Wedelolactone, a Component from Eclipta prostrata (L.) L., Inhibits the Proliferation and Migration of Head and Neck Squamous Cancer Cells through the AhR Pathway.

BACKGROUND: Ecliptae prostrata (L.) L. has been widely used in East Asia with reported biological activities, including anti-cancer properties. OBJECTIVES: We aimed to investigate the effect of ethyl acetate extract of Ecliptae prostrata (L.) L. (EAE) and its component wedelolactone on the proliferation and migration of head and neck squamous cancer cells. METHODS: The proliferation of human SCC-4 and mouse CU110-1 tongue squamous carcinoma cells was assessed using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) method. Scratch wound assays were performed to assess cell migration rates. The levels of Ecadherin and vimentin were used as markers of the epithelial-to-mesenchymal transition (EMT). AhR, CYP1A1, and CYP1B1 levels were examined to uncover the mechanism of inhibition of cell migration by wedelolactone. RESULTS: We found that EAE and wedelolactone decreased the proliferation of human SCC-4 cells and mouse CU110-1 cells at doses of EAE at > 25 µg/ml and wedelolactone at > 6.25 µg/ml. Similarly, both EAE and wedelolactone produced inhibitory effects against migration, but the effective doses that significantly inhibited migration were lower than those affecting proliferation. Wedelolactone below 12.5 µg/ml inhibited the epithelial-to-mesenchymal transition (EMT) with increased expression of E-cadherin and decreased expression of vimentin in SCC-4 and CU110-1 cells. Further analysis showed wedelolactone inhibited the expression of AhR and its downstream target molecules CYP1A1 and CYP1B1 in both squamous carcinoma cells at the same doses inhibiting cell migration. The addition of benzo (a)pyrene [B(a)P], an agonist of AhR, stimulated migration, especially in the CU110-1 cells with reported cancer stem cell-like characteristics. Instructively, B(a)P reversed the inhibitory effects of wedelolactone on AhR expression and cell migration, suggesting that wedelolactone antagonizes cell migration through the AhR pathway. Moreover, the higher activity of EAE and wedelolactone against the migration of cancer stem-like CU110-1 cells relative to SCC-4 cells suggests selective activity against cancer stem cells. CONCLUSION: Our study identifies wedelolactone as a major active component of Ecliptae prostrata (L.) L. with promising anti-cancer properties against head and neck squamous cancer cells.

Three new biflavonoids from the branches and leaves of Cephalotaxus oliveri and their antioxidant activity.

Three new biflavonoids, named oliveriflavones A-C (1-3), together with two known flavonoids (quercetin (4) and rutin (5)), were isolated from the endangered plant Cephalotaxus oliveri. The chemical structures of these compounds were elucidated by comprehensive spectroscopic methods including NMR, HRESIMS, IR, UV, and CD spectra. Compounds 1-5 were first isolated from the genus Cephalotaxus. All the compounds were tested for their antioxidant activity. Compounds 4 and 5 showed excellent activity with IC50 values of 0.03 ± 0.06 µM and 0.02 ± 0.10 µM, respectively.

Three new alkaloids isolated from the stem tuber of Pinellia pedatisecta.

The present study was designed to determine the chemical constituents of the stem tuber of Pinellia pedatisecta. The chemical constituents were isolated and purified by various chromatographic techniques, and their structures were elucidated on the basis of physicochemical properties and spectral data. Three new alkaloids (compounds 1, 2, and 3) were obtained and identified as 9-((5-methoxypyridin-2-yl)methyl)-9H-purin-6-amine (1), 4-(2-(2, 5-dioxopyrrolidin-1-yl)ethyl)phenyl acetate (2), and N-(9-((5-methoxypyridin-2-yl)methyl)-9H-purin-6-yl)acetamide (3). These compounds were evaluated for their cytotoxicity against human cervical cancer HeLa cells. Compounds 1 and 3 significantly inhibited the proliferation of HeLa cells with IC50 values being 3.02 ± 0.54 and 7.16 ± 0.62 µmol·L-1, respectively.

The inhibitory effect of Aconiti Sinomontani Radix extracts on the proliferation and migration of human synovial fibroblast cell line SW982.

