Phloridzin Acts as an Inhibitor of Protein-Tyrosine Phosphatase MEG2 Relevant to Insulin Resistance.

Inhibition of the megakaryocyte protein tyrosine phosphatase 2 (PTP-MEG2, also named PTPN9) activity has been shown to be a potential therapeutic strategy for the treatment of type 2 diabetes. Previously, we reported that PTP-MEG2 knockdown enhances adenosine monophosphate activated protein kinase (AMPK) phosphorylation, suggesting that PTP-MEG2 may be a potential antidiabetic target. In this study, we found that phloridzin, isolated from Ulmus davidiana var. japonica, inhibits the catalytic activity of PTP-MEG2 (half-inhibitory concentration, IC50 = 32 ± 1.06 µM) in vitro, indicating that it could be a potential antidiabetic drug candidate. Importantly, phloridzin stimulated glucose uptake by differentiated 3T3-L1 adipocytes and C2C12 muscle cells compared to that by the control cells. Moreover, phloridzin led to the enhanced phosphorylation of AMPK and Akt relevant to increased insulin sensitivity. Importantly, phloridzin attenuated palmitate-induced insulin resistance in C2C12 muscle cells. We also found that phloridzin did not accelerate adipocyte differentiation, suggesting that phloridzin improves insulin sensitivity without significant lipid accumulation. Taken together, our results demonstrate that phloridzin, an inhibitor of PTP-MEG2, stimulates glucose uptake through the activation of both AMPK and Akt signaling pathways. These results strongly suggest that phloridzin could be used as a potential therapeutic candidate for the treatment of type 2 diabetes.

Bioactivity-based analysis and chemical characterization of anti-inflammatory compounds from Curcuma zedoaria rhizomes using LPS-stimulated RAW264.7 cells.

Inflammation is not only a self-defense response of the innate immune system, but also the pathogenesis mechanism of multiple diseases such as arthritis, neurodegeneration, and cancer. Curcuma zedoaria Roscoe (Zingiberaceae), an indigenous plant of India, has been used traditionally in Ayurveda and folk medicine. As part of our ongoing efforts to screen traditional medicinal plants exhibiting pharmacological potential and to characterize the compounds involved, we examined the anti-inflammatory effects of the MeOH extract of C. zedoaria rhizomes using lipopolysaccharide (LPS)-stimulated RAW264.7 murine macrophage cells and found that MeOH extract inhibited the synthesis of nitric oxide (NO) in a dose-dependent manner (IC50: 23.44 ±â€¯0.77 µg/mL). In our efforts to characterize the compounds responsible for these anti-inflammatory effects, bioactivity-guided fractionation of the MeOH extract and chemical investigation of its active hexane-soluble fraction led to the successful isolation of five sesquiterpenes (1-5), the structures of which were elucidated by NMR spectroscopic analysis and LC/MS analysis. Among them, curcuzedoalide (5) exhibited potent inhibitory effects on NO synthesis (IC50: 12.21 ±â€¯1.67 µM) and also suppressed pre-inflammatory protein expression of iNOS and COX-2. Curcuzedoalide (5) was thus determined to be a contributor to the anti-inflammatory effect of C. zedoaria rhizomes and could be a potential candidate for therapeutic applications.

Chemical constituents of the root bark of Ulmus davidiana var. japonica and their potential biological activities.

