Dichloromethane fractions of Calystegia soldanella induce S­phase arrest and apoptosis in HT­29 human colorectal cancer cells.

Calystegia soldanella is a halophyte and a perennial herb that grows on coastal sand dunes worldwide. Extracts from this plant have been previously revealed to have a variety of bioactive properties in humans. However, their effects on colorectal cancer cells remain poorly understood. In the present study, the potential biological activity of C. soldanella extracts in the colorectal cancer cell line HT­29 was examined. First, five solvent fractions [n­hexane, dichloromethane (DCM), ethyl acetate, n­butanol and water] were obtained from the crude extracts of C. soldanella through an organic solvent extraction method. In particular, the DCM fraction was demonstrated to exert marked dose­ and time­dependent inhibitory effects according to results from the cell viability assay. Data obtained from the apoptosis assay suggested that the inhibition of HT­29 cell viability induced by DCM treatment was attributed to increased apoptosis. The apoptotic rate was markedly increased in a dose­dependent manner, which was associated with the protein expression levels of apoptosis­related proteins, including increased Fas, Bad and Bax, and decreased pro­caspase­8, Bcl­2, Bcl­xL, pro­caspase­9, pro­caspase­7 and pro­caspase­3. A mitochondrial membrane potential assay demonstrated that more cells became depolarized and the extent of cytochrome c release was markedly increased in a dose­dependent manner in HT­29 cells treated with DCM. In addition, cell cycle analysis confirmed S­phase arrest following DCM fraction treatment, which was associated with decreased protein expression levels of cell cycle­related proteins, such as cyclin A, CDK2, cell division cycle 25 A and cyclin dependent kinase inhibitor 1. Based on these results, the present study suggested that the DCM fraction of the C. soldanella extract can inhibit HT­29 cell viability whilst inducing apoptosis through mitochondrial membrane potential regulation and S­phase arrest. These results also suggested that the DCM fraction has potential anticancer activity in HT­29 colorectal cells. Further research on the composition of the DCM fraction is warranted.

Calystegia soldanella Extract Exerts Anti-Oxidative and Anti-Inflammatory Effects via the Regulation of the NF-κB/Nrf-2 Pathways in Mouse Macrophages.

Plant polyphenols are widely used to treat various inflammatory diseases, owing to their ability to suppress reactive oxygen species production and the expression of inflammatory cytokines. Herein, we investigated phenolic compounds from Calystegia soldanella using UPLC Q-TOF MS/MS and their antioxidative and anti-inflammatory activities were analyzed. The C. soldanella ethyl acetate fraction (CsEF) had the strongest antioxidative activity, given its high polyphenol compound content. It also exhibited anti-inflammatory effects, inhibiting the production of inflammatory cytokines such as NO, PGE2, IL-1ß, IL-6, and TNF-α in LPS-stimulated mouse macrophages. CsEF activated the nuclear transcription factor Nrf-2, thereby upregulating antioxidant enzymes such as HO-1 and NQO-1 and inhibiting NF-κB expression, which in turn, suppressed the expression of COX-2, iNOS, and inflammatory cytokines, ultimately exerting anti-inflammatory effects. Further, UPLC-Q-TOF-MS/MS was used to analyze the polyphenol compound contents in CsEF. The quercetin glycosides isoquercitrin and quercitrin were the primary flavonoid compounds, while the caffeic acid derivatives, chlorogenic acid and dicaffeoylquinic acid, were the primary phenolic acids. Thus, C. soldanella, which had only a limited use thus far as a medicinal plant, may serve as a natural medicinal resource for treating inflammatory diseases.

The protective effects of ethanolic extract of Clematis terniflora against corticosterone-induced neuronal damage via the AKT and ERK1/2 pathway.

