry., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Ivy gourd (Coccinia grandis L. Voigt) is a tropical plant widely distributed throughout Asia, Africa, and the Pacific Islands. The anti-obesity property of this plant has been claimed but still remains to be scientifically proven. We therefore investigated the effects of ivy gourd leaf, stem, and root on adipocyte differentiation by employing cell culture model., Methods: Dried roots, stems, and leaves of ivy gourd were separately extracted with ethanol. Each extract was then applied to 3T3-L1 pre-adipocytes upon induction with a mixture of insulin, 3-isobutyl-1-methylxanthine, and dexamethasone, for anti-adipogenesis assay. The active extract was further fractionated by a sequential solvent partitioning method, and the resulting fractions were examined for their abilities to inhibit adipogenesis in 3T3-L1 cells. Differences in the expression of adipogenesis-related genes between the treated and untreated cells were determined from their mRNA and protein levels., Results: Of the three ivy gourd extracts, the root extract exhibited an anti-adipogenic effect. It significantly reduced intracellular fat accumulation during the early stages of adipocyte differentiation. Together with the suppression of differentiation, expression of the genes encoding PPARγ, C/EBPα, adiponectin, and GLUT4 were down-regulated. Hexane-soluble fraction of the root extract also inhibited adipocyte differentiation and decreased the mRNA levels of various adipogenic genes in the differentiating cells., Conclusions: This is the first study to demonstrate that ivy gourd root may prevent obesity based mainly on the ability of its active constituent(s) to suppress adipocyte differentiation in vitro. Such an inhibitory effect is mediated by at least down-regulating the expression of PPARγ-the key transcription factor of adipogenesis in pre-adipocytes during their early differentiation processes.

Objective: Crude ethanolic extract of Peucedanum japonicum Thunb (PJT) has been identified as an antiobesity and antidiabetic candidate. The aim of this study was to assess the mechanisms related to the antiobesity activity of partially purified PJT ethanol extract in vitro., Methods: PJT ethanol extract (EE) was partitioned into hexane phase (HP) and residual water phase. The effect of PJT extracts on triacylglycerol (TG) content, lipid metabolism-related gene expression, and energy expenditure was assessed in vitro in 3T3-L1, HepG2, and C2C12 cells. Furthermore, the active components in PJT extracts were partially profiled by high-performance liquid chromatography (HPLC) analysis., Results: The HP significantly down-regulated lipogenic gene expressions in hepatocytes, inhibited TG accumulation, and decreased the size of 3T3-L1 adipocytes. Moreover, the inhibition of lipid accumulation was at maximum during the adipocyte maturation stage. Furthermore, we found an increase in fat hydrolysis by HP in 3T3-L1. In C2C12 myotubes, the HP tended to enhance energy expenditure. HPLC analysis demonstrated that hydrophobic compounds available in the HP were responsible for antiobesity, which corresponded to the latter peaks on the HPLC chromatogram of EE., Conclusion: Our study demonstrated that the HP plays a crucial role in regulating lipid metabolism-related gene expressions and energy expenditure in vitro. The results thus provide insight on the activity of HP involved in suppressing obesity and its chemical entities., (Copyright © 2014 Elsevier Inc. All rights reserved.)

