Anneslea fragrans Wall. ameliorates ulcerative colitis via inhibiting NF-κB and MAPK activation and mediating intestinal barrier integrity.

ETHNOPHARMACOLOGICAL RELEVANCE: Anneslea fragrans Wall. is traditionally used as a folk medicine in treating indigestion, fever, dysentery, diarrhea, and liver inflammation in China, Vietnam and Cambodia. However, its anti-inflammatory activity and mechanism under a safety therapeutic dose as well as the main chemical components have not yet been fully investigated. AIM OF THE STUDY: This study aimed to explore the therapeutic effect and possible molecular mechanisms of aqueous-methanol extract (AFE) of A. fragrans leaves on dextran sodium sulfate (DSS)-induced ulcerative colitis (UC) mice and illustrate its potent anti-inflammatory chemical compounds. MATERIALS AND METHODS: The AFE was obtained and then analyzed by high performance liquid chromatography (HPLC). Phytochemical investigation on the AFE was carried out to isolate and characterize its major components. The acute toxicity test was performed to provide the safety information of AFE. Subsequently, the protective effect of AFE on DSS-induced UC was evaluated by physiological changes, histopathological and immunohistochemical analysis, and the expressions of antioxidant enzyme, pro-inflammatory cytokines and anti-inflammatory cytokines. The expressions of target proteins in nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) were determined by western blot analysis. The tight junction (TJ) proteins in colon tissue were performed by immunohistochemical technique for evaluating the intestinal barrier integrity. RESULTS: HPLC guided isolation of AFE resulted into two dihydrochalcones, which were elucidated as vacciniifolin (1) and confusoside (2). Acute toxicity evaluation revealed that median lethal dose (LD50) of AFE was greater than 5000 mg/kg. Furthermore, AFE significantly attenuated ulcerative colitis symptoms, suppressed myeloperoxidase activity, and increased the expression of superoxide dismutase and glutathione. AFE treatment could also reduce the levels of tumor necrosis factor-α, interleukin-1ß, and interleukin-6 and increase the levels of interleukin-4 and interleukin-10 in colon tissues and serum of DSS-induced UC mice. In addition, AFE significantly increased the expression of zonula occludens-1, occludin and claudin-1, and inhibited the phosphorylation of target protein of the NF-κB and MAPK signaling pathways in colon tissue. CONCLUSION: Dihydrochalcone glycosides are the major chemical constituents in AFE. AFE ameliorated DSS-induced UC in mice by inhibiting the inflammatory response via modulation of NF-κB and MAPK pathways and maintaining the intestinal barrier function, indicating that the plant A. fragrans could be used as a therapeutic candidate for ulcerative colitis.

Anticandidal Potential of Stem Bark Extract from Schima superba and the Identification of Its Major Anticandidal Compound.

Plant-derived extracts are a promising source of new drugs. Schima superba is traditionally used in China for heat clearing, detoxification, and treatment of furuncles. In this study, the anticandidal properties and mechanism of action of S. superba (SSE) were explored using a stem bark extract. SSE possessed high polyphenol and saponin contents of 256.6 ± 5.1 and 357.8 ± 31.5 µg/mg, respectively. A clear inhibition zone was observed for C. albicans growth through the disc diffusion method and the 50% inhibition of C. albicans by SSE was 415.2 µg/mL. Transcriptomic analysis in C. albicans treated with different doses of SSE was conducted through RNA-seq. Average values of 6068 genes and 20,842,500 clean reads were identified from each sample. Among these samples, 1680 and 1956 genes were differentially expressed genes (DEGs) from the SSE treatments of 0.2 and 0.4 mg/mL, respectively. C. albicans growth was inhibited by the changes in gene expression associated with the cell wall and membrane composition including the regulation of chitin degradation and ergosterol biosynthesis. This result could be reflected in the irregularly wrinkled morphology of the ruptured cell as revealed through SEM analysis. ESI-MS and NMR analyses revealed that the major compound purified from SSE was sasanquasaponin III and the 50% inhibition of C. albicans was 93.1 µg/mL. In summary, the traditional Chinese medicine S. superba can be applied as an anticandidal agent in complementary and alternative medicine.

Sasanquasaponin ΙΙΙ from Schima crenata Korth induces autophagy through Akt/mTOR/p70S6K pathway and promotes apoptosis in human melanoma A375 cells.

