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Si-Wu Tang (SWT), a traditional Chinese formula, is commonly used for treating female diseases, such as relief of menstrual discomfort and climacteric syndrome. The aim of this study was to explore the synergistic effects between each herb in SWT on menstrual disorder patterns. Estradiol regulation and antioxidative effects were indicators that ameliorated menstrual disorder patterns and the total polyphenol and polysaccharide contents were quality markers. According to relationships of bioactivity and phytochemical contents, we discuss the effects of each herb in SWT. In a testosterone-treated MCF-7 cell model, Rehmannia glutinosa and catalpol significantly increased the estradiol content and aromatase upregulation in cell culture. We suggest that catalpol is an aromatase promoter in SWT, and R. glutinosa is a major actor. In terms of the antioxidant activity, pentagalloylglucose, gallic acid, and ferulic acid had stronger antioxidative effects than other compounds. We suggest that the antioxidative ability depends on polyphenols, and Paeonia lactiflora is a major contributor. Based on the prescribing principle of traditional Chinese medicine (TCM) theory, we suggest that R. glutinosa in SWT act as an aromatase promoter in the role of sovereign for ameliorating menstrual disorder patterns. As P. lactiflora has the strongest antioxidant effects and can prevent ROS damage ovarian; therefore, P. lactiflora could help R. glutinosa work as a minister for menstrual disorder patterns and R. glutinosa and P. lactiflora are a herbal pair in SWT.

Acne is a common skin condition observed in adolescents. Nutmeg (Myristica fragrans Houtt) (MF) is a well-known traditional Chinese medicine, its major toxic components, safrole and myristicin, are rich in essential oils. Essential oils of MF (MFO) were extracted by hydrodistillation, the residue was extracted using 50% methanol (MFE-M). The minimum inhibitory concentration (MIC) of MFE-M against Cutibacterium acnes and Staphylococcus aureus was 0.64 mg. Four compounds were obtained from MFE-M: myristicin (1), (+)-erythro-&Delta, 8&prime, 7S,8R- dihydroxy-3,3,5&prime, trimethoxy-8-O-4&prime, neolignan (2), (+)-erythro-&Delta, 8&rsquo, 7-hydroxy-3,4,3&rsquo, 5&rsquo, tetramethoxy 8-O-4-neolignan (3), and erythro-&Delta, 7-acetoxy-3,4,3&prime, 5&prime, tetramethoxy-8-O-4&prime, neolignan (4). Compound 2 exerted the strongest antimicrobial activity, with MICs of 6.25 and 3.12 &mu, g/mL against C. acnes and S. aureus, respectively. Moreover, 2 inhibited NO, PGE2, iNOS, and COX-2 levels in RAW 264.7 cells induced by LPS or heat-killed C. acnes, NO production at 50% inhibitory concentrations (IC50) was 11.07 and 11.53 &mu, g/mL, respectively. Myristicin and safrole content was higher in MFO than in MFE-M. MFO and MFE-M caused no skin irritation after a single topical application in Wistar rats. MFE-M, with low safrole and myristicin content, did not cause skin irritation and exhibited an anti-acne effect, moreover, 2 was identified as the active substance. Therefore, MFE-M could be employed to develop anti-acne compounds for use in cosmetics.

Background The root major proteins of sweet potato trypsin inhibitors (SPTIs) or named sporamin, estimated for 60 to 80% water-soluble proteins, exhibited many biological activities. The human low-density lipoprotein (LDL) showed to form in vivo complex with endogenous oxidized alpha-1-antitrypsin. Little is known concerning the interactions between SPTIs and LDL in vitro. Results The thiobarbituric-acid-reactive-substance (TBARS) assays were used to monitor 0.1 mM Cu2+-mediated low-density lipoprotein (LDL) oxidations during 24-h reactions with or without SPTIs additions. The protein stains in native PAGE gels were used to identify the bindings between native or reduced forms of SPTIs or soybean TIs and LDL, or oxidized LDL (oxLDL). It was found that the SPTIs additions showed to reduce LDL oxidations in the first 6-h and then gradually decreased the capacities of anti-LDL oxidations. The protein stains in native PAGE gels showed more intense LDL bands in the presence of SPTIs, and 0.5-h and 1-h reached the highest one. The SPTIs also bound to the oxLDL, and low pH condition (pH 2.0) might break the interactions revealed by HPLC. The LDL or oxLDL adsorbed onto self-prepared SPTIs-affinity column and some components were eluted by 0.2 M KCl (pH 2.0). The native or reduced SPTIs or soybean TIs showed different binding capacities toward LDL and oxLDL in vitro. Conclusion The SPTIs might be useful in developing functional foods as antioxidant and nutrient supplements, and the physiological roles of SPTIs-LDL and SPTIs-oxLDL complex in vivo will investigate further using animal models.

Pomegranate (Punica granatum) is a heath with abundant hydrolysable tannin in its peel. This study demonstrated that POMx (70% acetone extract of pomegranate peels) significantly reduced interleukin (IL)-1β-induced inducible nitric oxide synthase, cyclooxygenase-2, and matrix metalloproteinase-13 protein expression in primary rat chondrocytes (PRCs). In the type II collagenase-induced osteoarthritis rat model, POMx (150 mg/kg) recovered the weight-bearing ratio changes in experimental animals, suggesting the alleviation of knee-related inflammation and nociception. Therefore, the major compound, punicalagin, was isolated from POMx. Punicalagin also significantly reduced IL-1β-induced inflammatory factors in PRCs and exerted significant antiosteoarthritis effects in the vivo model after 28-d treatment (concentration: 0.50 mg/kg). However, the IC50 values of POMx and punicalagin against prostaglandin E2 production were 83.2 and 36.0 μg/mL, respectively, and the concentration of punicalagin in POMx was 19.1%. Altogether, POMx can be used to develop functional foods for knee-related diseases, and punicalagin can act as an active ingredient.

