OBJECTIVE: To investigate the inhibitory effect of genistein on activation of hepatic stellate cells (HSCs) and the role of the autophagy pathway regulated by PPAR-γ in mediating this effect. METHODS: Cultured HSC-T6 cells were exposed to different concentrations of genistein for 48 h, and HSC activation was verified by detecting the expressions of -SMA and 1(I) collagen; autophagy activation in the cells was determined by detecting the expressions of LC3-II and p62 using Western blotting. The autophagy inhibitor 3-MA was used to confirm the role of autophagy in genistein-induced inhibition of HSC activation. A PPAR-γ inhibitor was used to explore the role of PPAR-γ in activating autophagy in the HSCs. RESULTS: Genistein at concentrations of 5 and 50 μmol/L significantly inhibited the expressions of -SMA and 1(I) collagen ( < 0.05), markedly upregulated the expressions of PPAR-γ and the autophagy-related protein LC3-II ( < 0.05) and significantly down-regulated the expression of the ubiqutin-binding protein p62 ( < 0.05) in HSC-T6 cells. The cells pretreated with 3-MA prior to genistein treatment showed significantly increased protein expressions of -SMA and 1(I) collagen compared with the cells treated with genistein only ( < 0.05). Treatment with the PPAR-γ inhibitor obviously lowered the expression of LC3-II and enhanced the expression p62 in genistein-treated HSC-T6 cells, suggesting the activation of the autophagy pathway. CONCLUSIONS PPAR-γ- regulated autophagy plays an important role in mediating genistein-induced inhibition of HSC activation .
Anticarcinogenic Agents ; pharmacology ; Autophagy ; Collagen Type I ; Genistein ; pharmacology ; Hepatic Stellate Cells ; Humans ; PPAR gamma ; physiology

Capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide) is the main component in hot peppers, including red chili peppers, jalapenos, and habanero, belonging to the genus Capsicum. Capsaicin is a potent antioxidant that interferes with free radical activities. In the present study, the possible protective effect of capsaicin was studied against methyl methanesulphonate (MMS) induced toxicity in third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg. The third instar was allowed to feed on the diet having different doses of capsaicin and MMS separately and in combination. The results suggested that the exposure of third instar larvae to the diet having MMS alone showed significant hsp70 expression as well as tissue DNA and oxidative damage, whereas the larvae feed on the diet having MMS and capsaicin showed a decrease in the toxic effects for 48-h of exposure. In conclusion, capsaicin showed a dose-dependent decrease in the toxic effects induced by MMS in the third instar larvae of transgenic Drosophila melanogaster.
Acetylcholinesterase ; metabolism ; Animals ; Animals, Genetically Modified ; Anticarcinogenic Agents ; pharmacology ; Capsaicin ; pharmacology ; DNA Damage ; drug effects ; Drosophila melanogaster ; drug effects ; Larva ; drug effects ; Methyl Methanesulfonate ; antagonists & inhibitors

Mangosteen (Garcinia mangostana Linn.) is a well-known tropical tree indigenous to Southeast Asia. Its fruit's pericarp abounds with a class of isoprenylated xanthones which are referred as mangostins. Numerous in vitro and in vivo studies have shown that mangostins and their derivatives possess diverse pharmacological activities, such as antibacterial, antifungal, antimalarial, anticarcinogenic, antiatherogenic activities as well as neuroprotective properties in Alzheimer's disease (AD). This review article provides a comprehensive review of the pharmacological activities of mangostins and their derivatives to reveal their promising utilities in the treatment of certain important diseases, mainly focusing on the discussions of the underlying molecular targets/pathways, modes of action, and relevant structure-activity relationships (SARs). Meanwhile, the pharmacokinetics (PK) profile and recent toxicological studies of mangostins are also described for further druggability exploration in the future.
Animals ; Anti-Infective Agents ; pharmacology ; Anticarcinogenic Agents ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Antioxidants ; pharmacology ; Cardiovascular Agents ; pharmacology ; Fruit ; chemistry ; Garcinia mangostana ; chemistry ; Humans ; Neuroprotective Agents ; pharmacology ; Phytotherapy ; Plant Extracts ; pharmacology ; Protective Agents ; pharmacology ; Xanthones ; pharmacology

To study the role and molecular mechanism of epigallocatechin gallate (EGCG) in reversing drug-resistance to 5-fluorouracil (5-FU) in gastric cancer drug-resistant cell line SGC-7901/5-FU.

