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Sample records for cell line hepg2

  1. Selection of scFvs specific for the HepG2 cell line using ribosome ...

    Indian Academy of Sciences (India)

    In order to isolate specific scFvs, recognizing HepG2 negative selection on a normal hepatocyte line WRL-68 was carried out before three rounds of positive selection on HepG2. After three rounds of panning, cell enzyme-linked immunosorbent assay (ELISA) showed that one of the scFvs had high affinity for the HepG2 cell ...

  2. Selection of scFvs specific for the HepG2 cell line using ribosome ...

    Indian Academy of Sciences (India)

    Madhsudhan

    was transcribed and translated in vitro using a rabbit reticulocyte lysate system. In order to isolate specific scFvs, recognizing HepG2 negative selection on a normal hepatocyte line WRL-68 was carried out before three rounds of positive selection on HepG2. After three rounds of panning, cell enzyme-linked immunosorbent ...

  3. Synergistic cytotoxicity and mechanism of caffeine and lysozyme on hepatoma cell line HepG2

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    Yang, Hongchao; Li, Jingjuan; Cui, Lin; Ren, Yanqing; Niu, Liying; Wang, Xinguo; Huang, Yun; Cui, Lijian

    2018-03-01

    The influences of caffeine, lysozyme and the joint application of them on the hepatoma cell line HepG2 proliferation inhibition and cell apoptosis were observed by 3-(4, 5-dimethyl-2-thiazyl)-2, 5-diphenyl-2H-tetrazolium bromide assay and Hoechst 33342, which showed the proliferation inhibition rate of the joint application on HepG2 cells was 47.21%, significantly higher than caffeine or lysozyme, and the joint application promoted the apoptosis of HepG2 cells obviously. Van't Hoff classical thermodynamics formula, the Föster theory of non-radiation energy transfer and fluorescence phase diagram were used to manifest that the process of lysozyme binding to caffeine followed a two-state model, which was spontaneous at low temperature driven by enthalpy change, and the predominant intermolecular force was hydrogen bonding or Van der Waals force to stabilize caffeine-lysozyme complex with the distance 5.86 nm. The attenuated total reflection-Fourier transform infrared spectra indicated that caffeine decreased the relative contents of α-helix and β-turn, which inferred the structure of lysozyme tended to be "loose". Synchronous fluorescence spectra and ultraviolet spectra supported the above conclusion. The amino acid residues in the cleft of lysozyme were exposed and electropositivity was increased attributing to the loose structure, which were conducive to increasing caffeine concentration on the HepG2 cell surface by electrostatic interaction to show synergistic effect. The great quantities of microvilli on the liver cancer cell membrane surface, is beneficial for the lysozyme-caffeine compound to aggregate on cell surface to increase the concentration of caffeine to play stronger physiological role by electrostatic effect.

  4. Synthesis and Evaluation of New Pyrazoline Derivatives as Potential Anticancer Agents in HepG-2 Cell Line

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    Weijie Xu

    2017-03-01

    Full Text Available Cancer is a major public health concern worldwide. Adverse effects of cancer treatments still compromise patients’ quality of life. To identify new potential anticancer agents, a series of novel pyrazoline derivatives were synthesized and evaluated for cytotoxic effects on HepG-2 (human liver hepatocellular carcinoma cell line and primary hepatocytes. Compound structures were confirmed by 1H-NMR, mass spectrometry, and infrared imaging. An in vitro assay demonstrated that several compounds exerted cytotoxicity in the micromolar range. Benzo[b]thiophen-2-yl-[5-(4-hydroxy-3,5-dimethoxy-phenyl-3-(2-hydroxy-phenyl-4,5-dihydo-pyrazol-1-yl]-methanone (b17 was the most effective anticancer agent against HepG-2 cells owing to its notable inhibitory effect on HepG-2 with an IC50 value of 3.57 µM when compared with cisplatin (IC50 = 8.45 µM and low cytotoxicity against primary hepatocytes. Cell cycle analysis and apoptosis/necrosis evaluation using this compound revealed that b17 notably arrested HepG-2 cells in the G2/M phase and induced HepG-2 cells apoptosis. Our findings indicate that compound b17 may be a promising anticancer drug candidate.

  5. Synergistic complex from plants Solanaceae exhibits cytotoxicity for the human hepatocellular carcinoma cell line HepG2.

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    Schwarzlin, Romina; Pušenjak, Nika; Makuc, Damjan; Križman, Mitja; Vovk, Irena; Plavec, Janez; Švajger, Urban

    2016-10-18

    It had been demonstrated that sugars from various plants can act as potent agents, which induce apoptosis of cancer cells. Using HPLC, we fractionated a mixture of two plant extracts from the plant family Solanaceae, namely Capsicum chinense and the plant family Amaryllidaceae namely Allium sativum. We evaluated the effect of different fractions on apoptosis of HepG2 cell line. The most effective fraction was further studied to determine its molecular composition using mass spectrometry (MS) and NMR. We further evaluated the effect of determined molecular composition found in the selected fraction by using a mixture of commercially available substances, which were found in the fraction and tested its pro-apoptotic effect on HepG2 cells. To get some insight into potential apoptotic mechanisms we studied caspase-3 activity and mitochondrial integrity in treated cells. Out of 93 fractions obtained by HPLC from the plant extract we found HPLC fraction 10 (10 min elution) was the most effective. MS and NMR studies revealed high presence of cellobiose together with vitamin C, sulphur (S) and trace amounts of selenium (Se). HPLC fraction 10 triggered apoptosis of HepG2 within 3 h in the 0.01-1.0 mg/mL concentration range. Furthermore, a mixture of pure cellobiose, vitamin C, S and Se (complex cellobiose/C/S/Se) had a very similar capacity in inducing apoptosis of HepG2 cells compared to HPLC fraction 10. Complex cellobiose/C/S/Se was capable of inducing caspase-3 activity and led to loss of mitochondrial integrity. The capacity of cellobiose alone to induce apoptosis of HepG2 was approximately 1000-fold lower compared to complex cellobiose/C/S/Se. In this study we present the highly synergistic effect of a unique complex consisting of cellobiose, vitamin C, sulphur and selenium on triggering the apoptosis of human hepatocellular carcinoma (HepG2) cell line.

  6. Garcinia dulcis Fruit Extract Induced Cytotoxicity and Apoptosis in HepG2 Liver Cancer Cell Line

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    Mohd Fadzelly Abu Bakar

    2015-01-01

    Full Text Available Garcinia dulcis or locally known in Malaysia as “mundu” belongs to the family of Clusiaceae. The study was conducted to investigate the anticancer potential of different parts of G. dulcis fruit extracts and their possible mechanism of action in HepG2 liver cancer cell line. MTT assay showed that the peel, flesh, and seed extracts of G. dulcis induced cytotoxicity in HepG2 cell line with IC50 values of 46.33 ± 4.51, 38.33 ± 3.51, and 7.5 ± 2.52 µg/mL, respectively. The flesh extract of G. dulcis induced cell cycle arrest at sub-G1 (apoptosis phase in a time-dependent manner. Staining with Annexin V-FITC and propidium iodide showed that 41.2% of the cell population underwent apoptosis after 72 hours of exposure of the HepG2 cell line to G. dulcis flesh extract. Caspase-3 has been shown to be activated which finally leads to the death of HepG2 cell (apoptosis. GC-MS analysis showed that the highest percentage of compound identified in the extract of G. dulcis flesh was hydroxymethylfurfural and 3-methyl-2,5-furandione, together with xanthones and flavonoids (based on literature, could synergistically contribute to the observed effects. This finding suggested that the flesh extract of G. dulcis has its own potential as cancer chemotherapeutic agent against liver cancer cell.

  7. Oroxylin A induced apoptosis of human hepatocellular carcinoma cell line HepG2 was involved in its antitumor activity

    International Nuclear Information System (INIS)

    Hu Yang; Yang Yong; You Qidong; Liu Wei; Gu Hongyan; Zhao Li; Zhang Kun; Wang Wei; Wang Xiaotang; Guo Qinglong

    2006-01-01

    We previously reported that wogonin, a flavonoid compound, was a potent apoptosis inducer of human hepatoma SMMC-7721 cells and murine sarcoma S180 cells. In the present study, the effect of oroxylin A, one wogonin structurally related flavonoid isolated from Scutellariae radix, on human hepatocellular carcinoma cell line HepG2 was examined and molecular mechanisms were also investigated. Oroxylin A inhibited HepG2 cell proliferation in a concentration- and time-dependent manner measured by MTT-assay. Treatment with an apoptosis-inducing concentration of oroxylin A caused typical morphological changes and apoptotic blebbing in HepG2 cells. DNA fragmentation assay was used to examine later apoptosis induced by oroxylin A. FACScan analysis revealed a dramatic increase in the number of apoptotic and G 2 /M phase arrest cells after oroxylin A treatment. The pro-apoptotic activity of oroxylin A was attributed to its ability to modulate the concerted expression of Bcl-2, Bax, and pro-caspase-3 proteins. The expression of Bcl-2 protein and pro-caspase-3 protein was dramatically decreased after treatment with oroxylin A. These results demonstrated that oroxylin A could effectively induce programmed cell death and suggested that it could be a promising antitumor drug

  8. Cytotoxic and apoptotic effects of six herbal plants against the human hepatocarcinoma (HepG2 cell line

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    Nonpunya Apiyada

    2011-10-01

    Full Text Available Abstract Background Six plants from Thailand were evaluated for their cytotoxicity and apoptosis induction in human hepatocarcinoma (HepG2 as compared to normal African green monkey kidney epithelial cell lines. Methods Ethanol-water crude extracts of the six plants were tested with neutral red assay for their cytotoxicity after 24 hours of exposure to the cells. Apoptotic induction was tested in the HepG2 cells with diamidino-2-phenylindole staining. DNA fragmentation, indicative of apoptosis, was analyzed with agarose gel electrophoresis. Alkylation, indicative of DNA damage, was also evaluated in vitro by 4-(4'-nitrobenzyl pyridine assay. Results The extract of Pinus kesiya showed the highest selectivity (selectivity index = 9.6 and potent cytotoxicity in the HepG2 cell line, with an IC50 value of 52.0 ± 5.8 μg/ml (mean ± standard deviation. Extract of Catimbium speciosum exerted cytotoxicity with an IC50 value of 55.7 ± 8.1 μg/ml. Crude extracts from Glochidion daltonii, Cladogynos orientalis, Acorus tatarinowii and Amomum villosum exhibited cytotoxicity with IC50 values ranging 100-500 μg/ml. All crude extracts showed different alkylating abilities in vitro. Extracts of P. kesiya, C. speciosum and C. orientalis caused nuclei morphological changes and DNA laddering. Conclusion The extracts of C. speciosum, C. orientalis and P. kesiya induced apoptosis. Among the three plants, P. kesiya possessed the most robust anticancer activity, with specific selectivity against HepG2 cells.

  9. Effect of bixin and norbixin on the expression of cytochrome P450 in HepG2 cell line.

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    Matuo, Míriam Cristina Sakuragui; de Oliveira Takamoto, Rafael Teruiti; Kikuchi, Irene Satiko; de Jesus Andreoli Pinto, Terezinha

    2013-08-01

    Bixin and norbixin are the main components of annatto, which is extracted from Bixa orellana and largely used as natural colorant in the food and pharmaceutical industries. Annatto can enhance CYP1A and CYP2B activity in rats; however, the inducer effect has not been investigated in human cell lines. In this study, the ability of bixin and norbixin to induce the cytochrome P450 (CYP) enzymes was assessed in HepG2 human hepatoma cell line. HepG2 cells were treated with bixin and norbixin, and the expression of the CYP genes quantified by real-time reverse transcription polymerase chain reaction (RT-PCR). Expression of CYP1A1 and CYP1A2 was significantly increased by bixin treatment, while CYP2B6, 2C9, 2E1 and 3A4 were unaffected. Cells were treated with norbixin showed no inducer effect. The results suggest that the inducer potential of annatto is attributed to bixin, but not to norbixin, despite their similarities in molecular structure. © 2013 International Federation for Cell Biology.

  10. Portulaca oleracea Seed Oil Exerts Cytotoxic Effects on Human Liver Cancer (HepG2) and Human Lung Cancer (A-549) Cell Lines.

