Sheng-Yang Wang

Ethyl caffeate suppresses NF-κB activation and its downstream inflammatory mediators, iNOS, COX-2, and PGE2 in vitro or in mouse skin

Ethyl caffeate, a natural phenolic compound, was isolated from Bidens pilosa, a medicinal plant popularly used for treating certain inflammatory syndromes. The purpose of this study was to investigate the structural activity, and the anti‐inflammatory functions and mechanism(s) of ethyl caffeate. Ethyl caffeate was found to markedly suppress the lipopolysaccharide (LPS)‐induced nitric oxide (NO) production (IC50=5.5 μg ml−1), mRNA and protein expressions of inducible nitric oxide synthase (iNOS), and prostaglandin E2 (PGE2) production in RAW 264.7 macrophages. Transient gene expression assays using human cox‐2 promoter construct revealed that ethyl caffeate exerted an inhibitory effect on cox‐2 transcriptional activity in 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐treated MCF‐7 cells. Immunohistochemical studies of mouse skin demonstrated that TPA‐induced COX‐2 expression was significantly inhibited by ethyl caffeate with a superior effect to that of celecoxib, a nonsteroidal anti‐inflammatory drug. The phosphorylation and degradation of inhibitor κB (IκB) and the translocation of nuclear transcription factor‐κB (NF‐κB) into the nucleus, as well as the activation of mitogen‐activated protein kinases (MAPKs) induced by LPS in macrophages, were not affected by ethyl caffeate. Ethyl caffeate, however, could inhibit NF‐κB activation by impairing the binding of NF‐κB to its cis‐acting element. These results suggest that ethyl caffeate suppresses iNOS and COX‐2 expressions partly through the inhibition of the NF‐κB·DNA complex formation. Structure–activity relationship analyses suggested that the catechol moiety and α,β‐unsaturated ester group in ethyl caffeate are important and essential structural features for preventing NF‐κB·DNA complex formation. This study provides an insight into the probable mechanism(s) underlying the anti‐inflammatory and therapeutic properties of ethyl caffeate.

Shikonins, Phytocompounds from Lithospermum erythrorhizon, Inhibit the Transcriptional Activation of Human Tumor Necrosis Factor α Promoter in Vivo

Tumor necrosis factor α (TNF-α) contributes to the pathogenesis of both acute and chronic inflammatory diseases and has been a target for the development of new anti-inflammatory drugs. Shikonins, the naphthoquinone pigments present in the root tissues of Lithospermum erythrorhizon Sieb. et Zucc. (Boraginaceae), have been reported to exert anti-inflammatory effects both in vitro and in vivo. In this study, we evaluated the effects of shikonin and its derivatives on the transcriptional activation of human TNF-α promoter in a gene gun-transfected mouse skin system by using a luciferase reporter gene assay. The crude plant extract of L. erythrorhizon as well as derived individual compounds shikonin, isobutyryl shikonin, acetyl shikonin, dimethylacryl shikonin and isovaleryl shikonin showed significant dose-dependent inhibition of TNF-α promoter activation. Among the tested compounds, shikonin and isobutyryl shikonin exhibited the highest inhibition of TNF-α promoter activation and also showed significant suppression of transgenic human TNF-α mRNA expression and protein production. We demonstrated that shikonin-inhibitory response was retained in the core TNF-α promoter region containing the TATA box and a 48-bp downstream sequence relative to the transcription start site. Further our results indicated that shikonin suppressed the basal transcription and activator-regulated transcription of TNF-α by inhibiting the binding of transcription factor IID protein complex (TATA box-binding protein) to TATA box. These in vivo results suggest that shikonins inhibit the transcriptional activation of the human TNF-α promoter through interference with the basal transcription machinery. Thus, shikonins may have clinical potential as anti-inflammatory therapeutics.

Flavokawain B, a novel chalcone from Alpinia pricei Hayata with potent apoptotic activity: Involvement of ROS and GADD153 upstream of mitochondria-dependent apoptosis in HCT116 cells

