Fermentation with mono- and mixed cultures of Lactobacillus plantarum and L. casei enhances the phytochemical content and biological activities of cherry silverberry (Elaeagnus multiflora Thunb.) fruit.

BACKGROUND: Lactic acid fermentation has been widely used to improve the nutritional and functional properties of food products. Cherry silverberry (Elaeagnus multiflora Thunb.) is considered as an invasive plant species with known medicinal and functional properties. In this study, improvement of the biological activity and health benefits of cherry silverberry fruit through lactic acid fermentation was investigated. RESULTS: Extracts of cherry silverberry fruits fermented by pure cultures of Lactobacillus plantarum KCTC 33131 and L. casei KCTC 13086 exhibited favorable physicochemical properties and enhanced phytochemical content, antioxidant properties (DPPH radical scavenging activity, reducing power, superoxide dismutase-like property and hydrogen peroxide scavenging activity) and α-glucosidase and tyrosinase enzyme inhibitory activity as compared with unfermented fruits. Despite a decrease in the specific phenolic acid contents among the fermented samples, the cherry silverberry fruit fermented by mixed cultures of L. plantarum and L. casei contained superior total polyphenols (3.78 ± 0.22 mg GAE g-1 ) and total (0.66 ± 0.12 mg QE g-1 ) and individual flavonoid contents in comparison with fruits fermented by single cultures and unfermented ones. Multivariate analysis also showed strong association among total phytochemical contents and biological activities. CONCLUSIONS: This work has elucidated the effect of fermentation with L. plantarum KCTC 33131 and L. casei KCTC 13086 on the improvement of the physicochemical properties and biological activity of cherry silverberry fruit. It also revealed the potential application of fermented cherry silverberry in the production of food materials beneficial for health. © 2020 Society of Chemical Industry.

Solid state fermentation process with Aspergillus kawachii enhances the cancer-suppressive potential of silkworm larva in hepatocellular carcinoma cells.

BACKGROUND: Mulberry silkworm larvae (Bombyx mori) are known as the oldest resource of food and traditional medicine. Although silkworm larvae have been reported to treat various chronic diseases, the effect of fermentation by microorganisms improving the biological activities of silkworm larvae was not reported. In the present study, fermented silkworm larvae was developed via solid-state fermentation with Aspergillus kawachii and investigated its anti-cancer activity in human hepatocellular carcinoma cells. METHODS: We investigated the anti-cancer effects of unfermented (SEE) and fermented silkworm larva ethanol extract (FSEE) on HepG2 human hepatocellular carcinoma cells as well as compared changes in free amino acid, fatty acid, and mineral contents. Anti-cancer activities were evaluated by SRB staining, cell cycle analysis, Annexin V staining, Hoechst staining, DNA fragmentation analysis and western blot analysis. Fatty acid, free amino acid and mineral contents of SEE and FSEE were determined by gas chromatography, amino acid analyzer and flame atomic absorption spectrophotometer, respectively. RESULTS: Compared with SEE, treatment with FSEE resulted in apoptotic cell death in HepG2 cells characterized by G0/G1 phase cell cycle arrest, DNA fragmentation, and formation of apoptotic bodies. Furthermore, FSEE significantly up-regulated pro-apoptotic as well as down-regulated anti-apoptotic proteins in HepG2 cells. However, an equivalent concentration of SEE did not induce cell cycle arrest or apoptosis in HepG2 cells. Moreover, fermentation process by Aspergillus kawachii resulted in enhancement of fatty acid contents in silkworm larvae, whereas amino acid and mineral contents were decreased. CONCLUSION: Collectively, this study demonstrates that silkworm larvae solid state-fermented by Aspergillus kawachii strongly potentiates caspase-dependent and -independent apoptosis pathways in human hepatocellular carcinoma cells by regulating secondary metabolites.

Kochia scoparia seed extract suppresses VEGF-induced angiogenesis via modulating VEGF receptor 2 and PI3K/AKT/mTOR pathways.