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    Al-Sheddi, Ebtesam Saad; Farshori, Nida Nayyar; Al-Oqail, Mai Mohammad; Musarrat, Javed; Al-Khedhairy, Abdulaziz Ali; Siddiqui, Maqsood Ahmed

    2015-01-01

    Portulaca oleracea (Family: Portulacaceae), is well known for its anti-inflammatory, antioxidative, anti- bacterial, and anti-tumor activities. However, cytotoxic effects of seed oil of Portulaca oleracea against human liver cancer (HepG2) and human lung cancer (A-549) cell lines have not been studied previously. Therefore, the present study was designed to investigate the cytotoxic effects of Portulaca oleracea seed oil on HepG2 and A-549 cell lines. Both cell lines were exposed to various concentrations of Portulaca oleracea seed oil for 24h. After the exposure, percentage cell viability was studied by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT), neutral red uptake (NRU) assays, and cellular morphology by phase contrast inverted microscopy. The results showed a concentration-dependent significant reduction in the percentage cell viability and an alteration in the cellular morphology of HepG2 and A-549 cells. The percentage cell viability was recorded as 73%, 63%, and 54% by MTT assay and 76%, 61%, and 50% by NRU assay at 250, 500, and 1000 μg/ml, respectively in HepG2 cells. Percentage cell viability was recorded as 82%, 72%, and 64% by MTT assay and 83%, 68%, and 56% by NRU assay at 250, 500, and 1000 μg/ml, respectively in A-549 cells. The 100 μg/ml and lower concentrations were found to be non cytotoxic to A-549 cells, whereas decrease of 14% and 12% were recorded by MTT and NRU assay, respectively in HepG2 cells. Both HepG2 and A-549 cell lines exposed to 250, 500, and 1000 μg/ ml of Portulaca oleracea seed oil lost their normal morphology, cell adhesion capacity, become rounded, and appeared smaller in size. The data from this study showed that exposure to seed oil of Portulaca oleracea resulted in significant cytotoxicity and inhibition of growth of the human liver cancer (HepG2) and human lung cancer (A-549) cell lines.

  11. Proteome analysis of the effects of sorafenib on human hepatocellular carcinoma cell line HepG2.

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    Suo, Aili; Zhang, Mingxin; Yao, Yu; Zhang, Lingmin; Huang, Chen; Nan, Kejun; Zhang, Wanggang

    2012-09-01

    Sorafenib is a multi-target oral anticancer drug used as first-line treatment for patients with advanced human hepatocellular carcinoma (HCC). But the exact mechanism of sorafenib involved in HCC treatment is not clear yet. In this study, a comparative proteomic approach was performed to identify novel sorafenib-related proteins in HCC. Proteomes of HepG2 cells treated with sorafenib and the control (without sorafenib) were obtained by two-dimensional differential gel electrophoresis. Comprehensive analysis of proteins was focused on total protein spots to filtrate the different protein spots between the two groups. The differentially expressed proteins were identified by peptide mass fingerprinting with high-performance liquid chromatography-tandem mass spectrometry. Then, Western blot and immunohistochemistry were used to verify the expression of some candidate proteins. Results indicated that 19 protein spots were differentially expressed with significant changes, including 6 up-regulated proteins and 13 down-regulated proteins. It was confirmed by Western blot that expressions of Annexin A1 and cyclophilin A were down-regulated in sorafenib-treated HCC cell lines. Immunohistochemical study revealed their oncogenic role in HCC tissues. These observations might be novel findings leading to bring new insights into the exact mechanism of sorafenib and identify possible therapeutic targets.

  12. Tyramine-O-sulfate is produced and secreted by human hepatoma cells, line HepG2

    Energy Technology Data Exchange (ETDEWEB)

    Liu, M.C.; Yu, S.; Suiko, M.

    1987-05-01

    Human hepatoma cells, line HepG2, were metabolically labeled with (/sup 35/S)sulfate. The spent medium separated following 24 hr labeling was subjected to ultrafiltration using an Amicon Centricon unit. The filtrate obtained was analyzed by a two-dimensional separation procedure combining high-voltage electrophoresis and thin-layer chromatography. The autoradiograph taken from the cellulose thin-layer plate following the analysis revealed the presence of tyramine-O-(/sup 35/)sulfate in addition to tyrosine-O-(/sup 35/)sulfate. Using adenosine, 3'-phosphate, 5'-phospho(/sup 35/S)sulfate as the sulfate donor, it was shown that tyramine was actively sulfated to form tyramine-O-(/sup 35/S)sulfate as catalyzed by the sulfotransferase(s) present in dog liver homogenate. Attempts to decarboxylate tyrosine-O-sulfate to tyramine-O-sulfate using intrinsic p-tyrosine decarboxylase present in dog liver homogenate, however, were unsuccessful. Employing purified Streptococcus faecalis tyrosine decarboxylase, it was shown that L-tyrosine was actively decarboxylated to tyramine, whereas tyrosine-O-sulfate could not serve as a substrate.

  13. Tyramine-O-sulfate is produced and secreted by human hepatoma cells, line HepG2

    International Nuclear Information System (INIS)

    Liu, M.C.; Yu, S.; Suiko, M.

    1987-01-01

    Human hepatoma cells, line HepG2, were metabolically labeled with [ 35 S]sulfate. The spent medium separated following 24 hr labeling was subjected to ultrafiltration using an Amicon Centricon unit. The filtrate obtained was analyzed by a two-dimensional separation procedure combining high-voltage electrophoresis and thin-layer chromatography. The autoradiograph taken from the cellulose thin-layer plate following the analysis revealed the presence of tyramine-O-[ 35 ]sulfate in addition to tyrosine-O-[ 35 ]sulfate. Using adenosine, 3'-phosphate, 5'-phospho[ 35 S]sulfate as the sulfate donor, it was shown that tyramine was actively sulfated to form tyramine-O-[ 35 S]sulfate as catalyzed by the sulfotransferase(s) present in dog liver homogenate. Attempts to decarboxylate tyrosine-O-sulfate to tyramine-O-sulfate using intrinsic p-tyrosine decarboxylase present in dog liver homogenate, however, were unsuccessful. Employing purified Streptococcus faecalis tyrosine decarboxylase, it was shown that L-tyrosine was actively decarboxylated to tyramine, whereas tyrosine-O-sulfate could not serve as a substrate

  14. microRNA-mediated resistance to hypoglycemia in the HepG2 human hepatoma cell line

    International Nuclear Information System (INIS)

    Ueki, Satomi; Murakami, Yuko; Yamada, Shoji; Kimura, Masaki; Saito, Yoshimasa; Saito, Hidetsugu

    2016-01-01

    It is generally accepted that the energy resources of cancer cells rely on anaerobic metabolism or the glycolytic system, even if they have sufficient oxygen. This is known as the Warburg effect. The cells skillfully survive under hypoglycemic conditions when their circumstances change, which probably at least partly involves microRNA (miRNA)-mediated regulation. To determine how cancer cells exploit miRNA-mediated epigenetic mechanisms to survive in hypoglycemic conditions, we used DNA microarray analysis to comprehensively and simultaneously compare the expression of miRNAs and mRNAs in the HepG2 human hepatoma cell line and in cultured normal human hepatocytes. The hypoglycemic condition decreased the expression of miRNA-17-5p and -20a-5p in hepatoma cells and consequently upregulated the expression of their target gene p21. These regulations were also confirmed by using antisense inhibitors of these miRNAs. In addition to this change, the hypoglycemic condition led to upregulated expression of heat shock proteins and increased resistance to caspase-3-induced apoptosis. However, we could not identify miRNA-mediated regulations, despite using comprehensive detection. Several interesting genes were also found to be upregulated in the hypoglycemic condition by the microarray analysis, probably because of responding to this cellular stress. These results suggest that cancer cells skillfully survive in hypoglycemic conditions, which frequently occur in malignancies, and that some of the gene regulation of this process is manipulated by miRNAs. The online version of this article (doi:10.1186/s12885-016-2762-7) contains supplementary material, which is available to authorized users

  15. Hypocholesterolemic mechanism of phenolics-enriched extract from Moringa oleifera leaves in HepG2 cell lines

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    Peera Tabboon

    2016-04-01

    Full Text Available Previous studies have demonstrated the hypolipidemic activity of Moringa oleifera (MO leaves via lowering serum levels of cholesterol, but the mechanism of action is unknown. In this study, we demonstrated the hypocholesterolemic mechanism of a phenolics-enriched extract of Moringa oleifera leaf (PMO in HepG2 cells. When compared to the control treatment, PMO significantly decreased total intracellular cholesterol, inhibited the activity of HMG CoA reductase in a dosedependent manner and enhanced LDL receptor binding activity. Moreover, PMO also significantly increased the genetic expressions of HMG CoA reductase and LDL receptor.

  16. Effect of Genistein and 17-β Estradiol on the Viability and Apoptosis of Human Hepatocellular Carcinoma HepG2 cell line

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    Masumeh Sanaei

    2017-01-01

    Full Text Available Background: One of the most lethal cancers is hepatocellular carcinoma (HCC. Genistein (GE is a choice compound for treatment of certain types of cancer. Phytoestrogens are plant derivatives that bear a structural similarity to 17-β estradiol (E2 and act in a similar manner. They are a group of lipophillic plant compounds with tumorigenic and antitumorigenic effects. E2 has stimulatory and inhibitory effects on cancer cell lines. This study was designed to investigate the antiproliferative and apoptotic effects of GE and E2 on the HCC HepG2 cell line. Materials and Methods: HepG2 cells were cultured and treated with various concentrations of GE and E2 and then 3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphenyl-2H-tetrazolium bromideand flow cytometry assay were performed to determine cell viability and apoptosis. Results: GE and E2 induced apoptosis and inhibited cell growth significantly. Reduction of cell viability by 50% required 20 μM E2 for E2-treatment groups and 20 μMGE for GE-treatment groups. The percentage of the GE-treated apoptotic cells was reduced by about 35%, 42%, and 47% (P < 0.001 and that of E2-treated groups 34%, 39%, and 42% (P < 0.001 after 24, 48, and 72 h, respectively. Conclusions: Our experimental work clearly demonstrated that GE and E2 exhibited significant antiproliferative and apoptotic effects on human HCC HepG2 cells.

  17. Saponins isolated from Asparagus induce apoptosis in human hepatoma cell line HepG2 through a mitochondrial-mediated pathway

    Science.gov (United States)

    Ji, Y.; Ji, C.; Yue, L.; Xu, H.

    2012-01-01

    Objective Many scientific studies have shown that Asparagus officinalis has an antitumour effect and enhances human immunity, but the active components and the antitumour mechanisms are unclear. We investigated the effects of saponins isolated from Asparagus on proliferation and apoptosis in the human hepatoma cell line HepG2. Methods HepG2 cells were treated with varying concentrations of Asparagus saponins at various times. Using mtt and flow cytometry assays, we evaluated the effects of Asparagus saponins on the growth and apoptosis of HepG2 cells. Transmission electron microscopy was used to observe the morphology of cell apoptosis. Confocal laser scanning microscopy was used to analyze intracellular calcium ion concentration, mitochondrial permeability transition pore (mptp), and mitochondrial membrane potential (mmp). Spectrophotometry was applied to quantify the activity of caspase-9 and caspase-3. Flow cytometry was used to investigate the levels of reactive oxygen species (ros) and pH, and the expressions of Bcl2, Bax, CytC, and caspase-3, in HepG2 cells. Results Asparagus saponins inhibited the growth of HepG2 cells in a dose-dependent manner. The median inhibitory concentration (IC50) was 101.15 mg/L at 72 hours. The apoptosis morphology at 72 hours of treatment was obvious, showing cell protuberance, concentrated cytoplasm, and apoptotic bodies. The apoptotic rates at 72 hours were 30.9%, 51.7%, and 62.1% (for saponin concentrations of 50 mg/L, 100 mg/L, 200 mg/L). Treatment with Asparagus saponins for 24 hours increased the intracellular level of ros and Ca2+, lowered the pH, activated intracellular mptp, and decreased mmp in a dose-dependent manner. Treatment also increased the activity of caspase-9 and caspase-3, downregulated the expression of Bcl2, upregulated the expression of Bax, and induced release of CytC and activation of caspase-3. Conclusions Asparagus saponins induce apoptosis in HepG2 cells through a mitochondrial-mediated and caspase

  18. Wharton’s Jelly-derived Mesenchymal Stem Cells can Differentiate into Hepatocyte-like Cells by HepG2 Cell Line Extract

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    Maryam Borhani-Haghighi

    2015-03-01

    Full Text Available Background: Wharton’s jelly is an unlimited source of stem cells that can be used in cell therapy and tissue engineering without any ethical concern. It has been revealed the cell-free extract could be effective to induce cell differentiation. The objective of this study was to induce Wharton’s jelly-derived mesenchymal stem cells (MSCs into hepatocyte-like cells by premeabilization of the cells in the presence of HepG2 cell line extract. Methods: MSCs were isolated from the umbilical cord, CD marker profile and their differentiation potential into adipogenic and osteogenic lineages were determined. The cells were then, permeabilized by streptolysin O in the presence of HepG cell extract. The treated cells were cultured for 17 days. The cell phenotype was evaluated and the hepatocyte specific markers were detected by immunofluorescence and immunocytochemistry. The Periodic Acid Schiff (PAS reaction and the cellular uptake of indocyanine green were performed to evaluate the functional behavior of the differentiated cells. Results: The phenotype of extract-treated MSCs changed into a round or polygonal cells with few short processes and they could express high level of albumin, cytokeratin 18 and 19. The MSCs also could store glycogen and uptake and release indocyanine green. Conclusion: We demonstrated for the first time that Wharton’s jelly-derived MSCs could differentiate into hepatocyte-like cells by premeabilization of them in the presence of HepG2 cell extract. This study suggests a feasible method to differentiate MSCs into functional hepatocyte-like cells.