METHODSDrug-resistance gastric cancer cell line (SGC-7901/5-FU) was established by high doses of repeated impact joint drug concentration increment methods. The cell viability of the parent cell line and the drug-resistance cell line were determined by standard MTT assay. Cell survival rate of drug-resistance was calculated by the formula [(Aof the treatment group / Aof the control group) × 100%]. Cell half inhibitory concentration (IC) and resistance index (RI) were calculated by the Graphpad prime 6.0 software(RI=ICvalue of drug-resistance cells / ICvalue of parent cells). The apoptosis rate of SGC-7901/5-FU cells was quantified by flow cytometry after staining with annexin-V and PI. Western blot was used to detect the protein expression of drug-resistance-related proteins (ABCG2, P-gp, MDR-1 and GST-π) and apoptosis-related proteins (PARP, Survivin, Bax and bcl-2).

RESULTSICvalue was significantly increased in drug-resistant cells compared with parental cells [(64.7±3.9) mg/L and (4.1±0.3) mg/L, respectively, t=26.46, P=0.000], and the RI was 15.6. Proliferation activity in the drug-resistant cells was higher than that in parental cells at different 5-FU concentrations (all P<0.05). In drug-resistant cells, the ICvalue of 5-FU combined with EGCG group obviously decreased compared with 5-FU group [(7.3±0.1) mg/L and (63.1±1.4) mg/L respectively, t=40.84, P=0.000], and the RI was 0.12. Proliferation activity in drug-resistant cells was significantly decreased after EGCG treatment at different 5-FU concentrations (all P<0.05). Cell apoptosis rates in control group, 5-FU group, EGCG group and 5-FU combined with EGCG group were (3.0±1.0)%, (7.0±1.3)%, (6.0±1.2)% and (18.0±1.4)%, while apoptosis rate in 5-FU combined with EGCG group was significantly higher than those of other 3 groups(F=129.5, P=0.000). Western blot revealed that after EGCG treatment, the expression levels of drug-resistance-related proteins (ABCG2, P-gp, MDR-1 and GST-π) in the drug-resistant cell line SGC-7901/5-FU decreased significantly; the expression levels of apoptosis marker protein PARP and pro-apoptotic protein Bax increased significantly; and the expression levels of anti-apoptotic protein Survivin and Bcl-2 decreased significantly (all P<0.05).

CONCLUSIONEGCG can reduce the resistance of gastric cancer resistant cell line SGC-7901/5-FU, whose role may be via the inhibition of the expression of drug-resistance-related proteins, and the elevation of the protein expression ratio of PARP/Survivin and Bax/Bcl-2.

Anticarcinogenic Agents ; pharmacology ; Apoptosis ; Apoptosis Regulatory Proteins ; Catechin ; analogs & derivatives ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; Cell Survival ; Drug Resistance, Neoplasm ; Fluorouracil ; pharmacology ; Humans ; Stomach Neoplasms ; drug therapy ; pathology ; bcl-2-Associated X Protein

OBJECTIVEChemopreventive approach with natural products, particularly plants and plant-derived ones, is receiving increasing attention for their effective role against cancer without any palpable side effects. In this study, efficacy of ethanolic extract of Ruta graveolens (RG) on skin melanoma cells (A375) in vitro and on 7,12-dimethylbenz(a)anthracene (DMBA)-induced skin cancer in vivo has been tested in Swiss albino mice.

METHODSStudies on cell viability, apoptosis and autophagy induction were conducted in vitro. To check apoptosis, assays like alteration in mitochondrial membrane potential, annexin V-fluorescein isothiocyanate/propidium iodide assay and immunoblot were performed. Fluorescence microscopic and immunoblot assays were performed to confirm autophagy induction. The effects of RG were determined by evaluating body weight, tumor incidence, tumor volume and tumor burden in mice. Enzymatic and non-enzymatic antioxidant status was assessed. The role of some relevant signaling proteins was also analyzed.

RESULTSRG caused death of A375 cells through induction of caspase 3-mediated apoptosis and Beclin-1-associated autophagy. Moreover, RG administration (75 mg/kg body weight) which showed no acute or chronic toxicity, showed significant reduction in the skin tumor burden of DMBA-painted mice. RG also demonstrated potent anti-lipid peroxidative and antioxidant functions during the course of skin cancer induction by DMBA.

CONCLUSIONChemopreventive potential of RG was demonstrated from overall results of this study, indicating its possible use in therapeutic formulation of an effective drug to treat skin cancer.