This paper describes the existence of temperature threshold in isoprene emission from tropical trees for the first time. Isoprene emission capacity of Ficus virgata leaves kept outdoors were measured over a period of 20 d in the wintertime. When the plants experienced the low temperature below 12 °C in the daytime, isoprene emission was completely suppressed, and re-activated by elevated temperature above the threshold of 12 °C. Photosynthesis is also decreased similarly, but too much smaller extent compared to the reduction observed for isoprene emission. The cut off level of accumulated temperature to shut off the isoprene emission was roughly estimated to be 300 degree/d. The shut off in the isoprene emission by the low temperature was confirmed experimentally by use of phytotron with tropical trees of F. virgata and F. septica. Isoprene emission of both species ceased at 12 °C, and re-activated by raising the temperature above the threshold level. Photosynthesis was lowered slightly as was the case for the field observation, and no close association between photosynthesis and isoprene emission was noted. The expression of isoprene synthase gene showed much greater variation after exposure to the cold temperature compared to those involved in photosynthesis. These observations therefore suggested the existence of putative direct connection between perception of temperature and isoprene emission in tropical trees, which may merit further investigation to estimate the net output of isoprene from subtropical or tropical forest., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The lignan arctigenin (ARG) from the herb Arctium lappa L. possesses anti-cancer activity, however the mechanism of action of ARG has been found to vary among tissues and types of cancer cells. The current study aims to gain insight into the ARG mediated mechanism of action involved in inhibiting proliferation and inducing apoptosis in lung adenocarcinoma cells. This study also delineates the cancer cell specificity of ARG by comparison with its effects on various normal cell lines. ARG selectively arrested the proliferation of cancer cells at the G0/G1 phase through the down-regulation of NPAT protein expression. This down-regulation occurred via the suppression of either cyclin E/CDK2 or cyclin H/CDK7, while apoptosis was induced through the modulation of the Akt-1-related signaling pathway. Furthermore, a GSH synthase inhibitor specifically enhanced the cytotoxicity of ARG against cancer cells, suggesting that the intracellular GSH content was another factor influencing the susceptibility of cancer cells to ARG. These findings suggest that specific cytotoxicity of ARG against lung cancer cells was explained by its selective modulation of the expression of NPAT, which is involved in histone biosynthesis. The cytotoxicity of ARG appeared to be dependent on the intracellular GSH level., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

Scope: Several recent studies have demonstrated that phospholipids (PLs) supplementation can modulate the function of cultured-immune cells. Furthermore, dietary PLs have been shown to ameliorate inflammatory processes and immune responses in arthritic and diabetic murine models, respectively. Thus, the aim of this study was to examine the immune-modulating activities of dietary soybean PLs in mice, with particular emphasis on the immune cell functions., Methods and Results: Mice were fed semisynthetic diets for 6 weeks, which contained either 7% soybean oil or 5% soybean oil plus 2% of either PL: phosphatidylcholine (PC), phosphatidylinositol (PI), or phosphatidylserine (PS). Production of concanavalin A (Con A)-induced proinflammatory cytokines was significantly decreased in the splenocytes isolated from mice fed PI compared to other lipids. Supplementation of the diet with PI, but not with the other lipids, significantly suppressed the proinflammatory cytokine serum levels and the development of Con A-induced liver damages., Conclusion: These observations suggest that dietary PI influenced immune functions, resulting in the prevention of pathogenesis and development of the liver injury in mice., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Various flavonoids obtained from the genus Cirsium have been reported to exhibit beneficial effects on health. The present study evaluated the antiobesity effects of Cirsium brevicaule A. GRAY leaf (CL) by using 3T3-L1 cells and C57BL/6 mice that were fed a high-fat diet (HFD)., Methods: Dried CL powder was serially extracted with solvents of various polarities, and these extracts were tested for antiadipogenic activity using 3T3-L1 adipocytes. Mice were fed experimental HFD supplemented with dried CL powder for 4 wk. Lipid levels and mRNA levels of genes related to lipid metabolism were determined in 3T3-L1 adipocytes and the white adipose tissue (WAT) and liver of mice fed on a HFD., Results: Treatment of 3T3-L1 adipocytes with a hexane extract of CL significantly reduced cellular lipid accumulation and expression of the fatty acid synthase (FASN) gene. Dietary CL reduced the serum levels of non-esterified fatty acids in HFD-fed mice. Significant decreases in subcutaneous WAT weight and associated FASN gene expression were observed in the mice fed the experimental CL diet. Dietary CL also reduced the hepatic lipid and serum levels of a hepatopathic indicator in the HFD-fed mice. A significant reduction in mRNA levels of FASN and HMG-CoA reductase were observed in the livers of the CL-diet group. Dietary CL, on the other hand, increased in the hepatic mRNA levels of genes related to β-oxidation, namely peroxisome proliferator-activated receptor α, calnitine palmitoyltrasferase 1A, and uncoupling protein 2. Expression of the insulin receptor gene was also significantly increased in the livers of mice-fed the CL diet., Conclusions: The present study therefore demonstrated that CL suppresses lipid accumulation in the WAT and liver partly through inhibiting mRNA levels of FASN gene and enhancing the lipolysis-related gene expression.