  19. Camel milk triggers apoptotic signaling pathways in human hepatoma HepG2 and breast cancer MCF7 cell lines through transcriptional mechanism.

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    Korashy, Hesham M; Maayah, Zaid H; Abd-Allah, Adel R; El-Kadi, Ayman O S; Alhaider, Abdulqader A

    2012-01-01

    Few published studies have reported the use of crude camel milk in the treatment of stomach infections, tuberculosis and cancer. Yet, little research was conducted on the effect of camel milk on the apoptosis and oxidative stress associated with human cancer. The present study investigated the effect and the underlying mechanisms of camel milk on the proliferation of human cancer cells using an in vitro model of human hepatoma (HepG2) and human breast (MCF7) cancer cells. Our results showed that camel milk, but not bovine milk, significantly inhibited HepG2 and MCF7 cells proliferation through the activation of caspase-3 mRNA and activity levels, and the induction of death receptors in both cell lines. In addition, Camel milk enhanced the expression of oxidative stress markers, heme oxygenase-1 and reactive oxygen species production in both cells. Mechanistically, the increase in caspase-3 mRNA levels by camel milk was completely blocked by the transcriptional inhibitor, actinomycin D; implying that camel milk increased de novo RNA synthesis. Furthermore, Inhibition of the mitogen activated protein kinases differentially modulated the camel milk-induced caspase-3 mRNA levels. Taken together, camel milk inhibited HepG2 and MCF7 cells survival and proliferation through the activation of both the extrinsic and intrinsic apoptotic pathways.

  20. Camel Milk Triggers Apoptotic Signaling Pathways in Human Hepatoma HepG2 and Breast Cancer MCF7 Cell Lines through Transcriptional Mechanism

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    Hesham M. Korashy

    2012-01-01

    Full Text Available Few published studies have reported the use of crude camel milk in the treatment of stomach infections, tuberculosis and cancer. Yet, little research was conducted on the effect of camel milk on the apoptosis and oxidative stress associated with human cancer. The present study investigated the effect and the underlying mechanisms of camel milk on the proliferation of human cancer cells using an in vitro model of human hepatoma (HepG2 and human breast (MCF7 cancer cells. Our results showed that camel milk, but not bovine milk, significantly inhibited HepG2 and MCF7 cells proliferation through the activation of caspase-3 mRNA and activity levels, and the induction of death receptors in both cell lines. In addition, Camel milk enhanced the expression of oxidative stress markers, heme oxygenase-1 and reactive oxygen species production in both cells. Mechanistically, the increase in caspase-3 mRNA levels by camel milk was completely blocked by the transcriptional inhibitor, actinomycin D; implying that camel milk increased de novo RNA synthesis. Furthermore, Inhibition of the mitogen activated protein kinases differentially modulated the camel milk-induced caspase-3 mRNA levels. Taken together, camel milk inhibited HepG2 and MCF7 cells survival and proliferation through the activation of both the extrinsic and intrinsic apoptotic pathways.

  1. Cytotoxic Activity of Origanum Vulgare L. on Hepatocellular Carcinoma cell Line HepG2 and Evaluation of its Biological Activity

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    Hazem S. Elshafie

    2017-08-01

    Full Text Available The potential of plant essential oils (EOs in anticancer treatment has recently received many research efforts to overcome the development of multidrug resistance and their negative side effects. The aims of the current research are to study (i the cytotoxic effect of the crude EO extracted from Origanum vulgare subsp hirtum and its main constituents (carvacrol, thymol, citral and limonene on hepatocarcinoma HepG2 and healthy human renal cells HEK293; (ii the antibacterial and phytotoxic activities of the above EO and its main constituents. Results showed that cell viability percentage of treated HepG2 by EO and its main constituents was significantly decreased when compared to untreated cells. The calculated inhibition concentration (IC50 values for HepG2 were lower than healthy renal cells, indicating the sort of selectivity of the studied substances. Citral is not potentially recommended as an anticancer therapeutic agent, since there are no significant differences between IC50 values against both tested cell lines. Results showed also that oregano EO and its main constituents have a significant antibacterial activity and a moderate phytotoxic effect. The current research verified that oregano EO and its main constituents could be potentially utilized as anticancer therapeutic agents.

  2. Protective potential [correction of potencial] of Euphorbia hirta against cytotoxicity induced in hepatocytes and a HepG2 cell line.

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    Brindha, D; Saroja, S; Jeyanthi, G P

    2010-01-01

    Medicinal plants play a key role in human health care. Frustration over the side effects of allopathic drugs has driven the medical world to take asylum in the plant kingdom for the treatment of various ailments. Euphorbia hirta belonging to the family of Euphorbiacae has been reported to possess antibacterial, antiviral, and anticancer activity. The aim of the present study was to investigate the protective effect of E. hirta against antitubercular drug-induced cytotoxicity in freshly isolated hepatocytes. The extent of cytotoxicity of the plant extracts was also analyzed using human liver derived HepG2 cell line by estimating the viability of cells (MTT assay). The alcoholic plant extract normalized the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), triacylglycerol (TAG), cholesterol, total protein, albumin, total and direct bilirubin, which were altered due to antitubercular drug intoxication. A dose-dependent increase in percent viability was observed when antitubercular drug exposed HepG2 cells were treated with different concentrations of plant extracts (125, 250, 500 and 1000 microg/mL) which were compared with a standard hepatoprotective drug silymarin. The highest percentage viability of HepG2 was observed at a concentration of 1000 microg/mL. The results suggest that E. hirta exerts protection against antitubercular drug-induced cytotoxicity in this vitro model system.

  3. Functional characterization of E2F3b in human HepG2 liver cancer cell line.

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    Lu, Yujia; Li, Wei

    2018-04-01

    E2F3 is a transcription factor that has been shown to be overexpressed in hepatocellular carcinoma (HCC). It is well-known that the E2F3 gene encodes two proteins E2F3a and E2F3b. Therefore, the functions of the two distinct isoforms need to be clarified separately. To characterize the function of E2F3b in HCC, the effects of ectopic expression of E2F3b on cell proliferation, cell cycle, apoptosis and gene expression were investigated. E2F3b promoted G1/S phase transition and markedly increased cell proliferation, but had minor effect on apoptosis. Microarray analyses identified 366 differentially expressed genes (171 upregulated and 195 downregulated) in E2F3b- overexpressing cells. Differential expression of 16 genes relevant to cell cycle and cell proliferation were further verified by real-time PCR. Six genes, including CDC2, CCNE1, ARF, MAP4K2, MUSK, and PAX2 were confirmed to be upregulated by more than twofold; one gene, CCNA2 was validated to be downregulated by more than twofold. We also confirmed that E2F3b increased the protein levels of both cyclin E and Arf but did not affect cyclin D1 protein. These results suggest that E2F3b functions as an important promoter for cell proliferation and plays important roles in transcriptional regulation in HepG2 liver cancer cells. © 2017 Wiley Periodicals, Inc.

  4. Toxicity profiles of four metals and 17 xenobiotics in the human hepatoma cell line HepG2 and the protozoa Tetrahymena pyriformis--a comparison.

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    Rudzok, Susanne; Krejči, Sebastian; Graebsch, Carolin; Herbarth, Olf; Mueller, Andrea; Bauer, Mario

    2011-04-01

    We performed an interspecies comparison for the human hepatoma cell line HepG2 and the eukaryotic single cell organism Tetrahymena pyriformis (T. pyriformis) for 17 xenobiotics with diverse structures and four metals. The cytotoxicity was assessed by four different cell viability assays (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide reduction (MTT), neutral red uptake (NRU), resazurin dye (AlamarBlue), 5-carboxyfluorescein diacetate acetoxymethyl ester (CFDA-AM)) for the HepG2 and by cell count and MTT for T. pyriformis. For HepG2 cells, the results revealed interassay variations depending on the compound. The highest assay conformity was found for the metal Hg(2+) and the fungicide prochloraz. The AlamarBlue assay was the most sensitive assay according to low-effect concentrations. By contrast, the NRU assay was comprised of more frequent whole concentration response relationships and was more susceptible to EC(50). For T. pyriformis the EC(50) values of the two applied assays displayed a high conformity (R(2) = 0.97). Comparing the EC(50) values obtained by the MTT assay for the two cell models, a direct correlation was absent for the xenobiotics and only present for the metals (Cd(2+), Cu(2+), and Ni(2+)). Moreover, the protozoa T. pyriformis displayed a 20 times higher sensitivity than the cell line. The highest interspecies difference of three log degrees was obtained for the polycyclic aromatic hydrocarbon fluoranthene. In addition, a correlation of the EC(50) values and octanol-water partition coefficient (log K(OW)) of the xenobiotics was performed. No correlation was found for HepG2, and a weak one for T. pyriformis. Interestingly, the interspecies difference of logarithmized EC(50) correlated positive with the log K(OW) (R(2) = 0.65). In conclusion, to obtain reliable evidence for human cytotoxicity, more than one viability/cytotoxicity assay had to be applied for cell lines. Second, the human hepatoma cell line was less affected by the

  5. Identification of MicroRNAs Involved in Growth Arrest and Apoptosis in Hydrogen Peroxide-Treated Human Hepatocellular Carcinoma Cell Line HepG2

    Directory of Open Access Journals (Sweden)

    Yuan Luo

    2016-01-01

    Full Text Available Although both oxidative stress and microRNAs (miRNAs play vital roles in physiological and pathological processes, little is known about the interactions between them. In this study, we first described the regulation of H2O2 in cell viability, proliferation, cycle, and apoptosis of human hepatocellular carcinoma cell line HepG2. Then, miRNAs expression was profiled after H2O2 treatment. The results showed that high concentration of H2O2 (600 μM could decrease cell viability, inhibit cell proliferation, induce cell cycle arrest, and finally promote cell apoptosis. Conversely, no significant effects could be found under treatment with low concentration (30 μM. miRNAs array analysis identified 131 differentially expressed miRNAs (125 were upregulated and 6 were downregulated and predicted 13504 putative target genes of the deregulated miRNAs. Gene ontology (GO analysis revealed that the putative target genes were associated with H2O2-induced cell growth arrest and apoptosis. The subsequent bioinformatics analysis indicated that H2O2-response pathways, including MAPK signaling pathway, apoptosis, and pathways in cancer and cell cycle, were significantly affected. Overall, these results provided comprehensive information on the biological function of H2O2 treatment in HepG2 cells. The identification of miRNAs and their putative targets may offer new diagnostic and therapeutic strategies for liver cancer.

  6. Measuring and modeling of binary mixture effects of pharmaceuticals and nickel on cell viability/cytotoxicity in the human hepatoma derived cell line HepG2

    International Nuclear Information System (INIS)

    Rudzok, S.; Schlink, U.; Herbarth, O.; Bauer, M.

    2010-01-01

    The interaction of drugs and non-therapeutic xenobiotics constitutes a central role in human health risk assessment. Still, available data are rare. Two different models have been established to predict mixture toxicity from single dose data, namely, the concentration addition (CA) and independent action (IA) model. However, chemicals can also act synergistic or antagonistic or in dose level deviation, or in a dose ratio dependent deviation. In the present study we used the MIXTOX model (EU project ENV4-CT97-0507), which incorporates these algorithms, to assess effects of the binary mixtures in the human hepatoma cell line HepG2. These cells possess a liver-like enzyme pattern and a variety of xenobiotic-metabolizing enzymes (phases I and II). We tested binary mixtures of the metal nickel, the anti-inflammatory drug diclofenac, and the antibiotic agent irgasan and compared the experimental data to the mathematical models. Cell viability was determined by three different methods the MTT-, AlamarBlue (registered) and NRU assay. The compounds were tested separately and in combinations. We could show that the metal nickel is the dominant component in the mixture, affecting an antagonism at low-dose levels and a synergism at high-dose levels in combination with diclofenac or irgasan, when using the NRU and the AlamarBlue assay. The dose-response surface of irgasan and diclofenac indicated a concentration addition. The experimental data could be described by the algorithms with a regression of up to 90%, revealing the HepG2 cell line and the MIXTOX model as valuable tool for risk assessment of binary mixtures for cytotoxic endpoints. However the model failed to predict a specific mode of action, the CYP1A1 enzyme activity.