Canine babesiosis is an important tick-borne disease worldwide. The prevalence varies between regions and countries; however, the incidence of tick infection is associated with the status of preventive tick control measures by the owner. To date, no studies have investigated the incidence of canine babesiosis and the condition of tick prevention in Taiwan. Therefore, the true risk of babesiosis could be underestimated in dogs that are not receiving tick prophylaxis. Samples were collected at 51 hospitals around Taiwan from 265 dogs not receiving regular tick prophylaxis. Diagnostic real-time PCR was performed, and 28 dogs (10.6%) were positive for Babesia spp., including B. gibsoni (26/28) and B. vogeli (2/28). Thirty-nine dogs (14.7%) were seropositive to B. gibsoni. Take the real-time PCR positive as the Babesia infected case, the positive and negative predictive value of serological assay were 64.1% and 98.7%, respectively. The seropositivity of B. gibsoni was significantly associated with real-time PCR positivity for Babesia spp. and vice versa (p 0.001). The odds of seropositive representing real-time PCR positivity was 132.7 times greater than the seronegative (OR: 132.731, 95% CI 35.683-493.728). Risk factors in the population identified included: dogs with a short-haired coat; intact dogs; dogs from multi-dog households; dogs with more than 10 ticks and fleas on the skin; dogs that go outdoors more than 9 times per week; and dogs with an abnormal blood test result that included anemia, thrombocytopenia, and leukopenia. However, the dogs were not tested for other co-infections, therefore, these hematological risk factors should be carefully interpreted and confirmed by further diagnostic tests. In conclusion, when dogs present with abnormal blood test results and share the risk factors listed above, babesiosis should be seriously considered and followed up with molecular and serological testing. The serological assay used in this study can provide valuable information in diagnosing babesiosis in dogs in Taiwan.

Maternal immune activation (MIA) increases the risk of autism spectrum disorder (ASD) in offspring. Microbial dysbiosis is associated with ASD symptoms. However, the alterations in the brain–gut–microbiota axis in lipopolysaccharide (LPS)-induced MIA offspring remain unclear. Here, we examined the social behavior, anxiety-like and repetitive behavior, microbiota profile, and myelination levels in LPS-induced MIA rat offspring. Compared with control offspring, MIA male rat offspring spent less time in an active social interaction with stranger rats, displayed more anxiety-like and repetitive behavior, and had more hypomyelination in the prefrontal cortex and thalamic nucleus. A fecal microbiota analysis revealed that MIA offspring had a higher abundance of Alistipes, Fusobacterium, and Ruminococcus and a lower abundance of Coprococcus, Erysipelotrichaies, and Actinobacteria than control offspring, which is consistent with that of humans with ASD. The least absolute shrinkage and selection operator (LASSO) method was applied to determine the relative importance of the microbiota, which indicated that the abundance of Alistipes and Actinobacteria was the most relevant for the profile of defective social behavior, whereas Fusobacterium and Coprococcus was associated with anxiety-like and repetitive behavior. In summary, LPS-induced MIA offspring showed an abnormal brain–gut–microbiota axis with social behavior deficits, anxiety-like and repetitive behavior, hypomyelination, and an ASD-like microbiota profile.

Objective Cryptotanshinone (CPT) and dihydrotanshinone (DHT) are diterpenoid anthraquinone compounds extracted from traditional Chinese herbal medicine (TCM). Recent studies have shown that CPT regulates the signal transduction pathways via microRNA (miRNA) alterations. However, few studies have investigated the role of DHT in miRNA alterations affecting cell-signaling pathways. This study aimed to investigate the miRNA alterations and post-transcriptional regulation activities of DHT in comparison to CPT. Methods HepG2 and HT-29 cells were treated with DHT or CPT for 72 h. MiRNA, transcription factor encoding mRNA, and downstream gene expression were determined using real-time quantitative PCR. Protein expression was analyzed using western blotting. Results The results revealed that CPT and DHT targeted cell proliferation and apoptosis signaling pathways via miR-15a-5p, miR-27a-5p, miR-100−5p, and miR-200a-5p alterations.In silico target predictions showed that downregulation of epidermal growth factor receptor (EGFR) mRNA expression by DHT might also suppress the expression of STAT family proteins and lead to anti-proliferation effects. We also found that, compared to CPT, DHT might possess higher potency in cell growth regulation via multi-miRNA and transcription factor alterations. Conclusion This study revealed that CPT and DHT targeted cell proliferation and apoptosis signaling pathways via alterations in miRNAs and transcription factors. In addition, the findings of this study suggest that DHT is more potent than CPT in cancer chemopreventive activities. Therefore, DHT at a low dose is a TCM compound with less toxic side effects and may contribute to the development of natural medicine as a potential cancer chemopreventive agent.

