Lie-Fen Shyur

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.

Metabolomics for phytomedicine research and drug development

Metabolomics, including both targeted and global metabolite profiling strategies, is fast becoming the approach of choice across a broad range of sciences including systems biology, drug discovery, molecular and cell biology, and other medical and agricultural sciences. New analytical and bioinformatics technologies and techniques are continually being created or optimized, significantly increasing the crossdisciplinary capabilities of this new biology. The metabolomes of medicinal plants are particularly a valuable natural resource for the evidence-based development of new phytotherapeutics and nutraceuticals. Comparative metabolomics platforms are evolving into novel technologies for monitoring disease development, drug metabolism, and chemical toxicology. An efficient multidisciplinary marriage of these emerging metabolomics techniques with agricultural biotechnology will greatly benefit both basic and applied medical research.

Crystal Structure of a Natural Circularly Permuted Jellyroll Protein: 1,3-1,4-β-d-Glucanase from Fibrobacter succinogenes

The 1,3-1,4-beta-D-glucanase from Fibrobacter succinogenes (Fsbeta-glucanase) is classified as one of the family 16 glycosyl hydrolases. It hydrolyzes the glycosidic bond in the mixed-linked glucans containing beta-1,3- and beta-1,4-glycosidic linkages. We constructed a truncated form of recombinant Fsbeta-glucanase containing the catalytic domain from amino acid residues 1-258, which exhibited a higher thermal stability and enzymatic activity than the full-length enzyme. The crystal structure of the truncated Fsbeta-glucanase was solved at a resolution of 1.7A by the multiple wavelength anomalous dispersion (MAD) method using the anomalous signals from the seleno-methionine-labeled protein. The overall topology of the truncated Fsbeta-glucanase consists mainly of two eight-stranded anti-parallel beta-sheets arranged in a jellyroll beta-sandwich, similar to the fold of many glycosyl hydrolases and carbohydrate-binding modules. Sequence comparison with other bacterial glucanases showed that Fsbeta-glucanase is the only naturally occurring circularly permuted beta-glucanase with reversed sequences. Structural comparison shows that the engineered circular-permuted Bacillus enzymes are more similar to their parent enzymes with which they share approximately 70% sequence identity, than to the naturally occurring Fsbeta-glucanase of similar topology with 30% identity. This result suggests that protein structure relies more on sequence identity than topology. The high-resolution structure of Fsbeta-glucanase provides a structural rationale for the different activities obtained from a series of mutant glucanases and a basis for the development of engineered enzymes with increased activity and structural stability.

Deoxyelephantopin, a novel multifunctional agent, suppresses mammary tumour growth and lung metastasis and doubles survival time in mice.

Background and purpose:  Elephantopus scaber L. (Asteraceae) is a traditional herbal medicine with anti‐cancer effects. We evaluated the in vitro and in vivo efficacy of a major sesquiterpene lactone constituent of E. scaber, deoxyelephantopin (DET), against mammary adenocarcinoma and the underlying molecular mechanism.

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.

Genomics and proteomics of immune modulatory effects of a butanol fraction of echinacea purpurea in human dendritic cells

BackgroundEchinacea spp. extracts and the derived phytocompounds have been shown to induce specific immune cell activities and are popularly used as food supplements or nutraceuticals for immuno-modulatory functions. Dendritic cells (DCs), the most potent antigen presenting cells, play an important role in both innate and adaptive immunities. In this study, we investigated the specific and differential gene expression in human immature DCs (iDCs) in response to treatment with a butanol fraction containing defined bioactive phytocompounds extracted from stems and leaves of Echinacea purpurea, that we denoted [BF/S+L/Ep].ResultsAffymetrix DNA microarray results showed significant up regulation of specific genes for cytokines (IL-8, IL-1β, and IL-18) and chemokines (CXCL 2, CCL 5, and CCL 2) within 4 h after [BF/S+L/Ep] treatment of iDCs. Bioinformatics analysis of genes expressed in [BF/S+L/Ep]-treated DCs revealed a key-signaling network involving a number of immune-modulatory molecules leading to the activation of a downstream molecule, adenylate cyclase 8. Proteomic analysis showed increased expression of antioxidant and cytoskeletal proteins after treatment with [BF/S+L/Ep] and cichoric acid.ConclusionThis study provides information on candidate target molecules and molecular signaling mechanisms for future systematic research into the immune-modulatory activities of an important traditional medicinal herb and its derived phytocompounds.