Context: Kochia scoparia (L.) Schrad (Amaranthaceae), known as a traditional medicine in China, Japan and Korea, is reported to have various biological activities. However, K. scoparia seed extract (KSE) functional roles on angiogenesis and prostate cancer inhibition have not been elucidated. Objective: This study elucidates the effects of KSE on vascular endothelial growth factor (VEGF)-induced angiogenesis in human umbilical vein endothelial cells (HUVECs) and inhibition of proliferation in prostate cancer cells. Materials and methods: HUVECs were treated with 10-20 µg/mL of KSE and 20-50 ng/mL of VEGF for 12-72 h. Anti-angiogenesis properties of KSE were determined by wound healing, trans-well, tube formation, rat aortic ring assay and western blotting. Prostate cancer and normal cells were incubated with 10-250 µg/mL of KSE for 24 h, and cell viability was measured by SRB assay. Phenolic compounds in KSE were analyzed using a HPLC-PDA system. Results: IC50 for cell viability of HUVECs, LNCaP, PC-3, RC-58T and RWPE-1 by KSE were 30.64, 89.25, 123.41, 141.62 and >250 µg/mL, respectively. Treatment with KSE (20 µg/mL) significantly suppressed VEGF-induced migration, invasion and capillary-like structure formation of HUVECs and microvessel sprouting from rat aortic rings. In addition, KSE down-regulated PI3K/AKT/mTOR levels and phosphorylation of VEGF receptor 2 in HUVECs. 3-OH-tyrosol (1.63 mg/g) and morin hydrate (0.17 mg/g) were identified in KSE. Conclusions: KSE inhibits angiogenesis in HUVECs as well as proliferation in human prostate cancer cells, suggesting KSE may be useful herbal medicine for preventing progression of prostate cancer and angiogenesis.

Scopoletin Supplementation Ameliorates Steatosis and Inflammation in Diabetic Mice.

Scopoletin is a bioactive component in many edible plants and fruits. This study investigated the effects of scopoletin on hepatic steatosis and inflammation in a high-fat diet fed type 1 diabetic mice by comparison with metformin. Scopoletin (0.01%, w/w) or metformin (0.5%, w/w) was provided with a high-fat diet to streptozotocin-induced diabetic mice for 11 weeks. Both scopoletin and metformin lowered blood glucose and HbA1c , serum ALT, TNF-α and IL-6 levels, glucose intolerance, and hepatic lipid accumulation compared with the diabetic control group. Scopoletin or metformin down-regulated hepatic gene expression of triglyceride (Pparg, Plpp2, and Dgat2) and cholesterol (Hmgcr) synthesis as well as inflammation (Tlr4, Myd88, Nfkb1, Tnfa, and Il6), while it up-regulated Cyp7a1 gene. Hepatic PPARγ and DGAT2 protein levels were also down-regulated in scopoletin or metformin group compared with the control group. Scopoletin or metformin also inhibited hepatic fatty acid synthase and phosphatidate phosphohydrolase activities. These results suggest that scopoletin protects against diabetes-induced steatosis and inflammation by inhibiting lipid biosynthesis and TLR4-MyD88 pathways. Copyright © 2017 John Wiley & Sons, Ltd.

Anti-adipogenic and anti-diabetic effects of cis-3',4'-diisovalerylkhellactone isolated from Peucedanum japonicum Thunb leaves in vitro.

Peucedanum japonicum Thunb is a medicinal plant belonging to the family Umbelliferae. This study evaluated the anti-diabetic and anti-obesity effects of cis-3',4'-diisovalerylkhellactone (cDIVK) isolated from Peucedanum japonicum Thunb leaves. cDIVK (30 and 50µM) effectively inhibited adipocyte differentiation and fat accumulation, whereas it stimulated glucose uptake compared with the control in 3T3-L1 cells. cDIVK significantly increased AMPK activation and suppressed protein and mRNA expression of major adipogenic transcriptional factors such as C/EBPα, PPARγ and SREBP-1c in 3T3-L1 cells. In addition, cDIVK had potential α-glucosidase inhibitory activity. These results indicated that cDIVK may act as a natural dual therapeutic agent for diabetes and obesity.

Production of novel vinegar having antioxidant and anti-fatigue activities from Salicornia herbacea L.

