The Modulation of Phospho-Extracellular Signal-Regulated Kinase and Phospho-Protein Kinase B Signaling Pathways plus Activity of Macrophage-Stimulating Protein Contribute to the Protective Effect of Stachydrine on Acetaminophen-Induced Liver Injury.

Stachydrine, a prominent bioactive alkaloid derived from Leonurus heterophyllus, is a significant herb in traditional medicine. It has been noted for its anti-inflammatory and antioxidant characteristics. Consequently, we conducted a study of its hepatoprotective effect and the fundamental mechanisms involved in acetaminophen (APAP)-induced liver injury, utilizing a mouse model. Mice were intraperitoneally administered a hepatotoxic dose of APAP (300 mg/kg). Thirty minutes after APAP administration, mice were treated with different concentrations of stachydrine (0, 2.5, 5, and 10 mg/kg). Animals were sacrificed 16 h after APAP injection for serum and liver tissue assays. APAP overdose significantly elevated the serum alanine transferase levels, hepatic pro-inflammatory cytokines, malondialdehyde activity, phospho-extracellular signal-regulated kinase (ERK), phospho-protein kinase B (AKT), and macrophage-stimulating protein expression. Stachydrine treatment significantly decreased these parameters in mice with APAP-induced liver damage. Our results suggest that stachydrine may be a promising beneficial target in the prevention of APAP-induced liver damage through attenuation of the inflammatory response, inhibition of the ERK and AKT pathways, and expression of macrophage-stimulating proteins.

Synergistic effect of Aloe vera flower and Aloe gel on cutaneous wound healing targeting MFAP4 and its associated signaling pathway: In-vitro study.

ETHNOPHARMACOLOGICAL RELEVANCE: Aloe vera (L.) Burm. f. (Liliaceae family) is a well-known traditional medicinal plant, that has been used to treat a variety of illnesses, for decades ranging from cancer to skin disorders including wounds. It has been included in the traditional and herbal healthcare systems of many cultures around the world, as well as the pharmacopeia of different countries. Several in vitro and in vivo studies have also confirmed its potential antioxidant, anti-inflammatory, and wound-healing activities, etc. in the consistency of its historical and traditional uses. However, most studies to date are based on the A. vera gel and latex including its wound-healing effects. Very few studies have been focused on its flower, and rarely with its effects on cutaneous wound healing and its molecular mechanism. AIM OF THE STUDY: To the best of our knowledge, this is the first study to report on the synergistic effect of the A. vera flower (AVF) and Aloe gel (PAG) on cutaneous wound-healing, as well as revealing its molecular mechanism targeting microfibril-associated glycoprotein 4 (MFAP4) and its associated signaling pathway. METHODS: To investigate the synergistic effect of A. vera flower and Aloe gel in cutaneous wound healing, cell viability, and cell migration, as well proliferation assay was performed. This was followed by quantitative real-time polymerase chain reaction and Western blot analyses in wounded conditions to check the effects of this mixture on protein and mRNA levels in normal human dermal fibroblast (NHDF) cells. Moreover, small interfering RNA (siRNA) -mediated knockdown of MFAP4 in NHDF cells was performed followed by migration assay and cell cycle analysis, to confirm its role in cutaneous wound healing. Additionally, HaCaT cells were included in this study to evaluate its migratory and anti-inflammatory effects. RESULTS: Based on our obtained results, the PAG and AVF mixture synergistically induced the proliferation, migration, and especially ECM formation of NHDF cells by enhancing the expression of MFAP4. Other extracellular components associated with MFAP4 signaling pathway, such as fibrillin, collagen, elastin, TGF ß, and α-SMA, also increased at both the protein and mRNA levels. Subsequently, this mixture initiated the phosphorylation of the extracellular signal-regulated kinase (ERK) and AKT signaling pathways, and the S-phase of the cell cycle was also slightly modified. Also, the mixture induced the migration of HaCaT cells along with the suppression of inflammatory cytokines. Moreover, the siRNA-mediated knockdown highlighted the crucial role of MFAP4 in cutaneous wound healing in NHDF cells. CONCLUSION: This study showed that the mixture of PAG and AVF has significant wound healing effects targeting MFAP4 and its associated signaling pathway. Additionally, MFAP4 was recognized as a new potential biomarker of wound healing, which can be confirmed by further in vivo studies.

