Effects of Imatinib Combined With Everolimus on Mouse Pituitary Tumor Cell AtT-20.

Context: Pituitary adenoma is a clinical syndrome in which excessive production of pituitary corticotropin (ACTH). For ACTH tumor cells, researchers know little about the influence of the cell-cycle process on ACTH production and cell proliferation. Some research has shown that imatinib can induce apoptosis of tumor cells. Objective: The study intended to explore the effects and molecular mechanisms of imatinib combined with everolimus on AtT-20 cells in AtT-20 mouse pituitary tumors. Design: The research team performed a laboratory study using murine corticotropin tumor AtT-20 cells. Setting: The study took place at the Department of Neurosurgery at Renmin Hospital of the Hubei University of Medicine in Shiyan, Hubei, China. Intervention: The research team cultured the cells in AtT-20-cell-specific medium containing 100 µg/mL of streptomycin, 100 U/mL of penicillin, and 10% fetal bovine serum at 37°C and 5% CO2. The team divided the cells into a control group, a normal culture without the drug, and an intervention group, incubated for 24 hours with 1 µM of imatinib and 3 µM of everolimus when the cells grew to 40% confluence. Outcome Measures: The research team: (1) determined the effects of the combined drugs on cell viability using a methyl thiazolyl tetrazolium (MTT) assay; (2) detected the cell's mitochondrial membrane potential and LDH leakage using "sytox blue, 5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide," CBIC2(3) or JC-1, and lactate dehydrogenase (LDH) assay kits, respectively; (3) detected AtT-20 cell apoptosis using a "terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labeling" (TUNEL) kit; (4) analyzed the expression of protein kinase B (p-Akt), cAMP-response element binding protein (p-CREB), p27, p53, and cyclin E using a Western blot test; (5) detected the mRNA expression of opioid melanin procorticotropin (POMC)), caspase-3, and pituitary tumor transforming gene 1 (PTTG1) using reverse transcription-polymerase chain reaction (RT-PCR); (6) measure the concentration of adreno-cortico-tropic-hormone (ACTH) in the supernatant using an enzyme-linked immunoassay (ELISA) kit; and (7) assessed the cell cycle distribution using flow cytometry. Results: No differences existed in cell viability between the groups at the baseline (0 h) of the culture period (P > .05). Compared to the control group, the intervention group's: (1) cell viability was significantly lower at 4, 8, and 12 hours and postintervention at 16 hours (P < .001); (2) LDH concentration was significantly higher (P < .001); (3) mitochondrial membrane potential was significantly lower (P < .001); (4) apoptosis rate of TUNEL was significantly higher (P < .001 ); (5) expression of p-Akt, p-CREB phosphorylation, and cyclin E was significantly lower (P < .001), (6) expression of p27 and p53 protein was significantly higher (P < .001); (7) mRNA expression of POMC and PTTG1 were significantly lower (P < .001); (8) mRNA expression of caspase-3 was significantly higher (P < .001); (9) concentration of ACTH was lower (P < .001); and (10) percentage of cells in the G0/G1 phase was significantly higher, while the percentage of cells in the S phase was significantly lower (P < .05). Conclusions: Imatinib combined with everolimus can affect the AtT-20 cell cycle through the signaling pathway of the phosphatidylin-ositol-3-kinase (PI3K)/Akt/ protein kinase A (PKA) system and can inhibit cell proliferation and induce cell apoptosis. Therefore, Imatinib and everolimus may be an effective combination of candidates for drugs for mouse pituitary tumor.

Potential use of kiwifruit extract for treatment of melanoma.

Skin cancers are the most common cancers in the world and among the different types of skin cancers, melanoma is the deadliest and incidence is rising. Previous studies have shown promising in vitro and human evidence of kiwifruit exhibiting anti-cancer effects. This study was designed to investigate if kiwifruit extract (KE) has any effect on CRL-11147 melanoma cancer cells and to investigate the possible mechanisms behind the results. The effects of KE on CRL-11147 melanoma cell survival, proliferation, and apoptosis was investigated using clonogenic survival assay, cell proliferation, and caspase-3 activity kits. Potential anti-tumor molecular mechanisms were elucidated using RT-PCR and IHC. Addition of KE decreased CRL-11147 cell colonies percentages indicated by a decreased optical density value of cancer cells when compared to control. Furthermore, treatment with KE increased relative caspase-3 activity in cancer cells, which indicated increased apoptosis of cancer cells. The anti-proliferative effect of KE on cancer cells corresponded with decreased expression of the pro-proliferative molecule Cyclin E and CDK4, while increased expression of the pro-apoptotic molecule TRAILR1 corresponded with the pro-apoptotic effect. KE decreases CRL-11147 melanoma cell growth via downregulation of Cyclin E and CDK4 and upregulation in TRAILR1. Our study suggests a potential use for KE in treatment of melanoma.

