Chamaecyparis lawsoniana and Its Active Compound Quercetin as Ca2+ Inhibitors in the Contraction of Airway Smooth Muscle.

The Cupressaceae family includes species considered to be medicinal. Their essential oil is used for headaches, colds, cough, and bronchitis. Cedar trees like Chamaecyparis lawsoniana (C. lawsoniana) are commonly found in urban areas. We investigated whether C. lawsoniana exerts some of its effects by modifying airway smooth muscle (ASM) contractility. The leaves of C. lawsoniana (363 g) were pulverized mechanically, and extracts were obtained by successive maceration 1:10 (w:w) with methanol/CHCl3. Guinea pig tracheal rings were contracted with KCl, tetraethylammonium (TEA), histamine (HIS), or carbachol (Cch) in organ baths. In the Cch experiments, tissues were pre-incubated with D-600, an antagonist of L-type voltage-dependent Ca2+ channels (L-VDCC) before the addition of C. lawsoniana. Interestingly, at different concentrations, C. lawsoniana diminished the tracheal contractions induced by KCl, TEA, HIS, and Cch. In ASM cells, C. lawsoniana significantly diminished L-type Ca2+ currents. ASM cells stimulated with Cch produced a transient Ca2+ peak followed by a sustained plateau maintained by L-VDCC and store-operated Ca2+ channels (SOCC). C. lawsoniana almost abolished this last response. These results show that C. lawsoniana, and its active metabolite quercetin, relax the ASM by inhibiting the L-VDCC and SOCC; further studies must be performed to obtain the complete set of metabolites of the extract and study at length their pharmacological properties.

Carbon stock projection for four major forest plantation species in Japan.

Carbon sequestration via afforestation and forest growth is effective for mitigating global warming. Accurate and robust information on forest growth characteristics by tree species, region, and large-scale land-use change is vital and future prediction of forest carbon stocks based on this information is of great significance. These predictions allow exploring forestry practices that maximize carbon sequestration by forests, including wood production. Forest inventories based on field measurements are considered the most accurate method for estimating forest carbon stocks. Japan's national forest inventories (NFIs) provide stand volumes for all Japanese forests, and estimates from direct field observations (m-NFIs) are the most reliable. Therefore, using the m-NFI from 2009 to 2013, we selected four major forest plantation species in Japan: Cryptomeria japonica, Chamaecyparis obtusa, Pinus spp., and Larix kaempferi and presented their forest age-carbon density function. We then estimated changes in forest carbon stocks from the past to the present using the functions. Next, we investigated the differences in the carbon sequestration potential of forests, including wood production, between five forestry practice scenarios with varying harvesting and afforestation rates, until 2061. Our results indicate that, for all four forest types, the estimates of growth rates and past forest carbon stocks in this study were higher than those considered until now. The predicted carbon sequestration from 2011 to 2061, assuming that 100 % of harvested carbon is retained for a long time, twice the rate of harvesting compared to the current rate, and a 100 % afforestation rate in harvested area, was three to four times higher than that in a scenario with no harvesting or replanting. Our results suggest that planted Japanese forests can exhibit a high carbon sequestration potential under the premise of active management, harvesting, afforestation, and prolonging the residence time of stored carbon in wood products with technology development.

Functional identification of specialized diterpene synthases from Chamaecyparis obtusa and C. obtusa var. formosana to illustrate the putative evolution of diterpene synthases in Cupressaceae.

Chamaecyparis obtusa and C. obtusa var. formosana of the Cupressaceae family are well known for their fragrance and excellent physical properties. To investigate the biosynthesis of unique diterpenoid compounds, diterpene synthase genes for specialized metabolite synthesis were cloned from C. obtusa and C. obtusa var. formosana. Using an Escherichia coli co-expression system, eight diterpene synthases (diTPSs) were characterized. CoCPS and CovfCPS are class II monofunctional (+)-copalyl diphosphate synthases [(+)-CPSs]. Class I monofunctional CoLS and CovfLS convert (+)-copalyl diphosphate [(+)-CPP] to levopimaradiene, CoBRS, CovfBRS1, and CovfBRS3 convert (+)-CPP to (-)-beyerene, and CovfSDS converts (+)-CPP to (-)-sandaracopimaradiene. These enzymes are all monofunctional diterpene syntheses in Cupressaceae family of gymnosperm, and differ from those in Pinaceae. The discovery of the enzyme responsible for the biosynthesis of tetracyclic diterpene (-)-beyerene was characterized for the first time. Diterpene synthases with different catalytic functions exist in closely related species within the Cupressaceae family, indicating that this group of monofunctional diterpene synthases is particularly prone to the evolution of new functions and development of species-specific specialized diterpenoid constituents.