6,7,4'-Trihydroxyflavone inhibits osteoclast formation and bone resorption in vitro and in vivo.

The balance between the osteoblasts and the osteoclasts is important for the maintenance of the skeleton of the human body. The osteoclasts absorb bone after differentiated into polymorphonuclear cells by the fusion of monocytes/macrophages. We have found that 6,7,4'-Trihydroxyflavone (THF), a compound from the heartwood of Dalbergia Odorifera inhibits receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, actin ring formation, and bone resorption in RAW 264.7 cells and bone marrow macrophage. THF significantly inhibited the c-Jun-N-terminal kinase signaling pathway without affecting extracellular signal-regulated kinase, p38, and AKT signaling. Moreover, THF inhibited the expression of c-Fos, nuclear factor-activated T cells cytoplasm 1, cathepsin K, and c-src by RANKL. We used a lipopolysaccharide (LPS)-induced bone loss model in mice. Consequently, bone volume per tissue volume, trabecular number's reduction was recovered in THF-treated mice, and trabecular separation's augmentation was also attenuated by THF administration. In summary, THF inhibits RANKL-induced osteoclast differentiation by MAPK signaling pathway and inhibits bone resorption by destroying the actin ring in mature osteoclasts. THF also prevented LPS-induced bone loss in a mice model. Thus, THF may be useful in the treatment of bone diseases associated with excessive osteoclast differentiation and bone resorption.

Triacanthine exerts antitumor effects on bladder cancer in vitro and in vivo.

BACKGROUND: Numerous studies have focused on solvent extracts from locust trees (Gleditsia spp.), which contain diverse bioactive components including saponins, flavonoids, and alkaloids. However, because of the undefined nature of such phytochemicals, their clinical application as chemotherapeutic agents has often been limited. PURPOSE: This study aimed to evaluate the anti-oncogenic activity of triacanthine, an alkaloid obtained from Gleditsia triacanthos L. STUDY DESIGN: The anti-oncogenicity of triacanthine in vitro was evaluated via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, cell-counting kit-8 assay (CCK-8 assay), flow cytometry, imunoblot, migration and invasion assays, zymography, and electrophoretic mobility shift assay in the human bladder carcinoma cell line EJ. The in vivo efficacy of triacanthine was evaluated via oral administration to EJ-xenografted BALB/c nude mice. To identify the side effects of triacanthine, cisplatin was also administered and an acute toxicity test was performed. RESULTS: Triacanthine significantly inhibited EJ cell proliferation (IC50 600 µM). Flow cytometry analysis revealed that cells were arrested in the G1 phase, and apoptotic cells accumulated in sub-G1 phase in a dose-dependent manner. Triacanthine inhibited the G1-S transition by deterring complex formation between cyclin-dependent kinases and cyclins, thereby up-regulating cell cycle inhibitors p21WAF1 and p27KIP1. In addition, triacanthine induced a caspase-dependent extrinsic pathway of apoptosis and autophagy. Early responsive kinases, extracellular signal-regulated kinase (ERK) and Janus kinase (JNK) were up-regulated by triacanthine. Triacanthine-mediated inhibition of the migratory and invasive potential of EJ cells was attributed to reduction of matrix metalloproteinase (MMP)-9 due to suppression of binding activities of the transcription factors activator protein (AP)-1, specificity protein (Sp)-1, and nuclear factor (NF)-κB. In an in vivo study, triacanthine significantly limited growth of xenografted tumors. Interestingly, while cisplatin resulted in significant weight loss after a 5-mg/kg dose, triacanthine did not cause weight loss, behavioral abnormalities, altered biochemical parameters, or tissue staining. A single oral dose acute-toxicity test (triacanthine 2,000 mg/kg) produced no adverse cytotoxic effects via blood biochemical tests and tissue-organ staining. CONCLUSION: To our knowledge, this is the first systematic evaluation of the anti-oncogenic activity of triacanthine. Therefore, we believe that our findings may guide the development of novel chemotherapeutic agents for bladder cancers.

