Pongapin and Karanjin,furanoflavanoids of Pongamia pinnata,induce G2/M arrest and apoptosis in cervical cancer cells by differential reactive oxygen species modulation,DNA damage,and nuclear factor kappa‐light‐chain‐enhancer of activated B cell signaling

In this study, the antitumor activity of two furanoflavanoid derivatives, Pongapin and Karanjin, was evaluated in comparison with Plumbagin, a plant‐derived polyphenol with proven antitumor activity. The compounds differentially inhibit the growth of different cancer cell lines (most effective on HeLa cells), with very low inhibitory effect on the growth of normal mouse embryonic fibroblast cell line. Pongapin like Plumbagin could significantly increase the intracellular reactive oxygen species (ROS) in the HeLa cells by stabilization of nuclear factor of kappa light polypeptide gene enhancer in B‐cells inhibitor (I‐κB) expression and reduction of nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) expression. In contrast, Karanjin could decrease ROS level by inhibition of I‐κB degradation resulting restriction of NF‐κB nuclear translocation. Pongapin and Plumbagin significantly increased DNA damage‐induced p53 expression and p21 nuclear expression. However, Karanjin treatment showed low DNA damage with increased p53 expression. The compounds induced G2/M arrest and increase in SubG1 population, indicating induction of apoptosis. Apoptosis was further validated by acridine orange/ethidium bromide dual staining and terminal deoxynucleotidyl transferase dUTP nick‐end labeling assay in HeLa cells after treatment with the compounds. The compounds induced caspase‐dependent apoptosis through induction of Bax/Bcl‐2 ratio either through increased expression of Bax by Pongapin and Plumbagin or low expression of Bcl‐2 by Karanjin. Thus, Pongapin and Karanjin may be potential natural anticancer agents in the future, like Plumbagin.     

Matrine triggers colon cancer cell apoptosis and G0/G1 cell cycle arrest via mediation of microRNA‐22

Matrine (MAT) is an alkaloid in the dried roots of Sophora flavescens. The antitumor activity has been testified in colon cancer. Howbeit, the latent mechanism is still indistinct. The research probed the antitumor mechanism of MAT in colon cancer cells. MAT (0.25, 0.5, 0.75, 1, and 1.25 mM) was utilized to stimulate SW480 and SW620 cells for 24, 48, and 72 hr. Cell viability, apoptosis, cell cycle, and the correlative proteins were assessed via Cell Counting Kit‐8, flow cytometry, and Western blot. microRNA‐22 (miR‐22) in MAT‐treated or miR‐22‐silenced cells was estimated via real‐time quantitative polymerase chain reaction. The functions of miR‐22 inhibition were reassessed. Western blot was conducted for quantifying β‐catenin, MEK, and ERK. Luciferase reporter assay was done for confirming the targeting relationship between miR‐22 and ERBB3 or MECOM. MAT prohibited cell viability, accelerated apoptosis, and triggered cells cycle stagnation at G0/G1 phase. Additionally, miR‐22 was elevated by MAT; meanwhile, the influences of MAT were all inverted by miR‐22 inhibitor. MAT enhanced the expression of miR‐22, thereby obstructing Wnt/β‐catenin and MEK/ERK pathways. miR‐22 had a potential to target mRNA 3'UTR of ERBB3 and MECOM. These discoveries manifested that MAT could evoke colon cancer cell apoptosis and G0/G1 cell cycle arrest via elevating miR‐22.     

1,7‐Bis(4‐hydroxyphenyl)‐4‐hepten‐3‐one from Betula platyphylla induces apoptosis by suppressing autophagy flux and activating the p38 pathway in lung cancer cells

Betula platyphylla (BP) is frequently administered in the treatment of various human diseases, including cancers. This study was undertaken to investigate the pharmacological function of the active components in BP and the underlying mechanism of its chemotherapeutic effects in human lung cancer cells. We observed that BP extracts and 1,7‐bis(4‐hydroxyphenyl)‐4‐hepten‐3‐one (BE1), one of the components of BP, effectively decreased the cell viability of several lung cancer cell lines. BE1‐treated cells exhibited apoptosis induction and cell cycle arrest at the G2/M phase. Further examination demonstrated that BE1 treatment resulted in suppression of autophagy, as evidenced by increased protein expression levels of both LC3 II and p62/SQSTM1. Interestingly, the pharmacological induction of autophagy with rapamycin remarkably reduced the BE1‐induced apoptosis, indicating that apoptosis induced by BE1 was associated with autophagy inhibition. Our data also demonstrated that BE1 exposure activated the p38 pathway resulting in regulation of the pro‐apoptotic activity. Taken together, we believe that BE1 is a potential anticancer agent for human lung cancer, which exerts its effect by enhancing apoptosis via regulating autophagy and the p38 pathway.     