Polygonum multiflorum Thunb. is a traditional herbal medicine that is rich in polyphenols. The major compound, 2,3,5,4′-tetrahydroxystilbene-2-O-β-d-glucoside (THSG) has many pharmacological activities, such as antioxidative and free radical-scavenging properties, and the abilities to reduce hyperlipidemia, prevent lipid peroxidation, and protect the cardiovascular system. In this study, the anti-osteoarthritis (OA) effects of THSG were explored using in vitro and in vivo models. THSG inhibited nitric oxide (NO) and prostaglandin E2 (PGE2) production and inducible NO synthase (iNOS) and cyclooxygenase-2 expressions by lipopolysaccharide-stimulated RAW 264.7 cells. On the other hand, THSG inhibited PGE2 production and iNOS and matrix metalloproteinase-13 expressions by interleukin-1β-stimulated primary rat chondrocytes. Through a mono-iodoacetate-induced rat OA model assay, THSG reduced paw edema and improved the weight-bearing distribution. Therefore, THSG has anti-inflammatory activity and could be applied as a lead compound for the development as an OA drug.

Autoimmune hypophysitis (AH) is a chronic inflammatory disease characterized by infiltration of T and B lymphocytes in the pituitary gland. The mechanisms through which infiltrating lymphocytes cause disease remain unknown. Using a mouse model of AH we assessed whether T lymphocytes undergo activation in the pituitary gland. Infiltrating T cells co-localized with dendritic cells in the pituitary and produced increased levels of interferon-γ and interleukin-17 upon stimulation in vitro. Assessing proliferation of CD3- and B220-postive lymphocytes by double immunohistochemistry (PCNA-staining) and flow cytometry (BrdU incorporation) revealed that a discrete proportion of infiltrating T cells and B cells underwent proliferation within the pituitary parenchyma. This proliferation persisted into the late disease stage (day 56 post-immunization), indicating the presence of a continuous generation of autoreactive T and B cells within the pituitary gland. T cell proliferation in the pituitary was confirmed in patients affected by autoimmune hypophysitis. In conclusion, we show that pituitary-infiltrating lymphocytes proliferate in situ during AH, providing a previously unknown pathogenic mechanism and new avenues for treatment.

The stem with hook of Uncaria rhynchophylla (Chinese herbal name Gou-Teng) is a traditional Chinese medicine that has been ethnopharmacologically used to extinguish wind and clean interior heat. Rhynchophylline (RHY), a tetracyclic oxindole alkaloid isolated from U. rhynchophylla, displays significant antineuroinflammatory effects. However, there is no evidence to indicate that rhynchophylline can cross the blood–brain barrier and be detected in the brain. In this study, an in vivo microdialysis sampling method coupled with UPLC/MS/MS was employed for the continuous simultaneous monitoring of unbound RHY in rat blood and brain. The precursor ion product ion transition at m/z 385.2 160.0 for rhynchophylline was monitored. A calibration curve gave good linearity (r > 0.996) over the concentration range from 0.5 to 1000 ng/mL. The results demonstrated that rhynchophylline could be detected in the brain and plasma from 15 min to 6 h after its administration (1 or 10 mg/kg, i.v.). All the pharmacokinetic parameters of rhynchophylline in the brain and plasma were obtained. These results show that rhynchophylline can cross the blood–brain barrier and they provide useful clinical information. Copyright © 2014 John Wiley & Sons, Ltd.

To determine the effects of co-ingesting caffeine (CAF) and carbohydrate (CHO) on high-intensity intermittent sprints (HIS) performance and physiological responses. Twelve active males underwent 4 interventions at least 7 days apart in a randomized, double-blind, placebo-controlled, balanced trial. A meal contained 65 % CHO was provided 2 h before the HIS test. Participants ingested the placebo (PLA) or CAF (6 mg kg−1 BW) 1 h before taking an HIS test, and ingested a PLA or CHO solution (0.8 g kg−1 BW) before undergoing the testing protocol. The HIS protocol comprised ten sets of 5 × 4-s sprints on a cycle ergometer with a 2-min recovery between each set. There was no significant difference between peak power output and mean power output between trials (p > 0.05). Compared with PLA, CAF + CHO resulted in a 5.2 % reduction in total work, corresponding to a 24.7–25.7 % increase in fatigue at the end stage of the HIS. The administration of CAF + CHO supplementation also resulted in an 11.1 % increase in blood lactate, and elevated blood glucose concentrations throughout HIS testing compared with PLA (p

Acne is a common skin condition with sebum overproduction, hyperkeratosis, Propionibacterium acnes (P. acnes) and Staphylococcus aureus, and inflammation. Punica granatum (pomegranate) is well-known for its anti-inflammatory effects; however, few studies have discussed the anti-acne effects of pomegranate. In this study, we found that pomegranate extract (PG-E) significantly reduced P. acnes-induced edema in Wistar rat ears. Therefore, an evaluation platform using multiple pathogenic mechanisms of acne was established to explore the anti-acne effects of pomegranate. Results showed that PG-E inhibited bacterial growth and lipase activity. Through a bioguided-fractionation-isolation system, four hydrolysable tannins, punicalagin (1), punicalin (2), strictinin A (3), and granatin B (4), were isolated. Compounds 1 and 2 had greater anti-bacterial activities and anti-testosterone-induced HaCaT proliferative effects than the others. Compounds 1, 3, and 4 displayed lipase inhibitory effects. Compound 4 decreased cyclooxygenase-2 expression and downregulated prostaglandin E2 production in heat-killed P. acnes-treated RAW 246.7 cells. In conclusion, PG-E is abundant in hydrolysable tannins that display multiple anti-acne capacities, including anti-bacterial, anti-lipase, anti-keratinocyte proliferation, and anti-inflammatory actions. Hence, PG-E has great potential in the application of anti-acne and skin-care products, and punicalagin (1), the most effective component in PG-E, can be employed as a quality control marker.