A galactolipid possesses novel cancer chemopreventive effects by suppressing inflammatory mediators and mouse B16 melanoma.

Crassocephalum rabens (Asteraceae) is a popular anti-inflammatory folk medicine and food supplement. We investigated the cancer chemopreventive bioactivity of C. rabens phytocompounds in vitro and in vivo using cell- and gene-based bioassays and a mouse B16 melanoma model. The bioactive glyceroglycolipid 1,2-di-O-alpha-linolenoyl-3-O-beta-galactopyranosyl-sn-glycerol (dLGG) that was identified from C. rabens was found in vitro and in vivo to be a potent nitric oxide (NO) scavenger. dLGG treatment inhibited both mRNA and protein expression of inducible NO synthase and cyclooxygenase-2 (COX-2) in murine macrophages and inhibited COX-2 gene transcription in 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated B16 cells. In immunohistochemical studies, dLGG inhibited TPA-induced expression of COX-2 and nitration of proteins in mouse skin. dLGG could also significantly inhibit lipopolysaccharide-induced prostaglandin E(2) production in murine macrophages. Furthermore, dLGG prevented nuclear translocation of cytoplasmic nuclear factor-kappaB (NF-kappaB) by suppressing IkappaBalpha phosphorylation and degradation. Structure-activity relationship study by electrophoretic mobility shift assay indicated that the dilinolenoylglycerol moiety in dLGG is the essential structural feature preventing NF-kappaB.DNA complex formation. A dLGG-enriched extract from C. rabens (10 mg/kg) markedly suppressed B16 melanoma growth in C57BL/6J mice following i.p. administration, an effect comparable with that of cisplatin, a cancer chemotherapeutic drug. This study shows the detailed molecular mechanism(s) underlying the anti-inflammatory and tumor-suppressive effects of a natural galactolipid.

Modulatory effects of Echinacea purpurea extracts on human dendritic cells : A cell-and gene-based study

Echinacea spp. are popularly used as an herbal medicine or food supplement for enhancing the immune system. This study shows that plant extracts from root [R] and stem plus leaf [S+L] tissues of E. purpurea exhibit opposite (enhancing vs inhibitory) modulatory effects on the expression of the CD83 marker in human dendritic cells (DCs), which are known as professional antigen-presenting cells. We developed a function-targeted DNA microarray system to characterize the effects of phytocompounds on human DCs. Down-regulation of mRNA expression of specific chemokines (e.g., CCL3 and CCL8) and their receptors (e.g., CCR1 and CCR9) was observed in [S+L]-treated DCs. Other chemokines and regulatory molecules (e.g., CCL4 and CCL2) involved in the c-Jun pathway were found to be up-regulated in [R]-treated DCs. This study, for the first time, demonstrates that E. purpurea extracts can modulate DC differentiation and expression of specific immune-related genes in DCs.

Phytoagents for Cancer Management: Regulation of Nucleic Acid Oxidation, ROS, and Related Mechanisms

Accumulation of oxidized nucleic acids causes genomic instability leading to senescence, apoptosis, and tumorigenesis. Phytoagents are known to reduce the risk of cancer development; whether such effects are through regulating the extent of nucleic acid oxidation remains unclear. Here, we outlined the role of reactive oxygen species in nucleic acid oxidation as a driving force in cancer progression. The consequential relationship between genome instability and cancer progression highlights the importance of modulation of cellular redox level in cancer management. Current epidemiological and experimental evidence demonstrate the effects and modes of action of phytoagents in nucleic acid oxidation and provide rationales for the use of phytoagents as chemopreventive or therapeutic agents. Vitamins and various phytoagents antagonize carcinogen-triggered oxidative stress by scavenging free radicals and/or activating endogenous defence systems such as Nrf2-regulated antioxidant genes or pathways. Moreover, metal ion chelation by phytoagents helps to attenuate oxidative DNA damage caused by transition metal ions. Besides, the prooxidant effects of some phytoagents pose selective cytotoxicity on cancer cells and shed light on a new strategy of cancer therapy. The “double-edged sword” role of phytoagents as redox regulators in nucleic acid oxidation and their possible roles in cancer prevention or therapy are discussed in this review.