Apoptotic effect of lambertianic acid through AMPK/FOXM1 signaling in MDA‐MB231 breast cancer cells

Though lambertianic acid (LA) was known to exert antitumor effect in liver and prostate cancers, its underlying anticancer mechanism is never reported in breast cancers so far. Thus, in this study, apoptotic mechanism of LA was elucidated in MDA‐MB‐231 breast cancer cells. Here, LA increased cytotoxicity in MCF‐7 and MDA‐MB‐231 cells; enhanced sub‐G1 population, G2/M arrest, and cleaved poly(ADP‐ribose) polymerase; activated phosphorylation of AMP‐activated protein kinase (AMPK)/acetyl‐CoA carboxylase pathway; and also suppressed phosphorylation of AKT and the expression of forkhead box M1 (FOXM1), X‐linked inhibitor of apoptosis protein, B‐cell lymphoma 2, and CyclinB1 in MDA‐MB‐231 cells. Furthermore, AMPK inhibitor compound C reversed the effect of LA on FOXM1, Cyclin B1, and cleaved poly(ADP‐ribose) polymerase in MDA‐MB‐231 cells. Notably, immunoprecipitation revealed that LA disturbed the direct binding of AKT and FOXM1 in MDA‐MB‐231 cells. Overall, these findings suggest that LA‐induced apoptosis is mediated via activation of AMPK and inhibition of AKT/FOXM1 signaling pathway.     

Activation of Caspase‐9/3 and Inhibition of Epithelial Mesenchymal Transition are Critically Involved in Antitumor Effect of Phytol in Hepatocellular Carcinoma Cells

This study was designed to investigate the antitumor mechanism of Phytol in hepatocellular carcinomas including Huh7 and HepG2 cells in association with caspase dependent apoptosis and epithelial mesenchymal transition (EMT) signaling. Phytol significantly suppressed the viability of Huh7 and HepG2 cells. Also, Phytol significantly increased the sub G1 population and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling (TUNEL) positive cells in a concentration dependent manner in Huh7 and HepG2 cells. Consistently, Phytol cleaved poly (adenosine diphosphate‐ribose) polymerase (PARP), activated caspase‐9/3, and Bax attenuated the expression of survival genes such as Bcl‐2, Mcl‐1, and c‐Myc in Huh7 and HepG2 cells. Of note, Phytol also suppressed typical morphology change of EMT such as loss of cell adhesion and formation of fibroblast like mesenchymal cells in HepG2 cells. Furthermore, Phytol also reversed the loss of E‐cadherin and overexpression of p‐smad2/3, alpha‐smooth muscle actin, and Snail induced by EMT promoter transforming growth factor beta1 in HepG2 cells. Overall, our findings suggest that Phytol exerts antitumor activity via apoptosis induction through activation of caspas‐9/3 and inhibition of EMT in hepatocellular carcinoma cells as a potent anticancer candidate for liver cancer treatment. Copyright © 2015 John Wiley & Sons, Ltd.     

Reactive Oxygen Species‐Mediated Activation of AMP‐Activated Protein Kinase and c‐Jun N‐terminal Kinase Plays a Critical Role in Beta‐Sitosterol‐Induced Apoptosis in Multiple Myeloma U266 cells