Guan-Cheng Huang,1–3 Shih-Yu Chen,4 Po-Wei Tsai,5 Jerome G Ganzon,5 Chia-Jung Lee,6 Her-Shyong Shiah,7 Ching-Chiung Wang4,5 1School of Medicine, College of Medicine, Taipei Medical University, Taipei, 2Division of Hematology and Oncology, Department of Internal Medicine, Yuan’s General Hospital, 3Department of Leisure and Recreation Industry Management, College of Humanities and Management, Fooyin University, Kaohsiung, 4Graduate Institute of Pharmacognosy, College of Pharmacy, 5School of Pharmacy, College of Pharmacy, 6PhD Program for Clinical Drug Discovery of Chinese Herbal Medicine, College of Pharmacy, Taipei Medical University, 7Division of Hematology and Oncology, Department of Internal Medicine, Taipei Medical University Hospital, Taipei, Taiwan Abstract: Dang-Gui-Bu-Xue-Tang (DBT), a combination of Angelicae Sinensis Radix and Astragali Radix, is a widely used herbal decoction in traditional Chinese medicine primarily to promote or invigorate the “blood”. In this study, we explored this ancient formulation and provide evidence of its blood-toning properties. We used the improvement iron uptake as promote or invigorate the “blood” indicator. Ferritin formation of Caco-2 cells in vitro assay and diet-induced anemia (DIA) in rat model were used to prove its improvement iron uptake and ameliorating effects. Finally, the iron–DBT interactions were measured by iron-bind­ing assay. We first demonstrated DBT increased uptake of ferrous iron through the biosynthesis of ferritin by Caco-2 cells and determined which complementary treatment would provide optimum results. Thereafter, effects of the treatment on improving the bioavailability of absorbed iron in the form of hemoglobin (Hb) were established using a DIA-animal model. The results showed that DBT slightly improved Hb levels compared with the baseline Hb and pretreatment with DBT for 2 hours prior to supplementation with ferrous sulfate provided the greatest gain in Hb levels in DIA rats. However, DBT and ferrous sulfate were co-treated with Caco-2 cell or DIA rats, the ferritin formation and Hb levels both were decreased. In iron-bind­ing assay, the DBT extract influenced the free Fe(II) type in the FeSO4 solution. Therefore, we suggest that complementary treatment with DBT and iron supplementation can have a strong ameliorating effect on iron-deficiency anemia in clinical settings, but needs a 2-hour interval of DBT administration prior to ferrous sulfate treatment. Keywords: Dang-Gui-Bu-Xue-Tang, ferulic acid, iron uptake, ferritin, iron-deficiency anemia, herb–drug interaction

Wu-Chia-Pi medicated wine, composed nine Chinese medicines soaked in 35% alcohol, is widely used in Asia for its health-promoting functions. However, long-term consumption of alcohol could result in liver dysfunction. In this study, Wu-Chia-Pi solution (WCPS) and extract (WCPE) were prepared by modification of the principals given by the Committee on Chinese Medicine and Pharmacy in Taiwan. The aim of this study was to explore the protective effect of WCPS against carbon tetrachloride (CCl4)-induced liver injury and to clarify its active component(s). Antioxidative effects of the test samples were evaluated via MDA inhibition, catalase activity and DPPH-scavenging assays. HPLC was used to analysis the active components. Results showed that WCPS (1 and 5 mL/kg) significantly prevented CCl4-induced liver injury without chronic liver toxicity. Referring to the antioxidative activities, WCPE displayed significant MDA inhibitory and DPPH-scavenging activities with IC50 values of 0.91 ± 0.03 and 0.60 ± 0.04 mg/mL, respectively. Catalase activity was also enhanced by treatment of WCPE, acteoside and quercetin. Therefore, we suggest that acteoside and quercetin are the major contributors to the antioxidative and hepatoprotective activities of WCPS, and a possible mechanism could be mediated through reduction of oxidative stress.

Cantharidin, an active component in mylabris, is used in traditional Chinese medicine (TCM) to treat scabies and hepatoma, but accompanied by hemorrhagic cystitis. Evidence shows that cantharidin induces human bladder carcinoma cell death through COX-2 overexpression in vitro. In TCM, Scutellaria baicalensis is usually used to cure mylabris-induced hematuria. This work was undertaken to determine the mechanisms of cantharidin-induced rat hemorrhagic cystitis and explore the uroprotective effect of S. baicalensis. In vitro results showed cantharidin could induce cytotoxicity through prostaglandin (PG)E2 overproduction of T24 cells. Boiling-water extract of S. baicalensis (SB-WE) could significantly inhibit PGE2 production and COX-2 expression in lipo-polysaccharide-induced RAW 264.7 cells, indicating obvious anti-inflammatory abilities. In vivo results indicated that cantharidin caused rat hemorrhagic cystitis with hematuria via c-Fos and COX-2 overexpression. SB-WE was given orally to cantharidin-treated rats, whereby hematuria level, elevated PGE2 and COX-2 protein overexpression were significantly and dose-dependently inhibited by SB-WE. The anti-inflammatory components of SB-WE are baicalin and wogonin, whose contents were 200.95 ± 2.00 and 31.93 ± 0.26 μg/mg, respectively. In conclusion, cantharidin induces rat cystitis through c-Fos and COX-2 over-expression and S. baicalensis can prevent the resulting hematuria because of its anti-inflammatory effects.