Flavonoids synthesized from chalcone precursors in plants have been shown to possess cytotoxic activities with therapeutic potential. We have isolated the novel chalcone flavokawain B from Alpinia pricei Hayata, a plant native to Taiwan that is used in food and traditional Chinese medicine. Here, we report for the first time that flavokawain B significantly inhibits the growth of colon cancer cells and provide novel insight into the molecular mechanisms that underlie its apoptotic activity. Flavokawain B exerts its apoptotic action through ROS generation and GADD153 up-regulation, which lead to mitochondria-dependent apoptosis characterized by release of cytochrome c and translocation of Bak. The up-regulation of GADD153 in flavokawain B-treated HCT116 cells is associated with mitochondrial dysfunction and altered expression of Bcl-2 family members. Moreover, pretreatment with the ROS scavenger N-acetylcysteine abolishes flavokawain B-induced ROS generation, GADD153 up-regulation, and apoptosis. Similarly, RNAi-mediated gene silencing reduced flavokawain B-enhanced expression of GADD153 and apoptotic Bim, leading to diminished apoptosis. Interestingly, flavokawain B provokes G2/M accumulation as well as autophagy, in addition to apoptosis, suggesting that multiple pathways are activated in flavokawain B-mediated anticancer activity. Taken together, our data provide evidence for a molecular mechanism to explain the apoptotic activity of Alpinia plants, showing that flavokawain B acts through ROS generation and GADD153 up-regulation to regulate the expression of Bcl-2 family members, thereby inducing mitochondrial dysfunction and apoptosis in HCT116 cells.

Comparison of the antifungal activity of cadinane skeletal sesquiterpenoids from Taiwania (Taiwania cryptomerioides Hayata) heartwood.

Summary The antifungal activity of cadinane skeletal sesquiterpenoids from Taiwania (Taiwania cryptomerioides Hayata) heartwood is demonstrated. Using spectral analyses, the absolute structures of three main cadinanes, T-cadinol, T-muurolol, and α-cadinol, all isolated from Taiwania with HPLC, were identified. The amount of these cadinanes was also quantified using GC. The results showed that the total amount of cadinanes extracted from heartwood with n-C6H14 was 6.49 mg per kg of wood. This was much more than the essential oils collected by water distillation from leaves (0.04 mg/kg), sapwood (0.36 mg/kg), or heartwood (1.77 mg/kg). Moreover, results obtained from the antifungal assays demonstrated that αcadinol exhibited the highest antifungal index for both Coriolus versicolor and Laetiporus sulphureus, followed by T-cadinol and T-muurolol. As a matter of fact, α-cadinol completely inhibited the growth of C. versicolor and L. sulphureus at the level as low as 100 ppm. Further comparison of the molecular configuration of these cadinanes reveals that cadinane skeletal sesquiterpenoids with an equatorial hydroxyl group at C-9 and a trans configuration at the ring junction, such as the case for α-cadinol, exhibited the strongest antifungal activity.

Anti-inflammation activity of fruit essential oil from Cinnamomum insularimontanum Hayata

In this study, the fruit essential oil of Cinnamomum insularimontanum was prepared by using water distillation. Followed by GC-MS analysis, the composition of fruit essential oil was characterized. The main constituents of essential oil were alpha-pinene (9.45%), camphene (1.70%), beta-pinene (4.30%), limonene (1.76%), citronellal (24.64%), citronellol (16.78%), and citral (35.89%). According to the results obtained from nitric oxide (NO) inhibitory activity assay, crude essential oil and its dominant compound (citral) presented the significant NO production inhibitory activity, IC(50) of crude essential oil and citral were 18.68 and 13.18microg/mL, respectively. Moreover, based on the results obtained from the protein expression assay, the expression of IKK, iNOS, and nuclear NF-kappaB was decreased and IkappaBalpha was increased in dose-dependent manners, it proved that the anti-inflammatory mechanism of citral was blocked via the NF-kappaB pathway, but it could not efficiently suppress the activity on COX-2. In addition, citral exhibited a potent anti-inflammatory activity in the assay of croton oil-induced mice ear edema, when the dosage was 0.1 and 0.3mg per ear, the inflammation would reduce to 22% and 83%, respectively. The results presented that the fruit essential oil of C. insularimontanum and/or citral may have a great potential to develop the anti-inflammatory medicine in the future.

Antifungal compounds in the ethyl acetate soluble fraction of the extractives of Taiwania (Taiwania cryptomerioides Hayata) heartwood

Summary This study was to isolate and identify the antifungal compounds in the ethyl acetate soluble fraction of the methanol extractives of Taiwania (Taiwania cryptomerioides Hayata) heartwood and to examine their antifungal activity. Five compounds were obtained by open column chromatography and HPLC and based upon the results from Mass, 1H-NMR, and 13C-NMR analyses. Their structures were identified, namely ferruginol, helioxanthin, savinin, taiwanin C, and hinokiol. According to the results of antifungal test, the order of antifungal index of these compounds for Coriolus versicolor (L. ex Fr.) Quel. was ferruginol > taiwanin C > savinin > hinokiol. For Laetiporus sulphureus (B. ex Fr.) Bond. it was taiwanin C > savinin > ferruginol > hinokiol.