Chronic stress induces neuronal cell death, which can cause nervous system disorders including Parkinson's disease and Alzheimer's disease. In this study, we evaluated the neuroprotective effects of Clematis terniflora extract (CTE) against corticosterone-induced apoptosis in rat pheochromocytoma (PC12) cells, and also investigated the underlying molecular mechanisms. At concentrations of 300 and 500 µg/ml, CTE significantly decreased apoptotic cell death and mitochondrial damage induced by 200 µM corticosterone. CTE decreased the expression levels of endoplasmic reticulum (ER) stress proteins GRP78, GADD153, and mitochondrial damage-related protein BAD, suggesting that it downregulates ER stress evoked by corticosterone. Furthermore, our results suggested that these protective effects were mediated by the upregulation of p-AKT and p-ERK1/2, which are involved in cell survival signaling. Collectively, our results indicate that CTE can lessen neural damage caused by chronic stress. [BMB Reports 2018; 51(8): 400-405].

Pretreated Glehnia littoralis Extract Prevents Neuronal Death Following Transient Global Cerebral Ischemia through Increases of Superoxide Dismutase 1 and Brain-derived Neurotrophic Factor Expressions in the Gerbil Hippocampal Cornu Ammonis 1 Area.

BACKGROUND: Glehnia littoralis, as a traditional herbal medicine to heal various health ailments in East Asia, displays various therapeutic properties including antioxidant effects. However, neuroprotective effects of G. littoralis against cerebral ischemic insults have not yet been addressed. Therefore, in this study, we first examined its neuroprotective effects in the hippocampus using a gerbil model of transient global cerebral ischemia (TGCI). METHODS: Gerbils were subjected to TGCI for 5 min. G. littoralis extract (GLE; 100 and 200 mg/kg) was administrated orally once daily for 7 days before ischemic surgery. Neuroprotection was examined by neuronal nuclear antigen immunohistochemistry and Fluoro-Jade B histofluorescence staining. Gliosis was observed by immunohistochemistry for glial fibrillary acidic protein and ionized calcium-binding adapter molecule 1. For neuroprotective mechanisms, immunohistochemistry for superoxide dismutase (SOD) 1 and brain-derived neurotrophic factor (BDNF) was done. RESULTS: Pretreatment with 200 mg/kg of GLE protected pyramidal neurons in the cornu ammonis 1 (CA1) area from ischemic insult area (F = 29.770, P < 0.05) and significantly inhibited activations of astrocytes (F = 22.959, P < 0.05) and microglia (F = 44.135, P < 0.05) in the ischemic CA1 area. In addition, pretreatment with GLE significantly increased expressions of SOD1 (F = 28.561, P < 0.05) and BDNF (F = 55.298, P < 0.05) in CA1 pyramidal neurons of the sham- and ischemia-operated groups. CONCLUSIONS: Our findings indicate that pretreatment with GLE can protect neurons from ischemic insults, and we suggest that its neuroprotective mechanism may be closely associated with increases of SOD1 and BDNF expressions as well as attenuation of glial activation.

Pre­treatment with Chrysanthemum indicum Linné extract protects pyramidal neurons from transient cerebral ischemia via increasing antioxidants in the gerbil hippocampal CA1 region.

Chrysanthemum indicum Linné extract (CIL) is used in herbal medicine in East Asia. In the present study, gerbils were orally pre­treated with CIL, and changes of antioxidant enzymes including superoxide dismutase (SOD) 1 and SOD2, catalase (CAT) and glutathione peroxidase (GPX) in the hippocampal CA1 region following 5 min of transient cerebral ischemia were investigated and the neuroprotective effect of CIL in the ischemic CA1 region was examined. SOD1, SOD2, CAT and GPX immunoreactivities were observed in the pyramidal cells of the CA1 region and their immunoreactivities were gradually decreased following ischemia­reperfusion and barely detectable at 5 days post­ischemia. CIL pre­treatment significantly increased immunoreactivities of SOD1, CAT and GPX, but not SOD2, in the CA1 pyramidal cells of the sham­operated animals. In addition, SOD1, SOD2, CAT and GPX immunoreactivities in the CA1 pyramidal cells were significantly higher compared with the ischemia­operated animals. Furthermore, it was identified that pre­treatment with CIL protected the CA1 pyramidal cells in the CA1 region using neuronal nuclei immunohistochemistry and Fluoro­Jade B histofluorescence staining; the protected CA1 pyramidal cells were 67.5% compared with the sham­operated animals. In conclusion, oral CIL pre­treatment increased endogenous antioxidant enzymes in CA1 pyramidal cells in the gerbil hippocampus and protected the cells from transient cerebral ischemic insult. This finding suggested that CIL is promising for the prevention of ischemia­induced neuronal damage.