  7. Effects of PARP-1 inhibitors AG-014699 and AZD2281 on proliferation and apoptosis of human hepatoma cell line HepG2

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    DU Senrong

    2015-06-01

    Full Text Available ObjectiveTo observe the inhibitory and pro-apoptotic effects of two poly(ADP-ribose polymerase (PARP-1 inhibitors, AG-014699 and AZD2281, on human hepatoma HepG2 cells and preliminarily explore the mechanism by which AG-014699 induces HepG2 cell apoptosis, and to provide a new therapeutic target for hepatoma. MethodsThe effects of different concentrations of AG-014699 and AZD2281 on HepG2 cell proliferation were determined by MTT assay. The cell apoptosis rate was measured by flow cytometry. The expression levels of caspase-3 and caspase-8 were measured by Western Blot. Inter-group comparison was made by t test. ResultsBoth AG-014699 and AZD2281 suppressed HepG2 cell proliferation in a time- and dose-dependent manner. However, the sensitivity of HepG2 cells to the two PARP-1 inhibitors was different. The half-maximal inhibitory concentrations of AG-014699 and AZD2281 at 48 h determined by MTT assay were about 20 μmol/L and 400 μmol/L, respectively. Flow cytometry and Western blot were not used to evaluate the apoptosis of HepG2 cells exposed to AZD2281 to which these cells were not sensitive. HepG2 cell apoptosis could be induced by 10, 30, and 50 μmol/L AG-014699, and the highest apoptosis rate at 48 h was significantly higher than that of the control group (3100%±2.13% vs 09%±0013%, P<0.01. Compared with those in the control group, the protein levels of caspase-3 and caspase-8 in HepG2 cells after 48-h exposure to 30, and 50 μmol/L AG-014699 increased. ConclusionThe two PARP-1 inhibitors AG-014699 and AZD2281 can inhibit the proliferation of HepG2 cells, which showed different sensitivities to the two inhibitors. AG-014699 can induce HepG2 cell apoptosis by up-regulating the protein expression of caspase-3 and caspase-8.

  8. Cytotoxicity of hydro-alcoholic extracts of Cucurbita pepo and Solanum nigrum on HepG2 and CT26 cancer cell lines

    Science.gov (United States)

    Shokrzadeh, M.; Azadbakht, M.; Ahangar, N.; Hashemi, A.; Saeedi Saravi, S. S.

    2010-01-01

    Plants are used worldwide for the treatment of diseases, and novel drugs continue to be developed through research from plants. There are more than 20,000 species of plants used in traditional medicines, and these are all potential reservoirs for new drugs. Cucurbita pepo has been used in traditional folk medicine to treat cold and alleviate ache. Previous pharmacological tests have shown that it possesses antiviral, anti-inflammatory, and analgesic effects. Also, Solanum nigrum has been used as a diuretic and an antipyretic agent and it has also been used to cure inflammation, edema, mastitis and hepatic cancer. In this investigation, cytotoxicity of specific concentrations of hydro-alcoholic extracts of C. pepo and S. nigrum was studied on normal [Chinese hamster ovarian cells (CHO) and rat fibroblast] and cancer (HepG2 and CT26) cell lines. The cytotoxic effects and IC50 of the extracts on the selected cell lines were studied followed by colonogenic assay method. The results showed that IC50 of S. nigrum extract was significantly lower than that of the C. pepo extract on all four cell lines (P < 0.05). On the other hand, IC50 of S. nigrum extract was significantly higher than the extract of Taxus baccata and Cisplatin, herbal and chemical control positive anticancer compounds, respectively, on all four cell lines (P < 0.05). As a result, it is concluded that the extract of S. nigrum has almost similar cytotoxicity to the extract of T. baccata on cancer cells. PMID:20931075

  9. Bactericidal application and cytotoxic activity of biosynthesized silver nanoparticles with an extract of the red seaweed Pterocladiella capillacea on the HepG2 cell line.

    Science.gov (United States)

    El Kassas, Hala Yassin; Attia, Azza Ahmed

    2014-01-01

    Nano-biotechnology is recognized as offering revolutionary changes in various fields of medicine. Biologically synthesized silver nanoparticles have a wide range of applications. Silver nanoparticles (AgNPs) were biosynthesized with an aqueous extract of Pterocladiella (Pterocladia) capillacea, used as a reducing and stabilizing agent, and characterized using UV-VIS spectroscopy, Fourier Transform Infra red (FT-IR) spectroscopy, transmission electron microscopy (TEM) and energy dispersive analysis (EDX). The biosynthesized AgNPs were tested for cytotoxic activity in a human hepatocellular carcinoma (HepG2) cell line cultured in Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum, 1% antibiotic and antimycotic solution and 2 mM glutamine. Bacterial susceptibility to AgNPs was assessed with Staphylococcus aureus, Bacillus subtilis [Gram+ve] and Pseudomonas aeruginosa and Escherichia coli [Gram-ve]. The agar well diffusion technique was adopted to evaluate the bactericidal activity of the biosynthesized AgNPs using Ampicillin and Gentamicin as gram+ve and gram-ve antibacterial standard drugs, respectively. The biosynthesized AgNPs were 11.4±3.52 nm in diameter. FT-IR analysis showed that carbonyl groups from the amino acid residues and proteins could assist in formation and stabilization of AgNPs. The AgNPs showed potent cytotoxic activity against the human hepatocellular carcinoma (HepG2) cell line at higher concentrations. The results also showed that the biosynthesized AgNPs inhibited the entire panel of tested bacteria with a marked specificity towards Bacillus subtillus. Cytotoxic activity of the biosynthesized AgNPs may be due to the presence of alkaloids present in the algal extract. Our AgNPs appear more bactericidal against gram-positive bacteria (B. subtillus).

  10. Plasmatic concentration of organochlorine lindane acts as metabolic disruptors in HepG2 liver cell line by inducing mitochondrial disorder

    Energy Technology Data Exchange (ETDEWEB)

    Benarbia, Mohammed el Amine [LUNAM Université, Angers (France); Inserm 1063, Angers (France); Macherel, David [LUNAM Université, Angers (France); UMR 1345 IRHS, Angers (France); Faure, Sébastien; Jacques, Caroline; Andriantsitohaina, Ramaroson [LUNAM Université, Angers (France); Inserm 1063, Angers (France); Malthièry, Yves, E-mail: yves.malthiery@univ-angers.fr [LUNAM Université, Angers (France); Inserm 1063, Angers (France)

    2013-10-15

    Lindane (LD) is a persistent environmental pollutant that has been the subject of several toxicological studies. However, concentrations used in most of the reported studies were relatively higher than those found in the blood of the contaminated area residents and effects of low concentrations remain poorly investigated. Moreover, effects on cell metabolism and mitochondrial function of exposure to LD have received little attention. This study was designed to explore the effects of low concentrations of LD on cellular metabolism and mitochondrial function, using the hepatocarcinoma cell line HepG2. Cells were exposed to LD for 24, 48 and 72 h and different parameters linked with mitochondrial regulation and energy metabolism were analyzed. Despite having any impact on cellular viability, exposure to LD at plasmatic concentrations led to an increase of maximal respiratory capacity, complex I activity, intracellular ATP and NO release but decreased uncoupled respiration to ATP synthesis and medium lactate levels. In addition, LD exposure resulted in the upregulation of mitochondrial biogenesis genes. We suggest that, at plasmatic concentrations, LD acts as a metabolic disruptor through impaired mitochondrial function and regulation with an impact on cellular energetic metabolism. In addition, we propose that a cellular assay based on the analysis of mitochondria function, such as described here for LD, may be applicable for larger studies on the effects of low concentrations of xenobiotics, because of the exquisite sensitivity of this organelle. - Highlights: Our data clearly demonstrated in HepG2 cells that exposure at plasmatic low concentrations of LD were able to: • Impair mitochondrial function • Caused alteration on nucleo-mitochondrial cross-talk • Increase nitric oxide release and protein nitration • Impair cellular energetic metabolism and lipid accumulation.

  11. Evaluation of Synergetic Anticancer Activity of Berberine and Curcumin on Different Models of A549, Hep-G2, MCF-7, Jurkat, and K562 Cell Lines

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    Acharya Balakrishna

    2015-01-01

    Full Text Available Ayurvedic system of medicine is using Berberis aristata and Curcuma longa herbs to treat different diseases including cancer. The study was performed to evaluate the synergetic anticancer activity of Berberine and Curcumin by estimating the inhibition of the cell proliferation by cytotoxicity assay using MTT method on specified human cell lines (A549, Hep-G2, MCF-7, Jurkat, and K562. All the cells were harvested from the culture and seeded in the 96-well assay plates at seeding density of 2.0 × 104 cells/well and were incubated for 24 hours. Test items Berberine with Curcumin (1 : 1, Curcumin 95% pure, and Berberine 95% pure were exposed at the concentrations of 1.25, 0.001, and 0.5 mg/mL, respectively, and incubated for a period of 48 hours followed by dispensing MTT solution (5 mg/mL. The cells were incubated at 37 ± 1°C for 4 hours followed by addition of DMSO for dissolving the formazan crystals and absorbance was read at 570 nm. Separate wells were prepared for positive control, controls (only medium with cells, and blank (only medium. The results had proven the synergetic anticancer activity of Berberine with Curcumin inducing cell death greater percentage of >77% when compared to pure curcumin with <54% and pure Berberine with <45% on average on all cell line models.

  12. Metformin affects the features of a human hepatocellular cell line (HepG2) by regulating macrophage polarization in a co-culture microenviroment.

    Science.gov (United States)

    Chen, Miaojiao; Zhang, Jingjing; Hu, Fang; Liu, Shiping; Zhou, Zhiguang

    2015-11-01

    Accumulating evidence suggests an association between diabetes and cancer. Inflammation is a key event that underlies the pathological processes of the two diseases. Metformin displays anti-cancer effects, but the mechanism is not completely clear. This study investigated whether metformin regulated the microenvironment of macrophage polarization to affect the characteristics of HepG2 cells and the possible role of the Notch-signalling pathway. RAW264.7 macrophages were cultured alone or co-cultured with HepG2 cells and treated with metformin. We analysed classical (M1) and alternative (M2) gene expression in RAW264.7 cells using quantitative real-time polymerase chain reaction. Changes in mRNA and protein expressions of Notch signalling in both cell types were also detected using quantitative real-time polymerase chain reaction and Western-blotting analyses. The proliferation, apoptosis and migration of HepG2 cells were detected using Cell Titer 96 AQueous One Solution Cell Proliferation Assay (MTS) (Promega Corporation, Fitchburg, WI, USA), Annexin V-FITC/PI (7SeaPharmTech, Shanghai, China) and the cell scratch assay, respectively. Metformin induced single-cultured RAW264.7 macrophages with an M2 phenotype but attenuated the M2 macrophage differentiation and inhibited monocyte chemoattractant protein-1 (MCP-1) secretion in a co-culture system. The co-cultured group of metformin pretreatment activated Notch signalling in macrophages but repressed it inHepG2 cells. Co-culture also promoted the proliferation and migration of HepG2 cells. However, along with the enhanced apoptosis, the proliferation and the migration of HepG2 cells were remarkably inhibited in another co-culture system with metformin pretreatment. Metformin can skew RAW264.7 macrophages toward different phenotypes according to changes in the microenvironment, which may affect the inflammatory conditions mediated by macrophages, induce apoptosis and inhibit the proliferation and migration of HepG2

  13. Cytotoxicity and Antioxidant Activity of Zingiber Officinale and 6-Gingerol on HepG2 Cells

    OpenAIRE

    Hanif, Harliansyah; Azian Murad, Noor; Wan Ngah, Wan Zurinah; Anum Mohd Yusof, Yasmin

    2008-01-01

    The present study was designed to compare the effects of ethanolic extract of ginger (Zingiber officinale) and its phenolic component [6]-Gingerol on viability, antiproliferation and apoptotic levels of human hepatoma cell lines (HepG2) and its antioxidant activity. HepG2 cells were cultured in Eagles minimum essential medium (EMEM) and the percentage of cell cytotoxicity was evaluated by tetrazolium salt (MTS) assay. Antiproliferation and apoptotic levels were measured by 5Bromo-2deoxyuridin...

  14. Hexachlorobenzene induces cell proliferation, and aryl hydrocarbon receptor expression (AhR) in rat liver preneoplastic foci, and in the human hepatoma cell line HepG2. AhR is a mediator of ERK1/2 signaling, and cell cycle regulation in HCB-treated HepG2 cells.