BACKGROUND: Melanoma is a high fatality skin cancer which lacks effective drugs. Sasanquasaponin, an important sort of constituents in theaceae, has been demonstrated to have potent anti-tumor effect in breast cancer and hepatocellular carcinoma. As a sasanquasaponin, we speculate that Sasanquasaponin III (SQS III) isolated from Schima crenata Korth may also have anti-tumor activity. PURPOSE: This study aims to investigate whether SQS III has anti-melanoma activity and examine the underlying mechanisms of SQS III against melanoma. METHODS/STUDY DESIGNS: The anti-proliferative effect of SQS III was assessed by cells viability assay. Annexin V-FITC/PI double staining assay was utilized for detection of apoptosis. Mitochondrial membrane potential and reactive oxygen species (ROS) production were detected using JC-1 and DCFH-DA assay, respectively. Autophagy was monitored using transmission electron microscopy (TEM) and GFP-LC3 transfection fluorescence analysis. Autophagosome-lysosome fusion and lysosomal degradation were determined using a GFP-LC3 & LAMP1 co-localization assay and DQ-BSA staining. Proteins related to apoptosis and autophagy were analyzed by Western blotting. RESULTS: Our results demonstrated that the SQS III exhibited potent anti-cancer activity in A375 cells by inducing both apoptosis and autophagy. In melanoma cells treated with SQS III, caspases were activated and PARP was cleaved, proving the occurrence of apoptosis. Mechanistic studies indicated that the pro-apoptosis activity of SQS III was mediated by death receptor pathway and mitochondrial dysfunction which was induced by ROS accumulation and reversed by the ROS inhibitor N-acetyl-cysteine (NAC). In addition to triggering apoptosis, SQS III may also cause autophagy in melanoma cells. Our results demonstrated that SQS III induced up-regulated expression of GFP-LC3, autophagosome-lysosomal fusion and lysosomal degradation. Additionally, the ROS accumulation was also involved in the activation of autophagy. Meanwhile, it was also found that after SQS III treatment, the expression of LC3-II was up-regulated and the AKT/mTOR signaling pathway was inhibited. The autophagy inhibitor 3-MA converted cytotoxicity and apoptosis of SQS III in A375 cells, which indicated that autophagy promoted the SQS III-induced apoptosis. CONCLUSION: SQS III showed potent anti-cancer activity by inducing apoptosis and autophagy, which provides insights into its possible use as a therapy for melanoma.

Two new ent-kaurane diterpenes from the stems of Eurya chinensis.

Two new ent-kaurane diterpenes (1-2), together with five known analogs, were isolated from the stems of Eurya chinensis. The structures of new compounds were established by extensive analysis of mass spectrometric and 1D and 2D NMR spectroscopic data. Compound 3 exhibited noticeable anti-inflammatory activity as denoted by inhibiting LPS-induced nitric oxide (NO) production in RAW264.7 cells with an IC50 value of 7.82 µM. Compound 4 showed potent cytotoxic activity against human cancer cell lines NCI-H46, HepG2 and SW480 with IC50 values ranging from 7.45 to 8.54 µM.

Phenylacylphenol derivatives with anti-melanogenic activity from Stewartia pseudocamellia.

Three new phenylacylphenol derivatives, stewartianol (1), deoxystewartianol-4'-O-arabinoglucoside (2), and stewartianol-3-O-glucoside (3), along with nine known compounds, methylesculin (4), fraxoside (5), fraxetin (6), scopletin (7), (+)-dihydromyricetin (8), (+)-taxifolin-7-O-ß-D-glucose (9), (+)-taxifolin (10), (+)-dihydrokaempferol-7-O-ß-D-glucose (11), and 3-acetyl-ursolic acid (12), were isolated from the twigs of Stewartia pseudocamellia; commonly used as folk medicine in Korea. The structures of the isolated compounds were identified using spectroscopic analysis, including 1D, 2D NMR, MS and compared with published data. The compounds were tested for their anti-melanogenic activity in cultured murine B16 melanoma cells. Stewartianol (1) and stewartianol-3-O-glucoside (3) showed an inhibitory effect significantly on melanogenesis in a concentration-dependent manner.

Principles of Biomedical Agriculture Applied to the Plant Family Theaceae To Identify Novel Interventions for Cancer Prevention and Control.