Background: Various mushrooms have been used in folk medicine for the treatment of lifestyle diseases in eastern countries, and several compounds that modulate the immune system, lower blood lipid levels, and inhibit tumor and viral action have been isolated. The fruiting body of Panellus serotinus (Mukitake) is recognized in Japan as one of the most delicious edible mushrooms, and previous studies have demonstrated that the dietary intake of powdered whole Mukitake or Mukitake extracts prevents the development of non-alcoholic fatty liver disease (NAFLD) in leptin-resistant db/db mice. In the present study, we evaluated the effect of the Mukitake diet on the pathogenesis of metabolic disorders in leptin-deficient ob/ob mice., Results: After 4 weeks of feeding, hepatomegaly, hepatic lipid accumulation, and elevated hepatic injury markers in the serum were markedly alleviated in Mukitake-fed ob/ob mice compared with control mice. Moreover, the mild hyperlipidemia in control ob/ob mice was attenuated and the elevated atherogenic index was reduced in Mukitake-fed ob/ob mice. These effects were partly attributable to the suppression of hepatic lipogenic enzyme activity due to the Mukitake diet., Conclusion: The current results showed that Mukitake supplementation is beneficial for the alleviation of NAFLD and dyslipidemia in obese, diabetic ob/ob mice.

Although it has been hypothesized that autoimmune-like phenomena may play a critical role in the protective immune responses to both human and animal malaria, there are still no evidence-based data to support this view. In this study we demonstrate that the majority of anti-single stranded (ss) DNA autoantibody secreting B cells were confined to B220(+)CD21(+)CD23(-) cells and that these cells expanded significantly in the spleen of C57BL/6 mice infected with Plasmodium yoelii 17X non-lethal (PyNL). To determine the role of ssDNA-binding autoantibody secreting B cell responses in murine malaria, we conjugated generation 6 (poly) amidoamine dendrimer nanoparticles with ssDNA to deplete ssDNA-binding autoreactive B cells in vivo. Our data revealed that 55.5% of mice died after DNA-coated nanoparticle-mediated in vivo depletion of ssDNA-specific autoreactive B cells and subsequent challenge using PyNL. Adoptive transfer of B cells with ssDNA specificity to mice, followed by PyNL infection, caused a later appearance and inhibition of parasitemia. The possible mechanism by which the ssDNA-binding autoantibody secreting B cells is involved in the protection against murine malaria has also been demonstrated., (Copyright © 2012 Elsevier GmbH. All rights reserved.)

Various subsets of T-cell receptor (TCR) γδ⁺ T cells expressing distinct TCR-Vγ chains are found in many different anatomical locations. These TCR γδ⁺ T cells perform multiple functions as an essential part of the immune system. In particular, protection against malaria infection by TCR γδ⁺ T cells is of special interest. In the present study, we confirmed that the Vγ7⁺ γδ T cells, which are an unconventional subset usually localized within the intestine, are recruited to the liver and spleen at the late stage of malaria infection, and contribute to protection against malaria infection via a multifunctional approach in apoptosis inhibitor-deficient mice., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Natural killer (NK) T cells are well known to play important roles in both tumor rejection and the defense against infectious. Therefore, the antitumor potential of NKT cell-activating antigens have been the focus for the development of NKT cell-based immunotherapies. Up to now, several studies have revealed that the administrations of glycolipids (e.g. α-galactosylceramide) can successfully treat certain metastatic tumors. However, liver injuries appeared upon the application of these antigens. We previously examined the potential of using β-glucosylceramide (β-GlcCer) to inhibit tumor metastasis to the liver. The aim of this study was to determine the antimetastatic effects of β-GlcCer and its impact on the activation of NKT cells. Intraperitoneal administration of β-GlcCer enhanced the production of interferon-γ from hepatic lymphocytes containing NKT cells, and increased the cytotoxicity of hepatic lymphocytes against tumor cells. Moreover, β-GlcCer administration suppressed the hepatic metastasis of tumors in wild type (WT) mice, but not in CD1d (-/-) or Jα18 (-/-) mice. The drawback associated with the other glycolipids in liver injury was not noted in WT mice treated with the continuous daily administration of β-GlcCer for 2 weeks. The present study demonstrated that β-GlcCer treatment activates invariant NKT cells, thus resulting in the inhibition of tumor metastasis.