ETHNOPHARMACOLOGICAL RELEVANCE: Aconiti Sinomontani Radix is frequently used in the treatment of Bi syndrome in traditional Chinese medicine. Several reports indicate that Aconiti Sinomontani Radix has therapeutic effects for rheumatoid arthritis (RA). However, the cellular mode of action is still unclear. To investigate the effect of alkaloid extracts of Aconiti Sinomontani Radix on proliferation and migration of human synovial sarcoma SW982 cells as well as the molecular mechanism underlying. MATERIALS AND METHODS: SW982 cells were examined for proliferation by a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) method. Wound scratch assays were performed to assess the migrated rate of SW982 cells. Quantitative real-time PCR was used to measure the mRNA expression levels of Wnt5a, Runx2, MMP3, and Bmp2. Western blotting was used to measure the phosphorylated levels of JNK and NF-κB as well as the expression of MMP3. RESULTS: The alkaloid extract from Aconiti Sinomontani Radix (MQA) and MQB, which removed lappaconitine from MQA significantly inhibited the proliferation of SW982 in a dose-dependent manner. The proliferation inhibitory effect of MQB was more potent. Incubation with 10µg/ml MQB for 12, 24, and 36h inhibited the migration of SW982 cells by 83%, 58%, and 42%, respectively. Treatment with different concentrations of MQB for 24h inhibited mRNA expression of Wnt5a, Runx2, and MMP3, but Bmp2 mRNA expression was elevated by MQB. Further, MQB inhibited phosphorylation of JNK and NF-κB p65 as well as MMP3 expression by Western blotting analysis. CONCLUSION: The results showed that MQB inhibited proliferation and migration of SW982 cells possibly through suppressing Wnt5a-mediated JNK and NF-κB pathways. These results indicated that MQB might be an active extract of Aconiti Sinomontani Radix for targeting fibroblast-like synoviocytes (FLS) and be potential for RA therapy.

Anti-inflammatory and antioxidant activities of flavonoids from the flowers of Hosta plantaginea.

Hosta plantaginea was a traditional Chinese medicinal plant used to treat inflammation-related diseases with little scientific validation. Twelve flavonoids, including two new compounds namely plantanones A (1) and B (2), were isolated from the flowers of Hosta plantaginea. Their structures were elucidated by NMR and HRMS as well as comparison with literature data. All of the isolated compounds showed significant inhibitory activities against ovine COX-1 and COX-2 at a concentration of 50 µM, with inhibition ratios from 53.00% to 80.55% for COX-1 and from 52.19% to 66.29% for COX-2. Further detailed testing showed that compounds 1, 2, 4 and 12 inhibited the COX-1 and COX-2 enzymes with IC50 values 12.90-33.37 µM and 38.32-46.16 µM, respectively. Moreover, the antioxidant effects of these isolates against DPPH free radical-scavenging were also evaluated in vitro, and a tight structure-activity relationship was discussed. Our results suggested that the anti-inflammatory and antioxidant activities of H. plantaginea flowers are partly attributed to these flavonoids.

A microbial model of mammalian metabolism: biotransformation of 4,5-dimethoxyl-canthin-6-one using Cunninghamella blakesleeana CGMCC 3.970.

1. A filamentous fungus, Cunninghamella blakesleeana CGMCC 3.970, was applied as a microbial system to mimic mammalian metabolism of 4,5-dimethoxyl-canthin-6-one (1). Compound 1 belongs to canthin-6-one type alkaloids, which is a major bioactive constituent of a traditional Chinese medicine (the stems of Picrasma quassioides). 2. After 72 h of incubation in potato dextrose broth, 1 was metabolized to seven metabolites as follows: 4-methoxyl-5-hydroxyl-canthin-6-one (M1), 4-hydroxyl-5-methoxyl-canthin-6-one (M2), canthin-6-one (M3), canthin-6-one N-oxide (M4), 10-hydroxyl-4,5-dimethoxyl-canthin-6-one (M5), 1-methoxycarbonl-ß-carboline (M6), and 4-methoxyl-5-O-ß-D-glucopyranosyl-canthin-6-one (M7). 3. The structures of metabolites were determined using spectroscopic analyses, chemical methods, and comparison of NMR data with those of known compounds. Among them, M7 was a new compound. 4. The metabolic pathways of 1 were proposed, and the metabolic processes involved phase I (O-demethylation, dehydroxylation, demethoxylation, N-oxidation, hydroxylation, and oxidative ring cleavage) and phase II (glycosylation) reactions. 5. This was the first research on microbial transformation of canthin-6-one alkaloid, which could be a useful microbial model for producing the mammalian phase I and phase II metabolites of canthin-6-one alkaloids. 6. 1, M1-M5, and M7 are canthin-6-one alkaloids, whereas M6 belongs to ß-carboline type alkaloids. The strain of Cunninghamella blakesleeana can supply an approach to transform canthin-6-one type alkaloids into ß-carboline type alkaloids.