Aim of the study This study examined the modulating effects of Clematis chinensis Osbeck (Ranunculaeae) on pro-inflammatory and degradative mediators associated with inflammatory arthritis. Materials and methods Primary human chondrocytes (PHC) were stimulated with IL-1β or lipopolysaccharide (LPS) to induce the enhanced release of prostaglandin E 2 (PGE 2 ), metalloproteinase (MMP-3 and -13), and cyclooxygenase-2 (COX-2) protein expression. The 18 F-FDG microPET imaging system was used to evaluate the anti-arthritic effects of Clematis chinensis in vivo . Results The acetone extracted Clematis chinensis (CC6) contained the most total saponins compared to other solvent's extracts and showed significant and dose-dependent inhibitory effects on PGE 2 , MMP-3, -13, and COX-2 productions by LPS-stimulated PHC. Furthermore, CC6 also exerted inhibitory effects on 2- 18 F-fluoro-2-deoxy- d -glucose ( 18 F-FDG) uptake when assessed by positron emission tomography (PET) uptake in the joints and serum PGE 2 of rabbits with knee joints injected with LPS. Conclusion The results suggest the significant chondroprotective effects of Clematis chinensis are through its anti-inflammatory and MMPs inhibitory abilities. Meanwhile, we established a new analysis method to evaluate the Chinese herbal anti-arthritic effects.

In this study, well-known skin-care functional parameters were used, including UV absorption, DPPH-scavenging, NO-production-inhibition, tyrosinase-inhibition activity and anti-Staphylococcus aureus activity to measure the effects of the 70% acetone-extracts form 28 species of Lamiaceae plants. Further, the phytochemical contents were explored by total phenol (TP), total flavone (TF) and total coumarin (TC). The correlation between the skin-care effects and the phytochemical contents was analysed by non-parametric correlation analysis. Amongst the 70% acetone-extracts, Origanum majorana displayed the strongest DPPH-scavenging and tyrosinase-inhibitory effects and the richest phenol content. Based on the statistics results, the phytochemical contents were related with those parameters, such as: DPPH-scavenging effects vs. TP (R = 0.542), TF vs. TP (R = 0.613), and NO-inhibitory vs. anti-bacterial activities (R = 0.767). Moreover, each genus of Lamiaceae had different properties of skin-care effects. Form our research works, the Lamiaceae is good resources to develop skin-care cosmetics.

MicroRNAs (miRNAs), i.e. small non-coding RNA molecules (∼22 nt), can bind to one or more target sites on a gene transcript to negatively regulate protein expression, subsequently controlling many cellular mechanisms. A current and curated collection of miRNA-target interactions (MTIs) with experimental support is essential to thoroughly elucidating miRNA functions under different conditions and in different species. As a database, miRTarBase has accumulated more than 3500 MTIs by manually surveying pertinent literature after data mining of the text systematically to filter research articles related to functional studies of miRNAs. Generally, the collected MTIs are validated experimentally by reporter assays, western blot, or microarray experiments with overexpression or knockdown of miRNAs. miRTarBase curates 3576 experimentally verified MTIs between 657 miRNAs and 2297 target genes among 17 species. miRTarBase contains the largest amount of validated MTIs by comparing with other similar, previously developed databases. The MTIs collected in the miRTarBase can also provide a large amount of positive samples to develop computational methods capable of identifying miRNA-target interactions. miRTarBase is now available on http://miRTarBase.mbc.nctu.edu.tw/, and is updated frequently by continuously surveying research articles.

Inflammation can cause various physical dysfunctions. Punica granatum Linne (pomegranate), a high phenolic content fruit, is widely used as an antipyretic analgesic in Chinese culture. Pomegranate has shown potential nitric oxide (NO) inhibition in LPS-induced RAW 264.7 macrophage cells. Moreover, pomegranate (100 mg/kg) significantly decreased carrageenan-induced mice paw edema for 1, 3, 4, and 5 h. Therefore, column chromatography combined with in vitro bioassay-guided fractionation was used to isolate the active anti-inflammatory components from the pomegranate. Punicalagin (1), punicalin (2), strictinin A( 3), and granatin B (4) were obtained with yields of 0.093%, 0.015%, 0.003%, and 0.013%, respectively. All these hydrolysable tannins inhibited NO production and iNOS expression in RAW 264.7 cells. Among them, 4 showed the strongest iNOS and COX-2 inhibitory effects, and exhibited these effects in the inhibition of paw swelling and the PGE2 level in carrageenan-induced mice. Taken together, we suggest that 4 could be used as a standard marker for the anti-inflammatory effect of pomegranate. 2009 Elsevier Ltd. All rights reserved.