Although beta‐sitosterol has been well known to have anti‐tumor activity in liver, lung, colon, stomach, breast and prostate cancers via cell cycle arrest and apoptosis induction, the underlying mechanism of anti‐cancer effect of beta‐sitosterol in multiple myeloma cells was never elucidated until now. Thus, in the present study, the role of reactive oxygen species (ROS) in association with AMP‐activated protein kinase (AMPK) and c‐Jun N‐terminal kinase (JNK) pathways was demonstrated in beta‐sitosterol‐treated multiple myeloma U266 cells. Beta‐sitosterol exerted cytotoxicity, increased sub‐G₁ apoptotic population and activated caspase‐9 and ‐3, cleaved poly (ADP‐ribose) polymerase (PARP) followed by decrease in mitochondrial potential in U266 cells. Beta‐sitosterol promoted ROS production, activated AMPK, acetyl‐CoA carboxylase (ACC) and JNK in U266 cells. Also, beta‐sitosterol attenuated the phosphorylation of AKT, mammalian target of rapamycin and S6K, and the expression of cyclooxygenase‐2 and VEGF in U266 cells. Conversely, AMPK inhibitor compound C and JNK inhibitor SP600125 suppressed apoptosis induced by beta‐sitosterol in U266 cells. Furthermore, ROS scavenger N‐acetyl L‐cysteine attenuated beta‐sitosterol‐mediated sub‐G₁ accumulation, PARP cleavage, JNK and AMPK activation in U266 cells. Overall, these findings for the first time suggest that ROS‐mediated activation of cancer metabolism‐related genes such as AMPK and JNK plays an important role in beta‐sitosterol‐induced apoptosis in U266 multiple myeloma cells. Copyright © 2013 John Wiley & Sons, Ltd.     

Effect of Orostachys japonicus on cell growth and apoptosis in human hepatic stellate cell line LX2

Orostachys japonicus (O. japonicus), used to treat diseases such as various cancers, gastric ulcers, fever, hepatitis, arthritis, eczema, for hemostasis, and intoxication in folk medicine, has been an important constituent in many herbal formulae. We demonstrated that the water extract of O. japonicus led to growth inhibition of LX2 cells by inducing apoptosis through the caspase activation, related to the MAPK pathway. O. japonicus inhibited proliferation of LX2 cells in a dose- and time-dependent manner, increased the apoptosis fraction at cell cycle progression with an accompanying DNA fragmentation, and resulted in a significant decrease in Bcl-2 and an increase in Bax mRNA levels. Exposure of LX2 cells to O. japonicus induced caspase-3 activation, however when the LX2 cells were also treated with the pan-caspase inhibitor Z-VAD-FMK and the caspase-3 inhibitor Z-DEVE-FMK, apoptosis was blocked. O. japonicus inhibited anti-apoptotic Mcl-1 protein and MEK/ERK phosphorylation in LX2 cells. The results indicate that O. japonicus inhibits the cell growth of LX2 cells by inducing apoptosis through caspase activity. O. japonicus down-regulated Mcl-1 protein levels and inhibited the phosphorylation of MEK/ERK, suggesting that it mediates cell death in LX2 cells through the down-regulation of Mcl-1 protein via a MEK/ERK-independent pathway.     

Chemo‐preventive and therapeutic effect of the dietary flavonoid kaempferol: A comprehensive review

Kaempferol, a natural flavonoid present in several plants, possesses a wide range of therapeutic properties such as antioxidant, anticancer, and anti‐inflammatory. It has a significant role in reducing cancer and can act as a therapeutic agent in the treatment of diseases and ailments such as diabetes, obesity, cardiovascular diseases, oxidative stress, asthma, and microbial contamination disorders. Kaempferol acts through different mechanisms: It induces apoptosis (HeLa cervical cancer cells), decreases cell viability (G2/M phase), downregulates phosphoinositide 3‐kinase (PI3K)/AKT (protein kinase B) and human T‐cell leukemia/lymphoma virus‐I (HTLV‐I) signaling pathways, suppresses protein expression of epithelial‐mesenchymal transition (EMT)‐related markers including N‐cadherin, E‐cadherin, Slug, and Snail, and metastasis‐related markers such as matrix metallopeptidase 2 (MMP‐2). Accordingly, the aim of the present review is to collect information pertaining to the effective role of kaempferol against various degenerative disorders, summarize the antioxidant, anti‐inflammatory, anticancer, antidiabetic, and antiaging effects of kaempferol and to review the progress of recent research and available data on kaempferol as a protective and chemotherapeutic agent against several ailments.     