Non-alcoholic fatty liver disease (NAFLD) is emerging as the most common liver disease in industrialised countries. Various mushrooms have been used in Eastern folk medicine for the treatment of lifestyle diseases. We previously found that the dietary intake of powdered whole Panellus serotinus (Mukitake) alleviates NAFLD in obese, diabetic db/db mice. In the present study, we investigated the influence of Mukitake fractional extracts on the development of NAFLD in db/db mice. A significant reduction in the hepatic TAG content, macrovesicular hepatocytes and activities of key enzymes for de novo synthesis of the fatty acid was observed in both the water-soluble Mukitake extract (WE) diet and the ethanol-soluble Mukitake extract (EE) diet groups compared with the control diet group of the db/db mice. The serum level of monocyte chemoattractant protein-1 (MCP-1), which is known to exacerbate insulin resistance, was significantly decreased in the WE group. On the other hand, the serum level of adiponectin, which plays a protective role against the metabolic syndrome, was significantly increased in the EE group. Additionally, differential analysis between Mukitake and Shiitake, mycelia from the same family, using liquid chromatography time-of-flight MS technology revealed that only seven and five compounds exist in WE and EE from Mukitake, respectively. In conclusion, the present study demonstrated that Mukitake displays at least two different physiological actions that alleviate NAFLD: one through the reduction in inflammatory damage by its suppression in MCP-1 production and the other through an increase in level of serum adiponectin and the prevention of visceral fat accumulation.

Supplementation of the diet with Peucedanum japonicum Thunb (PJT) powder inhibits high-fat diet-induced obesity in mice. Either the fiber component or other bioactive components in the PJT powder may inhibit obesity. This study, therefore, was an attempt to identify the components, fiber or other phytochemicals of PJT that were responsible for the anti-obesity activity, and also studied the modulation of obesity-related gene expression in C57BL/6 mice. Animals were fed a modified-AIN76 diet supplemented with PJT powder or extracts of PJT in water, 50 % ethanol, or ethanol. Body weight gain, tissue weight, serum biochemical parameters, liver lipid concentrations, and gene expression in tissues were compared between the control and treatment groups. Of the extracts, the ethanol extract of PJT decreased fat accumulation and adipocyte size, reduced serum and liver triglyceride concentrations, and inhibited obesity. This finding clearly demonstrates the presence of anti-obesity phytochemicals in PJT. Ethanol extract of PJT inhibited lipase activity in vitro. Modulation of gene expression by PJT ethanol extract was largely similar to that by PJT powder in the hepatic and adipose tissues: RORC and PBEF1 were upregulated and DUSP1, INSIG2, and SERPINA12 were downregulated in the liver; FXRα and PPARγ were upregulated and PEG1/MEST, the size-marker of adipocytes, was downregulated in the adipose tissue. Furthermore, PJT ethanol extract increased the expression of the UCP3 gene in muscle. These results suggest that the anti-obesity phytochemicals in PJT lower lipid levels by inhibiting fat absorption and by modulating obesity-related gene expression in the liver, adipose tissue, and muscle.