9,10-Dimethyl-1,2-benzanthracene ; Animals ; Anticarcinogenic Agents ; pharmacology ; Apoptosis ; drug effects ; Autophagy ; drug effects ; Cell Line, Tumor ; DNA Damage ; Humans ; Melanoma ; drug therapy ; pathology ; Mice ; Phytotherapy ; Plant Extracts ; pharmacology ; Ruta ; Skin Neoplasms ; drug therapy ; pathology

Induction of apoptosis in target cells is a key mechanism by which chemotherapy promotes cell killing. The purpose of this study was to determine whether Indole-3-Carbinol (I3C) and Genistein in combination with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induce apoptosis in endometrial cancer cell (Ishikawa) and to assess apoptotic mechanism. The MTT assay and flow cytometry were performed to determine cell viability and cell cycle. The induction of apoptosis was measured by caspase-3 activity test, DNA fragmentation assay, annexin V binding assay and western blot analysis. There was no effect in cell growth inhibition and cell cycle progression alone or in two-combination. However, the treatment of I3C and Genistein followed by TRAIL showed significant cell death and marked increase in sub-G1 arrest. Three-combination treatment revealed elevated expression of DR4, DR5 and cleaved forms of caspase-3, caspase-8, PARP. The Flip was found down regulated. Moreover, increase in caspase-3 activity and DNA fragmentation indicated the induction of apoptosis. The results indicate that I3C and Genistein with TRAIL synergistically induced apoptosis via death receptor dependent pathway. Our findings might provide a new insight into the development of novel combination therapies against endometrial cancer.
Anticarcinogenic Agents/*pharmacology ; Apoptosis/*drug effects ; Caspase 3/metabolism ; Caspase 8/metabolism ; Cell Line, Tumor ; Drug Synergism ; Endometrial Neoplasms/metabolism/pathology ; Female ; G1 Phase Cell Cycle Checkpoints/drug effects ; Genistein/*pharmacology ; Humans ; Indoles/*pharmacology ; Poly(ADP-ribose) Polymerases/metabolism ; Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism ; TNF-Related Apoptosis-Inducing Ligand/*pharmacology

Gastrointestinal tract carcinoma is one of the leading causes of cancer-related death in China. Chemoprevention has been considered as a potential approach to control this type of disease. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that protects cells from oxidative/electrophilic stresses by activating the expression of a battery of cytoprotective genes through the antioxidant response element (ARE). Recently, Nrf2 has emerged as a novel target for chemoprevention. Several natural or synthetic chemicals, which activate Nrf2/ARE signaling pathway, have showed effect in animal models, and promises in many ongoing clinical trials. This review summarizes the recent findings on the regulation of Nrf2/ARE signaling pathway, and the developments in both preclinical and clinical studies.
Animals ; Anticarcinogenic Agents ; pharmacology ; Antioxidants ; metabolism ; Chemoprevention ; Digestive System Neoplasms ; genetics ; metabolism ; prevention & control ; Humans ; NF-E2-Related Factor 2 ; genetics ; metabolism ; Oxidative Stress ; drug effects ; Response Elements ; genetics ; Signal Transduction ; drug effects

Previously, we found that high doses of genistein show an inhibitory effect on uterine leiomyoma (UtLM) cell proliferation. In this study, using microarray analysis and Ingenuity Pathways Analysis(TM), we identified genes (up- or down-regulated, > or = 1.5 fold, P < or = 0.001), functions and signaling pathways that were altered following treatment with an inhibitory concentration of genistein (50 microg/ml) in UtLM cells. Downregulation of TGF-beta signaling pathway genes, activin A, activin B, Smad3, TGF-beta2 and genes related to cell cycle regulation, with the exception of the upregulation of the CDK inhibitor P15, were identified and validated by real-time RT-PCR studies. Western blot analysis further demonstrated decreased protein expression of activin A and Smad3 in genistein-treated UtLM cells. Moreover, we found that activin A stimulated the growth of UtLM cells, and the inhibitory effect of genistein was partially abrogated in the presence of activin A. Overexpression of activin A and Smad3 were found in tissue samples of leiomyoma compared to matched myometrium, supporting the contribution of activin A and Smad3 in promoting the growth of UtLM cells. Taken together, these results suggest that down-regulation of activin A and Smad3, both members of the TGF-beta pathway, may offer a mechanistic explanation for the inhibitory effect of a high-dose of genistein on UtLM cells, and might be potential therapeutic targets for treatment of clinical cases of uterine leiomyomas.
Activins/*genetics/metabolism/pharmacology ; Anticarcinogenic Agents/*pharmacology ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Cyclin-Dependent Kinase Inhibitor p15/genetics/metabolism ; Down-Regulation ; Female ; Genistein/*pharmacology ; Humans ; Leiomyoma/*metabolism ; Oligonucleotide Array Sequence Analysis ; Signal Transduction/drug effects ; Smad3 Protein/*genetics/metabolism ; Transforming Growth Factor beta/*genetics/metabolism ; Up-Regulation ; Uterine Neoplasms/*metabolism

BACKGROUNDPancreatic cancer is a devastating disease with the worst mortality rate. Therefore, a rational strategy for future drug development is critical. Genistein is a small, biologically active flavonoid that is found in high amounts in soy. This important compound supports a wide variety of biological activities, but is best known for its ability to inhibit cancer progression.