Betulinic acid attenuates liver fibrosis by inducing autophagy via the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway.

The present study was designed to investigate the effects of betulinic acid on human hepatic stellate cells in vitro and C57BL/6 mice in vivo, as well as the signaling pathways involved. In this study, we explored the effects of betulinic acid on expression of alpha smooth muscle actin and autophagy-related proteins. Betulinic acid reduced pathological damage associated with liver fibrosis, as well as serum platelet-derived growth factor and serum hydroxyproline levels. Furthermore, betulinic acid downregulated the expression of alpha smooth muscle actin and type I collagen in mouse liver and upregulated the expression of microtubule-associated protein light chain 3B and autophagy-related gene 7 at the gene and protein levels. LC3II expression was increased and alpha smooth muscle actin expression was decreased in betulinic acid-treated hepatic stellate cells. Interventions with bafilomycin A1 and mCherry-GFP-LC3 adenoviruses promoted the formation of autophagosomes in hepatic stellate cells and the development of autophagic flow. Our study found that mitogen-activated protein kinase/extracellular signal-regulated kinase may be involved in the effects of betulinic acid on liver fibrosis. The present study suggests that betulinic acid has anti-hepatic fibrosis activity by inducing autophagy and could serve as a promising new agent for treating hepatic fibrosis.

Targeting ERK signaling pathway by polyphenols as novel therapeutic strategy for neurodegeneration.

Numerous chemicals, such as phenolic compounds are strong radical scavengers, capable of alleviating oxidative stress induced neurodegeneration. Dietary antioxidants, especially flavonoids, are being considered as a promising approach to prevent or slow the pathological development of neurological illness and aging. One of the major advantage of natural products is that of their anti-amyloid effects over synthetic counterpart, however a healthy diet provides these beneficial natural substances as nutraceuticals. The extracellular-signal-regulated kinase (ERK) is one of the main pharmacological target of natural phenolic compounds, participating in several therapeutic effects. Mounting evidence revealed that numerous bioflavonoids, obtained from a variety of dietary fruits or plants as well as medicinal herbal sources, exhibit protective or therapeutic functions versus development of neurodegenerative diseases mainly through modulation of different compartments of ERK signaling pathway. Currently, there is remarkable interest in the beneficial effects of natural flavonoids to improve neural performance and prevent the onset and development of major neurodegenerative diseases. Natural products originated from medicinal plants, in particular antioxidants, have gained a great deal of attention due to their safe and non-toxic natures. Here, we summarized the effect of natural bioflavonoids on ERK signaling pathway and their molecular mechanism.

Antidepressant-like effects of standardized gypenosides: involvement of brain-derived neurotrophic factor signaling in hippocampus.