Onosma bracteata Wall. induces G0/G1 arrest and apoptosis in MG-63 human osteosarcoma cells via ROS generation and AKT/GSK3ß/cyclin E pathway.

Onosma bracteata Wall. (Boraginaceae), commonly known as "gaozaban" is a highly valuable medicinal herb, useful in the treatment of body swellings, abdominal pain, eye-related problems, fever, and urinary calculi. The present study was performed to investigate the antioxidant properties of extract/fractions, viz. ethanol (Obeth) extract, hexane (Obhex) fraction, chloroform (Obcl) fraction, ethyl acetate (Obea) fraction, butanol (Obbu) fraction, and aqueous (Obaq) fraction isolated from O. bracteata. Obea fraction showed stronger free radical quenching ability in various antioxidant assays, as compared to the other fractions. Obea fraction with effective free radical-scavenging properties was further evaluated for the antiproliferative activity against human osteosarcoma MG-63, human neuroblastoma IMR-32, and human lung cancer A549 cell lines using MTT assay. Obea fraction showed strong cytotoxicity with GI50 value of 88.56, 101.61, and 112.7 µg/ml towards MG-63, IMR-32, and A549 cells respectively. Mechanistic studies revealed that Obea fraction in osteosarcoma MG-63 cells increased reactive oxygen species (ROS) level and reduced mitochondrial membrane potential. In the presence of Obea, the cells were found to be arrested in the G0/G1 phase in a dose-dependent manner which is also confirmed by the enhancement in the early apoptotic cell population in flow cytometer analysis. Western blotting demonstrated the decrease in expression of p-NFκB, COX-2, p-Akt, and Bcl-xL, whereas upregulation was observed in the expression of GSK-3ß, p53, caspase-3, and caspase-9 proteins. RT-qPCR studies revealed downregulation of Bcl-2, cyclin E, CDK2, and mortalin gene expression and upregulation in the expression of p53 genes. The antioxidant and cytotoxic potential of Obea was attributed to the presence of catechin, kaempferol, onosmin A, and epicatechin, as revealed by HPLC analysis. This is the first report regarding the antiproliferative potential of O. bracteata against osteosarcoma.

Garcinone C suppresses colon tumorigenesis through the Gli1-dependent hedgehog signaling pathway.

BACKGROUND: Although garcinone C, a natural xanthone derivative identified in the pericarp of Garcinia mangostana, has been demonstrated to exert different health beneficial activities in oxidative stress and ß-amyloid aggregation, the role of garcinone C in colon tumorigenesis has not been investigated. In addition, aberrant Hedgehog (Hh) signaling activation is associated with tumorigenesis including colon cancer. Here, we hypothesized that garcinone C can prevent colon tumorigenesis through regulating the Hh signaling pathway. METHOD: Colony formation assay and flow cytometry were used to evaluate the effect of garcinone C on the proliferation and cell cycle progression of colon cancer cells. Protein expression of cell cycle related markers and Hh/Gli1 signaling mediators were determined. The regulatory effect of orally administered garcinone C on the Hh/Gli1 signaling pathway and colon tumorigenesis was evaluated in an azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced colon cancer animal model. RESULTS: Garcinone C suppressed the proliferation of colon cancer cells, induced G0/G1 cell cycle arrest, as well as regulated the expression of cell cycle-related markers such as cyclin D1, cyclin E, CDK6, and p21. Garcinone C inhibited the expression of Gli1, a key mediator of Hedgehog signaling, and protein kinase B (AKT) phosphorylation in Smo-independent colon cancer cells. In the AOM/DSS-induced colon tumorigenesis model, garcinone C significantly inhibited tumor development, regulated the expression of cell cycle markers and Gli1, and reduced AKT phosphorylation in colon tumor tissues, which is consistent with our in vitro results. CONCLUSION: Garcinone C can suppress colon tumorigenesis in vitro and in vivo through Gli1-dependent non-canonical Hedgehog signaling, suggesting that it may serve as a potent chemopreventive agent against colon tumorigenesis.

Myricetin inhibits endometriosis growth through cyclin E1 down-regulation in vitro and in vivo.