METHODSTranswell chamber assay was performed to determine the effect of genistein on the invasion of the human pancreatic cancer cell line Panc-1 induced by transforming growth factor-β1 (TGF-b1) in the different condition (5 ng/ml 24 hours and 10 ng/ml 48 hours); Reverse transcription-polymerase chain reaction (RT-PCR) was used to estimate the mRNA levels of urinary plasminogen activator (uPA), matrix metallopeptidase 2/9 (MMP-2/9), Smad4, E-Cadherin and Vimentin; Western blotting was used to detect the protein levels of uPA, E-Cadherin, ERK1/2, P38 and P-P38, and the activity of MMP-2/9 protein were detected by gelatin zymography assay method. Cells structure was observed and analyzed by microscopy.

RESULTSGenistein can inhibit effectively TGF-b1-induced invasion and metastasis in Panc-1 by Transwell assay, which is through regulating the mRNA and protein expression of uPA and MMP2, but not MMP9 by RT-PCR/Western blotting, and is positively correlated with the concentration of genistein. At the same time, genistein also could improve the progress of epithelial-mesenchymal transition (EMT) via morphology observation using light microscopy/transmission electron microscopy (TEM), which is mediated by the down-regulation of E-cadherin and the up-regulation of vimentin.

CONCLUSIONSTGF-b1 mediates EMT process via numerous intracellular signal transduction pathways. The potential molecular mechanisms are all or partly through Smad4-dependent and -independent pathways (p38 MAPK) to regulate the antitumor effect of genistein.

Anticarcinogenic Agents ; pharmacology ; Blotting, Western ; Cell Line, Tumor ; Epithelial-Mesenchymal Transition ; drug effects ; Genistein ; pharmacology ; Humans ; Microscopy, Electron, Transmission ; Pancreatic Neoplasms ; metabolism ; pathology ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; drug effects ; Smad4 Protein ; metabolism ; Transforming Growth Factor beta1 ; pharmacology

Sulforaphane (SFN) is a naturally occurring compound which is known to induce the phase II antioxidant genes via Nrf2 activation, although the underlying mechanism has not been fully elucidated. In this study, we investigated Nrf2 induction in response to SFN in human bronchial epithelial BEAS-2B cells and determined the signaling pathways involved in this process. SFN treatment reduced cell viability. Prior to cell death, intracellular reactive oxygen species (ROS) were generated at a high rate within a minute of commencing SFN treatment. Pretreatment with antioxidant N-acetylcysteine (NAC) blocked SFN-induced decrease in cell growth. Erk1/2 was activated within 30 min of SFN addition, whereas Akt phosphorylation did not significantly change until the first 8 hr after SFN treatment but then became substantially low until 48 hr. Inhibition of Erk1/2 phosphorylation attenuated SFN-induced loss of cell viability. Nrf2 protein levels in both nuclear and whole cell lysates were increased by SFN treatment, which was dependent on ROS production. Knockdown of Nrf2 with siRNA attenuated SFN-induced heme oxygenase-1 (HO-1) up-regulation. Induction of the Nrf2/HO-1 after SFN treatment was potently suppressed by pretreatment with NAC. Overall, our results indicate that SFN mediates antioxidative and antiproliferative responses by generating ROS in BEAS-2B cells.
Acetylcysteine/pharmacology ; Anticarcinogenic Agents/pharmacology ; Antioxidants/*pharmacology ; Bronchi/cytology/*drug effects/metabolism ; Cell Line ; Cell Proliferation/*drug effects ; Epithelial Cells/drug effects/metabolism ; Extracellular Signal-Regulated MAP Kinases/metabolism ; Free Radical Scavengers/pharmacology ; Heme Oxygenase-1/biosynthesis ; Humans ; NF-E2-Related Factor 2/biosynthesis/genetics ; Oxidative Stress/drug effects ; Proto-Oncogene Proteins c-akt/metabolism ; RNA Interference ; RNA, Small Interfering ; Reactive Oxygen Species/*metabolism ; Respiratory Mucosa/cytology/*drug effects/metabolism ; Signal Transduction/drug effects ; Thiocyanates/*pharmacology