Zingiber zerumbet Smith has long been used as a botanical medicine for anti-inflammation in Southeast Asia. In this paper, zerumbone (1), 3-O-methyl kaempferol (2), kaempferol-3-O-(2, 4-di-O-acetyl-α- l -rhamnopyranoside) (3), and kaempferol-3-O-(3,4-di-O-acetyl-α- l -rhamnopyranoside) (4) were isolated from the rhizome of Z. zerumbet. The inhibitory effects of these compounds on NO and PGE2 production from lipopolysaccharide (LPS)-induced RAW 264.7 macrophages were measured. Among them, 1 and 2 demonstrated potent inhibition of NO production, with respective IC50 values of 4.37 and 24.35 μM, and also significantly suppressed iNOS expression in a dose-dependent manner. However, 1 and 2 could inhibit PGE2 production only at high doses (20 and 40 μM, respectively), and COX-2 protein level was not affected. According to the in vitro study, 1 had greater anti-inflammatory effects than 2. Therefore, mice were administered with 1 (10 mg/kg) 1 h before carrageenan injection, and the oedema was significantly attenuated compared to the vehicle control. Mature rhizomes were richer in 1 and lower in moisture. We suggest that the economic cultivation period of Z. zerumbet is the 5th month after seeding when its functions as food and anti-inflammatory are maximum, because 1 is dramatically increased at that time.

The fresh rhizome of Zingiber zerumbet Smith (Zingiberaceae) is used as a food flavoring and also serves as a folk medicine as an antipyretic and for analgesics in Taiwan. Zerumbone, a monocyclic sesquiterpene was isolated from the rhizome of Z. zerumbet and is the major active compound. In this study, the anti-inflammatory and antinociceptive effects of zerumbone on arthritis were explored using in vitro and in vivo models. Results showed that zerumbone inhibited inducible nitric oxide (NO) synthase (iNOS), cyclooxygenase (COX)-2 expressions, and NO and prostaglandin E2 (PGE2) production, but induced heme oxygenase (HO)-1 expression in a dose-dependent manner in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. When zerumbone was co-treated with an HO-1 inhibitor (tin protoporphyrin (SnPP)), the NO inhibitory effects of zerumbone were recovered. The above results suggest that zerumbone inhibited iNOS and COX-2 through induction of the HO-1 pathway. Moreover, matrix metalloproteinase (MMP)-13 and COX-2 expressions of interleukin (IL)-1β-stimulated primary rat chondrocytes were inhibited by zerumbone. In an in vivo assay, an acetic acid-induced writhing response in mice was significantly reduced by treatment with zerumbone. Furthermore, zerumbone reduced paw edema and the pain response in a mono-iodoacetate (MIA)-induced rat osteoarthritis model. Therefore, we suggest that zerumbone possesses anti-inflammatory and antinociceptive effects which indicate zerumbone could be a potential candidate for osteoarthritis treatment.

Objective.Kawasaki disease (KD) is an acute febrile vasculitic syndrome in children. CD40 ligand (CD40L) has been implicated in certain types of vasculitis. We proposed that CD40L expression might be correlated with coronary artery lesions in KD.Methods.Blood samples were collected from 43 patients with KD before intravenous immunoglobulin (IVIG) treatment and 3 days afterward. Forty-three age-matched febrile children with various diseases were studied in parallel as controls. CD40L expression on T-cells and platelets were detected by flow cytometry, and soluble CD40L (sCD40L) levels were measured by enzyme-linked immunosorbent assay.Results.We found that CD40L expression on CD4+ T-cells was significantly higher in patients with KD than in the febrile control (FC) group (28.69 ± 1.17% vs 4.37 ± 0.36%). CD40L expression decreased significantly 3 days after IVIG administration (28.69 ± 1.17% vs 13.53 ± 0.55%). CD40L expression on platelets from patients with KD was also significantly higher than in the FC group (8.20 ± 0.41% vs 1.26 ± 0.12%) and decreased after IVIG therapy. sCD40L levels were also significantly higher in KD patients with those of FC (9.69 ± 0.45 ng/mL vs 2.25 ± 0.19 ng/mL) but were not affected by IVIG treatment 3 days afterward (9.69 ± 0.45 ng/mL vs 9.03 ± 0.32 ng/mL). More interesting, we found that in KD patients, CD40L expression on CD4+ T-cells and platelets but not on CD8+ T-cells or sCD40L was correlated with the occurrence of coronary artery lesions.Conclusions.CD40L might play a role in the immunopathogenesis of KD. IVIG therapy might downregulate CD40L expression, resulting in decrease of CD40L-mediated vascular damage in KD. This implicates that modulation of CD40L expression may benefit to treat KD vasculitis.

Objective: To assess nitric oxide (NO) production and different displays of NO synthase (NOS) isoforms in patients with Kawasaki disease (KD) before and after intravenous immunoglobulin (IVIG) treatment. Study design: Blood and urine samples were collected from patients with KD before IVIG treatment and 3 days afterward. Age-matched febrile patients with various diseases served as control patients. Plasma NOx (NO3- + NO2-) levels were measured as NO production. Real-time reverse transcriptase–polymerase chain reaction and immunohistochemical staining were used to detect differential NOS isoform expression in mononuclear leukocytes (MNCs). Results: Patients with KD (n = 40) had higher levels of NOx than the non-KD febrile control patients. The elevated NOx levels in patients with KD were significantly associated with the occurrence of coronary artery dilation (>3 mm). These elevated NOx levels significantly decreased after IVIG treatment (157.8 ± 15.9 μmol/L vs 57.0 ± 4.27 μmol/L, P