The effectiveness of postoperative intervention in patients after rhinoplasty: a meta-analysis.

Rhinoplasty is the most common facial plastic surgical procedure, and the occurrence of periorbital edema and ecchymosis is normal after rhinoplasty. The goal of this study was to perform a systematic review with meta-analysis of the efficacy of postoperative care of edema and ecchymosis following rhinoplasty. Two authors independently searched the databases (PubMed, SCOPUS, Embase, Web of Science, and the Cochrane database) from inception to September 2016. We included studies that compared postoperative care methods (intervention groups) with no treatment (control group) where the outcomes of interest were edema, ecchymosis, and satisfaction rate of patients on postoperative days. Sufficient data for meta-analysis were retrieved for 11 trials with a total of 627 patients. Eyelid edema and ecchymosis during the first 7 days postoperatively were statistically decreased in the arnica administration groups versus the control group. Eyelid edema and ecchymosis during the first 24 h postoperatively were statistically decreased in the cold compression group versus the control group. The ratio of patient satisfaction was statistically higher in the tapping application group than in the control group. However, the analysis indicated that surgeons had a significant tendency to decrease intranasal packing. The administration of arnica, cold compression, and tape could reduce eyelid edema and ecchymosis. Intranasal packing was associated with more adverse effects in terms of postoperative ecchymosis compared to non-packing. However, additional trials with thorough research methodologies should be conducted to confirm the results of this study.

Enzyme-treated Ecklonia cava extract inhibits adipogenesis through the downregulation of C/EBPα in 3T3-L1 adipocytes.

In this study, we examined the inhibitory effects of enzyme- treated Ecklonia cava (EEc) extract on the adipogenesis of 3T3-L1 adipocytes. The components of Ecklonia cava (E. cava) were first separated and purified using the digestive enzymes pectinase (Rapidase® X­Press L) and cellulase (Rohament® CL). We found that the EEc extract contained three distinct phlorotannins: eckol, dieckol and phlorofucofuroeckol-A. Among the phlorotannins, dieckol was the most abundant in the EEc extract at 16 mg/g. Then we examined the inhibitory effects of EEc extract treatment on differentiation­related transcription factors and on adipogenesis­related gene expression in vitro using 3T3-L1 adipocytes. 3T3­L1 pre­adipocytes were used to determine the concentrations of the EEc extract and Garcinia cambogia (Gar) extract that did not result in cytotoxicity. Glucose utilization and triglyceride (TG) accumulation in the EEc­treated adipocytes were similarly inhibited by 50 µg/ml EEc and 200 µg/ml Gar, and these results were confirmed by Oil Red O staining. Protein expression of adipogenesis differentiation­related transcription factors following treatment with the EEc extract was also examined. Only the expression of CCAAT/enhancer­binding protein (C/EBP)α was decreased, while there was no effect on the expression of C/EBPß, C/EBPδ, and peroxisome proliferator­activated receptor Î³ (PPARγ). Treatment with the EEc extract decreased the expression levels of adipogenesis­related genes, in particular sterol regulatory element binding protein­1c (SREBP­1c), adipocyte fatty acid binding protein (A­FABP), fatty acid synthase (FAS) and adiponectin. These results suggest that EEc extract treatment has an inhibitory effect on adipogenesis, specifically by affecting the activation of the C/EBPα signaling pathway and the resulting adipogenesis-related gene expression.