RATIONALE: Gypenosides have been reported to produce neuroprotective effects and increase monoamine neurotransmitter levels in the brain. OBJECTIVE: Considering that depression is involved in monoamine reduction, this study evaluated the antidepressant-like effects of gypenosides in mice exposed to chronic unpredictable mild stress (CUMS). METHODS: The sucrose preference test and forced swimming test were performed after administration of gypenosides (at 25, 50, or 100 mg/kg) for 4 weeks. Hippocampal brain-derived neurotrophic factor (BDNF) and its downstream targets were analyzed by western blot. Additionally, hippocampal neuronal proliferation was measured by immunohistochemistry. RESULTS: Four-week treatment with fluoxetine (20 mg/kg) and gypenosides (at either 50 or 100 mg/kg) increased sucrose preference and decreased the immobility time in mice exposed to CUMS. In addition, gypenosides (at either 50 or 100 mg/kg) also increased BDNF expression and neuronal proliferation in the hippocampus of CUMS animals. Further, we showed that treating CUMS mice with K252a, which is an inhibitor of the BDNF receptor TrkB, blocked the effects of gypenosides (100 mg/kg), including behavioral improvements, neuronal proliferation, and up-regulation of p-TrkB, p-ERK, and p-Akt proteins. CONCLUSIONS: This study demonstrates that gypenosides exhibit antidepressant-like effects in mice, which may be mediated by activation of the BDNF-ERK/Akt signaling pathway in the hippocampus.

Green tea (-)-epigallocatechin gallate induced growth inhibition of human placental choriocarcinoma cells.

This study investigated the pathways involved in the effect of green tea epigallocatechin gallate (EGCG) on mitogenesis in BeWo, JEG-3, and JAR placental choriocarcinoma cells. EGCG inhibited cell proliferation in dose-dependent and time-dependent manners, as indicated by the number of cells and incorporation of bromodeoxyuridine (BrdU). A catechin-specific effect of green tea was evident; EGCG was more effective than epicatechin, epicatechin gallate, and epigallocatechin in suppressing cell growth. When all three of the mitogen-activated protein kinase (MAPK) subfamilies, i.e., ERK, p38, and JNK, were examined, EGCG significantly increased levels of phospho-ERK1/2 (pERK1/2) and phospho-p38 (pp38) and did not alter the total protein levels of ERK1/2, p38 MAPK, JNK, and phospho-JNK. EGCG-induced increases in the levels of pERK1/2 and pp38 proteins were prevented by pre-treatment with specific inhibitors of ERK1/2 MAPK and p38 MAPK, respectively. These inhibitors also suppressed EGCG-induced decreases in both cell number and BrdU incorporation. Moreover, pre-treatment with an AMP-activated protein kinase (AMPK) inhibitor prevented the actions of EGCG on proliferation and AMPK phosphorylation. These data suggest that EGCG mediates choriocarcinoma cell growth via the AMPK, ERK, and p38 pathways, but not JNK pathway.

Giganteaside D induces ROS-mediated apoptosis in human hepatocellular carcinoma cells through the MAPK pathway.

PURPOSE: Every year, almost one million individuals are diagnosed with hepatocellular carcinoma (HCC) worldwide and more than 690,000 patients die of it. At present, most therapeutic anti-HCC agents are not effective, which is due to the appearance of chemo-resistance and/or toxic side effects. Therefore, it is imperative to find novel more effective anti-HCC agents. Here, we evaluated the effect of giganteaside D (GD), an oleanolic acid saponin from P. scabiosaefolia, on the growth and apoptosis of HCC cells. METHODS AND RESULTS: Using MTT and clonogenic assays, we found that GD exhibited a significant growth inhibitory effect on the HCC-derived cell lines HepG2 and Bel-7402. In addition, we found that GD induced mitochondria-mediated apoptosis in these HCC-derived cells, as indicated by a decreased mitochondrial potential, activation of Caspase-9 and Caspase-3, cleavage of PARP and release of Cytochrome C from the mitochondria. Besides, we found that GD stimulated the generation of reactive oxygen species (ROS) and that blockage of ROS attenuated the GD-induced mitochondria-mediated apoptosis. Additionally, we found that GD treatment led to a decrease in phosphorylated Erk (p-Erk) and triggered the generation of p-JNK, both components of the mitogen-activated protein kinase (MAPK) signaling pathway. Inhibition of Erk or JNK by specific inhibitors or siRNAs augmented or attenuated the cytotoxic and apoptotic effects of GD. CONCLUSIONS: From our results we conclude that GD can induce ROS-mediated apoptosis in HCC-derived cells through the MAPK pathway. This observation may open up avenues to explore the future use of GD as a HCC chemotherapeutic agent.