Background Caffeine (CAF) has been shown to improve performance during early phase of repeated sprint exercise; however some studies show that CAF also increases the magnitude of physical stress represented by augmented blood lactate, glucose, and cortisol concentrations during latter phase of repeated sprint exercise. No studies have investigated the efficacy of combined carbohydrate (CHO) and CAF consumption during repeated sprint exercise (RSE) in female athletes. Thus, the purpose of this study was to investigate the effects of CAF with CHO supplementation on RSE and agility. Methods Eleven female athletes completed four experimental trials performed 7 d apart in a double-blind, randomized, and counter-balanced crossover design. Treatments included CAF + PLA (placebo), CAF + CHO, PLA + CHO, and PLA + PLA. Participants ingested capsules containing 6 mg · kg−1 of CAF or PLA 60-min prior to RSE, and 0.8 g · kg−1 of CHO solution or PLA immediately before the RSE, which consisted of ten sets of 5 × 4-s sprints on the cycle ergometer with 20-s active recovery. The agility T-test (AT-test) was performed before and after the RSE. Blood samples were acquired to assess glucose, lactate, testosterone, and cortisol. Results During Set 6 of RSE, peak power and mean power were significantly higher in PLA + CHO than those in CAF + PLA and PLA + PLA, respectively (p .05). Blood lactate and glucose concentrations were significantly higher under CAF + CHO, CAF + PLA, and PLA + CHO versus PLA + PLA (p .05). Conclusions Findings indicate that CAF + PLA or CAF + CHO ingestion did not improve repeated sprint performance with short rest intervals or agility. However, CHO ingested immediately prior to exercise provided a small but significant benefit on RSE performance in female athletes.

Nowadays herbal medicines of skin-whitening cosmetics are popular with women. We attempted to find the whitening activity compounds present in many herbal medicines used for this purpose and discuss their mechanisms in melanin biosynthesis. The 70% acetone extracts of 10 kinds of herbs were investigated for their mushroom tyrosinase activity inhibition. Among these 10 extracts, Chinese galls showed inhibitory activity against tyrosinase, with a 50% inhibitory concentration (IC(50)) value of 22 microg/ml. In a B16 mouse melanoma cell culture assay, Chinese galls dose-dependently inhibited melanin biosynthesis. Using ultraviolet A (UVA) or alpha-melanocyte-stimulating hormone (alpha-MSH) to stimulate B16 cells after Chinese gall treatment, the melanin biosynthesis of B16 cells was inhibited in a dose-dependent manner. The active compounds of Chinese galls were isolated by column chromatography, and the melanin biosynthesis inhibition in B16 melanoma cells was measured. Three gallotannins, 2,3,4,6-tetra-O-galloyl-D-glucopyranose, 1,2,3,6-tetra-O-galloyl-beta-D-glucopyranose, and 1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranose, were isolated from Chinese gall extract, and their IC(50) values of tyrosinase inhibition activity were 54, 30, and 15 muM, respectively. By the mushroom tyrosinase inhibition kinetics assay, the three gallotannins were all determined to be non-competitive inhibitors. These results indicated that Chinese galls inhibit melanin biosynthesis, associated with hyperpigmentation and can be used as skin-whitening cosmetics for skin care.

Indomethacin is used as an anti-inflammatory drug and a nonselective cyclooxygenase inhibitor. When indomethacin in methanol was photo-irradiated with an Hg lamp, methyl ester, ethyl ester, and gamma-lactone derivatives of indomethacin were produced. In the present study, we found that the methyl ester derivative of indomethacin (M-IN) could more potently inhibit prostaglandin E(2) (PGE(2)) and nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX 2) protein expression from lipopolysaccharide (LPS)-stimulated RAW 264.7 cells than indomethacin, similar to the effect of a non-steroidal anti-inflammatory drugs (NSAID). On the other hand, the results showed that M-IN with an IC(50) value maintained at 36.9 microg/ml for 12 h exhibited stronger cytotoxicity than ethyl ester, gamma-lactone derivatives of indomethacin, and indomethacin in promyelocytic leukemia HL-60 cells. Moreover, a series of biochemical analyses determined that M-IN caused apoptotic bodies, DNA fragmentation, and enhanced PARP and pro-caspase 3 degradation in HL-60 cells. These above results indicate that the photosynthesized product, M-IN, had stronger anti-inflammatory effects in LPS-stimulated RAW 264.7 cells and cytotoxicity effects in HL-60 cells than the parent drug, indomethacin.

Triflamp, a metalloproteinase isolated from Trimeresurus flavo-viridis, inhibits heterotypic adhesion between platelets and neutrophils. Coincubation studies demonstrate that direct interaction of triflamp with neutrophils is sufficient to inhibit the formation of neutrophil-platelet complexes. Its anti-adhesive effect is in a concentration- and incubation time-dependent manner. Triflamp reduces the expression of P-selectin glycoprotein ligand-1 (PSGL-1) on neutrophils and glycoprotein (GP) Ibalpha on platelets as probed by flow cytometry and Western blot. Moreover, triflamp disrupts P-selectin-mediated adhesion by cleaving PSGL-1 from the neutrophil surface. There are obvious differences regarding PSGL-1 proteolysis by triflamp and cathepsin G. Besides the NH2-terminus of PSGL-1, other sites are truncated by triflamp. The inhibitory effect of triflamp on PSGL-1 expression was prevented by pretreatment with a metalloproteinase inhibitor, phenanthroline. However, triflamp-treated platelets fully keep the ability for binding to PAF- or fMLP-stimulated neutrophils. Our results indicate that degradation of platelet GPIb alpha by triflamp does not interfere with neutrophil-platelet adhesion. Its effect on neutrophil PSGL-1 appears to be a critical factor for its inhibition on neutrophil-platelet interaction.