Endometriosis is a benign gynecological condition prevalent among reproductive-aged women. Although active research and studies have been carried out to discover new drugs, surgery and hormone therapy are still the gold standard for endometriosis treatment. Nowadays, various flavonoids are considered long-term supplements for different diseases. Myricetin, a flavonol, has antiproliferative, anti- or pro-oxidant, and anticancer effects in gynecological diseases. Here, we reveal for the first time, to our knowledge, the antigrowth effects of myricetin in endometriosis. Myricetin inhibited cell proliferation and cell cycle progression of human VK2/E6E7 and End1/E6E7 cells and induced apoptosis, with the loss of mitochondrial membrane potential and accumulation of reactive oxygen species and calcium ions. Additionally, myricetin decreased the activation of AKT and ERK1/2 proteins, whereas it induced p38 activation in both cell lines. Moreover, myricetin decreased lesion size in the endometriosis mouse model via Ccne1 inhibition. Thus, myricetin has antiproliferative effects on endometriosis through cell cycle regulation.

Icariside II inhibits tumorigenesis via inhibiting AKT/Cyclin E/ CDK 2 pathway and activating mitochondria-dependent pathway.

Cervical cancer contributes largely in women cancer-related mortality. Herein, Icariside II, a flavonoid extracted from edible and pharmaceutical plant Epimedium brevicornum Maxim, exhibited significant anticancer activity on cervical cancer. At first, it was observed that Icariside II inhibited Hela cell proliferation at IC50 (9.2 µM) and the growth of Hela-originated xenografts in BALB/c nude mice. Next, we studied the underlying mechanisms of Icariside II from the aspects of cell growth and cell death. As for cell growth, Icariside II arrested cell cycle at G0/G1 phase through AKT/Cyclin E/CDK 2 from transcriptional and translational levels. As for cell death, Flow Cytometry and Immunofluorescence showed that Icariside II promoted cell death in a dose-dependet manner. And, Icariside II turned to activate the mitochondria-dependent pathway Caspase 9/Caspase 3 much more significantly than death receptor pathway Caspase 8/Caspase 3. Taken together, Icariside II presented anticancer effect on cervical cancer both in vitro and in vivo. Our study provides the evidence that Icariside II can be used as a suitable novel agent in cervical cancer treatment.

Adjuvant therapy with γ-tocopherol-induce apoptosis in HT-29 colon cancer via cyclin-dependent cell cycle arrest mechanism.

Resistance to chemotherapy with 5-fluorouracil (5-FU) in patients with colorectal cancer (CRC) is the major obstacle to reach the maximum efficiency of CRC treatment. Combination therapy has emerged as a novel anticancer strategy. The present study evaluates the cotreatment of γ-tocopherol and 5-FU in enhancing the efficacy of chemotherapy against HT-29 colon cancer cells. Cytotoxic effect of this combination was examined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and a synergistic effect was evaluated by a combination index technique. Nuclear morphology was studied via 4',6-diamidino-2-phenylindole staining and flow cytometric assays were conducted to identify molecular mechanisms of apoptosis and cell cycle progression. We investigated the expression of Cyclin D1, Cyclin E, Bax, and Bcl-2 by a quantitative real-time polymerase chain reaction. The IC50 values for 5-FU and γ-tocopherol were 21.8 ± 2.5 and 14.4 ± 2.6 µM, respectively, and also this combination therapeutic increased the percentage of apoptotic cells from 35% ± 2% to 40% ± 4% (P < .05). Furthermore, incubation HT-29 colon cells with combined concentrations of two drugs caused significant accumulation of cells in the subGsubG1 phase. Our results presented the combination therapy with 5-FU and γ-tocopherol as a novel therapeutic approach, which can enhance the efficacy of chemotherapy.

Ganoderiol F purified from Ganoderma leucocontextum retards cell cycle progression by inhibiting CDK4/CDK6.