The Precious Potential of the Sacred Tree Chamaecyparis obtusa (Siebold & Zucc.) Endl. as a Source of Secondary Metabolites with Broad Biological Applications.

Chamaecyparis obtusa (Siebold & Zucc.) Endl., which belongs to the Cupressaceae family, occurs naturally in North America and Asia, especially in Korea, Taiwan and Japan, where it is an evergreen, coniferous, sacred, ethnic tree. It has many useful varieties that are widespread throughout the world and grown for decorative purposes. It is most commonly used as an ornamental plant in homes, gardens or parks. It is also widely used in many areas of the economy; for example, its wood is used in architecture as well as furniture production. In addition, oil extracted from Chamaecyparis obtusa is increasingly used in cosmetology for skin care. Due to its wide economic demand, mainly in Japan, it represents the largest area of plantation forest. Despite this, it is on the red list of endangered species. Its use in ethnopharmacology has led to more and more research in recent years in an attempt to elucidate the potential mechanisms of its various biological activities, such as antimicrobial, antioxidant, anticancer, antidiabetic, antiasthmatic, anti-inflammatory, antiallergic, analgesic and central nervous system effects. It has also been shown that Chamaecyparis obtusa can be used as an insect repellent and an ingredient in plant disease treatment. This thesis provides a comprehensive review of the biological studies to date, looking at different areas of the economic fields of potential use of Chamaecyparis obtusa.

Anti-Inflammatory Effect of Chamaecyparis obtusa (Siebold & Zucc.) Endl. Leaf Essential Oil.

Chamaecyparis obtusa (Siebold & Zucc.) Endl. (C. obtusa) belongs to the Cupressaceae family and is native to East Asian regions. Essential oils extracted from the leaves, bark, branches, and roots of C. obtusa have both aesthetic and medicinal properties and are thus widely used. However, detailed analyses of the active ingredients of C. obtusa extract are lacking. In this study, the sabinene content in the hydro-distillation of C. obtusa leaf essential oil (COD) was analyzed using GC-MS, and the anti-inflammatory effect of COD was compared with that of pure sabinene. Cell viability was evaluated by MTT assay, and nitric oxide (NO) production was measured using Griess reagent. Relative mRNA and protein levels were analyzed using RT-qPCR and western blot, and secreted cytokines were analyzed using a cytokine array kit. The results showed that both COD and sabinene inhibited the expression of inducible nitric oxide synthase (iNOS) and the phosphorylation of c-Jun N-terminal kinase (JNK) and p38 in lipopolysaccharide (LPS)-induced RAW 264.7 cells. COD and sabinene also reduced the production of pro-inflammatory cytokines interleukin (IL)-1ß, IL-6, IL-27, IL-1 receptor antagonist (IL-1ra), and granulocyte-macrophage colony-stimulating factor (GM-CSF). The anti-inflammatory mechanisms of COD and sabinene partially overlap, as COD was shown to inhibit MAPKs and the JAK/STAT axis, and sabinene inhibited MAPKs, thereby preventing LPS-induced macrophage activation.

The component of the Chamaecyparis obtusa essential oil and insecticidal activity against Tribolium castaneum (Herbst).