This study examined the antiobese activity of Peucedanum japonicum Thunb (PJT) in mice. In the first experiment, 4-week-old C57BL/6 mice were fed seven different diets containing 15% corn oil and 0-20% PJT powder for 4 weeks. Feeding the 10% and 20% PJT diet suppressed the body weight gain and the accumulation of abdominal and subcutaneous fats. PJT reduced serum and liver levels of triglyceride and serum levels of leptin in a dose-dependent manner. PJT intake decreased the proportion of saturated fatty acids and increased polyunsaturated and n-3 fatty acids in the liver. To obtain more insight into the antiobese activity of PJT, its effect on lipid absorption and enzyme activities related to lipid metabolism was studied in the second experiment. There was an increased faecal excretion of triglyceride in mice fed 5% and 10% PJT diets. Fatty acid synthase activity was decreased while carnitine palmitoyltransferase activity was increased by 10% PJT intake. These findings pointed to the usefulness of PJT for the development of a safe natural agent to reduce obesity or body weight for the first time. The rationale for the lipid lowering mechanism of PJT and the candidate compound responsible for the observations have also been discussed., (Copyright © 2010 John Wiley & Sons, Ltd.)

The present study describes the effect of Peucedanum japonicum Thunb (PJT) intake on the expression of obesity-related genes in mice fed a high-fat diet. To explore the mechanism underlying the effect of PJT, This study focused on the expression of genes, especially those related to obesity and metabolism syndrome, in the liver and adipose tissues. In agreement with our previous observations, intake of 10 % PJT for 4 weeks significantly reduced serum triglyceride (TG), leptin, abdominal fat, and adipocyte size. PJT also significantly increased fecal excretion of TG, decreased that of bile acid, and tended to increase the fecal excretion of total cholesterol. Microarray analysis was used to monitor changes in 324 metabolic syndrome-related genes in the liver. Statistically significant upregulation of PPP1R10, RORC, and PBEF1 genes and downregulation of DUSP1, INSIG2, and SERPINA12 genes were noted and confirmed by real-time RT-PCR. These changes were indicative of increased fatty acid oxidation in the maintenance of lipid homeostasis and insulin sensitivity in the livers of PJT-fed mice. PJT increased the expression of PPARγ, FXRα, DGAT1, and ATGL genes, suggesting an enhancement of adipocyte differentiation and normalization of functionality of adipose tissue.

The effects of a hot water extract and fractional extracts from rabbiteye blueberry (Vaccinium ashei reade) leaves (BBL) on lipid metabolism were studied in obese Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Feeding the hot water extract and fractional extracts from BBL alleviated hepatic triglyceride accumulation in the rats. Additionally, feeding with the flavonol glycoside (FG) and proanthocyanidin (PA) fractions lowered serum cholesterol levels in the obese rats. The results from measurements of the hepatic enzyme activity indicate that the hypolipidemic effects of the hot water extract and the PA fraction might be attributable to enhanced lipolysis in the liver. The reduced serum levels of C-reactive protein, an inflammatory cytokine, by the chlorogenic acid + rutin fraction and FG fraction might be associated with alleviating the metabolic abnormalities in obese rats. These results indicate that the BBL extracts, and especially FG and PA, exerted hypolipidemic effects on obese OLETF rats and suggest that an infusion of BBL can be useful as a dietary hypolipidemic component.

Nonalcoholic fatty liver disease (NAFLD) is emerging as the most common liver disease in industrialized countries. Thus, the discovery of food components that would ameliorate NAFLD is of interest. Various mushrooms have been used in folk medicine for the treatment of lifestyle diseases in eastern countries and several compounds that modulate immune system, lower blood lipid levels, inhibit tumor and viral action have been isolated from them. In this study, we tested whether feeding Panellus serotinus (Mukitake) to db/db mice protects them from hepatic injury. After 4 weeks of feeding, hepatomegaly, hepatic triglyceride accumulation and elevated hepatic injury markers in serum were markedly alleviated in Mukitake-fed db/db mice compared with control mice. These effects were partly attributable to the enhancement of lipolytic enzyme activity and the suppression of lipogenic enzyme activities due to the Mukitake diet. The severe hyperinsulinemia in control db/db mice tended to attenuate in Mukitake-fed mice due to an enhanced production of adiponectin, which improves insulin sensitivity. Moreover, production of monocyte chemoattractant protein 1 (MCP1), an inflammatory cytokine, was markedly suppressed by the Mukitake diet. In addition, water-soluble extracts of Mukitake powder showed an inhibitory effect on inhibitor of kappaB (IkappaB) kinase (IKK) beta, whose activation is required for nuclear factor kappaB (NFkappaB)-mediated inflammatory response. We speculate that the development and progression of NAFLD was prevented by the reduction of MCP1 production through the interference in the IKKbeta-NFkappaB signaling pathway in Mukitake-fed db/db mice., (Copyright 2010 Elsevier Inc. All rights reserved.)