This study was designed to purify molecules possess anti-cancer cell activity from the fruit body of Ganoderma leucocontextum. Bio-activity-guided purification and chromatographic separation of Ganoderma leucocontextum extract led to the enrichment of bioactive fractions and isolation of a single compound. The purified compound was identified as Ganoderiol F, which induced cancer cell death. In the in vivo experiments, we founded ethanol extract and ethyl acetate fraction inhibited tumor growth in the mice injected with 4T1 cells. We found that Ganoderiol F-mediated suppression of breast cancer cell viability occurred through cell cycle arrest. Ganoderiol F down-regulated expression of cyclin D, CDK4, CDK6, cyclin E and CDK2 and inhibited cell cycle progression arresting the cells in G1 phase. In addition, Ganoderiol F up-regulated pro-apoptotic Foxo3, down-regulated anti-apoptotic c-Myc, Bcl-2 and Bcl-w leading to apoptosis in human breast cancer cells MDA-MB-231. These results showed that c-Myc, cyclin D-CDK4/CDK6 and cyclin E-CDK2 are the central components of Ganoderiol F regulation of cell cycle progression. Hence Ganoderiol F may serve as a potential CDK4/CDK6 inhibitor for breast cancer therapy. Abbreviations: GLE: Ganoderma leucocontextum ethanol extract; GLEA: Ganoderma leucocontextum ethyl acetate fraction; GLPE: Ganoderma leucocontextum petroleum ether fraction; RP-HPLC: reversed-phase high-performance liquid chromatograph; DMEM: Dulbecco's modified Eagle's medium; FBS: fetal bovine serum; PAGE: polyacrylamide gel electrophoresis.

Shaping of Drosophila Neural Cell Lineages Through Coordination of Cell Proliferation and Cell Fate by the BTB-ZF Transcription Factor Tramtrack-69.

Cell diversity in multicellular organisms relies on coordination between cell proliferation and the acquisition of cell identity. The equilibrium between these two processes is essential to assure the correct number of determined cells at a given time at a given place. Using genetic approaches and correlative microscopy, we show that Tramtrack-69 (Ttk69, a Broad-complex, Tramtrack and Bric-à-brac - Zinc Finger (BTB-ZF) transcription factor ortholog of the human promyelocytic leukemia zinc finger factor) plays an essential role in controlling this balance. In the Drosophila bristle cell lineage, which produces the external sensory organs composed by a neuron and accessory cells, we show that ttk69 loss-of-function leads to supplementary neural-type cells at the expense of accessory cells. Our data indicate that Ttk69 (1) promotes cell cycle exit of newborn terminal cells by downregulating CycE, the principal cyclin involved in S-phase entry, and (2) regulates cell-fate acquisition and terminal differentiation, by downregulating the expression of hamlet and upregulating that of Suppressor of Hairless, two transcription factors involved in neural-fate acquisition and accessory cell differentiation, respectively. Thus, Ttk69 plays a central role in shaping neural cell lineages by integrating molecular mechanisms that regulate progenitor cell cycle exit and cell-fate commitment.

Lycopene treatment inhibits activation of Jak1/Stat3 and Wnt/ß-catenin signaling and attenuates hyperproliferation in gastric epithelial cells.

Helicobacter pylori (H pylori) colonizes the human stomach and increases the risk of gastric diseases including gastric cancer. H pylori increases reactive oxygen species (ROS), which activate Janus-activator kinase 1 (Jak1)/signal transducers and activators of transcription 3 (Stat3) in gastric epithelial cells. ROS mediate hyperproliferation, a hallmark of carcinogenesis, by activating Wnt/ß-catenin signaling in various cells. Lycopene is a potent antioxidant exhibiting anticancer effects. We hypothesized that lycopene may inhibit H pylori-induced hyperproliferation by suppressing ROS-mediated activation of Jak1/Stat3 and Wnt/ß-catenin signaling, and ß-catenin target gene expression in gastric epithelial cells. We determined cell viability, ROS levels, and the protein levels of phospho- and total Jak1/Stat3, Wnt/ß-catenin signaling molecules, Wnt-1, lipoprotein-related protein 5, and ß-catenin target oncogenes (c-Myc and cyclin E) in H pylori-infected gastric epithelial AGS cells. The Jak1/Stat3 inhibitor AG490 served as the control treatment. The significance of the differences among groups was calculated using the 1-way analysis of variance followed by Newman-Keuls post hoc tests. The results show that lycopene reduced ROS levels and inhibited Jak1/Stat3 activation, alteration of Wnt/ß-catenin multiprotein complex molecules, expression of c-Myc and cyclin E, and cell proliferation in H pylori-infected AGS cells. AG490 similarly inhibited H pylori-induced cell proliferation, alteration of Wnt/ß-catenin multiprotein complex molecules, and oncogene expression. H pylori increased the levels of Wnt-1 and its receptor lipoprotein-related protein 5; this increase was inhibited by either lycopene or AG490 in AGS cells. In conclusion, lycopene inhibits ROS-mediated activation of Jak1/Stat3 and Wnt/ß-catenin signaling and, thus, oncogene expression in relation to hyperproliferation in H pylori-infected gastric epithelial cells. Lycopene might be a potential and promising nutrient for preventing H pylori-associated gastric diseases including gastric cancer.