Ursolic acid promotes apoptosis,autophagy, and chemosensitivity in gemcitabine‐resistant human pancreatic cancer cells

Gemcitabine (GEM) resistance in pancreatic adenocarcinoma mediated by the receptor for advanced glycation end products (RAGE) has been demonstrated. Therefore, investigating the safety and the potential of new auxiliary methods for pancreatic cancer treatment is urgent. Ursolic acid (UA), a natural pentacyclic triterpenoid found in apple peels, rosemary, and thyme, has been reported to have anticancer capacity. This study aimed to reveal the underlying mechanisms of UA in cell death and drug enhancement, especially in GEM‐resistant pancreatic cancer cells. First, GEM‐resistant cells (MIA Paca‐2^GEMR cells) were established by incrementally increasing GEM culture concentrations. UA treatment reduced cell viability through cell cycle arrest and endoplasmic reticulum (ER) stress, resulting in apoptosis and autophagy in a dose‐dependent manner in MIA Paca‐2 and MIA Paca‐2^GEMR cells. High RAGE expression in MIA Paca‐2^GEMR cells was suppressed by UA treatment. Interestingly, knocking down RAGE expression showed similar UA‐induced effects in both cell lines. Remarkably, UA had a drug‐enhancing effect by decreasing cell viability and increasing cell cytotoxicity when combined with GEM treatment. In conclusions, UA triggered ER stress, subsequently regulating apoptosis‐ and autophagy‐related pathways and increasing GEM chemosensitivity in pancreatic cancer cells by inhibiting the expression of RAGE.     

Yuk-Hap-Tang induces apoptosis by intervening mn-SOD in human cervical carcinoma HeLa cells

Yuk-Hap-Tang (YHT) induces cell death in human cervical carcinoma HeLa cells. Caspase-3, -6 and -9 were markedly activated in HeLa cells treated with YHT. The preferred substrate for caspase-3 cysteine protease, PARP, was cleaved to its 85-kDa cleavage product. YHT increased the amount of the anti-apoptotic protein, Bcl-2, and the pro-apoptotic protein, Bax. Although p53 has been reported to accumulate in cancer cells in response to anticancer agents, the p53 expression level was not changed in HeLa cells treated with YHT. Manganese (Mn)-TBAP, a mitochondria-specific SOD mimetic agent and NAC/GSH (N-acetyl cysteine/ reduced glutathione) reduced the YHT-induced cytotoxicity and decreased the number of the YHT-induced apoptotic cells. Furthermore, YHT reduced the expression of Mn-SOD protein and its activity in HeLa cells. The data demonstrate that YHT induces the apoptosis of human cervical carcinoma HeLa cells by intervening Mn-SOD.     

Tanshinone IIA Induces Autophagic Cell Death via Activation of AMPK and ERK and Inhibition of mTOR and p70 S6K in KBM‐5 Leukemia Cells

Although tanshinone IIA (Tan IIA) from Salviae miltiorrhizae was known to induce apoptosis in various cancers, its underlying mechanism of autophagic cell death was not reported yet. Thus, in the present study, the molecular mechanism of autophagic cell death by Tan IIA was investigated in KBM‐5 leukemia cells. Tan IIA significantly increased the expression of microtubule‐associated protein light chain 3 (LC3) II as a hallmark of autophagy in western blotting and immunofluorescence staining. Tan IIA augmented the phosphorylation of adenosine monophosphate‐activated protein kinase (AMPK) and attenuated the phosphorylation of mammalian target of rapamycin (mTOR) and p70 S6K in a dose‐dependent manner. Conversely, autophagy inhibitor 3‐methyladenine partly reversed the cytotoxicity and the phosphorylation of AMPK, mTOR and p70 S6K induced by Tan IIA in KBM‐5 leukemia cells. In addition, Tan IIA dramatically activated the extracellular signal regulated kinase (ERK) signaling pathway including Raf, ERK and p90 RSK in a dose‐dependent and time‐dependent manner. Consistently, ERK inhibitor PD184352 suppressed LC3‐II activation induced by Tan IIA, whereas PD184352 and PD98059 did not affect poly (ADP‐ribose) polymerase cleavage and sub‐G1 accumulation induced by Tan IIA in KBM‐5 leukemia cells. Furthermore, Tan IIA could induce autophagy via LC3‐II activation in various cancer cells such as prostate (PC‐3), multiple myeloma (U266), lung (NCI‐H460), and breast (MDA‐MB‐231) cells. Overall, these findings suggest that Tan IIA induces autophagic cell death via activation of AMPK and ERK and inhibition of mTOR and p70 S6K in KBM‐5 cells as a potent natural compound for leukemia treatment. Copyright © 2013 John Wiley & Sons, Ltd.     