BACKGROUND: Salicornia herbacea L. is a halophyte that grows in salt marshes and contains significant amounts of salts and minerals. Because it is known as a folk medication to treat diseases, various processed products such as powder, globular type of powder, laver and extract have been developed. However, it is difficult to process as a drink because of its high salinity. In the present study, glasswort vinegar (GV) containing high amounts of organic acids and minerals was developed via two-step fermentation with unpolished rice substrates and investigated its antioxidant and anti-fatigue activities. RESULTS: GV showed various free radical scavenging effects, reducing power, oxidized-LDL inhibition and superoxide dismutase-like activities. Compared with the control group (orally administered 7 g kg(-1) distilled water), the GV supplementation group showed increased running endurance and had higher glycogen accumulation in liver and muscles of rats exhausted by exercise. Furthermore, the GV-administered group demonstrated significantly elevated lactate and ATP metabolism, promoting enzyme activities such as muscle creatine kinase and lactate dehydrogenase, whereas serum fatigue biomarkers such as ammonia, lactate and inorganic acid were markedly decreased. CONCLUSION: These results indicate that GV can be used as a functional food for the development of a dietary beverage to alleviate fatigue.

Sensitization of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-resistant primary prostate cancer cells by isoegomaketone from Perilla frutescens.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is currently in clinical trials as a cancer treatment due to its ability to induce apoptosis selectively in cancer cells. Nevertheless, the risk of developing resistance warrants the development of sensitizers that can overcome resistance to TRAIL. In this study, isoegomaketone (1) acted as a synergistic TRAIL sensitizer by mediating up-regulation of DR5 expression in primary prostate cancer RC-58T/h/SA#4 cells. Combined with 1, TRAIL exhibited enhanced apoptotic activity in a human prostate cancer cell line designated RC-58T/h/SA#4, as indicated by increases in annexin V-positive and sub-G1 cell populations as well as condensation of chromatin or apoptotic bodies. Combined treatment also activated caspases-8, -9, and -3; increased the protein levels of Bax, AIF, and cytosolic cytochrome c; and induced PARP cleavage while reducing Bcl-2 protein expression. Human recombinant DR5 Fc chimera efficiently attenuated 1-induced apoptosis, thereby demonstrating the critical role of DR5 in 1-mediated apoptotic cell death. Furthermore, DR5 expression induced by 1 was mediated via a ROS-independent pathway that required CHOP and p53. Overall, these findings provide evidence that 1 potentiates TRAIL-mediated apoptosis through up-regulation of DR5 via a ROS-independent pathway. This suggests that 1 has potential for increasing the effectiveness of prostate cancer therapy with TRAIL.

Induction of apoptosis by 2,3-dehydrosilybin via a caspase-dependent pathway in human HeLa cells.

The aim of this study was to investigate the mechanisms involved in the apoptosis of HeLa cells due to 2,3-dehydrosilybin (DHS) treatment. DHS treatment over 24 h significantly inhibited cell viability and induced apoptosis in a dose-dependent manner. It also triggered the cleavage of caspase-8, caspase-9, caspase-3, and PARP, and significantly increased caspase-3 activity in a dose-dependent manner. Moreover, it triggered the depolarization of the mitochondrial membrane potential (Δψm), the release of cytochrome c into the cytosol, the cleavage of Bid, and the downregulation of Bcl-2 in a dose-dependent manner. Furthermore, z-VAD-fmk (a pan-caspase inhibitor) and z-IETD-fmk (a specific caspase-8 inhibitor) abolished the DHS-induced activation of the caspase-8, -9, and -3, cleavage of PARP, the depolarization of Δψm, the release of cytochrome c, the cleavage of Bid, and the downregulation of Bcl-2. Taken together, these results suggest that DHS-induced apoptosis is mediated by a caspase-dependent pathway in human HeLa cells.

Antioxidant activity and blood alcohol concentration lowering effect of fermented Hovenia dulcis fruit vinegar.

In this study, Hovenia dulcis fruit fermented vinegar (HFV) was produced by the two-step fermentation of the H. dulcis fruit. The bioactivities before and after fermentation were compared. During the two-stage fermentation, the highest total acidity (4.99%) in the H. dulcis fruit extract juice was determined to be 16°Bx. During fermentation, the acetic acid content increased from 54.45 to 5404.30 mg%, and the fructose level in the HFV decreased from 130.68 to 54.91 mg%. The levels of DPPH and ABTS·+ free radicals scavenging activities, reducing power, hydrogen peroxide scavenging and ß-carotene bleaching activities were found to be increased in HFV as compared to before fermentation. Furthermore, the serum alcohol and acetaldehyde levels were reduced significantly in HFV compared to before fermentation. This study shows that HFV enhances the antioxidant and alcohol degradation activities and can potentially be used as a functional drink to prevent hangovers.