    Science.gov (United States)

    de Tomaso Portaz, Ana Clara; Caimi, Giselle Romero; Sánchez, Marcela; Chiappini, Florencia; Randi, Andrea S; Kleiman de Pisarev, Diana L; Alvarez, Laura

    2015-10-02

    Hexachlorobenzene (HCB) is a widespread environmental pollutant, and a liver tumor promoter in rodents. Depending on the particular cell lines studied, exposure to these compounds may lead to cell proliferation, terminal differentiation, or apoptosis. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is involved in drug and xenobiotic metabolism. AhR can also modulate a variety of cellular and physiological processes that can affect cell proliferation and cell fate determination. The mechanisms by which AhR ligands, both exogenous and endogenous, affect these processes involve multiple interactions between AhR and other signaling pathways. In the present study, we examined the effect of HCB on cell proliferation and AhR expression, using an initiation-promotion hepatocarcinogenesis protocol in rat liver and in the human-derived hepatoma cell line, HepG2. Female Wistar rats were initiated with a single dose of 100 mg/kg of diethylnitrosamine (DEN) at the start of the experiment. Two weeks later, daily dosing of 100 mg/kg HCB was maintained for 10 weeks. Partial hepatectomy was performed 3 weeks after initiation. The number and area of glutathione S-transferase-P (GST-P)-positive foci, in the rat liver were used as biomarkers of liver precancerous lesions. Immunohistochemical staining showed an increase in proliferating cell nuclear antigen (PCNA)-positive cells, along with enhanced AhR protein expression in hepatocytes within GST-P-positive foci of (DEN HCB) group, when compared to DEN. In a similar manner, Western blot analysis demonstrated that HCB induced PCNA and AhR protein expression in HepG2 cells. Flow cytometry assay indicated that the cells were accumulated at S and G2/M phases of the cell cycle. HCB increased cyclin D1 protein levels and ERK1/2 phosphorylation in a dose-dependent manner. Treatment of cells with a selective MEK1 inhibitor, prevented HCB-stimulatory effect on PCNA and cyclinD1, indicating that these effects

  15. PCBP-1 regulates alternative splicing of the CD44 gene and inhibits invasion in human hepatoma cell line HepG2 cells

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    Ge Changhui

    2010-04-01

    Full Text Available Abstract Background PCBP1 (or alpha CP1 or hnRNP E1, a member of the PCBP family, is widely expressed in many human tissues and involved in regulation of transcription, transportation process, and function of RNA molecules. However, the role of PCBP1 in CD44 variants splicing still remains elusive. Results We found that enforced PCBP1 expression inhibited CD44 variants expression including v3, v5, v6, v8, and v10 in HepG2 cells, and knockdown of endogenous PCBP1 induced these variants splicing. Invasion assay suggested that PCBP1 played a negative role in tumor invasion and re-expression of v6 partly reversed the inhibition effect by PCBP1. A correlation of PCBP1 down-regulation and v6 up-regulation was detected in primary HCC tissues. Conclusions We first characterized PCBP1 as a negative regulator of CD44 variants splicing in HepG2 cells, and loss of PCBP1 in human hepatic tumor contributes to the formation of a metastatic phenotype.

  16. Synthesis, characterization and dose dependent antimicrobial and anti-cancerous activity of phycogenic silver nanoparticles against human hepatic carcinoma (HepG2 cell line

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    N. Supraja

    2016-10-01

    Full Text Available In the present study silver nanoparticles (AgNPs were successfully synthesized using aqueous extract of sea weed, Gracilaria corticata. The aqueous callus extract (5% treated with 1 mM silver nitrate solution resulted in the formation of AgNPs and the surface plasmon resonance (SPR of the formed AgNPs was recorded at 405 nm using UV-Visible spectrophotometer. The molecules involved in the formation of AgNPs were identified by Fourier transform infrared spectroscopy (FT-IR, surface morphology was studied by using scanning electron microscopy (SEM, and X-ray diffraction spectroscopy (XRD was used to determine the crystalline structure. SEM micrograph clearly revealed the size of the AgNPs was in the range of 20–55 nm with spherical, hexagonal in shape and poly-dispersed nature. High positive Zeta potential (22.9 mV of formed AgNPs indicates the stability and XRD pattern revealed the crystal structure of the AgNPs by showing the Bragg’s peaks corresponding to (111, (200, (220 planes of face-centered cubic crystal phase of silver. The synthesized AgNPs exhibited effective anticancerous activity (at doses 6.25 and 12.5 µg/ml of AgNPs against human hepatic carcinoma cell line (HepG2.

  17. Camel Milk Triggers Apoptotic Signaling Pathways in Human Hepatoma HepG2 and Breast Cancer MCF7 Cell Lines through Transcriptional Mechanism

    OpenAIRE

    Korashy, Hesham M.; Maayah, Zaid H.; Abd-Allah, Adel R.; El-Kadi, Ayman O. S.; Alhaider, Abdulqader A.

    2012-01-01

    Few published studies have reported the use of crude camel milk in the treatment of stomach infections, tuberculosis and cancer. Yet, little research was conducted on the effect of camel milk on the apoptosis and oxidative stress associated with human cancer. The present study investigated the effect and the underlying mechanisms of camel milk on the proliferation of human cancer cells using an in vitro model of human hepatoma (HepG2) and human breast (MCF7) cancer cells. Our results showed t...

  18. Human Sulfatase-1 Improves the Effectiveness of Cytosine Deaminase Suicide Gene Therapy with 5-Fluorocytosine Treatment on Hepatocellular Carcinoma Cell Line HepG2 In Vitro and In Vivo

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    Xiao-Ping Yang

    2015-01-01

    Full Text Available Background: Human sulfatase-1 (Hsulf-1 is an endosulfatase that selectively removes sulfate groups from heparan sulfate proteoglycans (HSPGs, altering the binding of several growth factors and cytokines to HSPG to regulate cell proliferation, cell motility, and apoptosis. We investigated the role of combined cancer gene therapy with Hsulf-1 and cytosine deaminase/5-fluorocytosine (CD/5-FC suicide gene on a hepatocellular carcinoma (HCC cell line, HepG2, in vitro and in vivo. Methods: Reverse transcription polymerase chain reaction and immunohistochemistry were used to determine the expression of Hsulf-1 in HCC. Cell apoptosis was observed through flow cytometry instrument and mechanism of Hsulf-1 to enhance the cytotoxicity of 5-FC against HCC was analyzed in HCC by confocal microscopy. We also establish a nude mice model of HCC to address the effect of Hsulf-1 expression on the CD/5-FC suicide gene therapy in vivo. Results: A significant decrease in HepG2 cell proliferation and an increase in HepG2 cell apoptosis were observed when Hsulf-1 expression was combined with the CD/5-FC gene suicide system. A noticeable bystander effect was observed when the Hsulf-1 and CD genes were co-expressed. Intracellular calcium was also increased after HepG2 cells were infected with the Hsulf-1 gene. In vivo studies showed that the suppression of tumor growth was more pronounced in animals treated with the Hsulf-1 plus CD than those treated with either gene therapy alone, and the combined treatment resulted in a significant increase in survival. Conclusions: Hsulf-1 expression combined with the CD/5-FC gene suicide system could be an effective treatment approach for HCC.

  19. Gene Network Analysis of Glucose Linked Signaling Pathways and Their Role in Human Hepatocellular Carcinoma Cell Growth and Survival in HuH7 and HepG2 Cell Lines

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    Emmanuelle Berger

    2015-01-01

    Full Text Available Cancer progression may be affected by metabolism. In this study, we aimed to analyze the effect of glucose on the proliferation and/or survival of human hepatocellular carcinoma (HCC cells. Human gene datasets regulated by glucose were compared to gene datasets either dysregulated in HCC or regulated by other signaling pathways. Significant numbers of common genes suggested putative involvement in transcriptional regulations by glucose. Real-time proliferation assays using high (4.5 g/L versus low (1 g/L glucose on two human HCC cell lines and specific inhibitors of selected pathways were used for experimental validations. High glucose promoted HuH7 cell proliferation but not that of HepG2 cell line. Gene network analyses suggest that gene transcription by glucose could be mediated at 92% through ChREBP in HepG2 cells, compared to 40% in either other human cells or rodent healthy liver, with alteration of LKB1 (serine/threonine kinase 11 and NOX (NADPH oxidases signaling pathways and loss of transcriptional regulation of PPARGC1A (peroxisome-proliferator activated receptors gamma coactivator 1 target genes by high glucose. Both PPARA and PPARGC1A regulate transcription of genes commonly regulated by glycolysis, by the antidiabetic agent metformin and by NOX, suggesting their major interplay in the control of HCC progression.

  20. silver nanoparticles on liver cancer cells (HepG2

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    Ahmed I. El-Batal

    2018-01-01

    Full Text Available This study demonstrates a novel approach for the synthesis of silver nanoparticles (AgNPs against human liver cancer cell line (HepG2 using prodigiosin pigment isolated from Serratia marcescens. It further investigates the influence of various parameters such as initial pH, temperature, silver nitrate (AgNO 3 concentration, and prodigiosin concentration on stability and optical properties of synthesized prodigiosin AgNPs. Highly stable, spherical prodigiosin-conjugated AgNPs were synthesized with a mean diameter of 9.98 nm using a rapid one-step method. The cytotoxic activity investigated in the present study indicated that prodigiosin and prodigiosin-conjugated AgNPs possessed a strong cytotoxic potency against human liver cancer. The In silico molecular docking results of prodigiosin and prodigiosin-conjugated AgNPs are congruent with the In vitro studies and these AgNPs can be considered as good inhibitors of mitogen-activated protein kinase 1 (MEK kinases. The study opened the possibility of using prodigiosin-conjugated AgNPs to increase the efficiency of liver cancer treatment.

  1. Cadmium, lead, and arsenic contamination in paddy soils of a mining area and their exposure effects on human HEPG2 and keratinocyte cell-lines.

    Science.gov (United States)

    Xue, Shengguo; Shi, Lizheng; Wu, Chuan; Wu, Hui; Qin, Yanyan; Pan, Weisong; Hartley, William; Cui, Mengqian

    2017-07-01

    A mining district in south China shows significant metal(loid) contamination in paddy fields. In the soils, average Pb, Cd and As concentrations were 460.1, 11.7 and 35.1mgkg -1 respectively, which were higher than the environmental quality standard for agricultural soils in China (GB15618-1995) and UK Clea Soil Guideline Value. The average contents of Pb, Cd and As in rice were 5.24, 1.1 and 0.7mgkg -1 respectively, which were about 25, 4.5 or 2.5 times greater than the limit values of the maximum safe contaminant concentration standard in food of China (GB 2762-2012), and about 25, 10 or 1 times greater than the limit values of FAO/WHO standard. The elevated contents of Pb, Cd and As detected in soils around the factories, indicated that their spatial distribution was influenced by anthropogenic activity, while greater concentrations of Cd in rice appeared in the northwest region of the factories, indicating that the spatial distribution of heavy metals was also affected by natural factors. As human exposure around mining districts is mainly through oral intake of food and dermal contact, the effects of these metals on the viability and MT protein of HepG2 and KERTr cells were investigated. The cell viability decreased with increasing metal concentrations. Co-exposure to heavy metals (Pb+Cd) increased the metals (Pb or Cd)-mediated MT protein induction in both human HepG2 and KERTr cells. Increased levels of MT protein will lead to greater risk of carcinogenic manifestations, and it is likely that chronic exposure to metals may increase the risk to human health. Nevertheless, when co-exposure to two or more metals occur (such as As+Pb), they may have an antagonistic effect thus reducing the toxic effects of each other. Metal contaminations in paddy soils and rice were influenced by anthropogenic activity; metal co-exposure induced MT protein in human cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. In vitro antitumor efficacy of berberine: solid lipid nanoparticles against human HepG2, Huh7 and EC9706 cancer cell lines

    Science.gov (United States)

    Meng, Xiang-Ping; Wang, Xiao; Wang, Huai-ling; Chen, Tong-sheng; Wang, Yi-fei; Wang, Zhi-ping

    2016-03-01

    Hepatocarcinoma and esophageal squamous cell carcinomas threaten human life badly. It is a current issue to seek the effective natural remedy from plant to treat cancer due to the resistance of the advanced hepatocarcinoma and esophageal carcinoma to chemotherapy. Berberine (Ber), an isoquinoline derivative alkaloid, has a wide range of pharmacological properties and is considered to have anti-hepatocarcinoma and antiesophageal carcinoma effects. However its low oral bioavailability restricts its wide application. In this report, Ber loaded solid lipid nanoparticles (Ber-SLN) was prepared by hot melting and then high pressure homogenization technique. The in vitro anti-hepatocarcinoma and antiesophageal carcinoma effects of Ber-SLN relative to efficacy of bulk Ber were evaluated. The particle size and zeta potential of Ber-SLN were 154.3 ± 4.1 nm and -11.7 ± 1.8 mV, respectively. MTT assay showed that Ber-SLN effectively inhibited the proliferation of human HepG2 and Huh7 and EC9706 cells, and the corresponding IC50 value was 10.6 μg/ml, 5.1 μg/ml, and 7.3 μg/ml (18.3μg/ml, 6.5μg/ml, and 12.4μg/ml μg/ml of bulk Ber solution), respectively. These results suggest that the delivery of Ber-SLN is a promising approach for treating tumors.

  3. Anti-hepatocarcinoma effects of berberine-nanostructured lipid carriers against human HepG2, Huh7, and EC9706 cancer cell lines

    Science.gov (United States)

    Meng, Xiang-Ping; Fan, Hua; Wang, Yi-fei; Wang, Zhi-ping; Chen, Tong-sheng

    2016-10-01

    Hepatocarcinoma and esophageal squamous cell carcinomas threaten human life badly. It is a current issue to seek the effective natural remedy from plant to treat cancer due to the resistance of the advanced hepatocarcinoma and esophageal carcinoma to chemotherapy. Berberine (Ber), an isoquinoline derivative alkaloid, has a wide range of pharmacological properties and is considered to have anti-hepatocarcinoma and antiesophageal carcinoma effects. However its low oral bioavailability restricts its wide application. In this report, Ber loaded nanostructured lipid carriers (Ber-NLC) was prepared by hot melting and then high pressure homogenization technique. The in vitro anti-hepatocarcinoma and antiesophageal carcinoma effects of Ber-NLC relative to efficacy of bulk Ber were evaluated. The particle size and zeta potential of Ber-NLC were 189.3 +/- 3.7 nm and -19.3 +/- 1.4 mV, respectively. MTT assay showed that Ber-NLC effectively inhibited the proliferation of human HepG2 and Huh7 and EC9706 cells, and the corresponding IC50 value was 9.1 μg/ml, 4.4 μg/ml, and 6.3 μg/ml (18.3μg/ml, 6.5μg/ml, and 12.4μg/ml μg/ml of bulk Ber solution), respectively. These results suggest that the delivery of Ber-NLC is a promising approach for treating tumors.