Tribolium castaneum (Herbst) is a worldwide grain storage pest controlled by chemical control methods of phosphine fumigation, which results in many hazards, damages human health, makes pests resistant to pesticides, and pollutes the environment. In recent years, the popularity of botanical insecticides has continued to rise, and plant essential oils (EO) are considered potential alternatives for developing insecticides. In the current study, we selected the Chamaecyparis obtusa EO to determine its insecticidal effects and component analysis on T. castaneum. Through gas chromatography-ion mobility spectrometry (GC-IMS) technology, cedrol was the most obvious compound in the signal peak of the volatile components detected in the C. obtusa EO. The results of the bioassay showed that the C. obtusa EO had certain contact activity against T. castaneum, and the LD50 was 52.54 µg/adult. At three concentrations (0.41,1.62, 2.83 uL/cm2), the repellent rates of C. obtusa EO against T. castaneum were all above 80% at 15, 30, 60, and 120 min, respectively, indicating that the repellent effect was strong. Meanwhile, the C. obtusa EO exhibited fumigant toxicity against T. castaneum with LC50 values of 7.09 µg/L air. In addition, C. obtusa EO significantly increased the activity of AChE, CarE, POD, CAT, T-SOD, and chitinase in T. castaneum. Finally, the mechanism of C. obtusa EO on T. castaneum adults was explored based on transcriptome sequencing. We found that the DEGs focused on the chitin metabolic process and some aging genes in T. castaneum. Therefore, C. obtusa EO could be used as potential eco-friendly candidates for stored grain pest management.

Pharmacological anti-tumor effects of natural Chamaecyparis obtusa (siebold & zucc.) endl. Leaf extracts on breast cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Chamaecyparis obtusa (C. obtusa, cypress species) is a plant that grows mainly in the temperate Northern Hemisphere and has long been used as a traditional anti-inflammatory treatment in East Asia. C. obtusa contains phytoncides, flavonoids, and terpenes, which have excellent anti-cancer effects and have been reported to prevent the progression of various cancers. However, the detailed mechanisms underlying the anti-cancer effects of C. obtusa extracts are unknown. AIM OF THE STUDY: We sought to confirm the anti-cancer effects of C. obtusa leaf extracts and to reveal the mechanism of action, with the possibility of its application in the treatment or prevention of cancer. MATERIAL &METHODS: The cytotoxicity of C. obtusa leaf extracts was confirmed using an MTT assay. Intracellular changes in protein levels were measured by immunoblotting, and mRNA levels were measured with qRT-PCR. Wound healing assay and transwell migration assay were used to evaluate the metastatic potential of breast cancer cells. The extract-induced apoptosis was observed using IncuCyte Annexin V Red staining analysis. A syngeneic breast cancer mouse model was established by injecting 4T1-Luc mouse breast cancer cells into the fat pad of female BALB/c mice, and the extract was administered orally. Luciferin solution was injected intraperitoneally to assess primary tumor development and metastasis by bioluminescence. RESULTS: C. obtusa leaf extracts were extracted with boiling water, 70% EtOH, and 99% EtOH. Among the extracts, the 99% EtOH extract of C. obtusa leaf (CO99EL) most clearly inhibited the tyrosine phosphorylation of Signal Transducer and Activator of Transcription 3 (pY-STAT3) in MDA-MB-231 breast cancer cells at a concentration of 25 and 50 µg/mL. In addition, CO99EL strongly inhibited not only endogenous pY-STAT3 levels but also IL-6-induced STAT3 activation in various types of cancer cells, including breast cancer. CO99EL inhibited metastatic potential by downregulating the expression of N-cadherin, fibronectin, TWIST, MMP2, and MMP9 in MDA-MB-231 breast cancer cells. CO99EL also induced apoptotic cell death by increasing cleaved caspase-3 and decreasing anti-apoptotic proteins Bcl-2 and Bcl-xL. In an in vivo syngeneic breast cancer mouse model, 100 mg/kg CO99EL suppressed tumor growth and induced apoptosis of cancer cells. Moreover, CO99EL significantly inhibited lung metastasis from primary breast cancer. CONCLUSIONS: Our study demonstrated that 100 mg/kg CO99EL has potent anti-tumor effects against breast cancer, thus suggesting that 100 mg/kg CO99EL has potential applications in the treatment and prevention of breast cancer.

Biological Activity and Component Analyses of Chamaecyparis obtusa Leaf Extract: Evaluation of Antiwrinkle and Cell Protection Effects in UVA-Irradiated Cells.