Plant materials from the family Theaceae have been used for over a thousand years as integral components within the food systems of many globally distributed cultures and to treat a variety of human ailments. These markedly different uses remain of considerable interest in the 21st century. This perspective draws heavily from the agricultural and biomedical literature published using plant materials from the genus Camellia. Our objective is to provide a rationale and framework for broadening the scope of investigation of genera and species within Theaceae beyond Camellia sinensis to accelerate the development of a new generation of Theaceae-based pharmaceuticals/nutraceuticals and the more general enhancement of the food supply with Theaceae-containing products that affect the development of chronic diseases such as cancer. This will require a concerted effort to systematically capitalize on the rapidly growing knowledge of germplasm resources within Theaceae using metabolomic profiling in combination with in vivo and in vitro approaches. The successful translation of this research into products that affect human health will be facilitated by recognition of the agronomic factors that are critical in making hot water infusions generically referred to as tea as well as food products containing ground leaf powders.

Triterpenoid saponins from the root bark of Schima superba and their cytotoxic activity on B16 melanoma cell line.

Eight new oleanane-type triterpenoid saponins, schisusaponins A-H, along with eight known triterpenoid saponins, were isolated from the root bark of Schima superb (Theaceae). Their structures were elucidated on the basis of extensive spectroscopic analyses and chemical methods. The cytotoxicity of the new compounds against B16 melanoma cells was assessed. Among the isolated new saponins, schisusaponins C and E showed more potent effects (with IC50 values of 10.08 and 10.89 µM) than vinblastine (with an IC50 value of 19.48 µM).

The stimulatory effects of Stewartia koreana extract on the proliferation and migration of fibroblasts and the wound healing activity of the extract in mice.

Stewartia koreana (S. koreana) has been used in the treatment of inflammatory diseases, such as acute gastroenteritis and aches, in Korean folk medicine and has been reported to have a number of biological activities, such as anti-inflammatory activity and the promotion of angiogenesis. In this study, we aimed to determine the effects of S. koreana extract (SKE) and its components on dermal fibroblast growth and migration, and to investigate the wound healing activity of the extract in mice. In vitro experiments revealed that the numbers of SKE-treated cells increased by approximately 2.5-­ and 3.7-fold with 50 and 100 µg/ml of SKE, respectively. 5-bromo-2'-deoxy-uridine (BrdU) incorporation was also increased in the SKE-treated cells by 2.3-fold. SKE promoted the migration of human skin fibroblasts and, among the isolated compounds, hyperin increased the proliferation and migration of the fibroblasts to almost the same degree as SKE. Western blot analysis demonstrated that SKE stimulated the MEK/ERK1/2 and PI3K/Akt signaling pathways. In in vivo experiments, the SKE-treated wound lesions of mice decreased by approximately 7% in diameter after 2 days of treatment with SKE compared with the wound lesions on the 1st day of the experiment. On the 9th day of treatment, the diameter of the lesions was further reduced by approximately 83% in the SKE-treated wound areas compared with the wound areas on the 1st day of treatment. Our results demonstrate that methanol extracts of S. koreana leaves promote the proliferation and migration of skin fibroblasts and possess effective wound healing activity through the activation of the MEK/ERK1/2 and PI3K/Akt signaling pathways. Hyperin was identified as an active compound responsible for the stimulation of fibroblast growth and migration.

The comparison of DPPH-scavenging capacity and anti-inflammatory effects of phenolic compounds isolated from the stems of Stewartia koreana Nakai.

The Stewartia koreana Nakai (SK) had been used in oriental traditional medicine as a remedy for acute gastroenteritis, liver diseases, quadriplegia and pain. The antioxidant activity guided isolation 80% methyl extract from stems of SK yielded eight phenolic compounds. We evaluated the anti-oxidative and anti-inflammatory effects of these compounds via assays of 1,1-diphenyl-2-picrylhydazyl (DPPH) radicals and inhibition of nitric oxide (NO) production in lipopolysaccharide-stimulated RAW 264.7 macrophage cells. The results demonstrated that syringaresinol (6) exhibited significant DPPH radical-scavenging activity and inhibitory effects on NO production compared with its positive controls, ascorbic acid and L-NMMA, respectively.

Cytotoxic triterpenoid saponins from the stems of Gordonia longicarpa.

Nine new triterpenoid saponins named longicarposides A-I (1-9), together with three known saponins (10-12), were isolated from the stems of Gordonia longicarpa. The structures of the saponins were elucidated by a combination of 1D and 2D NMR techniques, mass spectrometry, and chemical methods. They were characterized to be oleanane-type saponins with sugar moieties linked to C-3 of the aglycone. Cytotoxic activities of these saponins were evaluated against five human tumor cell lines (HCT-8, Bel-7402, BGC-823, A549, and A2780) by using the MTT in vitro assay. Compounds 5, 7, 8, 10, and 11 exhibited potent cytotoxic activity with IC50 values of 1.42-8.42 µM, while 6, 9, and 12 showed selective cytotoxic activity toward the tested cell lines.