Effects of DA-9701, a novel prokinetic agent, on phosphorylated extracellular signal-regulated kinase expression in the dorsal root ganglion and spinal cord induced by colorectal distension in rats.

BACKGROUND/AIMS: DA-9701, a standardized extract of Pharbitis Semen and Corydalis Tuber, is a new prokinetic agent that exhibits an analgesic effect on the abdomen. We investigated whether DA-9701 affects visceral pain induced by colorectal distension (CRD) in rats. METHODS: A total of 21 rats were divided into three groups: group A (no CRD+no drug), group B (CRD+no drug), and group C (CRD+DA-9701). Expression of pain-related factors, substance P (SP), c-fos, and phosphorylated extracellular signal-regulated kinase (p-ERK) in the dorsal root ganglion (DRG) and spinal cord was determined by immunohistochemical staining and Western blotting. RESULTS: The proportions of neurons in the DRG and spinal cord expressing SP, c-fos, and p-ERK were higher in group B than in group A. In the group C, the proportion of neurons in the DRG and spinal cord expressing p-ERK was lower than that in group B. Western blot results for p-ERK in the spinal cord indicated a higher level of expression in group B than in group A and a lower level of expression in group C than in group B. CONCLUSIONS: DA-9701 may decrease visceral pain via the downregulation of p-ERK in the DRG and spinal cord.

Extracellular signal­regulated kinase inhibition is required for methanol extract of Smilax china L.­induced apoptosis through death receptor 5 in human oral mucoepidermoid carcinoma cells.

Smilax china L., a well­known Chinese traditional medicine, has been used as an anti­inflammatory, anti­cancer and analgesic agent, but its role has not yet been fully elucidated in oral mucoepidermoid carcinoma (MEC). The present study focused on addressing the anticancer activity and molecular mechanism of methanol extract of Smilax china L. (MESC) in MC­3 human oral MEC cells. The results indicated that MESC inhibited cell growth and induced apoptosis in MC­3 cells. These observations were found to correlate with increases in truncated BH3 interacting­domain death agonist and B­cell lymphoma 2 (Bcl­2) interacting mediator of cell death, but not Bcl­2 homologous antagonist killer, Bcl­2­associated X protein, Bcl­2, B­cell lymphoma­extra large and induced myeloid leukemia cell differentiation protein levels. MESC also damaged the mitochondrial membrane potential, cleaved caspase­8 protein and increased death receptor 5 (DR5) protein levels by enhancing the stability of DR5 protein. Furthermore, MESC affected the phosphorylation of extracellular signal­regulated kinase (ERK) only, and did not affect c­Jun N­terminal kinase or p38 phosphorylation. Co­treatment with MESC and an ERK inhibitor (PD98059) significantly increased the expression of DR5 to induce apoptosis in MC­3 cells. Therefore, these results suggest that MESC may induce apoptosis via the ERK pathway and may be a potential anticancer drug candidate against human oral MEC.

The HMGB1/RAGE inflammatory pathway promotes pancreatic tumor growth by regulating mitochondrial bioenergetics.

Tumor cells require increased adenosine triphosphate (ATP) to support anabolism and proliferation. The precise mechanisms regulating this process in tumor cells are unknown. Here, we show that the receptor for advanced glycation endproducts (RAGE) and one of its primary ligands, high-mobility group box 1 (HMGB1), are required for optimal mitochondrial function within tumors. We found that RAGE is present in the mitochondria of cultured tumor cells as well as primary tumors. RAGE and HMGB1 coordinately enhanced tumor cell mitochondrial complex I activity, ATP production, tumor cell proliferation and migration. Lack of RAGE or inhibition of HMGB1 release diminished ATP production and slowed tumor growth in vitro and in vivo. These findings link, for the first time, the HMGB1-RAGE pathway with changes in bioenergetics. Moreover, our observations provide a novel mechanism within the tumor microenvironment by which necrosis and inflammation promote tumor progression.