Background and Objectives: Chamaecyparis obtusa (C. obtuse) extract has been used as a folk medicinal remedy in East Asian countries to alleviate inflammation and prevent allergies. Active oxygen causes skin aging and leads to skin cell and tissue damage. Extensive research has been conducted to control active oxygen generation to prevent skin aging. We evaluated the antioxidant activity and antiwrinkle effect of C. obtusa extract to determine its potential as a cosmetic material. Materials and Methods: The antioxidant activity of a 70% ethanol extract of C. obtusa (COE 70) and a water extract of C. obtusa (COW) was determined using 2,2-diphenyl-1-picrylhydrazy (DPPH) scavenging, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS+) scavenging, superoxide dismutase-like activity, xanthine oxidase inhibition, and ferric-reducing antioxidant power assays. The effective concentration of the extracts was determined using the methyl thiazolyl tetrazolium assay to evaluate their toxicity. The effects of COE 70 on the production of matrix metalloproteinases (MMPs) and procollagen, and expression of activated cytokines, interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α), in UVA-irradiated fibroblasts were determined using quantitative real-time PCR. Additionally, quercitrin, amentoflavone, hinokiflavone, and myricetin concentrations in COE 70 were determined using high-pressure high-performance liquid chromatography. Results: COE 70 had higher polyphenol and flavonoid concentrations than COW and exhibited an excellent antioxidant effect. COE 70 suppressed UVA-induced fibroblast death by 21.3% at 25 µg/mL. It also increased MMP-1, MMP-3, TNF-α, and IL-6 mRNA levels at 5-25 µg/mL compared with those in control UVA-irradiated fibroblasts. Moreover, mRNA levels of collagen type I and superoxide dismutase significantly increased, indicating the antiwrinkle and anti-inflammatory effects of the extract. Among the COE 70 components, quercitrin concentration was the highest; hence, quercitrin could be an active ingredient. Conclusions: COE 70 could be used as a natural antioxidant and antiwrinkle agent.

Essential Oils Composition and Biological Activity of Chamaecyparis obtusa, Chrysopogon nigritanus and Lavandula coronopifolia Grown Wild in Sudan.

Generally, there are scant data about the constituents and eventually the biological activity of essential oils (EOs) from aromatic plants that grow naturally in Sudan. The present study aimed to determine the chemical composition, and antioxidant and enzyme inhibitory activities of EO extracted from the fruit of Chamaecyparis obtusa (Siebold and Zucc.) Endl. (family Cupressaceae), root of Chrysopogon nigritanus (Benth.) Veldkampis (family Poaceae) and aerial part of Lavandula coronopifolia Poir (family Lamiaceae). The fruit of C. obtusa contained only monoterpenes, mainly hydrogenated ones, with α-pinene (69.07%) as the major component. Oxygenated sesquiterpenes comprised the highest content of the C. nigritanus root EO with cedr-8-en-15-ol (28.69%) as the major constituent while aerial parts of L. coronopifolia contained both monoterpenes and sesquiterpenes and the oxygenated monoterpene lavandulol (26.56%) as dominant compounds. The EO of the root of C. nigritanus significantly displayed (p < 0.05) the highest anti-DPPH radical, Fe3+- and Cu2+-reducing and metal-chelating activities, while that of C. obtusa fruit significantly exerted (p < 0.05) the best anti-ABTS radical and total antioxidant activity. The two EOs significantly exhibited (p < 0.05) the highest anti-acetylcholinesterase and -butyrylcholinesterase activities, respectively, while EO of L. coronopifolia was the only oil to show a considerable inhibitory effect against the tyrosinase and α-glucosidase enzymes. In conclusion, EOs from these three plants could be natural agents with promising functional properties for food, cosmetics, and pharmaceutical applications.

Intra-annual fluctuation in morphology and microfibril angle of tracheids revealed by novel microscopy-based imaging.