Promotion of direct angiogenesis in vitro and in vivo by Puerariae flos extract via activation of MEK/ERK‐, PI3K/Akt/eNOS‐, and Src/FAK‐dependent pathways

Puerariae flos has been used for oriental herbal medicine; however, its angiogenic effect has not been elucidated. We found that the extract from Puerariae flos (PFE) increased in vitro angiogenic events, such as endothelial cell proliferation, migration, and tube formation, as well as in vivo neovascularization. These events were followed by the activation of multiple signal modulators, such as extracellular signal‐regulated kinase (ERK), Akt, endothelial nitric oxide synthase (eNOS), nitric oxide production, p38, Src, and focal adhesion kinase (FAK), without increasing vascular endothelial growth factor (VEGF) expression. Inhibition of ERK, Akt, and eNOS suppressed PFE‐induced angiogenic events, and inhibition of p38 and Src activities blocked PFE‐induced endothelial cell migration. PFE did not affect the expression of intracellular adhesion molecule‐1 and vascular cell adhesion molecule‐1 and transendothelial permeability, which are involved in the adverse effects of the well‐known angiogenic inducer VEGF. These results suggest that PFE directly stimulates angiogenesis through the activation of MEK/ERK‐, phosphatidylinositol 3‐kinase/Akt/eNOS‐, and Src/FAK‐dependent pathways, without altering VEGF expression, vascular inflammation, and permeability in vitro and in vivo and may be used as a therapeutic agent for ischemic disease and tissue regeneration. Copyright © 2009 John Wiley & Sons, Ltd.     

Comparison of the effect of oligonol,a new lychee fruit‐derived low molecular form of polyphenol,and epigallocatechin‐3‐gallate on lipolysis in rat primary adipocytes

Several polyphenols have been shown to be beneficial in preventing the accumulation of body fat in mammals. This paper reports that adding oligonol, a lychee fruit‐derived low‐molecular form of polyphenol mixture, has a greater effect on lipolysis in primary adipocytes compared with tea (?)‐epigallocatechin‐3‐gallate (EGCG) alone, accompanied by a significant increase in activation of extracellular signalling‐related kinase 1/2 (ERK1/2). However, neither phosphorylation of mitogen‐activated protein kinase 1/2 (MEK1/2), a molecule upstream of ERK1/2, nor the level of heme oxygenase‐1 (HO‐1), a molecule downstream of ERK1/2 was significantly changed between oligonol and EGCG, although the addition of oligonol and EGCG significantly increased the levels of phosphorylated MEK1/2 and HO‐1 compared with the non‐treated control cells. These results suggest that the coordinated direct effect of mixed polyphenol, which comprises oligonol, on ERK1/2 plays a key role in a greater lipolytic response to oligonol than EGCG alone. Copyright © 2010 John Wiley & Sons, Ltd.     

Gleditsia sinensis thorn extract inhibits human colon cancer cells: the role of ERK1/2, G2/M‐phase cell cycle arrest and p53 expression

The thorns of Gleditsia sinensis are used as a medicinal herb in China and Korea. However, the mechanisms responsible for the antitumor effects of the water extract of Gleditsia sinensis thorns (WEGS) remain unknown. HCT116 cells treated with the WEGS at a dose of 800 μg/mL (IC₅₀) showed a significant decrease in cell growth and an increase in cell cycle arrest during the G2/M‐phase. G2/M‐phase arrest was correlated with increased p53 levels and down‐regulation of the check‐point proteins, cyclinB1, Cdc2 and Cdc25c. In addition, treatment with WEGS induced phosphorylation of extracellular signal‐regulated kinase (ERK), p38 MAP kinase and JNK (c‐Jun N‐terminal kinases). Moreover, inhibition of ERK by treatment of cells with the ERK‐specific inhibitor PD98059 blocked WEGS‐mediated p53 expression. Similarly, blockage of ERK function in the WEGS‐treated cells reversed cell‐growth inhibition and decreased cell cycle proteins. Finally, in vivo WEGS treatment significantly inhibited the growth of HCT116 tumor cell xenografts in nude mice with no negative side effects, including loss of body weight. These results describe the molecular mechanisms whereby the WEGS might inhibit proliferation of colon cancer both in vitro and in vivo, suggesting that WEGS has potential as an anticancer agent for the treatment of malignancies. Copyright © 2010 John Wiley & Sons, Ltd.     