Crude extract and solvent fractions of Calystegia soldanella induce G1 and S phase arrest of the cell cycle in HepG2 cells.

The representative halophyte Calystegia soldanella (L) Roem. et Schult is a perennial vine herb that grows in coastal dunes throughout South Korea as well as in other regions around the world. This plant has long been used as an edible and medicinal herb to cure rheumatic arthritis, sore throat, dropsy, and scurvy. Some studies have also shown that this plant species exhibits various biological activities. However, there are few studies on cytotoxicity induced by C. soldanella treatment in HepG2 human hepatocellular carcinoma cells. In this study, we investigated the viability of HepG2 cells following treatment with crude extracts and four solvent-partitioned fractions of C. soldanella. Of the crude extract and four solvent fractions tested, treatment with the 85% aqueous methanol (aq. MeOH) fraction resulted in the greatest inhibition of HepG2 cell proliferation. Flow cytometry showed that the 85% aq. MeOH fraction induced a G0/G1 and S phase arrest of the cell cycle progression. The 85% aq. MeOH fraction arrested HepG2 cells at the G0/G1 phase in a concentration-dependent manner, and resulted in decreased expression of cyclin D1, cyclin E, cyclin-dependent kinase (CDK)2, CDK4, CDK6, p21, and p27. Additionally, the 85% aq. MeOH fraction treatment also arrested HepG2 cells in the S phase, with decreased expression of cyclin A, CDK2, and CDC25A. Also, treatment with this fraction reduced the expression of retinoblastoma (RB) protein and the transcription factor E2F. These results suggest that the 85% aq. MeOH fraction exhibits potential anticancer activity in HepG2 cells by inducing G0/G1 and S phase arrest of the cell cycle.

Pre-treated Populus tomentiglandulosa extract inhibits neuronal loss and alleviates gliosis in the gerbil hippocampal CA1 area induced by transient global cerebral ischemia.

The genus Populus (poplar) belonging to the Salicaceae family has been used in traditional medicine, and its several species show various pharmacological properties including antioxidant and anti-inflammatory effects. No study regarding protective effects of Populus species against cerebral ischemia has been reported. Therefore, in the present study, we examined neuroprotective effects of ethanol extract from Populus tomentiglandulosa (Korea poplar) in the hippocampal cornu ammonis (CA1) area of gerbils subjected to 5 minutes of transient global cerebral ischemia. Pretreatment with 200 mg/kg of P. tomentiglandulosa extract effectively protected CA1 pyramidal neurons from transient global cerebral ischemia. In addition, glial fibrillary acidic protein immunoreactive astrocytes and ionized calcium binding adapter molecule 1 immunoreactive microglia were significantly diminished in the ischemic CA1 area by pretreatment with 200 mg/kg of P. tomentiglandulosa extract. Briefly, our results indicate that pretreatment with P. tomentiglandulosa extract protects neurons from transient cerebral ischemic injury and diminish cerebral ischemia-induced reactive gliosis in ischemic CA1 area. Based on these results, we suggest that P. tomentiglandulosa can be used as a potential candidate for prevention of ischemic injury.

Pyropia yezoensis glycoprotein regulates antioxidant status and prevents hepatotoxicity in a rat model of D-galactosamine/lipopolysaccharide-induced acute liver failure.

The present study aimed to investigate the effects of Pyropia yezoensis glycoprotein (PYGP) on hepatic antioxidative enzyme activity and mitogen-activated protein kinase (MAPK) phosphorylation in a rat model of D-galactosamine/lipopolysaccharide (D-GalN/LPS)-induced hepatotoxicity. Glutamic-oxaloacetic transaminase (GOT) and glutamic-pyruvic transaminase (GPT) were measured to determine the severity of hepatotoxicity. Treatment with D­GalN/LPS significantly increased the GOT, GPT and lipid peroxidation levels, and decreased the antioxidant capacity of the rats. Treatment with PYGP (150 and 300 mg/kg/body weight) decreased the levels of GOT, GPT and lipid peroxidation levels. The activities of antioxidative enzymes, including catalase, glutathione S­transferase and glutathione were upregulated following PYGP treatment. Furthermore, D­GalN/LPS­induced MAPK phosphorylation, and inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression were downregulated by PYGP. These results indicated that PYGP may exert hepatoprotective effects via the upregulation of antioxidative enzymes, and the downregulation of the MAPK signaling pathway and iNOS and COX-2 expression.