Effects of DA-9701, a Novel Prokinetic Agent, on Phosphorylated Extracellular Signal-Regulated Kinase Expression in the Dorsal Root Ganglion and Spinal Cord Induced by Colorectal Distension in Rats

BACKGROUND/AIMS: DA-9701, a standardized extract of Pharbitis Semen and Corydalis Tuber, is a new prokinetic agent that exhibits an analgesic effect on the abdomen. We investigated whether DA-9701 affects visceral pain induced by colorectal distension (CRD) in rats. METHODS: A total of 21 rats were divided into three groups: group A (no CRD+no drug), group B (CRD+no drug), and group C (CRD+DA-9701). Expression of pain-related factors, substance P (SP), c-fos, and phosphorylated extracellular signal-regulated kinase (p-ERK) in the dorsal root ganglion (DRG) and spinal cord was determined by immunohistochemical staining and Western blotting. RESULTS: The proportions of neurons in the DRG and spinal cord expressing SP, c-fos, and p-ERK were higher in group B than in group A. In the group C, the proportion of neurons in the DRG and spinal cord expressing p-ERK was lower than that in group B. Western blot results for p-ERK in the spinal cord indicated a higher level of expression in group B than in group A and a lower level of expression in group C than in group B. CONCLUSIONS: DA-9701 may decrease visceral pain via the downregulation of p-ERK in the DRG and spinal cord.

Inhibition of vascular smooth muscle cell proliferation by Gentiana lutea root extracts.

Gentiana lutea belonging to the Gentianaceae family of flowering plants are routinely used in traditional Serbian medicine for their beneficial gastro-intestinal and anti-inflammatory properties. The aim of the study was to determine whether aqueous root extracts of Gentiana lutea consisting of gentiopicroside, gentisin, bellidifolin-8-O-glucoside, demethylbellidifolin-8-O-glucoside, isovitexin, swertiamarin and amarogentin prevents proliferation of aortic smooth muscle cells in response to PDGF-BB. Cell proliferation and cell cycle analysis were performed based on alamar blue assay and propidium iodide labeling respectively. In primary cultures of rat aortic smooth muscle cells (RASMCs), PDGF-BB (20 ng/ml) induced a two-fold increase in cell proliferation which was significantly blocked by the root extract (1 mg/ml). The root extract also prevented the S-phase entry of synchronized cells in response to PDGF. Furthermore, PDGF-BB induced ERK1/2 activation and consequent increase in cellular nitric oxide (NO) levels were also blocked by the extract. These effects of extract were due to blockade of PDGF-BB induced expression of iNOS, cyclin D1 and proliferating cell nuclear antigen (PCNA). Docking analysis of the extract components on MEK1, the upstream ERK1/2 activating kinase using AutoDock4, indicated a likely binding of isovitexin to the inhibitor binding site of MEK1. Experiments performed with purified isovitexin demonstrated that it successfully blocks PDGF-induced ERK1/2 activation and proliferation of RASMCs in cell culture. Thus, Gentiana lutea can provide novel candidates for prevention and treatment of atherosclerosis.

Magnolol inhibits angiogenesis by regulating ROS-mediated apoptosis and the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells.