  4. The three-dimensional cultivation of the carcinoma cell line HepG2 in a perfused chip system leads to a more differentiated phenotype of the cells compared to monolayer culture

    Energy Technology Data Exchange (ETDEWEB)

    Altmann, B; Giselbrecht, S; Weibezahn, K-F; Welle, A; Gottwald, E [Forschungszentrum Karlsruhe, Institute for Biological Interfaces, 76344 Eggenstein-Leopoldshafen (Germany)], E-mail: eric.gottwald@ibg.fzk.de

    2008-09-01

    We describe a polymer chip with a grid-like architecture that it is intended for the three-dimensional cultivation of cells with an active nutrient and gas supply. The chip is typically made from polymethyl methacrylate or polycarbonate but can also be manufactured from biodegradable polymers, such as poly(lactic-co-glycolic acid). Different designs of the chip can be realized. In this study, we evaluated a chip with 506 microcontainers of the size of 300 x 300 x 300 {mu}m that are capable of housing up to 6 million cells, and its suitability as a tissue-specific culture system for the carcinoma cell line HepG2 instead of primary liver cells. Related to an earlier study, where we could show the principal suitability of the system for rat primary cells, we here investigated the system's suitability for the human carcinoma cell line HepG2. The carcinoma cells were used in two different types of chip-containing bioreactors. By confocal laser scanning microscopy, we could show that cellular integrity in the chip culture was maintained and that there were no signs of apoptosis as confirmed by the absence of K18 fragmentation. Gene expression analysis of some liver-specific genes revealed a significantly higher expression of the phase II metabolism genes uridine-diphosphate- glucosyl-transferase (UGT1A1) and glutathione-S-transferase (GST{pi}1) as a marker. Therefore, we conclude that by using a three-dimensional instead of a conventional monolayer culture system, hepatocellular carcinoma cells display a phenotype that resembles more closely the tissue of origin.

  5. Effects of defined mixtures of persistent organic pollutants (POPs) on multiple cellular responses in the human hepatocarcinoma cell line, HepG2, using high content analysis screening.

    Science.gov (United States)

    Wilson, Jodie; Berntsen, Hanne Friis; Zimmer, Karin Elisabeth; Frizzell, Caroline; Verhaegen, Steven; Ropstad, Erik; Connolly, Lisa

    2016-03-01

    Persistent organic pollutants (POPs) are toxic substances, highly resistant to environmental degradation, which can bio-accumulate and have long-range atmospheric transport potential. Most studies focus on single compound effects, however as humans are exposed to several POPs simultaneously, investigating exposure effects of real life POP mixtures on human health is necessary. A defined mixture of POPs was used, where the compound concentration reflected its contribution to the levels seen in Scandinavian human serum (total mix). Several sub mixtures representing different classes of POPs were also constructed. The perfluorinated (PFC) mixture contained six perfluorinated compounds, brominated (Br) mixture contained seven brominated compounds, chlorinated (Cl) mixture contained polychlorinated biphenyls and also p,p'-dichlorodiphenyldichloroethylene, hexachlorobenzene, three chlordanes, three hexachlorocyclohexanes and dieldrin. Human hepatocarcinoma (HepG2) cells were used for 2h and 48h exposures to the seven mixtures and analysis on a CellInsight™ NXT High Content Screening platform. Multiple cytotoxic endpoints were investigated: cell number, nuclear intensity and area, mitochondrial mass and membrane potential (MMP) and reactive oxygen species (ROS). Both the Br and Cl mixtures induced ROS production but did not lead to apoptosis. The PFC mixture induced ROS production and likely induced cell apoptosis accompanied by the dissipation of MMP. Synergistic effects were evident for ROS induction when cells were exposed to the PFC+Br mixture in comparison to the effects of the individual mixtures. No significant effects were detected in the Br+Cl, PFC+Cl or total mixtures, which contain the same concentrations of chlorinated compounds as the Cl mixture plus additional compounds; highlighting the need for further exploration of POP mixtures in risk assessment. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Inferring Toxicological Responses of HepG2 Cells from ...

    Science.gov (United States)

    Understanding the dynamic perturbation of cell states by chemicals can aid in for predicting their adverse effects. High-content imaging (HCI) was used to measure the state of HepG2 cells over three time points (1, 24, and 72 h) in response to 976 ToxCast chemicals for 10 different concentrations (0.39-200µM). Cell state was characterized by p53 activation (p53), c-Jun activation (SK), phospho-Histone H2A.x (OS), phospho-Histone H3 (MA), alpha tubulin (Mt), mitochondrial membrane potential (MMP), mitochondrial mass (MM), cell cycle arrest (CCA), nuclear size (NS) and cell number (CN). Dynamic cell state perturbations due to each chemical concentration were utilized to infer coarse-grained dependencies between cellular functions as Boolean networks (BNs). BNs were inferred from data in two steps. First, the data for each state variable were discretized into changed/active (> 1 standard deviation), and unchanged/inactive values. Second, the discretized data were used to learn Boolean relationships between variables. In our case, a BN is a wiring diagram between nodes that represent 10 previously described observable phenotypes. Functional relationships between nodes were represented as Boolean functions. We found that inferred BN show that HepG2 cell response is chemical and concentration specific. We observed presence of both point and cycle BN attractors. In addition, there are instances where Boolean functions were not found. We believe that this may be either

  7. In vitro transfection of the hepatitis B virus PreS2 gene into the human hepatocarcinoma cell line HepG2 induces upregulation of human telomerase reverse transcriptase

    International Nuclear Information System (INIS)

    Liu Hua; Luan Fang; Ju Ying; Shen Hongyu; Gao Lifen; Wang Xiaoyan; Liu Suxia; Zhang Lining; Sun Wensheng; Ma Chunhong

    2007-01-01

    The preS2 domain is the minimal functional unit of transcription activators that is encoded by the Hepatitis B virus (HBV) surface (S) gene. It is present in more than one-third of the HBV-integrates in HBV induced hepatocarcinoma (HCC). To further understand the functional role of PreS2 in hepatocytes, a PreS2 expression plasmid, pcS2, was constructed and stably transfected into HepG2 cells. We conducted growth curve and colony-forming assays to study the impact of PreS2 expression on cell proliferation. Cells transfected with PreS2 proliferated more rapidly and formed colonies in soft agar. PreS2 expressing cells also induced upregulation of human telomerase reverse transcriptase (hTERT) and telomerase activation by RT-PCR and the modified TRAP assay. Blocking expression of hTERT with antisense oligonuleotide reversed the growth rate in cells stably transfected with PreS2. Our data suggest that PreS2 may increase the malignant transformation of human HCC cell line HepG2 by upregulating hTERT and inducing telomerase activation

  8. Effects of defined mixtures of persistent organic pollutants (POPs) on multiple cellular responses in the human hepatocarcinoma cell line, HepG2, using high content analysis screening

    International Nuclear Information System (INIS)

    Wilson, Jodie; Berntsen, Hanne Friis; Zimmer, Karin Elisabeth; Frizzell, Caroline; Verhaegen, Steven; Ropstad, Erik; Connolly, Lisa

    2016-01-01

    Persistent organic pollutants (POPs) are toxic substances, highly resistant to environmental degradation, which can bio-accumulate and have long-range atmospheric transport potential. Most studies focus on single compound effects, however as humans are exposed to several POPs simultaneously, investigating exposure effects of real life POP mixtures on human health is necessary. A defined mixture of POPs was used, where the compound concentration reflected its contribution to the levels seen in Scandinavian human serum (total mix). Several sub mixtures representing different classes of POPs were also constructed. The perfluorinated (PFC) mixture contained six perfluorinated compounds, brominated (Br) mixture contained seven brominated compounds, chlorinated (Cl) mixture contained polychlorinated biphenyls and also p,p’-dichlorodiphenyldichloroethylene, hexachlorobenzene, three chlordanes, three hexachlorocyclohexanes and dieldrin. Human hepatocarcinoma (HepG2) cells were used for 2 h and 48 h exposures to the seven mixtures and analysis on a CellInsight™ NXT High Content Screening platform. Multiple cytotoxic endpoints were investigated: cell number, nuclear intensity and area, mitochondrial mass and membrane potential (MMP) and reactive oxygen species (ROS). Both the Br and Cl mixtures induced ROS production but did not lead to apoptosis. The PFC mixture induced ROS production and likely induced cell apoptosis accompanied by the dissipation of MMP. Synergistic effects were evident for ROS induction when cells were exposed to the PFC + Br mixture in comparison to the effects of the individual mixtures. No significant effects were detected in the Br + Cl, PFC + Cl or total mixtures, which contain the same concentrations of chlorinated compounds as the Cl mixture plus additional compounds; highlighting the need for further exploration of POP mixtures in risk assessment. - Highlights: • High content analysis (HCA) is a novel approach for determining toxicity of

  9. Flavonoids from Enicostema littorale blume enhances glucose uptake of cells in insulin resistant human liver cancer (HepG2) cell line via IRS-1/PI3K/Akt pathway.

    Science.gov (United States)

    Mokashi, Priyanka; Khanna, Aparna; Pandita, Nancy

    2017-06-01

    Diabetes mellitus has spread over the world with 347 million people affected. Insulin resistance is a main pathogenic event in Type 2 Diabetes Mellitus (T2DM) leading to a reduction in glucose uptake by peripheral tissue and increased hepatic glucose output. In this study, we have isolated four flavonoid rich fractions fraction A (FA), fraction B (FB), fraction C (FC) and fraction D (FD) from Enicostema littorale. All the fractions were preliminary screened for TLC fingerprinting, total flavonoid content. Total eight flavonoids were identified by LC/MS. Insulin resistant HepG2 (IR/HepG2) model was established by inducing insulin resistance in HepG2 cells to investigate the effect of these fractions on IR/HepG2 cell line for their glucose uptake. The results showed the significant dose dependant increase in glucose uptake of cells treated with FD. It showed significant activity at a concentration of 10μg/ml. The LC/MS results of FD demonstrated the presence of C-glycoside Swertisin which could be responsible for the effect. Further, to investigate the mechanism of action, gene expression for insulin receptor substrate 1 (IRS-1), protein kinase B (Akt-2) and glucose transporter 4 (GLUT-4) genes were evaluated by real time PCR. A significant upregulation of these genes was observed in FD treated samples, thereby indicating the enhancement of glucose uptake rate of cells via IRS-1/PI3K/Akt pathway. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  10. HepG2 human hepatocarcinomas cells sensitization by endogenous porphyrins

    Science.gov (United States)

    Vonarx-Coinsmann, Veronique; Foultier, Marie-Therese; de Brito, Leonor X.; Morlet, Laurent; Patrice, Thierry

    1995-03-01

    We assessed the ability of the human hepatocarcinoma cell line HepG2 to synthesize PpIX in vitro from exogenous ALA and analyzed ALA-induced toxicity and phototoxicity on this cell line. ALA induced a slight dose-dependent dark toxicity, with 79 and 66% cell survival respectively for ALA 50 and 100 mg/ml after 3-h incubation. Whereas the same treatment followed by laser irradiation (l equals 632 nm, 25 J/sq cm) induced dose-dependent phototoxicity, with 54 and 19% cell survival 24 h after PDT. Whatever the incubation time with ALA, a 3-h delay before light exposure was found optimal to reach a maximal phototoxicity. Photoproducts induced by porphyrin light irradiation absorbed light in the red spectral region at longer wavelengths than did the original porphyrins. The possible enhancement of PDT effects after ALA HepG2 cell incubation was investigated by irradiating cells successively with red light (l equals 632 nm) and light (l equals 650 nm). Total fluence was kept constant at 25 J/sq cm. Phototoxicity was lower when cells were irradiated for increased periods of l equals 650 nm light than with l equals 632 nm light alone. Any photoproducts involved had either a short life or were poorly photoreactive. HepG2 cells, synthesizing enzymes and precursors of endogenous porphyrin synthesis, represent a good in vitro model for experiments using ALA-PpIX-PDT.

  11. In Vitro Characterization and Evaluation of the Cytotoxicity Effects of Nisin and Nisin-Loaded PLA-PEG-PLA Nanoparticles on Gastrointestinal (AGS and KYSE-30), Hepatic (HepG2) and Blood (K562) Cancer Cell Lines.