Oenanthe Javanica Extract Protects Against Experimentally Induced Ischemic Neuronal Damage via its Antioxidant Effects.

BACKGROUND: Water dropwort (Oenanthe javanica) as a popular traditional medicine in Asia shows various biological properties including antioxidant activity. In this study, we firstly examined the neuroprotective effect of Oenanthe javanica extract (OJE) in the hippocampal cornus ammonis 1 region (CA1 region) of the gerbil subjected to transient cerebral ischemia. METHODS: Gerbils were established by the occlusion of common carotid arteries for 5 min. The neuroprotective effect of OJE was estimated by cresyl violet staining. In addition, 4 antioxidants (copper, zinc superoxide dismutase [SOD], manganese SOD, catalase, and glutathione peroxidase) immunoreactivities were investigated by immunohistochemistry. RESULTS: Pyramidal neurons in the CA1 region showed neuronal death at 5 days postischemia; at this point in time, all antioxidants immunoreactivities disappeared in CA1 pyramidal neurons and showed in many nonpyramidal cells. Treatment with 200 mg/kg, not 100 mg/kg, OJE protected CA1 pyramidal neurons from ischemic damage. In addition, 200 mg/kg OJE treatment increased or maintained antioxidants immunoreactivities. Especially, among the antioxidants, glutathione peroxidase immunoreactivity was effectively increased in the CA1 pyramidal neurons of the OJE-treated sham-operated and ischemia-operated groups. CONCLUSION: Our present results indicate that treatment with OJE can protect neurons from transient ischemic damage and that the neuroprotective effect may be closely associated with increased or maintained intracellular antioxidant enzymes by OJE.

Effect of Oenanthe Javanica Extract on Antioxidant Enzyme in the Rat Liver.

BACKGROUND: Oenanthe javanica (O. javanica) has been known to have high antioxidant properties via scavenging reactive oxygen species. We examined the effect of O. javanica extract (OJE) on antioxidant enzymes in the rat liver. METHODS: We examined the effect of the OJE on copper, zinc-superoxide dismutase (SOD1), manganese superoxide dismutase (SOD2), catalase (CAT), and glutathione peroxidase (GPx) in the rat liver using immunohistochemistry and western blot analysis. Sprague-Dawley rats were randomly assigned to three groups; (1) normal diet fed group (normal-group), (2) diet containing ascorbic acid (AA)-fed group (AA-group) as a positive control, (3) diet containing OJE-fed group (OJE-group). RESULTS: In this study, no histopathological finding in the rat liver was found in all the experimental groups. Numbers of SOD1, SOD2, CAT, and GPx immunoreactive cells and their protein levels were significantly increased in the AA-fed group compared with those in the normal-group. On the other hand, in the OJE-group, numbers of SOD1, SOD2, CAT, and GPx immunoreactive cells in the liver were significantly increased by about 190%, 478%, 685%, and 346%, respectively, compared with those in the AA-group. In addition, protein levels of SOD1, SOD2, CAT, and GPx in the OJE-group were also significantly much higher than those in the AA-group. CONCLUSION: OJE significantly increased expressions of SOD1 and SOD2, CAT, and GPx in the liver cells of the rat, and these suggests that significant enhancements of endogenous enzymatic antioxidants by OJE might be a legitimate strategy for decreasing oxidative stresses in the liver.

PTP1B inhibitory effect of alkyl p-coumarates from Calystegia soldanella.