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??OBJECTIVE To investigate the apoptosis of the auction and mechanism of the hepatoma BEL-7402 cells induced by the ginseng polysaccharides(GPS). METHODS The hematoma cells BEL-7402 were incubated with GPS, cell viability was measured by CCK8, cell cycle distribution was assessed by fluorescence-activated cell sorting(FACS), the cell morphological changes were traced with TUNEL and scanning electron microscope, TNFR1, BCL-2 and Bax protein expression and ERK phosphorylation are tested by Western blot. RESULTS CCK8 results showed that GPS inhibited cells growth of BEL-7402 in dose-dependent and time-dependent. Flow cytometry found that S phase arrest was increased upon GPS concentrations. Obviously apoptotic sub-g peak was also found. And the peak was gradually enhanced with the concentration increased. TUNEL staining and SEM results showed that GPS led to significant cell morphology changes in hepatoma cells BEL-7402.And the apoptosis effect was increased upon the GPS concentrations. Western blot showed that level of apoptosis related protein Bax was increased, the expression of apoptosis-antagonizing protein Bcl-2 was decreased and the expression of death receptor TNFR1 appeared with GPS concentrations increased gradually,raise ERK phosphorylation. CONCLUSION GPS can induce apoptosis of hepatoma cells BEL-7402 by the mitochondrial pathway and the death receptor dependent pathway and ERK pathway.     

Paris saponin VII induces cell cycle arrest and apoptosis by regulating Akt/MAPK pathway and inhibition of P‐glycoprotein in K562/ADR cells

Paris saponinVII (PSVII) is a steroidal saponin isolated from the roots and rhizomes of Trillium tschonoskii Maxim. We found that PSVII could inhibit the growth of adriamycin‐resistant human leukemia cells (K562/ADR) in a dose‐dependent manner. Furthermore, the molecular mechanism underlying the cytotoxicity and downregulation of P‐glycoprotein (P‐gp) expression by PSVII was clarified. PSVII significantly suppressed cell proliferation by cell cycle arrest in the G0/G1 phase, which was associated with an obvious decrease in cyclin B1/D1 and CDK2/4/6 protein expression. Moreover, PSVII could attenuate mitochondrial membrane potential, increase the expression of apoptosis‐related proteins, such as Bax and cytochrome c, and decrease the protein expression levels of Bcl‐2, caspase‐9, caspase‐3, PARP‐1, and p‐Akt. We also found that JNK, ERK1/2, and p38 were regulated by PSVII in K562/ADR cells. And further studies indicated that the decrease in the reactive oxygen species level inhibited intrinsic P‐gp expression. Therefore, PSVII‐induced apoptosis in K562/ADR cells was associated with Akt/MAPK and the inhibition of P‐gp. In addition, PSVII induced a robust autophagy in K562/ADR cells as demonstrated by the degradation of LC3‐I. These results provide a biochemical basis for possible clinical applications of PSVII in the treatment of leukemia.     

Activation of AMP‐activated Protein Kinase and Phosphorylation of Glycogen Synthase Kinase3 β Mediate Ursolic Acid Induced Apoptosis in HepG2 Liver Cancer Cells

Despite the antitumour effect of ursolic acid observed in several cancers, the underlying mechanism remains unclear. Thus, in the present study, the roles of AMP‐activated protein kinase (AMPK) and glycogen synthase kinase 3 beta (GSK3β) were examined in ursolic acid induced apoptosis in HepG2 hepatocellular carcinoma cells. Ursolic acid significantly exerted cytotoxicity, increased the sub‐G1 population and the number of ethidium homodimer and terminal deoxynucleotidyl transferase(TdT) mediated dUTP nick end labeling positive cells in HepG2 cells. Also, ursolic acid enhanced the cleavages of poly‐ADP‐ribose polymerase (PARP) and caspase3, attenuated the expression of astrocyte elevated gene (AEG1) and survivin in HepG2 cells. Interestingly, ursolic acid increased the phosphorylation of AMPK and coenzyme A carboxylase and also enhanced phosphorylation of GSK3β at inactive form serine 9, whereas ursolic acid attenuated the phosphorylation of AKT and mTOR in HepG2 cells. Conversely, AMPK inhibitor compound C or GSK3β inhibitor SB216763 blocked the cleavages of PARP and caspase 3 induced by ursolic acid in HepG2 cells. Furthermore, proteosomal inhibitor MG132 suppressed AMPK activation, GSK3β phosphorylation, cleaved PARP and deceased AEG‐1 induced by ursolic acid in HepG2 cells. Overall, our findings suggest that ursolic acid induced apoptosis in HepG2 cells via AMPK activation and GSK3β phosphorylation as a potent chemopreventive agent. Copyright © 2013 John Wiley & Sons, Ltd.     