Woody cells, such as tracheids, fibers, vessels, rays etc., have unique structural characteristics such as nano-scale ultrastructure represented by multilayers, microfibril angle (MFA), micro-scale anatomical properties and spatial arrangement. Simultaneous evaluation of the above indices is very important for their adequate quantification and extracting the effects of external stimuli from them. However, it is difficult in general to achieve the above only by traditional methodologies. To overcome the above point, a new methodological framework combining polarization optical microscopy, fluorescence microscopy, and image segmentation is proposed. The framework was tested to a model softwood species, Chamaecyparis obtusa for characterizing intra-annual transition of MFA and tracheid morphology in a radial file unit. According our result, this framework successfully traced the both characteristics tracheid by tracheid and revealed the high correlation (|r| > 0.5) between S2 microfibril angles and tracheidal morphology (lumen radial diameter, tangential wall thickness and cell wall occupancy). In addition, radial file based evaluation firstly revealed their complex transitional behavior in transition and latewood. The proposed framework has great potential as one of the unique tools to provide detailed insights into heterogeneity of intra and inter-cells in the wide field of view through the simultaneous evaluation of cells' ultrastructure and morphological properties.

Cloning and functional characterization of three sesquiterpene synthase genes from Chamaecyparis formosensis Matsumura.

Terpene synthase (TPS) analysis may contribute to a better understanding of terpenoids biosynthesis and the evolution of phylogenetic taxonomy. Chamaecyparis formosensis Matsumura is an endemic and valuable conifer of Taiwan. Its excellent wood quality, fragrance, and durability make it become the five precious conifers in Taiwan. In this study, three sesquiterpene synthase genes that belong to the TPS-d2 clade were isolated and characterized through in vitro reaction of recombinant protein and in vivo reaction of Escherichia coli heterologous expression system. The main product of Cf-GerA was germacrene A using GC/MS analysis, while the product of Cf-Aco and Cf-Gor were identified as acora-4(14),8-diene and (5R,6R,10S)-α-gorgonene by using NMR analysis. These are the first reported enzymes that biosynthesize acora-4(14),8-diene and (5 R,6 R,10 S)-α-gorgonene. Both sesquiterpene synthases may isomerize the farnesyl pyrophosphate substrate to nerolidyl pyrophosphate for further cyclization. Cf-Aco may catalyze 1,6-cyclization of nerolidyl cation while Cf-Gor may catalyze through an uncharged intermediate, isogermacrene A.

Late phase radiocesium dynamics in Fukushima forests post deposition.

The long term dynamics of radiocesium in typical forest ecosystems was studied in the radioactive contaminated areas in Fukushima Prefecture. Six observations sites located in Yamakiya Village (Kawamata Town; since 2014), Tsushima Village (Namie Town, since 2015), and Tomioka Town (since 2017) were monitored. The forests consisted of artificial plantations of Japanese cedar (Cryptomeria japonica) at Yamakiya Village, Tsushima Village, and Tomioka Town. Tsushima Village also had a natural mixed forest dominated by Japanese red pine (Pinus densiflora), and Tomioka Town had a young and a mature artificial plantation of Japanese cypress (Chamaecyparis obtuse). Concentrations of 137Cs were monitored in the samples collected from the main aboveground biomass compartments, fresh litterfall, forest litter, and soil. Concentrations of exchangeable forms of 137Cs and stable K were measured in soil samples. During the observation period, the litter radiocesium inventories at all sites decreased significantly to approximately 1% or less of the total ground deposition. Approximately 80% of the total radiocesium inventory is localized in the upper 5-cm layer of soil and there is little downward migration of radiocesium. At the sites with the longest monitoring series (Yamakiya and Tsushima), the radiocesium expectation depths and expectation mass depths were relatively constant at 2-3 cm and 5-6 kg m-2, respectively. Aboveground biomass compartments showed similar decreasing trends in radiocesium aggregated transfer factors, Tag, in the compartments that were exposed to atmospheric fallout in March 2011 (old foliage, small branches, and outer bark). The mean Tag in cedar stand compartments currently are in the range of 10-3-10-2 m2 kg-1 dw. However, the mean Tag and their dynamic trend significantly differed in the wood compartments of the cedar stands, which may indicate root uptake differences of orders of magnitude between observation sites. The difference in radiocesium concentration in wood between the sites becomes less pronounced when normalized by the ratio of exchangeable 137Cs/K in the soils.

Incomplete lineage sorting and local extinction shaped the complex evolutionary history of the Paleogene relict conifer genus, Chamaecyparis (Cupressaceae).