    Science.gov (United States)

    Goudarzi, Fariba; Asadi, Asadollah; Afsharpour, Maryam; Jamadi, Robab Hassanvand

    2018-02-22

    The aim of this study was an in vitro evaluation and comparison of the cytotoxic effects of free nisin and nisin-loaded PLA-PEG-PLA nanoparticles on gastrointestinal (AGS and KYSE-30), hepatic (HepG2), and blood (K562) cancer cell lines. To create this novel anti-cancer drug delivery system, the nanoparticles were synthesized and then loaded with nisin. Subsequently, their biocompatibility, ability to enter cells, and physicochemical properties, including formation, size, and shape, were studied using hemolysis, fluorescein isothiocyanate (FITC), Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), and scanning electron microscopy (SEM), respectively. Then, its loading efficiency and release kinetics were examined to assess the potential impact of this formulation for the nanoparticle carrier candidacy. The cytotoxicities of nisin and nisin-loaded nanoparticles were evaluated by using the MTT and Neutral Red (NR) uptake assays. Detections of the apoptotic cells were done via Ethidium Bromide (EB)/Acridine Orange (AO) staining. The FTIR spectra, SEM images, and DLS graph confirmed the formations of the nanoparticles and nisin-loaded nanoparticles with spherical, distinct, and smooth surfaces and average sizes of 100 and 200 nm, respectively. The loading efficiency of the latter nanoparticles was about 85-90%. The hemolysis test represented their non-cytotoxicities and the FITC images indicated their entrance inside the cells. An increase in the percentage of apoptotic cells was observed through EB/AO staining. These results demonstrated that nisin had a cytotoxic effect on AGS, KYSE-30, HepG2, and K562 cancer cell lines, while the cytotoxicity of nisin-loaded nanoparticles was more than that of the free nisin.

  12. Reduction of angiocidin contributes to decreased HepG2 cell ...

    African Journals Online (AJOL)

    Angiocidin protein production in HepG2 cells were reduced significantly by siRNA. When HepG2 cells were transfected with siRNA-angiocidin, these cells showed very low proliferation activity compared with control cells. Our study suggests that reduction of angiocidin may contribute to decreased proliferation activity in liver ...

  13. Protective effects of quercetin on nicotine induced oxidative stress in 'HepG2 cells'.

    Science.gov (United States)

    Yarahmadi, Amir; Zal, Fatemeh; Bolouki, Ayeh

    2017-10-01

    Nicotine is a natural component of tobacco plants and is responsible for the addictive properties of tobacco. Nicotine has been recognized to result in oxidative stress by inducing the generation of reactive oxygen species (ROS). The purpose of this work was to estimate the hepatotoxicity effect of nicotine on viability and on antioxidant defense system in cultures of HepG2 cell line and the other hand, ameliorative effect of quercetin (Q) as an antioxidant was analyzed. Nicotine induced concentration dependent loss in HepG2 cell line viability. The results indicated that nicotine decreased activity of superoxide dismutase (SOD) and glutathione reductase (GR) and increased activities of catalase (CAT) and glutathione peroxidase (GPx) and glutathione (GSH) content in the HepG2 cells. Q significantly increased activity of SOD, GR and GSH content and decreased activity of GPX in nicotine + Q groups. Our data demonstrate that Q plays a protective role against the imbalance elicited by nicotine between the production of free radicals and antioxidant defense systems, and suggest that administration of this antioxidant may find clinical application where cellular damage is a consequence of ROS.

  14. Selectivity of biopolymer membranes using HepG2 cells.

    Science.gov (United States)

    Lü, Dongyuan; Gao, Yuxin; Luo, Chunhua; Lü, Shouqian; Wang, Qian; Xu, Xianghong; Sun, Shujin; Wang, Chengzhi; Long, Mian

    2015-03-01

    Bioartificial liver (BAL) system has emerged as an alternative treatment to bridge acute liver failure to either liver transplantation or liver regeneration. One of the main reasons that the efficacy of the current BAL systems was not convincing in clinical trials is attributed to the lack of friendly interface between the membrane and the hepatocytes in liver bioreactor, the core unit of BAL system. Here, we systematically compared the biological responses of hepatosarcoma HepG2 cells seeded on eight, commercially available biocompatible membranes made of acetyl cellulose-nitrocellulose mixed cellulose (CA-NC), acetyl cellulose (CA), nylon (JN), polypropylene (PP), nitrocellulose (NC), polyvinylidene fluoride (PVDF), polycarbonate (PC) and polytetrafluoroethylene (PTFE). Physicochemical analysis and mechanical tests indicated that CA, JN and PP membranes yield high adhesivity and reasonable compressive and/or tensile features with friendly surface topography for cell seeding. Cells prefer to adhere on CA, JN, PP or PTFE membranes with high proliferation rate in spheriod-like shape. Actin, albumin and cytokeratin 18 expressions are favorable for cells on CA or PP membrane, whereas protein filtration is consistent among all the eight membranes. These results further the understandings of cell growth, morphology and spreading, as well as protein filtration on distinct membranes in designing a liver bioreactor.

  15. Cacao polyphenols influence the regulation of apolipoprotein in HepG2 and Caco2 cells.

    Science.gov (United States)

    Yasuda, Akiko; Natsume, Midori; Osakabe, Naomi; Kawahata, Keiko; Koga, Jinichiro

    2011-02-23

    Cocoa powder is rich in polyphenols, such as catechins and procyanidins, and has been shown to inhibit low-density lipoprotein (LDL) oxidation and atherogenesis in a variety of models. Human studies have also shown daily intake of cocoa increases plasma high-density lipoprotein (HDL) and decreases LDL levels. However, the mechanisms responsible for these effects of cocoa on cholesterol metabolism have yet to be fully elucidated. The present study investigated the effects of cacao polyphenols on the production of apolipoproteins A1 and B in human hepatoma HepG2 and intestinal Caco2 cell lines. The cultured HepG2 cells or Caco2 cells were incubated for 24 h in the presence of cacao polyphenols such as (-)-epicatechin, (+)-catechin, procyanidin B2, procyanidin C1, and cinnamtannin A2. The concentration of apolipoproteins in the cell culture media was quantified using an enzyme-linked immunoassay, and the mRNA expression was quantified by RT-PCR. Cacao polyphenols increased apolipoprotein A1 protein levels and mRNA expression, even though apolipoprotein B protein and the mRNA expression were slightly decreased in both HepG2 cells and Caco2 cells. In addition, cacao polyphenols increased sterol regulatory element binding proteins (SREBPs) and activated LDL receptors in HepG2 cells. These results suggest that cacao polyphenols may increase the production of mature form SREBPs and LDL receptor activity, thereby increasing ApoA1 and decreasing ApoB levels. These results elucidate a novel mechanism by which HDL cholesterol levels become elevated with daily cocoa intake.

  16. Study of Viral Vectors in a Three-dimensional Liver Model Repopulated with the Human Hepatocellular Carcinoma Cell Line HepG2

    OpenAIRE

    Hiller, Thomas; R?hrs, Viola; Dehne, Eva-Maria; Wagner, Anke; Fechner, Henry; Lauster, Roland; Kurreck, Jens

    2016-01-01

    This protocol describes the generation of a three-dimensional (3D) ex vivo liver model and its application to the study and development of viral vector systems. The model is obtained by repopulating the extracellular matrix of a decellularized rat liver with a human hepatocyte cell line. The model permits studies in a vascularized 3D cell system, replacing potentially harmful experiments with living animals. Another advantage is the humanized nature of the model, which is closer to human phys...

  17. Casearin D inhibits ERK phosphorylation and induces downregulation of cyclin D1 in HepG2 cells.

    Science.gov (United States)

    Ferreira-Silva, Guilherme Álvaro; Lages, Carla Carolina Lopes; Sartorelli, Patricia; Hasegawa, Flávia Rie; Soares, Marisi Gomes; Ionta, Marisa

    2017-02-01

    Cancer is a public health problem which represents the second cause of death in the world. In this framework, it is necessary to identify novel compounds with antineoplastic potential. Plants are an important source for discovering novel compounds with pharmacological potential. In this study, we aimed to investigate the antiproliferative potential of isolated compounds from Casearia sylvestris on tumor cell lines. Crude extract effectively reduced cell viability of 4 tumor cell lines (HepG2, A549, U251-MG, and HT-144) after 48h treatment. HepG2 and HT-144 were the most responsive cells. Three fractions (aqueous ethanol, n-hexane and ethyl acetate) were tested against HepG2 and HT-144 cells and we observed that compounds with antiproliferative activity were concentrated in n-hexane and ethyl acetate fractions. The casearins A, G and J were isolated from n-hexane fraction, while casearin D was obtained from ethyl acetate fraction. We demonstrated that casearin D significantly inhibited the clonogenic capacity of HepG2 cells after 24h exposure indicating its antiproliferative activity. In addition, G1/S transition cell cycle arrest in HepG2 cells was also observed. These effects are related, at least in part, to ability of the casearin D in reducing ERK phosphorylation and cyclin D1 expression levels. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. The Human NADPH Oxidase, Nox4, Regulates Cytoskeletal Organization in Two Cancer Cell Lines, HepG2 and SH-SY5Y

    Directory of Open Access Journals (Sweden)

    Simon Auer

    2017-05-01

    Full Text Available NADPH oxidases of human cells are not only functional in defense against invading microorganisms and for oxidative reactions needed for specialized biosynthetic pathways but also during the past few years have been established as signaling modules. It has been shown that human Nox4 is expressed in most somatic cell types and produces hydrogen peroxide, which signals to remodel the actin cytoskeleton. This correlates well with the function of Yno1, the only NADPH oxidase of yeast cells. Using two established tumor cell lines, which are derived from hepatic and neuroblastoma tumors, respectively, we are showing here that in both tumor models Nox4 is expressed in the ER (like the yeast NADPH oxidase, where according to published literature, it produces hydrogen peroxide. Reducing this biochemical activity by downregulating Nox4 transcription leads to loss of F-actin stress fibers. This phenotype is reversible by adding hydrogen peroxide to the cells. The effect of the Nox4 silencer RNA is specific for this gene as it does not influence the expression of Nox2. In the case of the SH-SY5Y neuronal cell line, Nox4 inhibition leads to loss of cell mobility as measured in scratch assays. We propose that inhibition of Nox4 (which is known to be strongly expressed in many tumors could be studied as a new target for cancer treatment, in particular for inhibition of metastasis.

  19. Estrogen receptor α and aryl hydrocarbon receptor cross-talk in a transfected hepatoma cell line (HepG2 exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin

    Directory of Open Access Journals (Sweden)

    Manuela Göttel

    2014-01-01

    Full Text Available The prototype dioxin congener 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD is known to exert anti-estrogenic effects via activation of the aryl hydrocarbon receptor (AhR by interfering with the regulation of oestrogen homeostasis and the estrogen receptor α (ERα signalling pathway. The AhR/ER cross-talk is considered to play a crucial role in TCDD- and E2-dependent mechanisms of carcinogenesis, though the concerted mechanism of action in the liver is not yet elucidated. The present study investigated TCDD's impact on the transcriptional cross-talk between AhR and ERα and its modulation by 17β-estradiol (E2 in the human hepatoma cell line HepG2, which is AhR-responsive but ERα-negative. Transient transfection assays with co-transfection of hERα and supplementation of receptor antagonists showed anti-estrogenic action of TCDD via down-regulation of E2-induced ERα signaling. In contrast, enhancement of AhR signaling dependent on ERα was observed providing evidence for increased cytochrome P450 (CYP induction to promote E2 metabolism. However, relative mRNA levels of major E2-metabolizing CYP1A1 and 1B1 and the main E2-detoxifying catechol-O-methyltransferase were not affected by the co-treatments. This study provides new evidence of a TCDD-activated AhR-mediated molecular AhR/ERα cross-talk mechanism at transcriptional level via indirect inhibition of ERα and enhanced transcriptional activity of AhR in HepG2 cells.

  20. Bioactive chemical constituents of Curcuma longa L. rhizomes extract inhibit the growth of human hepatoma cell line (HepG2

    Directory of Open Access Journals (Sweden)

    Abdel-Lateef Ezzat

    2016-09-01

    Full Text Available The present study was designed to identify the chemical constituents of the methanolic extract of Curcuma longa L. rhizomes and their inhibitory effect on a hepatoma cell line. The methanolic extract was subjected to GC-MS analysis to identify the volatile constituents and the other part of the same extract was subjected to liquid column chromatographic separation to isolate curcumin. The inhibition of cell growth in the hepatoma cell line and the cytopathological changes were studied. GC-MS analysis showed the presence of fifty compounds in the methanolic extract of C. longa. The major compounds were ar-turmerone (20.50 %, β-sesquiphellandrene (5.20 % and curcumenol (5.11 %. Curcumin was identified using IR, 1H and 13C NMR. The inhibition of cell growth by curcumin (IC50 = 41.69 ± 2.87 μg mL-1 was much more effective than that of methanolic extract (IC50 = 196.12 ± 5.25 μg mL-1. Degenerative and apoptotic changes were more evident in curcumin- treated hepatoma cells than in those treated with the methanol extract. Antitumor potential of the methanolic extract may be attributed to the presence of sesquiterpenes and phenolic constituents including curcumin (0.051 %, 511.39 μg g-1 dried methanol extract in C. longa rhizomes.