Phloroglucinol Protects INS‐1 Pancreatic β‐cells Against Glucotoxicity‐Induced Apoptosis

Decreasing numbers, and impaired function, of pancreatic β‐cells are key factors in the development of type 2 diabetes. This study was designed to investigate whether phloroglucinol protected pancreatic β‐cells against glucotoxicity‐induced apoptosis using a rat insulinoma cell line (INS‐1). High glucose treatment (30 mM) induced INS‐1 cell death; however, the level of glucose‐induced apoptosis was significantly reduced in cells treated with 100‐μM phloroglucinol. Treatment with 10–100‐μM phloroglucinol increased cell viability and decreased intracellular levels of reactive oxygen species, nitric oxide, and lipid peroxidation dose‐dependently in INS‐1 cells pretreated with high glucose. Furthermore, phloroglucinol treatment markedly reduced the protein expression of Bax, cytochrome c, and caspase 9, while increasing anti‐apoptotic Bcl‐2 protein expression. Cell death type was examined using annexin V/propidium iodide staining, revealing that phloroglucinol markedly reduced high glucose‐induced apoptosis. These results demonstrated that phloroglucinol could be useful as a potential therapeutic agent for the protection of pancreatic β‐cells against glucose‐induced apoptosis. Copyright © 2015 John Wiley & Sons, Ltd.     

Effects of Artemetin on Nitric Oxide Release and Protection against Peroxidative Injuries in Porcine Coronary Artery Endothelial Cells

Artemetin is one of the main components of Achillea millefolium L. and Artemisia absinthium, which have long been used for the treatment of various diseases. To date, however, available information about protective effects of their extracts on the cardiovascular system is scarce. Therefore, we planned to analyze the effects of artemetin on nitric oxide (NO) release and the protection exerted against oxidation in porcine aortic endothelial (PAE) cells. In PAE, we examined the modulation of NO release caused by artemetin and the involvement of muscarinic receptors, β2‐adrenoreceptors, estrogenic receptors (ER), protein‐kinase A, phospholipase‐C, endothelial‐NO‐synthase (eNOS), Akt, extracellular‐signal‐regulated kinases 1/2 (ERK1/2) and p38 mitogen activated protein kinase (p38 MAPK). Moreover, in cells treated with hydrogen peroxide, the effects of artemetin were examined on cell survival, glutathione (GSH) levels, apoptosis, mitochondrial membrane potential and transition pore opening. Artemetin increased eNOS‐dependent NO production by the involvement of muscarinic receptors, β2‐adrenoreceptors, ER and all the aforementioned kinases. Furthermore, artemetin improved cell viability in PAE that were subjected to peroxidation by counteracting GSH depletion and apoptosis and through the modulation of mitochondrial function. In conclusion, artemetin protected endothelial function by acting as antioxidant and antiapoptotic agent and through the activation of ERK1/2 and Akt. Copyright © 2015 John Wiley & Sons, Ltd.     

银杏内酯诱导嗜铬细胞瘤细胞表达低氧诱导因子-1α

 目的研究银杏内酯(ginkgolides,Gin)对神经生长因子(nerve growth factor,NGF)诱导分化的嗜铬细胞瘤(PC12)细胞的影响以及与其相关的信号通路。方法通过MTT比色法观察不同浓度Gin对PC12细胞活性的影响,RT-PCR和Western Blot法分析Gin对PC12细胞低氧诱导因子-1(αhypoxia inducible factor-1α,HIF-1α)表达以及MAPK信号通路的影响。结果一定浓度范围的Gin可促进PC12细胞的活性,在37.5 mg·L^-1作用24 h效果最明显。37.5 mg·L^-1 Gin单独处理PC12细胞24 h可诱导HIF-1αmRNA表达增强和蛋白水平上调。Gin还引起p-ERK水平的明显提高。Gin增加PC12细胞HIF-1α蛋白的稳定性存在一定的时间和剂量依赖关系,PD98059可部分抑制Gin的增强作用,金雀异黄素可完全阻断之;与此相对应的是,Gin以时间和剂量依赖方式诱导PC12细胞p-ERK水平的增高,PD98059和金雀异黄素均能完全阻断Gin的诱导作用。结论Gin可诱导PC12细胞表达HIF-1α,主要与MEK-ERK信号通路激活有关,可能是其促进分化的PC12生长的原因。