Magnolol, a neolignan from the traditional medicinal plant Magnolia obovata, has been shown to possess neuroprotective, anti-inflammatory, anticancer and anti-angiogenic activities. However, the precise mechanism of the anti-angiogenic activity of magnolol remains to be elucidated. In the present study, the anti-angiogenic effect of magnolol was evaluated in mouse embryonic stem (mES)/embryoid body (EB)-derived endothelial-like cells. The endothelial-like cells were obtained by differentiation from mES/EB cells. Magnolol (20 µM) significantly suppressed the transcriptional and translational expression of platelet endothelial cell adhesion molecule (PECAM), an endothelial biomarker, in mES/EB-derived endothelial-like cells. To further understand the molecular mechanism of the suppression of PECAM expression, signaling pathways were analyzed in the mES/EB-derived endothelial-like cells. Magnolol induced the generation of reactive oxygen species (ROS) by mitochondria, a process that was associated with the induction of apoptosis as determined by positive Annexin V staining and the activation of cleaved caspase-3. The involvement of ROS generation by magnolol was confirmed by treatment with an antioxidant, N-acetyl-cysteine (NAC). NAC inhibited the magnolol-mediated induction of ROS generation and suppression of PECAM expression. In addition, magnolol suppressed the activation of MAPKs (ERK, JNK and p38) and the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells. Taken together, these findings demonstrate for the first time that the anti-angiogenic activity of magnolol may be associated with ROS-mediated apoptosis and the suppression of the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells.

Effect of pollen from Typha angustata on hydrogen peroxide induced toxicity in osteoblastic MC3T3-E1 cells.

BACKGROUND: Typha angustata is a traditional Chinese medicine, commonly used for a variety of clinical disorders, including atherosclerosis and wound healing. In the present study, the protective effects of T. angustata pollen extract (TE) on the response of osteoblast to oxidative stress were evaluated. MATERIALS AND METHODS: Osteoblastic MC3T3-E1 cells were incubated with H(2)O(2) and/or TE, and markers of osteoblast function and oxidative damage were examined. RESULTS: TE treatment significantly (P < 0.05) reversed the cytotoxic effect of H(2)O(2) and this effect was blocked by ICI182780, suggesting that TE's effect might be partly involved in estrogen action. TE significantly (P < 0.05) increased collagen content, alkaline phosphatase activity, calcium deposition of osteoblasts in the presence of H(2)O(2) and these effects were blocked by rottlerin and PD98059, suggesting that the induction of differentiation by TE is associated with increased activation of protein kinase C and ERK. Moreover, H(2)O(2)-induced reduction of osteocalcin was significantly recovered in the presence of TE. Pretreatment with TE also decreased the increase in receptor activator of nuclear factor-kB ligand, malondialdehyde, and protein carbonyl induced by H(2)O(2). CONCLUSION: These results suggest that the pollen of T. angustata may be useful for the protection of H(2)O(2)-induced oxidative damage and dysfunction in osteoblasts.

Denbinobin, a phenanthrene from dendrobium nobile, inhibits invasion and induces apoptosis in SNU-484 human gastric cancer cells.

Dendrobium nobile is widely used as an analgesic, an antipyretic, and a tonic to nourish the stomach in traditional medicine. Mounting evidence suggests an antitumor activity of denbinobin, a major phenanthrene isolated from stems of Dendrobium nobile. The present study aimed to investigate the inhibitory effect of denbinobin on the invasive ability of human cancer cells. The cytotoxicity of denbonobin was examined in several human cancer cell lines including SK-Hep-1 hepato-carcinoma cells, SNU-484 gastric cancer cells, and HeLa cervix cancer cells. Because SNU-484 cells showed the lowest IC50 value, we examined the effect of denbinobin on the invasive ability of SNU-484 cells. The present study revealed, for the first time, that denbinobin inhibits the invasive phenotype of SNU-484 human gastric cancer cells in a dose-dependent manner. Expressions of matrix metalloproteinase (MMP)-2 and MMP-9 were significantly decreased by denbinobin, suggesting that MMP-2/-9 may be responsible for the anti-invasive activity of denbinobin. We also provide evidence that denbinobin induces apoptosis through down-regulation of Bcl-2 and an up-regulation of Bax. Taken together, this study demonstrates that denbinobin inhibits invasion and induces apoptosis in highly invasive SNU-484 human gastric cancer cells. Given that gastric cancer has been estimated to be one of the most common causes of cancer-related death among Asians and the major cause of death from gastric cancer is the metastatic spread of the disease, our findings may provide useful information regarding the application of denbinobin as a chemopreventive agent that could prevent or alleviate metastatic gastric cancer.