Novel phthalide derivatives identified from Ligusticum chuanxiong (Chuanxiong).

BACKGROUND: Ligusticum chuanxiong Hort. (Chuanxiong) is a well-known Chinese medicine, and studies on its chemical constituents are important for explaining its mechanism of action and quality control. This study aims to investigate the chemical constituents of the dried rhizome of. L. chuanxiong. METHODS: The dried rhizome of L. chuanxiong was extracted with 60 % ethanol, and the concentrated extract was isolated by silica gel, octadecyl silane, and Sephadex LH-20 columns, followed by preparative/semipreparative high-performance liquid chromatography (HPLC) to obtain the pure chemical constituents. The structures of the constituents were elucidated by HR-ESI-MS, UV, IR, 1D NMR, and 2D NMR methods. Enantiomeric separation was achieved by a chiral HPLC method. The absolute configuration was determined by the modified Mosher's method. RESULTS: Six novel phthalide derivatives, (+)/(-)-chuanxiongins A-F (1-6), together with four known phthalides (7-10) were isolated from Chuanxiong. All of the new compounds (1-6) were present as pairs of enantiomers. Enantiomeric separation of 1 was successfully achieved by HPLC on a chiral column. The absolute configuration of (-)-1 was determined by a modified Mosher's method. CONCLUSION: The six novel phthalide derivatives (1-6) isolated from Chuanxiong were phthalide fatty acid esters that were structurally analogous and characterized by fatty acid acylation at 6-OH or 7-OH.

Puberunine and puberudine, two new C18-diterpenoid alkaloids from Aconitum barbatum var. puberulum.

Two C(18)-diterpenoid alkaloids, puberunine (1) and puberudine (2), together with four other new alkaloids, including the first examples having ß-oriented substitution at C-3 and a rare chloro-substituent were isolated from Aconitum barbatum var. puberulum. Their structures were elucidated by spectroscopic methods. Puberunine and puberudine, which possess a unique rearranged E ring and an opened A ring, respectively, represent new subtypes of the C(18)-diterpenoid alkaloids. A plausible biosynthetic pathway of 1 and 2 was proposed.

Houttuynoids A-E, anti-herpes simplex virus active flavonoids with novel skeletons from Houttuynia cordata.

Houttuynoids A-E (1-5), a new type of flavonoid with houttuynin tethered to hyperoside, and their presumed biosynthetic precursor hyperoside (6) were isolated from the whole plant of Houttuynia cordata. Their structures were elucidated by analysis of 1D and 2D NMR. A hypothetical biogenetic pathway for houttuynoids A-E was proposed. Compounds 1-5 exhibited potent anti-HSV (herpes simplex viruses) activity.

Fas/FasL signaling allows extracelluar-signal regulated kinase to regulate cytochrome c release in oridonin-induced apoptotic U937 cells.

Previously, we found that human histocytic lymphoma U937 cells possessed high susceptibility to oridonin-induced cell death, but the molecular mechanisms in response to oridonin remain unclear. In this study, U937 cells showed susceptible to apoptosis induced by 27 microM oridonin and an agonistic anti-Fas IgM mAb (CH-11) (500 ng/ml) as a Fas-sensitized positive control. Caspase 8 inhibitor z-IETD, but neither caspase 1 inhibitor Ac-YVAD nor caspase 10 inhibitor z-AEVD, effectively blocked oridonin-induced cell death as well as DNA fragmentation. Western blot analysis showed the up-regulated expression of Fas, FasL, and FADD, and down-regulated expression of procaspase 8, suggesting that Fas/FasL pathway was activated in oridonin-induced cell apoptosis. Further, stimulation of U937 cells with oridonin and CH11 resulted in significant ERK MAPK activation. However, inhibition of ERK by PD98059 reversed oridonin-induced cell death as well as the activation of caspase 8, indicating that ERK-mediated control occured upstream of caspase 8. Simultaneously, ERK activation accounted for the release of cytochrome c, but failed to influence decreased Bcl-2 expression induced by oridonin. Taken together, these results suggest that Fas/FasL signaling pathway-mediated ERK activation sensitized U937 cells to mitochondrial pathway-mediated apoptosis induced by oridonin.