Celastrol inhibits growth and induces apoptotic cell death in melanoma cells via the activation ROS-dependent mitochondrial pathway and the suppression of PI3K/AKT signaling.

Celastrol has been reported to possess anticancer effects in various cancers; however, the precise mechanism underlying ROS-mediated mitochondria-dependent apoptotic cell death triggered by celastrol treatment in melanoma cells remains unknown. We showed that celastrol effectively induced apoptotic cell death and inhibited tumor growth using tissue culture and in vivo models of B16 melanoma. In addition to apoptotic cell death in B16 cells, several apoptotic events such as PARP cleavage and activation of caspase were confirmed. Pretreatment with caspase inhibitor modestly attenuated the celastrol-induced increase in PARP cleavage and sub-G1 cell population, implying that caspases play a partial role in celastrol-induced apoptosis. Moreover, ROS generation was detected following celastrol treatment. Blocking of ROS accumulation with ROS scavengers resulted in inhibition of celastrol-induced Bcl-2 family-mediated apoptosis, indicating that celastrol-induced apoptosis involves ROS generation as well as an increase in the Bax/Bcl-2 ratio leading to release of cytochrome c and AIF. Importantly, silencing of AIF by transfection of siAIF into cells remarkably attenuated celastrol-induced apoptotic cell death. Moreover, celastrol inhibited the activation of PI3K/AKT/mTOR signaling cascade in B16 cells. Our data reveal that celastrol inhibits growth and induces apoptosis in melanoma cells via the activation of ROS-mediated caspase-dependent and -independent pathways and the suppression of PI3K/AKT signaling.

Isoegomaketone induces apoptosis through caspase-dependent and caspase-independent pathways in human DLD1 cells.

Isoegomaketone (IK) is an essential oil component of Perilla frutescens (L.), but the mechanism by which IK induces apoptosis has never been studied. The purpose of this study was to elucidate the IK-induced apoptotic pathway in DLD1 human colon cancer cells. We observed that IK treatment over 24 h significantly inhibited cell viability in a dose-dependent manner. We also found that IK triggered cleavage of PARP. Moreover, IK treatment resulted in cleavage of caspase-8, -9, and -3 in a dose- and time-dependent manner. IK treatment also resulted in cleavage of Bid and translocation of Bax, and triggered the release of cytochrome c from the mitochondria to the cytoplasm. Furthermore, it resulted in the translocation of apoptosis inducing factor (AIF), a caspase-independent mitochondrial apoptosis factor, from the mitochondria into the nucleus. Overall, these results suggest that IK induces apoptosis through caspase-dependent and capase-independent pathways in DLD1 cells.

Apoptotic action of ursolic acid isolated from Corni fructus in RC-58T/h/SA#4 primary human prostate cancer cells.

Ursolic acid (3ß-hydroxy-urs-12-en-28-oic acid) is a major biological active component of Corni fructus that is known to induce apoptosis. However, the apoptotic mechanism of ursolic acid using primary malignant tumor (RC-58T/h/SA#4)-derived human prostate cells is not known. In the present study, ursolic acid significantly inhibited the growth of RC-58T/h/SA#4 cells in dose- and time-dependent manners. Ursolic acid induced cell death as evidenced by an increased proportion of cells in sub-G1 phase, the formation of apoptotic bodies, nuclear condensation, and DNA fragmentation. After ursolic acid treatment at concentrations above 40 µM, the activities of caspase-3, -8, and -9 were significantly increased compared that of control. Ursolic acid modulated the upregulation of Bax (pro-apoptotic) as well as the downregulation of Bcl-2 (anti-apoptotic). Ursolic acid also stimulated Bid cleavage, which indicates that the apoptotic action of caspase-8-mediated Bid cleavage leads to the activation of caspase-9. Thus, the apoptotic effect of ursolic acid was involved in extrinsic and intrinsic signaling pathways. In addition, ursolic acid increased the expression of the caspase-independent mitochondrial apoptosis factor (AIF) in RC-58T/h/SA#4 cells. The present results suggest that ursolic acid from Corni fructus activated apoptosis in RC-58T/h/SA#4 cells via both caspase-dependent and -independent pathways.