Sauchinone suppresses pro-inflammatory mediators by inducing heme oxygenase-1 in RAW264.7 macrophages.

Sauchinone, a biologically active lignan isolated from the roots of Saururus chinensis (LOUR.) BAILL. (Saururaceae), is reported to exert a variety of biological activities, such as hepatoprotective, anti-inflammatory actions and inhibitory effects on bone resorption. In this study, we investigated the effect of sauchinone in suppressing cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, leading to a reduction in COX-2-derived prostaglandin E(2) (PGE(2)) and iNOS-derived nitric oxide (NO) production in lipopolysaccharide (LPS) stimulated RAW264.7 macrophages. Present study also demonstrates the effects of sauchinone in inducing heme oxygenase-1 (HO-1) expression and an increase in heme oxygenase (HO) activity in RAW264.7 macrophages. The effects of sauchinone on LPS-induced PGE(2), NO, tumor necrosis factor-α (TNF-α) and interlukine-1ß (IL-1ß) production were partially reversed by the HO-1 inhibitor Tin protoporphyrin was also seen in this study. In addition, we found that treatment with extracellular signal-regulated kinase (ERK) inhibitor (PD98059) reduced sauchinone-induced HO-1 expression. Sauchinone also increased ERK phosphorylation. These results suggest that sauchinone inhibits pro-inflammatory mediators through expression of anti-inflammatory HO-1 via ERK pathway.

Extracellular signal-regulated kinase (ERK) inhibition decreases arginase activity and improves corpora cavernosal relaxation in streptozotocin (STZ)-induced diabetic mice.

INTRODUCTION: Increased arginase activity (AA) has been implicated in hypertension and diabetes-induced endothelial dysfunction by reducing L-arginine availability and nitric oxide production. Higher levels of active extracellular signal-regulated kinase (ERK) have been found in patients with erectile dysfunction (ED) compared to patients without it. Both ERK and arginase have been reported to affect the expression and activity of nitric oxide synthase (NOS) and consequently penile erection. Nevertheless, signaling pathways activated by ERK in the penis are not well known. AIM: We hypothesized that inhibition of ERK by ERK inhibitor PD98059 decreases AA and thus improves cavernosal relaxation in streptozotocin (STZ)-diabetic mice. METHODS: The AA, ERK, eNOS, and arginase I and II expressions were examined through Western blot, and functional response of cavernosal tissue were determined. Control and diabetic cavernosal tissues were pretreated with PD98059 (10(-5) M) and arginase inhibitor ((S)-(2-boronoethyl)-L-cysteine hydrochloride, [BEC]10(-4) M]). MAIN OUTCOME MEASURES: Diabetes increased AA significantly (twofold) over control mice and this effect was blocked by acute treatment with PD98059. Cavernosal strips from diabetic mice exhibited decreased relaxation (STZ-diabetic vs. control, respectively) to both the endothelium-dependent agonist acetylcholine (38.0 ± 5% vs. 82.5 ± 7%) and nitrergic stimulation (27 ± 2% vs. 76 ± 6%) by electrical field stimulation (EFS, 1-32 Hz). However, this impairment in cavernosal relaxation from diabetic mice was attenuated by treatment with PD98059 in nitrergic (27 ± 2% vs. 60 ± 4%) and endothelium-dependent relaxation responses (38.0 ± 5% vs. 67.5 ± 6%). Acute treatment with the arginase inhibitor BEC (10(-4) M) also improves EFS-induced relaxation in diabetic mice (31 ± 3% vs. 49 ± 2%). Moreover, vascular expression of activated ERK was increased in diabetic over control mice. CONCLUSION: These data suggest that ERK inhibition prevents elevation of penile AA and protects against ED caused by diabetes.