In this report, the PTP1B inhibitory effect of Calystegia soldanella was investigated. Bioassay-guided fractionation of the crude extracts revealed that the n-hexane fraction had the strongest PTP1B inhibitory effect. Nine known alkyl p-coumarates were isolated from the n-hexane fraction, and each compound was evaluated for its effect on PTP1B. All compounds effectively inhibited PTP1B activity. The IC50 values of the compounds were 3 (10.8 µg/mL) > 2 (15.5 µg/mL) > 7 (26.6 µg/mL) > 1 (37.0 µg/mL) > 8 (41.2 µg/mL) > 9 (43.4 µg/mL) > 5 (44.7 µg/mL) > 4 (> 50 µg/mL) > 6 (> 50 µg/mL).

Oenanthe javanica extract increases immunoreactivities of antioxidant enzymes in the rat kidney.

BACKGROUND: Oenanthe javanica is an aquatic perennial herb originated from East Asia. Nowadays, the effects of Oenanthe javanica have been proven in various disease models. Studies regarding the antioxidant effect of Oenanthe javanica in the kidney are still unclear. METHODS: This study was therefore performed to investigate the effect of the Oenanthe javanica extract (OJE) in the rat kidney using immunohistochemistry for antioxidant enzymes, copper, zinc-superoxide dismutase (SOD1), manganese superoxide dismutase (SOD2), catalase (CAT) and glutathione peroxidase (GPx). Sprague-Dawley rats were randomly assigned to three groups: (1) normal diet fed-group (normal-group), (2) diet containing ascorbic acid (AA)-fed group (AA-group) as a positive control, (3) diet containing OJE-fed group (OJE-group). AA and OJE were supplied during 28 days. RESULTS: The side-effects were not observed in all the groups. Immunoreactivities of SOD1, SOD2, CAT and GPx were easily detected in the distal tubules of the kidney, and their immunoreactivities in the AA-and OJE-groups were increased to about 1.4-1.5 times and 2 times, respectively, compared with those in the normal-group. CONCLUSION: OJE significantly increased expressions of SOD1 & 2, CAT and GPx immunoreactivities in the distal tubules of the rat kidney, and this finding suggests that significant enhancements of endogenous enzymatic antioxidants by OJE treatment may be a legitimate strategy for decreasing oxidative stresses in the kidney.

Neuroprotection via maintenance or increase of antioxidants and neurotrophic factors in ischemic gerbil hippocampus treated with tanshinone I.

BACKGROUND: Danshen (Radix Salvia miltiorrhizae) has been used as a traditional medicine in Asia for treatment of various microcirculatory disturbance related diseases. Tanshinones are mainly hydrophobic active components, which have been isolated from Danshen and show various biological functions. In this study, we observed the neuroprotective effect of tanshinone I (TsI) against ischemic damage in the gerbil hippocampal CA1 region (CA1) after transient cerebral ischemia and examined its neuroprotective mechanism. METHODS: The gerbils were divided into vehicle-treated-sham-group, vehicle-treated-ischemia-group, TsI-treated-sham-group, and TsI-treated-ischemia-group. TsI was administrated intraperitoneally three times (once a day for three days) before ischemia-reperfusion. The neuroprotective effect of TsI was examined using H&E staining, neuronal nuclei (NeuN) immunohistochemistry and Fluoro-Jade B staining. To investigate the neuroprotective mechanism of TsI after ischemia-reperfusion, immunohistochemical (IHC) and Western blotting analyses for Cu, Zn-superoxide dismutase (SOD1), Mn-superoxide dismutase (SOD2), brain-derived neurotrophic factor (BDNF) and insulin-like growth factor-I (IGF-I) were performed. RESULTS: Treatment with TsI protected pyramidal neurons from ischemia-induced neuronal death in the CA1 after ischemia-reperfusion. In addition, treatment with TsI maintained the levels of SOD1 and SOD2 as determined by IHC and Western blotting in the CA1 after ischemia-reperfusion compared with the vehicle-ischemia-group. In addition, treatment with TsI increased the levels of BDNF and IGF-I determined by IHC and Western blotting in the TsI-treated-sham-group compared with the vehicle-treated-sham-group, and their levels were maintained in the stratum pyramidale of the ischemic CA1 in the TsI-treated-ischemia-group. CONCLUSION: Treatment with TsI protects pyramidal neurons of the CA1 from ischemic damage induced by transient cerebral ischemia via the maintenance of antioxidants and the increase of neurotrophic factors.