Alginate oligosaccharide alleviates enterotoxigenic Escherichia coli-induced intestinal mucosal disruption in weaned pigs.

Alginate oligosaccharide (AOS) is a non-toxic, non-immunogenic, non-carcinogenic and biodegradable product generated by depolymerisation of alginate, and exhibits various salutary properties. The present study was designed to evaluate whether AOS supplementation could attenuate enterotoxigenic Escherichia coli (ETEC)-induced intestinal mucosal injury in weaned pigs. Twenty-four weaned pigs were randomly assigned to three treatments: (1) non-challenged control; (2) ETEC-challenged control; and (3) ETEC challenge + AOS treatment (100 mg kg-1). On day 12, pigs in the non-challenged group were orally infused with sterilised Luria-Bertani culture while pigs in other groups were orally infused with ETEC (2.6 × 1011 colony-forming units). At 3 days after the challenge, all pigs were orally administered d-xylose at 0.1 g per kg body weight and then euthanised 1 h later to obtain serum and intestinal mucosa samples. Our results showed that ETEC infection both reduced (P < 0.05) the villus height and proportion of epithelial cells in the S phase and elevated (P < 0.05) the percentage of total apoptotic epithelial cells in the jejunum and ileum; these deleterious effects caused by ETEC were alleviated (P < 0.05) by supplemental AOS. Meanwhile, AOS ingestion attenuated (P < 0.05) not only the up-regulated tumour necrosis factor receptor 1 (TNFR1), cysteinyl aspartate-specific protease-3 (caspase-3), -8 and -9 transcriptions, as well as the enhanced caspase activities (caspase-3, -8 and -9), but also the down-regulated cyclin E1 and cyclin-dependent kinase 2 (CDK2) transcriptions in jejunal and ileal mucosae, caused by the ETEC challenge. In conclusion, it is possible that the protective effects of AOS against ETEC-induced intestinal mucosal disruption in weaned pigs are associated with the restrained enterocyte death, by reducing both mitochondria-dependent and TNFR1-dependent apoptosis and the accelerated enterocyte proliferation, via enhancing the cyclin E-CDK2 complex formation.

Rosmarinic Acid, a Component of Rosemary Tea, Induced the Cell Cycle Arrest and Apoptosis through Modulation of HDAC2 Expression in Prostate Cancer Cell Lines.

Rosmarinic acid (RA), a main phenolic compound contained in rosemary which is used as tea, oil, medicine and so on, has been known to present anti-inflammatory, anti-oxidant and anti-cancer effects. Histone deacetylases (HDACs) are enzymes that play important roles in gene expression by removing the acetyl group from histone. The aberrant expression of HDAC in human tumors is related with the onset of human cancer. Especially, HDAC2, which belongs to HDAC class I composed of HDAC 1, 2, 3 and 8, has been reported to be highly expressed in prostate cancer (PCa) where it downregulates the expression of p53, resulting in an inhibition of apoptosis. The purpose of this study is to investigate the effect of RA in comparison with suberoylanilide hydroxamic acid (SAHA), an HDAC inhibitor used as an anti-cancer agent, on survival and apoptosis of PCa cell lines, PC-3 and DU145, and the expression of HDAC. RA decreased the cell proliferation in cell viability assay, and inhibited the colony formation and tumor spheroid formation. Additionally, RA induced early- and late-stage apoptosis of PC-3 and DU145 cells in Annexin V assay and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, respectively. In western blot analysis, RA inhibited the expression of HDAC2, as SAHA did. Proliferating cell nuclear antigen (PCNA), cyclin D1 and cyclin E1 were downregulated by RA, whereas p21 was upregulated. In addition, RA modulated the protein expression of intrinsic mitochondrial apoptotic pathway-related genes, such as Bax, Bcl-2, caspase-3 and poly (ADP-ribose) polymerase 1 (PARP-1) (cleaved) via the upregulation of p53 derived from HDAC2 downregulation, leading to the increased apoptosis of PC-3 and DU145 cells. Taken together, treatment of RA to PCa cell lines inhibits the cell survival and induces cell apoptosis, and it can be used as a novel therapeutic agent toward PCa.

Panaxydol Derived from Panax ginseng Inhibits G1 Cell Cycle Progression in Non-small Cell Lung Cancer via Upregulation of Intracellular Ca2+ Levels.