To optimize the adequate atherogenic diet composition for nutritional atherosclerosis studies utilizing B6.KOR-Apoe(shl) mice, we investigated the effect of dietary cholesterol, AIN formula, and dietary fats on the development of atherosclerosis. Cholesterol supplementation (0.15-2%) for 12 weeks resulted in a dose-dependent increase in the development of atherosclerosis. Furthermore, there were no significant differences in the degree of atherosclerosis between B6.KOR-Apoe(shl) mice fed a modified-AIN-76 diet and those fed a modified-AIN-93M diet containing corn oil or soybean oil for 10 and 12 weeks. The present experiment indicates that the adequate dietary level of cholesterol was 0.15%, and that further studies are necessary to determine the optimal level of various types of dietary oils for nutritional atherosclerosis experiments in B6.KOR-Apoe(shl) mice.

The halophytic Kandelia candel and Bruguiera gymnorrhiza are ideal model for studying the molecular mechanisms of salinity tolerance in mangrove plants. The correlation between mRNA expression of four oxidosqualene cyclase (OSC) genes namely, KcMS multifunctional terpenoid synthase and KcCAS cyloartenol synthase (K. candel), BgbAS beta-amyrin synthase and BgLUS lupeol synthase (B. gymnorrhiza) and salt concentration was examined. mRNA level of KcMS was increased with salt concentration in both roots and leaves of K. candel. Similarly, salt stress increased the mRNA levels of BgLUS and BgbAS in the root of B. gymnorrhiza. This result suggests that the function of terpenoids in root is associated with the salt stress. In contrast to these observations, the mRNA level of KcCAS was not modulated by salt stress in the roots, and decreased in the leaves. These results therefore suggest that the terpenoids but not phytosterols play an important role to cope with the salt stress in mangrove root. The content and proportion of beta-amyrin and lupeol increased with salinity in the root of K. candel and B. gymnorrhiza, providing additional evidence for the protective role of terpenoids. However, beta-amyrin and lupeol in B. gymnorrhiza leaves decreased with salt concentration, suggesting that the physiological significance for the terpenoids in the leaf may differ from that for the root.

In the present study, we investigated the effect of phenolic compounds (PCs) and policosanol of Kokuto, Okinawan noncentrifugal cane sugar, on the development of atherosclerosis. A total of 67 male Japanese quail were divided into eight dietary groups in trial 1. The dietary groups were fed the atherosclerotic diet (AD) containing 5% corn oil, 2% cholesterol, and 30% sucrose or seven different types of Kokuto. Dietary intakes of Kokuto notably prevented the development of atherosclerosis. Furthermore, there was a significant negative correlation between the serum radical scavenging activity and the degree of atherosclerosis in the dietary groups. In trial 2, a total of 63 Japanese quail were fed AD with sucrose, Kokuto, PC extracts from Kokuto, wax extracts from sugar cane, octacosanol, vitamin C, and vitamin E. As a result, the supplementation of the diet with Kokuto and PCs significantly reduced the development of atherosclerosis as compared with the ingestion of AD with sucrose. In conclusion, these findings suggest that, among various components of Kokuto, PCs play a central role for the prevention of experimental atherosclerosis in Japanese quail.