The root bark of Ulmus davidiana var. japonica (Ulmaceae), commonly known as yugeunpi, has been used as a traditional Korean medicine for the treatment of gastroenteric and inflammatory disorders. As part of continuing projects to discover bioactive natural products from traditional medicinal plants with pharmacological potential, phytochemical investigation of the root bark of this plant was carried out. This led to the successful isolation of a new chromane derivative (1) and 22 known compounds: catechin derivatives (2-5), megastigmane glycoside (6), dihydrochalcone glycosides (7 and 8), flavanone glycosides (9 and 10), coumarins (11 and 12), lignan derivatives (13-17), and phenolic compounds (18-23). The structure of the new compound (1) was determined with 1D and 2D NMR spectroscopy and HR-ESIMS, and its absolute configurations were achieved by chemical reactions and the gauge-including atomic orbital (GIAO) NMR chemical shifts calculations. All the isolated compounds were evaluated for their potential biological activities including neuro-protective, anti-neuroinflammatory, and anti-Helicobacter pylori activities. Among the isolates, compounds 1, 8, and 20 displayed stronger potency by causing a greater increase in the production and the activity of nerve growth factor (NGF) in C6 glioma cells (147.04 ±â€¯4.87, 206.27 ±â€¯6.70, and 143.70 ±â€¯0.88%, respectively), whereas compounds 11, 14, and 19 inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated murine microglial cells (IC50 of 18.72, 12.31, and, 21.40 µM, respectively). In addition, compounds 1, 11, 18, and 20 showed anti-H. pylori activity with MIC values of 25 or 50 µM against two strains of H. pylori 51 and 43504. These findings provide scientific evidence that supports the traditional usage of U. davidiana var. japonica root bark in the treatment of gastroenteric and inflammatory disorders.

Comparative evaluation of bioactive phytochemicals in Spinacia oleracea cultivated under greenhouse and open field conditions.

Various factors related to growing conditions can influence the nutritional quality of plants, including vegetable crops, especially the contents of health-promoting phytochemicals. In this study, the phytochemical contents of spinach (Spinacia oleracea) cultivated under greenhouse and open field conditions were comparatively analyzed using a metabolomic approach with Mass Profiler Professional (MPP) software. S. oleracea, which is one of the well-known leafy vegetables belonging to the family Chenopodiaceae, is cultivated worldwide. Although the nutritional value of spinach is high, the phytochemical contents of spinach cultivated under greenhouse and open field conditions have not been comparatively analyzed. Metabolomic analysis of the methanol (MeOH) extracts of greenhouse-cultivated spinach (GS) and open field-cultivated spinach (OFS) using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS), followed by principal component analysis (PCA) with MPP demonstrated the differential metabolite profiles of GS and OFS. The active compounds 1-3 were isolated and identified using LC-Q-TOF-MS-guided fractionation. Among these, 5,3',4'-trihydroxy-3-methoxy-6,7-methylenedioxyflavone 4'-glucuronide (2) exhibited growth-inhibitory activities against Helicobacter pylori 51. Distribution analysis of compound 2 revealed that the anti-H. pylori compound 2 is an OFS-specific bioactive phytochemical. This indicates that the phytochemical quality of OFS is better than that of GS. These findings will aid in providing vital data for vegetable processors, dieticians, nutritionists, and consumers to select optimal green leafy vegetables.

Bioactivity-based analysis and chemical characterization of cytotoxic compounds from a poisonous mushroom, Amanita spissacea, in human lung cancer cells in vitro.

As part of our systematic study on Korean toxic mushrooms, bioactivity-guided fractionation of the MeOH extract of Amanita spissacea (Amanitaceae) fruiting bodies and chemical investigation of its cytotoxic fractions led to the isolation of (9E)-8-oxo-9-octadecenoic acid (1), (10E)-9-oxo-10-octadecenoic acid (2), (9E)-8-oxo-9-octadecenoate methyl ester (3), (9Z)-9-octadecenoate-(2'S)-2',3'-dihydroxypropyl ester (4), (9Z)-9-octadecenoic acid (5), and palmitic acid (6). The structures of the isolates were elucidated by NMR spectroscopic analysis and LC/MS analysis. Among the isolated compounds, compounds 1 and 2 exhibited the most potent cytotoxic activity in all human lung cancer cell lines examined, with IC50 values ranging from 255.7 to 321.0 µM and 250.2 to 322.5 µM, respectively. The cytotoxicity of these compounds was also found to be mediated by apoptosis associated with caspase-3 activation. These findings provide experimental evidence suggesting the potential of A. spissacea as a promising natural source for the discovery of novel anticancer drug candidates.

Ulmus parvifolia Accelerates Skin Wound Healing by Regulating the Expression of MMPs and TGF-ß.