Triterpenoid glycosides from the stems of Gordonia kwangsiensis.

Eleven oleanane-type triterpenoid glycosides, named gordonsaponins A-K, were isolated from the stems of Gordonia kwangsiensis. Their structures were elucidated by spectroscopic and chemical methods. The cytotoxic activities of all eleven were evaluated against five human tumor cell lines (HCT-8, Bel-7402, BGC-823, A549, and A2780), with only one having activity against all tested cell lines, with IC50 values ranging from 0.1 to 2.41 µM.

Potential anti-inflammatory constituents of the stems of Gordonia chrysandra.

Eight new oleanane-type triterpenoid glycosides, gordonosides I-P (1-8), and two new phenolic glycosides (9 and 10) were isolated from the stems of Gordonia chrysandra. Their structures were elucidated by spectroscopic and chemical methods. In an in vitro bioassay, compound 1 showed a strong inhibitory effect on nitric oxide production in LPS-activated macrophages with an IC50 value of 0.14 µM.

Anti-inflammatory activity of a polyphenolic enriched extract of Schima wallichii bark.

In the present investigation, the anti-inflammatory activity of a polyphenolic enriched extract of Schima wallichii bark was evaluated in vitro using human peripheral blood mononuclear cells (PBMCs) and in vivo by carrageenan-induced paw oedema assay (acute study) and cotton pallet granuloma assay (chronic study). The extract exhibited significant inhibition of the production of tumour necrotic factor-α and interleukin-6 by PBMCs stimulated with lipopolysaccharide (LPS) in a concentration-dependent manner. The extract at the selected doses of 150 and 300 mg kg(-1) body weight p.o. exhibited significant dose-dependent anti-inflammatory responses, with 44.32 and 38.65% inhibition of inflammation in carrageenan-induced paw oedema and cotton pallet granuloma, respectively.

Two new 8-O-4'-type lignans from the stem of Schima superba and their cell growth inhibitory activities against human cancer cell lines.

Two new lignans (1 and 2) were isolated from the EtOH extracts of the stem of Schima superba, and elucidated as (7R,8S)-1-(3,4-dimethoxyphenyl)-2-O-(2-methoxy-4-omegahydroxypropylphenyl)propane-1,3-diol (1) and threo-1-(4-hydroxy-3-methoxyphenyl)-1-methoxy-2-{4-[1-formyl-(E)-vinyl]-2-methoxyphenoxy}-3-propanol (2) by spectral analysis. Compounds 1 and 2 showed cell growth inhibitory activity against HeLa, CNE, HepG-2, and HEp-2 cell lines. Compound 1 exhibited significant cytotoxicities with IC50 values of less than 4 µg/ml against all the tested cell lines.

Cytotoxic triterpenoid glycosides from the roots of Gordonia chrysandra.

Eight new oleanane triterpenoid glycosides, gordonosides A-H (1-8), were isolated from a 50% EtOH extract of the roots of Gordonia chrysandra. Their structures were determined by spectroscopic analysis, including 1D and 2D NMR and ESIMS, and by chemical methods. Among these substances, compounds 1, 3, 5, and 6 exhibited cytotoxic activity against several human cancer cell lines, with 3 being the most potent.

Acylated triterpenoid saponins from Schima noronhae and their cell growth inhibitory activity.

Two new acylated triterpenoid saponins were isolated from the branches of Schima noronhae by bioassay-guided purification. Their chemical structures were established on the basis of spectroscopic analysis and chemical means as 3-O-alpha-L-rhamnopyranosyl-(1-->4)-beta-D-galactopyranosyl-(1-->3)-[beta-D-glucopyranosyl-(1-->2)]-beta-D-glucuronopyranosyl 22-O-angeloyl-A1-barrigenol (1) and 3-O-alpha-L-rhamnopyranosyl-(1-->4)-beta-D-galactopyranosyl-(1-->3)-[beta-D-glucopyranosyl-(1-->2)]-beta-D-glucuronopyranosyl 22-O-angeloylerythrodiol (2). Compounds 1 and 2 showed cell growth inhibitory activity against both HeLa and DLD1 cells at a concentration of less than 10 microM.

Inhibitory effects of Stewartia koreana on osteoclast differentiation and bone resorption.