Panaxydol, a polyacetylenic compound derived from Panax ginseng has been reported to suppress the growth of cancer cells. However, the molecular mechanisms underlying cell cycle arrest by this compound in non-small cell lung cancer (NSCLC) are unknown. Our study found that panaxydol treatment induced cell cycle arrest at G1 phase in NSCLC cells. The cell cycle arrest was accompanied by down-regulation of the protein expression of cyclin-dependent kinase (CDK) 2, CDK4, CDK6, cyclin D1 and cyclin E, and decrease in the phosphorylation of retinoblastoma (Rb) protein. Furthermore, up-regulation of cyclin-dependent kinase inhibitor (CDKI) p21CIP1/WAF1 and p27KIP1 was observed in panaxydol-treated NSCLC cells. In addition, panaxydol also induced accumulation of intracellular Ca2+ ([Ca2+]i). (Acetyloxy)methyl 2-({2-[(acetyloxy)methoxy]-2-oxoethyl}[2-(2-{2-[bis({2-[(acetyloxy)methoxy]-2-oxoethyl})amino]phenoxy}ethoxy)phenyl]amino)acetate (BAPTA-AM), the Ca2+ chelator, attenuated not only panaxydol-induced accumulation of [Ca2+]i, but also G1 cell cycle arrest and decrease of CDK6 and cyclin D1 protein expression level. These results demonstrated that the anti-proliferative effects of panaxydol were caused by cell cycle arrest, which is closely linked to the up-regulation of [Ca2+]i and represents a promising approach for the treatment of lung cancer.

Curcumin inhibits cell proliferation and motility via suppression of TROP2 in bladder cancer cells.

Bladder cancer (BC) has become a serious health prob-lem and represents the second most commonly diagnosed urological tumor. Curcumin is a principal active natural component of turmeric and has long been used in Asia as a traditional herbal medicine. Curcumin suppresses cell growth in various types of cancer, including BC, by regulating numerous molecular signaling pathways. The human trophoblast cell surface antigen 2 (Trop2) belongs to the tumor-associated calcium signal transducer gene family. Trop2 has been described as a cancer driver and is deregulated in various types of cancer. However, whether Trop2 is involved in curcumin-induced BC cell inhibition remains to be elucidated. The present study hypothesized that Trop2 may be a promising target of curcumin in BC cells. It was found that Trop2 was closely involved in curcumin-induced cell proliferation suppression, mobility inhibition, apoptosis, and cell cycle arrest in BC cells. Curcumin decreased the expression of Trop2 and its downstream target cyclin E1, and increased the level of p27. The overexpression of Trop2 enhanced the oncogenic activity of BC cells, whereas downregulation of the expression of Trop2 suppressed cell proliferation and mobility, increased apoptosis, and sensitized BC cells to curcumin treatment. Therefore, Trop2 may be a promising target of curcumin in BC cells and the inhibition of Trop2 may be an important method for the therapeutic management of patients with BC.

Total saponins of Albiziae Cortex show anti-hepatoma carcinoma effects by inducing S phase arrest and mitochondrial apoptosis pathway activation.

ETHNOPHARMACOLOGICAL RELEVANCE: Albiziae Cortex (AC) is a widely used traditional medicine in China. It is possess various properties to treat insomnia, traumatic injuries, diuresis, sthenia, and confusion. Total saponins of Albiziae Cortex (TSAC) are the most abundant bioactive components of AC, which were reported to show significant anti-tumor effects in vivo and in vitro. But the underlying mechanism of TSAC remained to be revealed. AIM OF STUDY: In this study, we investigated the anti-hepatoma carcinoma effects and the potential mechanism of TSAC in vivo and in vitro. MATERIALS AND METHODS: We first purified TSAC from crude extracts and characterized the major bioactive compounds by high performance liquid chromatography (HPLC). Effects of TSAC on viability of various hepatoma carcinoma cell lines were measured by MTT. Inhibition on cell proliferation was analysed using colony formation assay. Cell cycle distribution was revealed by flow cytometry. The apoptotic cells were observed by Hoechst 33258 staining and acridine orange (AO)/ethidium bromide (EB) double staining. Microstructures of apoptotic cells were examined by Transmission electron microscopy (TEM). The mitochondrial membrane potential were determined by JC-1 staining. Western blot was used to investigate the effects of TSAC on apoptosis-related proteins, B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax), and S-phase related protein cyclin A, cyclin E and cyclin-dependent kinases 2 (CDK2). Effects on tumor growth was assessed by H22-bearing ICR mice. RESULTS: TSAC significantly decreased the hepatoma carcinoma cell viability and inhibited HepG2 cell colony formation in a concentration-dependent manner. We also found that TSAC inhibited HepG2 cell growth via induction of S phase arrest. Further study showed that TSAC significantly down-regulated the expressions of cyclin A, cyclin E and CDK2 in HepG2 cells. Meanwhile, TSAC could effectively induce mitochondria-dependent caspase apoptosis pathway activation. Furthermore, TSAC increased the expression of pro-apoptotic protein Bax and decreased the expression of anti-apoptotic protein Bcl-2. In vivo assay showed that the anti-tumor effects of TSAC were significantly augmented without increasing toxicity in H22-bearing ICR mice. CONCLUSION: TSAC could inhibit cell proliferation through inducing S phase arrest and activate cell apoptosis via mitochondria-dependent apoptosis pathway. Therefore, TSAC could be a promising agent in clinical trials for anti-hepatoma carcinoma treatment.