SummaryThe biologically active components fromViperidae venoms specifically affect cell-matrix interactions, and have been utilized for developing anti-adhesive therapy as anti-thrombotic and antiangiogenic agents. Utilizing platelet aggregometry coupled with flow cytometry, we found that a metalloproteinase isolated from Trimeresurus flavoviridis, termed triflamp, inhibited heterotypic adhesion between platelets and neutrophils in whole blood samples. Triflamp is a monomeric glycoprotein with a molecular weight of ∼28 kDa. Triflamp has a N-terminal amino acid sequence homologous to other venom metalloproteinases isolated from T. flavoviridis. The enzymatic activity of triflamp was inhibited by EDTA and phenanthroline but not by PMSF. Moreover, triflamp is a pure α-fibrinogenase. Studies aimed at determining the nature of triflamp in affecting platelets or neutrophils revealed a selective inhibitory activity to glycoprotein (GP) Ibα.-dependent platelet aggregation and PSGL-1-dependent neutrophil homotypic aggregation, indicating that its effects are rather specific. As judged by Western blotting, GPIbα. on platelets and PSGL-1 on neutrophils are the substrates of triflamp. In conclusion, we suggest the novel role of venom metalloproteinase from Viperidae affecting the blood cell-cell interactions, thus offering a potential approach for further exploration of anti-inflammatory agents.

Gastrodia elata Bl. (GE) is a traditional Chinese herb that is commonly used in Chinese communities to treat convulsive disorders such as epilepsy. The purpose of the present study was to determine the anticonvulsive and free radical activities of GE in rats. In vitro studies were conducted by using brain tissue from 6 male Sprague-Dawley (SD) rats treated with 120 μg/ml of kainic acid (KA), with or without the addition of various concentrations of GE. In vivo studies were conducted in a total of 30 male SD rats divided into 5 groups of 6 rats which were treated as follows: 1) the normal group received an intraperitoneal injection (i.p.) of PBS (Phosphate buffer saline, 1 ml/kg); 2) the control group received KA (12 mg/kg) i.p.; 3) the GE 1.0 group received oral administration of GE 1.0 g/kg 30 min prior to KA administration; 4) the GE 0.5 group reveived oral administration of GE 0.5 g/kg 30 min prior to KA administration; 5) the PH group reveived oral administration of phenytoin 20 mg/kg 30 min prior to KA administration. Seizures were verified by behavioral observations, electroencephalograph (EEG) and electromyography (EMG). Lipid peroxide levels in the rat brain, luminol chemiluminescence (CL) and lucigenin-CL in the peripheral blood were measured simultaneously after behavioral observations. The results indicate that GE administration significantly reduced KA-induced lipid peroxide levels in vitro. Oral administration of GE 1.0 g/kg and phenytoin 20 mg/kg significantly reduced counts of wet dog shakes (WSS), paw tremor (PT) and facial myoclonia (FM) in KA-treated rats. In addition, oral administration of GE 1.0 g/kg significantly delayed the onset of WDS, from 30 min in the control group to 46 min in the 0.5 g/kg group, and 63 min in the GE 1.0 g/kg group. A significantly reduced level of lipid peroxides in the rat brain was found in the GE 1.0 g/kg, 0.5 g/kg, and phenytoin 20 mg/kg groups. The GE 1.0 g/kg group showed significant reduction of luminol-CL and lucigenin-CL counts in the peripheral blood compared to the control group. The results of the present study demonstrate that GE has anticonvulsive and free radical scavenging activities. Further studies are needed to determine the clinical effectiveness of GE as an anticonvulsant in humans.

Tea tree oil (TTO), the essential oil of Melaleuca alternifolia L., is one of the most popular agents used in cosmetics. This study aimed to explore the correlations of components in TTO with its anti-acne activities and skin irritation. TTOs were isolated from the leaves (TTO-L), twigs, and branches of M. alternifolia by steam distillation, and the components analyzed by gas chromatography–mass spectrometry. Results showed that components of TTO-L satisfied the International Organization for Standardization (ISO) 4730 guidelines. TTO-L and its components, terpinen-4-ol, terpinolene, α-terpinene, and α-terpineol, had strong inhibitory activities against Propionibacterium acnes and Staphylococcus aureus . Moreover, six TTO formulas (DF-TTO) were designed according to ISO 4730 guidelines by adjusting the proportions of these four active components. All DF-TTO formulas showed a positive correlation between terpinen-4-ol concentration and anti- P. acnes activity. In the skin irritation assay, TTO-L, terpinen-4-ol, and 1,8-cineole did not cause significant skin irritation at 2% per site. In conclusion, terpinen-4-ol is the major active component responsible for TTO's antibacterial efficacy, while minor components in TTO also contributed to its efficacy. Moreover, we suggest that a concentration less than 5% is more suitable and safer for treating acne than higher concentrations.