Sanggenol L Induces Apoptosis and Cell Cycle Arrest via Activation of p53 and Suppression of PI3K/Akt/mTOR Signaling in Human Prostate Cancer Cells.

Prostate cancer is the most common cancer in Western countries. Recently, Asian countries are being affected by Western habits, which have had an important role in the rapid increase in cancer incidence. Sanggenol L (San L) is a natural flavonoid present in the root barks of Morus alba, which induces anti-cancer activities in ovarian cancer cells. However, the molecular and cellular mechanisms of the effects of sanggenol L on human prostate cancer cells have not been elucidated. In this study, we investigated whether sanggenol L exerts anti-cancer activity in human prostate cancer cells via apoptosis and cell cycle arrest. Sanggenol L induced caspase-dependent apoptosis (up-regulation of PARP and Bax or down-regulation of procaspase-3, -8, -9, Bid, and Bcl-2), induction of caspase-independent apoptosis (up-regulation of AIF and Endo G on cytosol), suppression of cell cycle (down-regulation of CDK1/2, CDK4, CDK6, cyclin D1, cyclin E, cyclin A, and cyclin B1 or up-regulation of p53 and p21), and inhibition of PI3K/Akt/mTOR signaling (down-regulation of PI3K, p-Akt, and p-mTOR) in prostate cancer cells. These results suggest the induction of apoptosis via suppression of PI3K/Akt/mTOR signaling and cell cycle arrest via activation of p53 in response to sanggenol L in prostate cancer cells.

Sanggenol L promotes apoptotic cell death in melanoma skin cancer cells through activation of caspase cascades and apoptosis-inducing factor.

Sanggenol L is one component of root bark of Morus alba. The molecular and cellular mechanisms of sanggenol L effects on melanoma cells are not well known. Recently, melanoma is the most common skin cancer with a high mortality rate not only in United States, but also in East Asia. Therefore, safe and effective treatments for melanoma treatment are required. In this study, we investigated whether or not sanggenol L possesses anti-cancer activity in human and mouse melanoma skin cancer cells. Sanggenol L treatment exerted significant cell growth inhibitory effects and inhibited colony formation capacity against B16, SK-MEL-2, and SK-MEL-28 melanoma skin cancer cells, whereas HaCaT human epithelial keratinocyte cells was unaffected by sanggenol L treatment. Sanggenol L treatment resulted in apoptotic cell death in melanoma skin cancer cells, which was characterized by accumulation of apoptotic cells, nuclear condensation, and apoptotic bodies. We also showed that sanggenol L treatment induced caspase-dependent apoptosis (up-regulation of Bax and cleaved-PARP or down-regulation of Bid, Bcl-2, procaspse-3, -8, and -9), induction of caspase-independent apoptosis (up-regulation of AIF and Endo G on cytosol) in melanoma skin cancer cells. These results suggest that sanggenol L induces caspase-dependent and -independent apoptosis in melanoma skin cancer cells.

Lupiwighteone induces caspase-dependent and -independent apoptosis on human breast cancer cells via inhibiting PI3K/Akt/mTOR pathway.

Breast cancer is one of the most common causes of mortality in women. Lupiwighteone has anticancer effects in prostate cancer cells and neuroblastoma cells. However, the molecular and cellular mechanisms of lupiwighteone effects on human breast cancer cells are not as well known. In the present study, we investigated the effects of lupiwighteone on the proliferation and apoptosis of two different human cancer cells; MCF-7, an estrogen receptor (ER)-positive human breast cancer cell, and MDA-MB-231, a triple negative human breast cancer cell. Lupiwighteone treatment decreased the viability of MCF-7 and MDA-MB-231 cells. Lupiwighteone treatment resulted in apoptotic cell death in breast cancer cells, which was characterized by DNA fragmentation, accumulation of apoptotic cells, and nuclear condensation. We also showed that treatment with lupiwighteone induced caspase-dependent apoptosis (up-regulation of caspase-3, -7, -8, -9, PARP, and Bax or down-regulation of Bid, Bcl-2), induction of caspase-independent apoptosis (up-regulation of AIF and Endo G on cytosol), and inhibition of the PI3K/Akt/mTOR signaling pathway (down-regulation of PI3K, p-Akt, and p-mTOR) in both MCF-7 and MDA-MB-231 cells. These results suggest that lupiwighteone induces caspase-dependent and -independent apoptosis in both breast cancer cell lines via inhibiting PI3K/Akt/mTOR pathway.