Circulating exosomal microRNAs reveal the mechanism of Fructus Meliae Toosendan-induced liver injury in mice.

The toxicological mechanisms of liver injury caused by most traditional Chinese medicine (TCM) remain largely unknown. Due to the unique features, exosomal microRNAs (miRNAs) are currently attracting major interests to provide further insights into toxicological mechanisms. Thus, taking Fructus Meliae Toosendan as an example of hepatoxic TCM, this study aimed to elucidate its hepatotoxicity mechanisms through profiling miRNAs in circulating exosomes of Fructus Meliae Toosendan water extract (FMT)-exposed mice. Biological pathway analysis of the 64 differentially expressed exosomal miRNAs (DEMs) showed that hepatic dysfunction induced by FMT likely related to apoptosis, mitochondrial dysfunction, and cell cycle dysregulation. Integrated analysis of serum exosomal DEMs and hepatic differentially expressed mRNAs further enriched oxidative stress and apoptosis related pathways. In vitro validation studies for omics results suggested that FMT-induced DNA damage was mediated by generating intracellular reactive oxygen species, leading to cell apoptosis through p53-dependent mitochondrial damage and S-phase arrest. Nrf2-mediated antioxidant response was activated to protect liver cells. Moreover, serum exosomal miR-370-3p, the most down-regulated miRNA involving in these pathways, might be the momentous event in aggravating cytotoxic effect of FMT by elevating p21 and Cyclin E. In conclusion, circulating exosomal miRNAs profiling could contribute to deepen the understanding of TCM-induced hepatotoxicity.

Ethyl acetate fraction from methanol extraction of Vitis thunbergii var. taiwaniana induced G0 /G1 phase arrest via inhibition of cyclins D and E and induction of apoptosis through caspase-dependent and -independent pathways in human prostate carcinoma DU145 cells.

Vitis thunbergii var. taiwaniana (VTT) is a wild grape native to Taiwan, belonging to the Vitaceae family and Vitis genus, and widely used as folk herbal medicine. It is traditionally used for the treatment of diarrhea, hypertension, neuroprotection, jaundice, and arthritis. We used the wild-collected VTT and sterilized them to establish the plant tissue culture, and then took the leaves for DNA sequencing to determine its original base. We use methanol to extract VTT in four different solvents: 1-butanol, n-hexane, ethyl acetate, and water. These four preliminary extracts were used to treat human prostate cancer DU145 cells in vitro. We use the flow cytometry to check the cell survival situation. Finally, we found the ethyl acetate layer roughing product (referred VTEA) in human prostate cancer apoptotic effects of cell line DU-145. In the present studies, we use the crude extract of VTT to examine whether or not it can induce apoptosis of DU145 cells in vitro. Viability assays for extracts of VTT treatment showed that it had dose-dependent effect on human prostate cancer DU145 cells. We also found that the extract of VTT induces time-dependent mitochondrial and intrinsic-dependent apoptosis pathways. The in vitro cytotoxic effects were investigated by cell cycle analysis and the determination of apoptotic DNA fragmentation in DU145 cells. The cell cycle analysis showed that extracts of VTT induced a significant increase in the number of cells in G0 /G1 phase. The extract of VTT induced chromatin changes and apoptosis of DU145 cells also were confirmed by DAPI and PI staining that were measured by fluorescence microscopy and flow cytometry, respectively. Finally, the expression of relevant proteins was analyzed by Western blot analysis. These results promoted us to further evaluate apoptosis associated proteins and elucidate the possible signal pathway in DU-145 cells after treated with the extract of VTT.