Inferring accurate biogeographic history of plant taxa with an East Asia (EA)-North America (NA) is usually hindered by conflicting phylogenies and a poor fossil record. The current distribution of Chamaecyparis (false cypress; Cupressaceae) with four species in EA, and one each in western and eastern NA, and its relatively rich fossil record, make it an excellent model for studying the EA-NA disjunction. Here we reconstruct phylogenomic relationships within Chamaecyparis using > 1400 homologous nuclear and 61 plastid genes. Our phylogenomic analyses using concatenated and coalescent approaches revealed strong cytonuclear discordance and conflicting topologies between nuclear gene trees. Incomplete lineage sorting (ILS) and hybridization are possible explanations of conflict; however, our coalescent analyses and simulations suggest that ILS is the major contributor to the observed phylogenetic discrepancies. Based on a well-resolved species tree and four fossil calibrations, the crown lineage of Chamaecyparis is estimated to have originated in the upper Cretaceous, followed by diversification events in the early and middle Paleogene. Ancestral area reconstructions suggest that Chamaecyparis had an ancestral range spanning both EA and NA. Fossil records further indicate that this genus is a relict of the "boreotropical" flora, and that local extinctions of European species were caused by global cooling. Overall, our results unravel a complex evolutionary history of a Paleogene relict conifer genus, which may have involved ILS, hybridization and the extinction of local species.

SSR individual identification system construction and population genetics analysis for Chamaecyparis formosensis.

Chamaecyparis formosensis is an endemic species of Taiwan, threatened from intensive use and illegal felling. An individual identification system for C. formosensis is required to provide scientific evidence for court use and deter illegal felling. In this study, 36 polymorphic simple sequence repeat markers were developed. By applying up to 28 non-linked of the developed markers, it is calculated that the cumulative random probability of identity (CPI) is as low as 1.652 × 10-12, and the identifiable population size is up to 60 million, which is greater than the known C. formosensis population size in Taiwan. Biogeographical analysis data show that C. formosensis from four geographic areas belong to the same genetic population, which can be further divided into three clusters: SY (Eastern Taiwan), HV and GW (Northwestern Taiwan), and MM (Southwestern Taiwan). The developed system was applied to assess the provenance of samples with 88.44% accuracy rate and therefore can serve as a prescreening tool to reduce the range required for comparison. The system developed in this study is a potential crime-fighting tool against illegal felling.

Transcriptome analysis of gibberellins and abscisic acid during the flooding response in Fokienia hodginsii.

Flooding is one of the main abiotic stresses suffered by plants. Plants respond to flooding stress through regulating their morphological structure, endogenous hormone biosynthesis, and genetic signaling transduction. We previously found that Fokienia hodginsii varieties originating from Gutian exhibited typical flooding tolerance traits compared to three other provenances (Yongzhou, Sanming, Nanping), expressed as increased height, longer diameter at breast height (DBH), and smaller branch angle. Herein, the changes in endogenous gibberellins (GA) and abscisic acid (ABA) contents were measured under flooding stress in F. hodginsii, and ABA was found to decrease, whereas GA increased with time. Furthermore, the GA and ABA contents of the varieties originating from Gutian and the three other provenances were measured, and the results indicated that F. hodginsii from Gutian could respond more rapidly to flooding stress. The transcriptomes of the varieties originating from Gutian and the other three provenances were compared using RNA sequencing to explore the underlying genetic mechanisms of the flood-resistant phenotypes in F. hodginsii. The results indicated that two flood-stress response genes (TRINITY_DN142_c0_g2 and TRINITY_DN7657_c0_g1) were highly related to both the ABA and GA response in F. hodginsii.

Secondary Metabolites with Antimicrobial Activities from Chamaecyparis obtusa var. formosana.