Protective Effect of Prunus mume Fermented with Mixed Lactic Acid Bacteria in Dextran Sodium Sulfate-Induced Colitis.

The fruit of Prunus mume (PM) is widely cultivated in East Asia, and it has been used as a folk medication for gastrointestinal disorders, e.g., diarrhea, stomach ache and ulceration. In this study, the pectinase-treated PM juice (PJ) was fermented with Lactobacillus strains containing fundamental organic acids and free amino acids. The PJ fermented with Lactobacillus plantarum and L. casei (FP) was investigated for its protective effect in dextran sodium sulfate (DSS)-induced colitis mice model. The administration of FP reduced lipid peroxidation and histopathological colitis symptoms, e.g., shortening of the colon length, depletion of mucin, epithelial injury and ulceration, in colonic tissues. The FP-supplemented group showed the alleviation of pro-inflammatory cytokines. Compared with the DSS control group, the supplementation of FP significantly reduced the levels of serum interferon-γ (IFN-γ), interleukin (IL)-1ß, IL-6, IL-12 and IL-17 as well as colonic tumor necrosis factor-α, IFN-γ, IL-12 and IL-17. Furthermore, the DSS-induced TUNEL-positive area was significantly reduced by the FP supplementation. These results show that the supplementation of FP fermented with mixed lactic acid bacteria, L. plantarum and L. casei, elucidated the protective effect in DSS-induced colitis mice. Hence, this study suggests that FP can be utilized as a natural therapeutic agent for colitis and intestinal inflammation.

Anti-Fatigue Effect of Prunus Mume Vinegar in High-Intensity Exercised Rats.

Nowadays, new types of vinegar have been developed using various raw materials and biotechnological processes. The fruit of Prunus mume has been extensively distributed in East Asia and used as a folk medication for fatigue. In this study, the Prunus mume vinegar (PV) was produced by a two-step fermentation and was evaluated for its anti-fatigue activity by C2C12 myoblasts and high-intensity exercised rats. The administration of PV significantly improved running endurance and glycogen accumulation in the liver and muscle of PV supplemented rats compared to sedentary and exercised control groups. In addition, PV supplementation elicited lower fatigue-related serum biomarkers, for instance, ammonia, inorganic phosphate, and lactate. PV administered rats exhibited higher lactate dehydrogenase activity and glutathione peroxidase activity, and lower creatine kinase activity and malondialdehyde levels. Furthermore, phenolic compounds in PV were identified using HPLC analysis. The phenolic acids analyzed in PV were protocatechuic acid, syringic acid, chlorogenic acid, and its derivates. These results indicate that the administration of PV with antioxidative property contributes to the improvement of fatigue recovery in exhausted rats. The findings of this study suggest that the PV containing various bioactive constituents can be used as a functional material against fatigue caused by high-intensity exercise.

Cultivated Orostachys japonicus extract inhibits VEGF-induced angiogenesis via regulation of VEGFR2 signaling pathway in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Orostachys japonicus A. Berger (O. japonicus), so-called Wa-song in Korea, a traditional food and medicine that grows on mountain rocks and roof tiles. Wa-song containing various phenolic compounds have been reported as a medicinal plant for prevention of fibrosis, cancer, inflammation, and oxidative damage. AIM OF THE STUDY: The present study was designed to examine the anti-angiogenic effects of cultivated Orostachys japonicus 70% ethanol extract (CE) in vascular endothelial growth factor (VEGF)-stimulated human umbilical vein endothelial cells (HUVECs). MATERIALS AND METHODS: CE was prepared with 70% ethanol. HUVECs, rat aortic rings, and matrigel plug in mice were treated with CE (10-20 µg/mL) and VEGF (20-50 ng/mL), and the anti-angiogenic activities of CE were analyzed by SRB, wound healing, trans-well invasion, capillary-like tubule formation, rat aortas, Western blot, and matrigel plug assay. Phenolic compounds in CE were analyzed using a high-performance liquid chromatography (HPLC)-PDA system. RESULTS: Treatment of CE (10-20 µg/mL) markedly suppressed proliferation of HUVECs in the presence (from 136.5% to 112.2%) or absence of VEGF (from 100.0% to 92.1%). The proliferation inhibitory effect of CE was caused by G0/G1 cell cycle arrest, and the decrease of CDK-2, CDK-4, Cyclin D1 and Cyclin E1. Furthermore, CE treatment showed significant angiogenesis inhibitory effects on motility, invasion and micro-vessel formation of HUVECs, rat aortic rings and subcutaneous matrigels under VEGF-stimulation condition. In HUVECs, CE-induced anti-angiogenic effect was regulated by inhibition of the PI3K/AKT/mTOR, MAPK/p38, MAPK/ERK, FAK-Src, and VEGF-VEGFR2 signaling pathways. CONCLUSION: This study demonstrated that CE might be used as a potential natural substance, multi-targeted angiogenesis inhibitor, functional food material.