Anti-inflammatory effect of tanshinone I in neuroprotection against cerebral ischemia-reperfusion injury in the gerbil hippocampus.

Tanshinone I (TsI) is an important lipophilic diterpene extracted from Danshen (Radix Salvia miltiorrhizae) and has been used in Asia for the treatment of cerebrovascular diseases such as ischemic stroke. In this study, we examined the neuroprotective effect of TsI against ischemic damage and its neuroprotective mechanism in the gerbil hippocampal CA1 region (CA1) induced by 5 min of transient global cerebral ischemia. Pre-treatment with TsI protected pyramidal neurons from ischemic damage in the stratum pyramidale (SP) of the CA1 after ischemia-reperfusion. The pre-treatment with TsI increased the immunoreactivities and protein levels of anti-inflammatory cytokines [interleukin (IL)-4 and IL-13] in the TsI-treated-sham-operated-groups compared with those in the vehicle-treated-sham-operated-groups; however, the treatment did not increase the immunoreactivities and protein levels of pro-inflammatory cytokines (IL-2 and tumor necrosis factor-α). On the other hand, in the TsI-treated-ischemia-operated-groups, the immunoreactivities and protein levels of all the cytokines were maintained in the SP of the CA1 after transient cerebral ischemia. In addition, we examined that IL-4 injection into the lateral ventricle did not protect pyramidal neurons from ischemic damage. In conclusion, these findings indicate that the pre-treatment with TsI can protect against ischemia-induced neuronal death in the CA1 via the increase or maintenance of endogenous inflammatory cytokines, and exogenous IL-4 does not protect against ischemic damage.

Capsosiphon fulvescens glycoprotein inhibits AGS gastric cancer cell proliferation by downregulating Wnt-1 signaling.

Previously, we examined various apoptosis pathways in the AGS gastric cancer cell line using Capsosiphon fulvescens glycoprotein (Cf-GP). In this study, we focused on the downregulation of the Wnt-1 signaling pathway and cell cycle arrest. Upregulation of the Wnt signaling pathway has been observed in various cancer cells. The Wnt signal ligand acts in both canonical and non-canonical pathways. Among them, Wnt-1 was dependent on the canonical pathway. Here, we show inhibition of Wnt-1 signaling, ß-catenin and transcription factors in AGS cells via Cf-GP. First, we examined the Frizzled receptor and Wnt-1 signal-related proteins including Axin, LRP, ß-catenin, APC and GSK-3ß. In addition, the expression levels of transcription factors Tcf/LEF were determined by western blot analysis and RT-PCR. Based on the data, we confirmed downregulation of the Wnt-1 signaling pathway by Cf-GP. Also, we determined the expression levels of cell cycle-related proteins cyclin D and c-myc, and looked for cell cycle arrest by cell cycle test analysis. We found that AGS cells arrested in the G0/G1 phase by Cf-GP. These results provide a mechanism of AGS cell inhibition through the downregulation of Wnt-1 signaling by Cf-GP.

Induction of apoptosis and the regulation of ErbB signaling by laminarin in HT-29 human colon cancer cells.