Seven new compounds, including one dimer novel skeleton, chamaecyformosanin A (1); three diterpenes, chamaecyformosanins B-D (2-4); one sesquiterpene, chamaecyformosanin E (5); and two monoterpenes, chamaecyformosanins F and G (6 and 7) were isolated from the methanol extract of the bark of Chamaecyparis obtusa var. formosana. Their structures were established by the mean of spectroscopic analysis and the comparison of NMR data with those of known analogues. Their structures were elucidated on the basis of physicochemical evidence, in-depth NMR spectroscopic analysis, and high-resolution mass spectrometry. Furthermore, the isolated compounds were subjected to an evaluation of their antimicrobial activity. Metabolites 1, 3, and 4 present antibacterial activities. It is worth mentioning that the chemical composition of the bark of C. obtusa var. formosana has never been studied in the past. This is the first time the barks from C. obtusa var. formosana were studied and two new skeleton compounds, 1 and 7, were obtained.

Drought affects the fate of non-structural carbohydrates in hinoki cypress.

Tree species that close stomata early in response to drought are likely to suffer from an imbalance between limited carbohydrate supply due to reduced photosynthesis and metabolic demand. Our objective was to clarify the dynamic responses of non-structural carbohydrates to drought in a water-saving species, the hinoki cypress (Chamaecyparis obtusa Sieb. et Zucc.). To this end, we pulse-labeled young trees with 13CO2 10 days after the beginning of the drought treatment. Trees were harvested 7 days later, early during drought progression, and 86 days later when they had suffered from a long and severe drought. The labeled carbon (C) was traced in phloem extract, in the organic matter and starch of all the organs, and in the soluble sugars (sucrose, glucose and fructose) of the most metabolically active organs (foliage, green branches and fine roots). No drought-related changes in labeled C partitioning between belowground and aboveground organs were observed. The C allocation between non-structural carbohydrates was altered early during drought progression: starch concentration was lower by half in the photosynthetic organs, while the concentration of almost all soluble sugars tended to increase. The preferential allocation of labeled C to glucose and fructose reflected an increased demand for soluble sugars for osmotic adjustment. After 3 months of a lethal drought, the concentrations of soluble sugars and starch were admittedly lower in drought-stressed trees than in the controls, but the pool of non-structural carbohydrates was far from completely depleted. However, the allocation to storage had been impaired by drought; photosynthesis and the sugar translocation rate had also been reduced by drought. Failure to maintain cell turgor through osmoregulation and to refill embolized xylem due to the depletion in soluble sugars in the roots could have resulted in tree mortality in hinoki cypress, though the total pool of carbohydrate was not completely depleted.

Chamaecyparis obtusa (Siebold & Zucc.) Endl. leaf extracts prevent inflammatory responses via inhibition of the JAK/STAT axis in RAW264.7 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Chamaecyparis obtusa (Siebold & Zucc.) Endl. (C. obtusa) has been used as folk medicine in East Asia and has been reported to alleviate inflammatory diseases. However, the detailed mechanisms for the anti-inflammatory effects of C. obtusa remain unclear. AIM OF THE STUDY: Although the anti-inflammatory mechanisms of natural products have been studied for decades, it is still important to identify the potential anti-inflammatory effects of natural sources. In this study, we investigated the anti-inflammatory effects and underlying mechanism of C. obtusa leaf extracts. MATERIAL &METHODS: The cell viability was determined by MTT and crystal violet staining. NO production in the supernatant was measured using Griess reagent. The cell lysates were analyzed by immunoblotting and RT-qPCR. Secreted cytokines were analyzed using ELISA kit and cytokine array kit. mRNA expression from the GSE9632 database set. Z-scores were calculated for each gene and visualized by heat map. RESULTS: Among the extracts of C. obtusa obtained with different extraction methods, the 99% ethanol leaf extract (CO99EL) strongly inhibited lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression and Janus kinase/signaling transducer and activator of transcription (JAK/STAT) phosphorylation in RAW264.7 cells. In addition, CO99EL strongly inhibited LPS-induced interleukin (IL)-1ß, IL-6, IL-27, and C-C motif chemokine ligand (CCL)-1 production and directly inhibited LPS-induced JAK/STAT phosphorylation in RAW264.7 cells. CONCLUSIONS: These findings demonstrate that CO99EL significantly prevents LPS-induced macrophage activation by inhibiting the JAK/STAT axis. Therefore, we suggest the use of C. obtusa extracts as therapeutic approach for inflammatory diseases.