Antitermitic activity of essential oils and components from Taiwania (Taiwania cryptomerioides)

Antitermitic activity of Taiwania (Taiwania cryptomerioides Hayata) against Coptotermes formosanus Shiraki was demonstrated in laboratory tests. Blocks of sapwood and heartwood from T. cryptomerioides exhibited antitermitic activity. Bioassays revealed that heartwood essential oil exhibited the highest antitermitic activity, followed by sapwood essential oil and then the n-C6H14soluble fraction when tested at 10 mg/g. The order of termite mortality of three compounds purified from n-C6H14soluble extracts of heartwood was cedrol > α-cadinol > ferruginol. The termite resistance of T. cryptomerioides wood can be attributed to the termiticidal activity of cedrol and α-cadinol.

The chalcone flavokawain B induces G2/M cell-cycle arrest and apoptosis in human oral carcinoma HSC-3 cells through the intracellular ROS generation and downregulation of the Akt/p38 MAPK signaling pathway.

Chalcones have been described to represent cancer chemopreventive food components that are rich in fruits and vegetables. In this study, we examined the anti-oral cancer effect of flavokawain B (FKB), a naturally occurring chalcone isolated from Alpinia pricei (shell gingers), and revealed its molecular mechanism of action. Treatment of human oral carcinoma (HSC-3) cells with FKB (1.25-10 μg/mL; 4.4-35.2 μM) inhibited cell viability and caused G(2)/M arrest through reductions in cyclin A/B1, Cdc2, and Cdc25C levels. Moreover, FKB treatment resulted in the induction of apoptosis, which was associated with DNA fragmentation, mitochondria dysfunction, cytochrome c and AIF release, caspase-3 and caspase-9 activation, and Bcl-2/Bax dysregulation. Furthermore, increased Fas activity and procaspase-8, procaspase-4, and procaspase-12 cleavages were accompanied by death receptor and ER-stress, indicating the involvement of mitochondria, death-receptor, and ER-stress signaling pathways. FKB induces apoptosis through ROS generation as evidenced by the upregulation of oxidative-stress markers HO-1/Nrf2. This mechanism was further confirmed by the finding that the antioxidant N-acetylcysteine (NAC) significantly blocked ROS generation and consequently inhibited FKB-induced apoptosis. Moreover, FKB downregulated the phosphorylation of Akt and p38 MAPK, while their inhibitors LY294002 and SB203580, respectively, induced G(2)/M arrest and apoptosis. The profound reduction in cell number was observed in combination treatment with FKB and Akt/p38 MAPK inhibitors, indicating that the disruption of Akt and p38 MAPK cascades plays a functional role in FKB-induced G(2)/M arrest and apoptosis in HSC-3 cells.

Antioxidant Activity of Abietane-Type Diterpenes from Heartwood of Taiwania cryptomerioides Hayata

Summary Nine abietane-type diterpenes were isolated from the heartwood of Taiwania cryptomerioides and their structures identified by spectral analyses. Among these nine compounds, six diterpenes were isolated for the first time from T. cryptomerioides, including 6,7-dehydroferruginol, 6α-hydroxysugiol, 5,6-dehydro-6-hydroxysugiol, 11-hydroxyferruginol, secoabietane dialdehyde and isohinokiol. We suggest that abietane-type diterpenes are the dominant diterpenes in the heartwood of T. cryptomerioides. A possible biosynthesis pathway is proposed. In addition, a 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay was performed to evaluate the antioxidant activity of these diterpenes. This study demonstrates that ferruginol exhibits the strongest antioxidant activity among the diterpenes isolated from T. cryptomerioides heartwood.

Antimite Activity of Essential Oils and Their Constituents from Taiwania cryptomerioides

Abstract Antimite activity of essential oils and their components obtained from Taiwania cryptomerioides Hayata heartwood against Dermatophagoides pteronyssinus (Trouessart) and Dermatophagoides farinae Hughes was investigated in this study. Results from antimite tests demonstrated that the essential oil extracted from T. cryptomerioides heartwood had miticidal activity against D. pteronyssinus and D. farinae with a mortality of 67.0 and 36.7% at the dosage of 12.6 μg/cm2 after 48 h. Alpha-cadinol possessed the strongest antimite activity compared with other components of the T. cryptomerioides heartwood essential oil. The rectified mortalities of D. pteronyssinus and D. farinae were 100% for α-cadinol at the dosage of 6.3 μg/cm2. The order of antimite activity of four dominant constituents was α-cadinol > T-muurolol > ferruginol > T-cadinol. Paired Student’s t-tests showed that there were significant differences between the rectified mortality of α-cadinol, T-muurolol, ferruginol and that of T-cadinol at the dosage of 6.3 μg/cm2 after 48 h.