Nutriproteomic Analysis of Hwangmaemok-Induced Antiangiogenic Effect Using Antibody-Arrayed Protein Chip Assay.

We investigated the antiangiogenic effects of Lindera obtusiloba Blume (Hwangmaemok, HMM), which is a plant in the Lauraceae family that is commonly used to treat colds and gastritis. Moreover, given that a recent study reported the inhibitory effects of HMM extract on cancer metastasis, we hypothesized that HMM extract might possess and antiangiogenic function. Thus, this study was conducted to investigate the effects of HMM extract on endothelial cell proliferation, migration, and neovascularization in chick chorioallantoic membrane (CAM), and investigated the molecular mechanism of antiangiogenesis using a ProteoChip-based proteomics technology. To examine the effects of HMM extracts on endothelial cell proliferation and migration, we conducted basic fibroblast growth factor (bFGF)-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration. To assess the molecular mechanism of the antiangiogenic effects of HMM extract, a ProteoChip-based forwarded phase antibody array was employed to identify the differential expression of cell cycle proteins in HMM-treated HUVECs. HMM extract inhibited bFGF-induced HUVEC proliferation and migration in a dose-dependent manner and CAM angiogenesis. The ProteoChip-based antibody microarray data showed upregulation of Nibrin/NBS1 and downregulation of Plk-1 and Cyclin E, which are involved in cell division and controlling the cell cycle in bFGF-induced HUVECs. These data suggest that HMM may be a potent antitumor medicinal herb. The present study demonstrates that the antiangiogenic effect of HMM may be due to suppression of endothelial cell proliferation and migration. Taken together, these results emphasize the potential to use HMM extract as a potent angiogenesis inhibitor to treat cancer.

10-Gingerol, a Phytochemical Derivative from "Tongling White Ginger", Inhibits Cervical Cancer: Insights into the Molecular Mechanism and Inhibitory Targets.

With the aim of evaluating anticancerous activities of 10-gingerol (10-G) against HeLa cells, it was purified and identified from "Tongling white ginger" by HSCCC, UPLC-TOF-MS/MS, and NMR analysis, respectively. 10-G inhibited the proliferation of HeLa cells at IC50 (29.19 µM) and IC80 (50.87 µM) with altered cell morphology, increased cytotoxicity, and arrested cell cycle in the G0/G1 phase. Most cell cycle related genes and protein expression significantly decreased, followed by a slight decrease in a few without affecting cyclin B1 and cyclin E1 (protein). Both death receptors significantly up-regulated and activated apoptosis indicators (caspase family). Furthermore, significant changes in mitochondria-dependent pathway markers were observed and led to cell death. 10-G led to PI3K/AKT inhibition and AMPK activation to induce mTOR-mediated cell apoptosis in HeLa cells. These results can be an asset to exploit 10-G with other medicinal plant derivatives for future applications.

[Insulin combined with selenium inhibit p38MAPK/CBP pathway and suppresses cardiomyocyte apoptosis in rats with diabetic cardiomyopathy].

Objective To investigate the effect of insulin in combination with selenium on p38-mitogen-activated protein kinase/CREB-binding protein (p38MAPK/CBP) pathway in rats with diabetic cardiomyopathy. Methods Fifty SD rats were randomly grouped into control group, diabetic cardiomyopathy (DCM) group, diabetic cardiomyopathy with insulin treatment (DCM-In) group, diabetic cardiomyopathy with selenium treatment (DCM-Se) group, and diabetic cardiomyopathy with insulin and selenium combination treatment (DCM-In-Se) group. Flow cytometry was used to analyze cell cycle. TUNEL staining was used to detect cardiomyocyte apoptosis. Western blotting was used to examine the levels of cyclin D1, cyclin E, Bax, Bcl-2, p38MAPK, p-p38MAPK, CBP and Ku70. Co-immunoprecipitation was used to examine the acetylation status of Ku70. Results Insulin in combination with selenium significantly inhibited cardiomyocyte apoptosis, increased Bcl-2 levels and decreased Bax, cyclin D1, cyclin E, p38MAPK, p-p38MAPK, CBP, Ku70 and acetylated Ku70 levels. Conclusion The combined treatment of insulin and selenium suppresses cardiomyocyte apoptosis by inhibiting p38MAPK/CBP pathway.