Laminarin, found in marine brown algae, is used as a carbohydrate reserve for phytoplankton; however, it is also used in traditional Chinese medicine, and has been shown to have several biological activities, including anticancer activities. In this study, we examined the mechanisms through which laminarin from Laminaria digitata induces apoptosis in HT-29 colon cancer cells, as well as the involvement of the ErbB signaling pathway. Cell viability assay revealed that laminarin induced cell death in a dose-dependent manner. Cell cycle analysis revealed that laminarin increased the percentage of cells in the sub-G1 and G2-M phase. Western blot analysis demonstrated that laminarin inhibited the heregulin-stimulated phosphorylation of ErbB2. A decrease in cellular proliferation was also observed; this was found to be dependent on ErbB, which activates c-Jun N-terminal kinase. These findings demonstrate the important role of the epidermal growth factor receptor in colon cancer tumorigenesis, and suggest the potential of laminarin as a bio-functional food with anticancer effects on human colon cancer.

Inhibition of AGS human gastric cancer cell invasion and proliferation by Capsosiphon fulvescens glycoprotein.

Seaweeds are increasingly being used as foodstuffs and therapeutics. Capsosiphon fulvescens (C. fulvescens) is a green sea alga which has demonstrated anti­tumor activity in various cancer cell lines. In cancer cells, homeostasis is not maintained, enabling mutations to develop and growth to continue unchecked. Overexpression of matrix metalloproteinase (MMP) and tight junction (TJ) proteins is important for cancer cell proliferation, invasion and metastasis. In addition, these proteins are closely associated with cell membrane permeability. In the current study, C. fulvescens glycoprotein (Cf­GP) was found to inhibit TJ proteins and invasion of AGS human gastric cancer cells. Cf­GP­mediated inhibition of cell proliferation and invasion was confirmed, as well as changes in TJ protein levels. In addition, MMP­2 and ­9 activities were inhibited, as indicated by increased transepithelial electrical resistance. Inhibition of MMP protein expression was also found to correlate with tissue inhibitor of metalloproteinase 1 and Cf­GP treatment, as revealed by western blot analysis and RT­PCR. In conclusion, these results indicate that Cf­GP inhibits cancer cell invasion and therefore demonstrates a potential therapeutic strategy to decrease cancer metastasis.

Anti-oxidative stress effect of red ginseng in the brain is mediated by peptidyl arginine deiminase type IV (PADI4) repression via estrogen receptor (ER) ß up-regulation.

AIM OF THE STUDY: Ginseng has been used as an anti-stress agent, and its active ingredient, ginsenoside, is similar in structure to estrogen. However, the effect of ginseng on the stressed brain is not completely understood. The aim of this study is to understand systematically how red ginseng (RG) affects gene expressions in the brain of immobilization (IMO) stressed mice to elucidate its underlying mechanism. MATERIALS AND METHODS: For in vivo experiments, mice were stressed by immobilization for 30, 45, or 60 min, and gene expression in the mice brain was analyzed by microarray and system biology. Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling (TUNEL) staining, and gene expression by Western blot or qPCR. For in vitro study, the SK-N-SH neuroblastoma cells were stressed by H2O2 exposure. The resultant cytotoxicity was measured by MTT assay, and gene expression by Western blot, ELISA, or qPCR. RESULTS: Microarray analysis of genes in IMO stressed mice brains showed that RG administration prior to IMO stress downregulated >40 genes including peptidyl arginine deiminase type 4 (PADI4). Interestingly, PADI4 was up-regulated by various stresses such as H2O2, acrylamide, and tunicamycin in neuroblastoma SK-N-SH cells but inhibited by RG. IMO stress and in vitro H2O2 stress depressed the estrogen receptor (ER)-ß expression but not ERα. However, RG treatment increased ERß expression both in vivo and in vitro. Comparative analysis regarding the networks by systems biology revealed that TNF-α plays a critical role in IMO stress, and the cell death associated network was much higher than other categories. Consistently, the IMO stress induced TNF-α and Cox-2 expressions, malondialdehyde (MDA), and cell death in the brain, whereas RG administration inhibited these inductions in vivo. siRNA and transient expression studies revealed that ERß inhibited the PADI4 expression. CONCLUSION: PADI4 could be used as an oxidative stress marker. RG seems to inhibit oxidative stress-inducible PADI4 by up-regulating ERß expression in the brain thus protecting brain cells from apoptosis.