Osteoclasts are responsible for bone lysis in several bone diseases such as osteoporosis and rheumatoid arthritis. Natural products from plants have been invaluable source in discovery of compounds for new therapies. In this study, we screened plant products for potential application to therapy for bone loss using a primary osteoclastogenesis culture system and found that extract of Stewartia koreana (SKE) had a strong inhibitory effect on osteoclast formation. To gain molecular insights, we examined the effect of SKE on signaling pathways and transcription factors stimulated by the osteoclast differentiation factor RANKL. SKE suppressed the induction of c-Fos and NFATc1 by RANKL. However, SKE did not inhibit NF-kappaB activation by RANKL. Among the MAPKs stimulated by RANKL, SKE significantly reduced the activation of ERK and p38. Therefore, the anti-osteoclastogenic effect of SKE is likely to be elicited by interference with RANKL signaling to ERK and p38, which mediate the induction of c-Fos and subsequently that of NFATc1. Consistent with the in vitro effect on osteoclast differentiation, SKE showed a great inhibitory effect on in vivo bone loss in LPS-challenged mice. Taken together, we demonstrated that SKE has inhibitory effects on osteoclast differentiation in vitro and confirmed its in vivo efficacy in prevention of inflammatory bone loss.

Methanol extracts of Stewartia koreana inhibit cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) gene expression by blocking NF-kappaB transactivation in LPS-activated RAW 264.7 cells.

Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) are involved in various pathophysiological processes such as inflammation and carcinogenesis. In a search for inhibitors of COX-2 and iNOS production we found that extracts of Stewartia koreana strongly inhibited NO and PGE2 production in LPS-treated macrophage RAW 264.7 cells. We have now shown that the mRNA and protein levels of iNOS and COX-2 are reduced by the Stewartia koreana extract (SKE). SKE inhibited expression of an NF-kappaB reporter gene in response to LPS, and gel mobility shift assays revealed that SKE reduced NF-kappaB DNA-binding activity. The extract also inhibited LPS-induced phosphorylation of IkappaB-alpha and nuclear translocation of p65. Administration of the extract reduced the symptoms of arthritis in a collagen-induced arthritic mouse model. These results indicate that Stewartia extracts contain potentially useful agents for preventing and treating inflammatory diseases.

Inhibitory effects of eutigosides isolated from Eurya emarginata on the inflammatory mediators in RAW264.7 cells.

The anti-inflammatory activity of Eurya emarginata (Thumb) Makino, of which leaves have been traditionally used to treat ulcers or diuretic in Jeju Island, has been investigated in the present study. Through the phytochemical study from the methanol extract of E. emaginiata, eutigosides B and C were isolated as the active components. Sseveral inflammatory markers including TNF-alpha, IL-1beta, IL-6, NO, iNOS, and COX-2 were examined. Eutigosides B and C potentially inhibited production of pro-inflammatory cytokines (IL-6 and TNF-alpha) in a dose-dependent manner. Additionally, the intracellular contents of iNOS protein were markedly decreased after treatment with eutigosides B and C. The inhibition of iNOS activity was correlated with the decrease in nitrite levels. These results suggest that eutigoside B and C from E. emarginata may have anti-inflammatory activity through the inhibition of pro-inflammatory cytokines (TNF-alpha and IL-6), iNOS and COX-2.

The cytotoxicity of eutigosides from Eurya emarginata against HL-60 promyelocytic leukemia cells.

Two phenolic glucosides, eutigoside B and eutigoside C were isolated from the fresh leaves of Eurya emarginata. These two phenolic glucosides exerted a significant inhibitory effect on the growth of HL-60 promyelocytic leukemia cells. Furthermore, when the HL-60 cells were treated with eutigoside C, several apoptotic characteristics such as DNA fragmentation, morphologic changes, and increase of the population of sub-G1 hypodiploid cells were observed. In order to understand the mechanism of apoptosis induction by eutigoside C, we examined the changes of Bcl-2 and Bax expression levels. The eutigoside C reduced Bcl-2 protein and mRNA levels, but slightly increased Bax protein and mRNA levels in a time-dependent manner. When we examined the activation of caspase-3, an effector of apoptosis, the eutigoside C increased the expression of active form (19-kDa) of caspase-3 and the increase of their activities was demonstrated by the cleavage of poly (ADP-ribose) polymerase, a substrate of caspase-3, to 85-kDa. The results suggest that the inhibitory effect of eutigoside C from E. emarginata on the growth of HL-60 appears to arise from the induction of apoptosis via the down-regulation of Bcl-2 and the activation of caspase.