Ulmus parvifolia is one of the medicinal plants used traditionally for treatment of wounds. We intended to investigate the wound healing effect of the powder of Ulmus parvifolia (UP) root bark in a mouse wound healing model. We also determined the mechanisms of effects of U. parvifolia in skin and skin wound healing effects using a keratinocyte model. Animal experiments showed that the wound lesions in the mice decreased with 200 mesh U. parvifolia root bark powder and were significantly reduced with treatment by UP, compared with those treated with Ulmus macrocarpa (UM). Results from in vitro experiments also revealed that UP extract promoted the migration of human skin keratinocytes. UP powder treatment upregulated the expression of the matrix metalloproteinase-2 and -9 protein and significantly increased transforming growth factor (TGF)-ß levels. We confirmed that topical administration of the bark powder exerted a significant effect on skin wound healing by upregulating the expression of MMP and transforming growth factor-ß. Our study suggests that U. parvifolia may be a potential candidate for skin wound healing including epidermal skin rejuvenation.

Vulpinic Acid Controls Stem Cell Fate toward Osteogenesis and Adipogenesis.

Vulpinic acid, a naturally occurring methyl ester of pulvinic acid, has been reported to exert anti-fungal, anti-cancer, and anti-oxidative effects. However, its metabolic action has not been implicated yet. Here, we show that vulpinic acid derived from a mushroom, Pulveroboletus ravenelii controls the cell fate of mesenchymal stem cells and preadipocytes by inducing the acetylation of histone H3 and α-tubulin, respectively. The treatment of 10T1/2 mesenchymal stem cells with vulpinic acid increased the expression of Wnt6, Wnt10a, and Wnt10b, which led to osteogenesis inhibiting the adipogenic lineage commitment, through the upregulation of H3 acetylation. By contrast, treatment with vulpinic acid promoted the terminal differentiation of 3T3-L1 preadipocytes into mature adipocytes. In this process, the increase in acetylated tubulin was accompanied, while acetylated H3 was not altered. As excessive generation of adipocytes occurs, the accumulation of lipid drops was not concentrated, but dispersed into a number of adipocytes. Consistently, the expressions of lipolytic genes were upregulated and inflammatory factors were downregulated in adipocytes exposed to vulpinic acid during adipogenesis. These findings reveal the multiple actions of vulpinic acid in two stages of differentiation, promoting the osteogenesis of mesenchymal stem cells and decreasing hypertrophic adipocytes, which can provide experimental evidence for the novel metabolic advantages of vulpinic acid.

(-)-Catechin-7-O-ß-d-Apiofuranoside Inhibits Hepatic Stellate Cell Activation by Suppressing the STAT3 Signaling Pathway.

Hepatic fibrosis is characterized by the abnormal deposition of extracellular matrix (ECM) proteins. During hepatic fibrogenesis, hepatic stellate cell (HSC) activation followed by chronic injuries is considered a key event in fibrogenesis, and activated HSCs are known to comprise approximately 90% of ECM-producing myofibroblasts. Here, we demonstrated that (-)-catechin-7-O-ß-d-apiofuranoside (C7A) significantly inhibited HSC activation via blocking the signal transducer and activator of transcription 3 (STAT3) signaling pathway. This is the first study to show the hepatic protective effects of C7A with possible mechanisms in vitro and in vivo. In our bioactivity screening, we figured out that the EtOH extract of Ulmusdavidiana var. japonica root barks, which have been used as a Korean traditional medicine, inhibited collagen synthesis in HSCs. Four catechins isolated from the EtOAc fraction of the EtOH extract were compared with each other in terms of reduction in collagen, which is considered as a marker of hepatic protective effects, and C7A showed the strongest inhibitory effects on HSC activation in protein and qPCR analyses. As a possible mechanism, we investigated the effects of C7A on the STAT3 signaling pathway, which is known to activate HSCs. We found that C7A inhibited phosphorylation of STAT3 and translocation of STAT3 to nucleus. C7A also inhibited expressions of MMP-2 and MMP-9, which are downstream genes of STAT3 signaling. Anti-fibrotic effects of C7A were evaluated in a thioacetamide (TAA)-induced liver fibrosis model, which indicated that C7A significantly inhibited ECM deposition through inhibiting STAT3 signaling. C7A decreased serum levels of aspartate amino transferase and alanine transaminase, which were markedly increased by TAA injection. Moreover, ECM-associated proteins and mRNA expression were strongly suppressed by C7A. Our study provides the experimental evidence that C7A has inhibitory effects on HSC activation after live injury and has preventive and therapeutic potentials for the management of hepatic fibrosis.