The Flavonol Isoquercitrin Promotes Mitochondrial-Dependent Apoptosis in SK-Mel-2 Melanoma Cell via the PI3K/AKT/mTOR Pathway.

Isoquercitrin (IQ), a major flavonol present in Prunus mume fruit, has gained much attention in recent studies because of its superior bioavailability and physiological effects. In this study, the anti-cancer mechanism of IQ against human melanoma, particularly its effect on the mitochondria-mediated apoptosis, was investigated. Treatment with IQ at 25 µM concentration effectively inhibited the proliferation of SK-MEL-2 skin cancer cells while the same concentration did not exhibit cytotoxicity against human keratinocytes HaCaT. Morphological analysis and clonogenic assay also showed that IQ can alter the growth and long-term survival of SK-MEL-2 cells. IQ also induced apoptosis in the melanoma cells as manifested in the nuclear morphology changes, DNA fragmentation, increase in the apoptosis rate (17.69% at 25 µM) and accumulation of sub-G1 cell cycle phase population (19.55% at 25 µM). Western blot analysis revealed the involvement of the mitochondrial apoptosis signaling pathway in the anti-cancer property of IQ. Treatment with IQ resulted in the decrease in the levels of procaspase-8 and -9, and Bcl-2 protein, and an increase in the expression of cleaved PARP and Bax. Moreover, AIF and Endo G protein expression increased, indicating a caspase-independent mitochondrial-mediated apoptosis. The anti-proliferative activity of IQ against SK-MEL-2 can also be attributed to the downregulation of the PI3K/AktmTOR signaling pathway. These findings showed that IQ can be developed into a chemopreventive therapeutic agent against the melanoma cells.

Ursolic acid enhances the cellular immune system and pancreatic beta-cell function in streptozotocin-induced diabetic mice fed a high-fat diet.

This study investigated the effects of ursolic acid on immunoregulation and pancreatic beta-cell function in type 1 diabetes fed a high-fat diet for 4 weeks. Male mice were divided into non-diabetic, diabetic control, and diabetic-ursolic acid (0.05%, w/w) groups, which were fed a high-fat (37% calories from fat). Diabetes was induced by injection of streptozotocin (200 mg/kg B.W., i.p.). Ursolic acid significantly improved blood glucose levels, glucose intolerance, and insulin sensitivity compared to the diabetic group. The plasma insulin and C-peptide concentrations were significantly higher in the diabetic-ursolic acid group than in the diabetic group. Ursolic acid significantly elevated the insulin levels with preservation of insulin staining of beta-cells in the pancreas. In splenocytes, concanavalin (Con) A-induced T-cell proliferation was significantly higher in the diabetic-ursolic acid group compared to the diabetic group, but liposaccharide (LPS)-induced B-cell proliferation did not differ between groups. Ursolic acid enhanced IL-2 and IFN-gamma production in response to Con A stimulation, whereas it inhibited TNF-alpha production in response to LPS stimulation. In this study, neither streptozotocin nor ursolic acid had effects on lymphocyte subsets. These results indicate that ursolic acid exhibits potential anti-diabetic and immunomodulatory properties by increasing insulin levels with preservation of pancreatic beta-cells and modulating blood glucose levels, T-cell proliferation and cytokines production by lymphocytes in type 1 diabetic mice fed a high-fat diet.