Bioactivity evaluations of betulin identified from the bark of Betula platyphylla var. japonica for cancer therapy.

Identification of bioactive natural products with anticancer activity as well as alleviating effects on chemotherapy-induced side effects has significant implications for cancer treatment. Betula platyphylla var. japonica, commonly known as Asian white birch, has been used in Chinese traditional medicine for a variety of purposes. In this study, the medicinal properties of betulin from B. platyphylla var. japonica useful for cancer management were investigated. LC/MS analysis revealed that betulin is a main chemical component of the EtOH extract of B. platyphylla var. japonica bark, and betulin was isolated from EtOH extract using an LC/MS-guided isolation method. Its structure was identified with 1H and 13C NMR spectroscopic data and LC/MS analysis and then compared to the previously reported spectroscopic and physical data. We first verified the cytotoxicity of betulin against three human lung adenocarcinoma cell lines, A549, H1264, and Calu-6, with IC50 values ranging from 18.7 to 39.6 µM. Regarding alleviation of side effects associated with anticancer chemotherapy, betulin ameliorated cisplatin-induced renal cell damage to 80% of the control value from the concentration of 5 µM. In addition, betulin showed anti-gastritis activity against ethanol-induced gastric damage in rats and notably reduced the gastric damage index compared to control in a concentration-dependent manner. These findings provide the first experimental evidence for potential use of B. platyphylla var. japonica as a functional food for cancer treatment that simultaneously alleviates the side effects of chemotherapy.

Bioactivity-guided isolation of ginsenosides from Korean Red Ginseng with cytotoxic activity against human lung adenocarcinoma cells.

BACKGROUND: Lung cancer is the leading cause of cancer-related death worldwide. In this study, we used a bioactivity-guided isolation technique to identify constituents of Korean Red Ginseng (KRG) with antiproliferative activity against human lung adenocarcinoma cells. METHODS: Bioactivity-guided fractionation and preparative/semipreparative HPLC purification were used with LC/MS analysis to separate the bioactive constituents. Cell viability and apoptosis in human lung cancer cell lines (A549, H1264, H1299, and Calu-6) after treatment with KRG extract fractions and constituents thereof were assessed using the water-soluble tetrazolium salt (WST-1) assay and terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, respectively. Caspase activation was assessed by detecting its surrogate marker, cleaved poly adenosine diphosphate (ADP-ribose) polymerase, using an immunoblot assay. The expression and subcellular localization of apoptosis-inducing factor were assessed using immunoblotting and immunofluorescence, respectively. RESULTS AND CONCLUSION: Bioactivity-guided fractionation of the KRG extract revealed that its ethyl acetate-soluble fraction exerts significant cytotoxic activity against all human lung cancer cell lines tested by inducing apoptosis. Chemical investigation of the ethyl acetatesoluble fraction led to the isolation of six ginsenosides, including ginsenoside Rb1 (1), ginsenoside Rb2 (2), ginsenoside Rc (3), ginsenoside Rd (4), ginsenoside Rg1 (5), and ginsenoside Rg3 (6). Among the isolated ginsenosides, ginsenoside Rg3 exhibited the most cytotoxic activity against all human lung cancer cell lines examined, with IC50 values ranging from 161.1 µM to 264.6 µM. The cytotoxicity of ginsenoside Rg3 was found to be mediated by induction of apoptosis in a caspase-independent manner. These findings provide experimental evidence for a novel biological activity of ginsenoside Rg3 against human lung cancer cells.