  1. Bioactive chemical constituents of Curcuma longa L. rhizomes extract inhibit the growth of human hepatoma cell line (HepG2).

    Science.gov (United States)

    Abdel-Lateef, Ezzat; Mahmoud, Faten; Hammam, Olfat; El-Ahwany, Eman; El-Wakil, Eman; Kandil, Sherihan; Abu Taleb, Hoda; El-Sayed, Mortada; Hassenein, Hanaa

    2016-09-01

    The present study was designed to identify the chemical constituents of the methanolic extract of Curcuma longa L. rhizomes and their inhibitory effect on a hepatoma cell line. The methanolic extract was subjected to GC-MS analysis to identify the volatile constituents and the other part of the same extract was subjected to liquid column chromatographic separation to isolate curcumin. The inhibition of cell growth in the hepatoma cell line and the cytopathological changes were studied. GC-MS analysis showed the presence of fifty compounds in the methanolic extract of C. longa. The major compounds were ar-turmerone (20.50 %), β-sesquiphellandrene (5.20 %) and curcumenol (5.11 %). Curcumin was identified using IR, 1H and 13C NMR. The inhibition of cell growth by curcumin (IC50 = 41.69 ± 2.87 μg mL-1) was much more effective than that of methanolic extract (IC50 = 196.12 ± 5.25 μg mL-1). Degenerative and apoptotic changes were more evident in curcumin- treated hepatoma cells than in those treated with the methanol extract. Antitumor potential of the methanolic extract may be attributed to the presence of sesquiterpenes and phenolic constituents including curcumin (0.051 %, 511.39 μg g-1 dried methanol extract) in C. longa rhizomes.

  2. Pooled human liver preparations, HepaRG, or HepG2 cell lines for metabolism studies of new psychoactive substances? A study using MDMA, MDBD, butylone, MDPPP, MDPV, MDPB, 5-MAPB, and 5-API as examples.

    Science.gov (United States)

    Richter, Lilian H J; Flockerzi, Veit; Maurer, Hans H; Meyer, Markus R

    2017-09-05

    Metabolism studies play an important role in clinical and forensic toxicology. Because of potential species differences in metabolism, human samples are best suitable for elucidating metabolism. However, in the case of new psychoactive substances (NPS), human samples of controlled studies are not available. Primary human hepatocytes have been described as gold standard for in vitro metabolism studies, but there are some disadvantages such as high costs, limited availability, and variability of metabolic enzymes. Therefore, the aim of our study was to investigate and compare the metabolism of six methylenedioxy derivatives (MDMA, MDBD, butylone, MDPPP, MDPV, MDPB) and two bioisosteric analogues (5-MAPB, 5-API) using pooled human liver microsomes (pHLM) combined with cytosol (pHLC) or pooled human liver S9 fraction (pS9) all after addition of co-substrates for six phase I and II reactions. In addition, HepaRG and HepG2 cell lines were used. Results of the different in vitro tools were compared to each other, to corresponding published data, and to metabolites identified in human urine after consumption of MDMA, MDPV, or 5-MAPB. Incubations with pHLM plus pHLC showed similar results as pS9. A more cost efficient model for prediction of targets for toxicological screening procedures in human urine should be identified. As expected, the incubations with HepaRG provided better results than those with HepG2 concerning number and signal abundance of the metabolites. Due to easy handling without special equipment, incubations with pooled liver preparations should be the most suitable alternative to find targets for toxicological screening procedures for methylenedioxy derivatives and bioisosteric analogues. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Synthesis, characterization of 1,2,4-triazole Schiff base derived 3d-metal complexes: Induces cytotoxicity in HepG2, MCF-7 cell line, BSA binding fluorescence and DFT study

    Science.gov (United States)

    Tyagi, Prateek; Tyagi, Monika; Agrawal, Swati; Chandra, Sulekh; Ojha, Himanshu; Pathak, Mallika

    2017-01-01

    Two novel Schiff base ligands H2L1 and H2L2 have been synthesized by condensation reaction of amine derivative of 1,2,4-triazole moiety with 2-hydroxy-4-methoxybenzaldehyde. Co(II), Ni(II), Cu(II) and Zn(II) of the synthesized Schiff bases were prepared by using a molar ratio of ligand:metal as 1:1. The structure of the Schiff bases and synthesized metal complexes were established by 1H NMR, UV-Vis, IR, Mass spectrometry and molar conductivity. The thermal stability of the complexes was study by TGA. Fluorescence quenching mechanism of metal complexes 1-4 show that Zn(II) and Cu(II) complex binds more strongly to BSA. In DFT studies the geometries of Schiff bases and metal complexes were fully optimized with respect to the energy using the 6-31 + g(d,p) basis set. The spectral data shows that the ligands behaves as binegative tridentate. On the basis of the spectral studies, TGA and DFT data an octahedral geometry has been assigned for Co(II), Ni(II), square planar for Cu(II) and tetrahedral for Zn(II) complexes. The anticancer activity were screened against human breast cancer cell line (MCF-7) and human hepatocellular liver carcinoma cell line (Hep-G2). Result indicates that metal complexes shows increase cytotoxicity in proliferation to cell lines as compared to free ligand.

  4. Study of Viral Vectors in a Three-dimensional Liver Model Repopulated with the Human Hepatocellular Carcinoma Cell Line HepG2

    Science.gov (United States)

    Hiller, Thomas; Röhrs, Viola; Dehne, Eva-Maria; Wagner, Anke; Fechner, Henry; Lauster, Roland; Kurreck, Jens

    2016-01-01

    This protocol describes the generation of a three-dimensional (3D) ex vivo liver model and its application to the study and development of viral vector systems. The model is obtained by repopulating the extracellular matrix of a decellularized rat liver with a human hepatocyte cell line. The model permits studies in a vascularized 3D cell system, replacing potentially harmful experiments with living animals. Another advantage is the humanized nature of the model, which is closer to human physiology than animal models. In this study, we demonstrate the transduction of this liver model with a viral vector derived from adeno-associated viruses (AAV vector). The perfusion circuit that supplies the 3D liver model with media provides an easy means to apply the vector. The system permits monitoring of the major metabolic parameters of the liver. For final analysis, tissue samples can be taken to determine the extent of recellularization by histological techniques. Distribution of the virus vector and expression of the delivered transgene can be analyzed by quantitative PCR (qPCR), Western blotting and immunohistochemistry. Numerous applications of the vector model in basic research and in the development of gene therapeutic applications can be envisioned, including the development of novel antiviral therapeutics, cancer research, and the study of viral vectors and their potential side effects. PMID:27805597

  5. A Novel Method to Improve the Anticancer Activity of Natural-Based Hydroxyapatite against the Liver Cancer Cell Line HepG2 Using Mesoporous Magnesia as a Micro-Carrier

    Directory of Open Access Journals (Sweden)

    Nasser S. Awwad

    2017-11-01

    Full Text Available Micro-carriers are the best known vehicles to transport different kinds of drugs to achieve high impact. In this study, mesoporous magnesium oxide has been harnessed as a micro-carrier to encapsulate the anticancer candidate drug natural-based cubic hydroxyapatite (HAP. HAP@MgO composites with different HAP loading (0–60 wt %, were prepared by a hydrothermal treatment method using triethanol amine as a template. The characterization of the prepared composites were achieved by using XRD, Raman spectroscopy, FTIR and SEM. Characterization data confirm the formation of sphere-like structures of MgO containing HAP particles. It was observed that the size of the spheres increased with HAP loading up to 40 wt %, then collapsed. Furthermore, the anticancer property of the prepared composites was evaluated against the HepG2 liver cancer cell line. The HAP@MgO composites exhibited higher activity than neat MgO or HAP. The 20 wt % of HAP was the optimum loading to control cell proliferation by inducing apoptosis. Apoptosis was determined by typical apoptotic bodies produced by the cell membrane.

  6. Cellular trafficking of thymosin beta-4 in HEPG2 cells following serum starvation.

    Directory of Open Access Journals (Sweden)

    Giuseppina Pichiri

    Full Text Available Thymosin beta-4 (Tβ4 is an ubiquitous multi-functional regenerative peptide, related to many critical biological processes, with a dynamic and flexible conformation which may influence its functions and its subcellular distribution. For these reasons, the intracellular localization and trafficking of Tβ4 is still not completely defined and is still under investigation in in vivo as well as in vitro studies. In the current study we used HepG2 cells, a human hepatoma cell line; cells growing in normal conditions with fetal bovine serum expressed high levels of Tβ4, restricted to the cytoplasm until 72 h. At 84 h, a diffuse Tβ4 cytoplasmic immunostaining shifted to a focal perinuclear and nuclear reactivity. In the absence of serum, nuclear reactivity was localized in small granules, evenly dispersed throughout the entire nuclear envelop, and was observed as earlier as at 48 h. Cytoplasmic immunostaining for Tβ4 in HepG2 cells under starvation appeared significantly lower at 48 h and decreased progressively at 72 and at 84 h. At these time points, the decrease in cytoplasmic staining was associated with a progressive increase in nuclear reactivity, suggesting a possible translocation of the peptide from the cytoplasm to the nuclear membrane. The normal immunocytochemical pattern was restored when culture cells submitted to starvation for 84 h received a new complete medium for 48 h. Mass spectrometry analysis, performed on the nuclear and cytosolic fractions of HepG2 growing with and without serum, showed that Tβ4 was detectable only in the cytosolic and not in the intranuclear fraction. These data suggest that Tβ4 is able to translocate from different cytoplasmic domains to the nuclear membrane and back, based on different stress conditions within the cell. The punctuate pattern of nuclear Tβ4 immunostaining associated with Tβ4 absence in the nucleoplasm suggest that this peptide might be localized in the nuclear pores, where it could

  7. Xanthorrhizol induced DNA fragmentation in HepG2 cells involving Bcl-2 family proteins

    International Nuclear Information System (INIS)

    Tee, Thiam-Tsui; Cheah, Yew-Hoong; Meenakshii, Nallappan; Mohd Sharom, Mohd Yusof; Azimahtol Hawariah, Lope Pihie

    2012-01-01

    Highlights: ► We isolated xanthorrhizol, a sesquiterpenoid compound from Curcuma xanthorrhiza. ► Xanthorrhizol induced apoptosis in HepG2 cells as observed using SEM. ► Apoptosis in xanthorrhizol-treated HepG2 cells involved Bcl-2 family proteins. ► DNA fragmentation was observed in xanthorrhizol-treated HepG2 cells. ► DNA fragmentation maybe due to cleavage of PARP and DFF45/ICAD proteins. -- Abstract: Xanthorrhizol is a plant-derived pharmacologically active sesquiterpenoid compound isolated from Curcuma xanthorrhiza. Previously, we have reported that xanthorrhizol inhibited the proliferation of HepG2 human hepatoma cells by inducing apoptotic cell death via caspase activation. Here, we attempt to further elucidate the mode of action of xanthorrhizol. Apoptosis in xanthorrhizol-treated HepG2 cells as observed by scanning electron microscopy was accompanied by truncation of BID; reduction of both anti-apoptotic Bcl-2 and Bcl-X L expression; cleavage of PARP and DFF45/ICAD proteins and DNA fragmentation. Taken together, these results suggest xanthorrhizol as a potent antiproliferative agent on HepG2 cells by inducing apoptosis via Bcl-2 family members. Hence we proposed that xanthorrhizol could be used as an anti-liver cancer drug for future studies.

  8. Xanthorrhizol induced DNA fragmentation in HepG2 cells involving Bcl-2 family proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tee, Thiam-Tsui, E-mail: thiamtsu@yahoo.com [School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor (Malaysia); Cheah, Yew-Hoong [School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor (Malaysia); Bioassay Unit, Herbal Medicine Research Center, Institute for Medical Research, Jalan Pahang, Kuala Lumpur (Malaysia); Meenakshii, Nallappan [Biology Department, Faculty of Science, Universiti Putra Malaysia, 43400 Serdang, Selangor (Malaysia); Mohd Sharom, Mohd Yusof; Azimahtol Hawariah, Lope Pihie [School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor (Malaysia)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer We isolated xanthorrhizol, a sesquiterpenoid compound from Curcuma xanthorrhiza. Black-Right-Pointing-Pointer Xanthorrhizol induced apoptosis in HepG2 cells as observed using SEM. Black-Right-Pointing-Pointer Apoptosis in xanthorrhizol-treated HepG2 cells involved Bcl-2 family proteins. Black-Right-Pointing-Pointer DNA fragmentation was observed in xanthorrhizol-treated HepG2 cells. Black-Right-Pointing-Pointer DNA fragmentation maybe due to cleavage of PARP and DFF45/ICAD proteins. -- Abstract: Xanthorrhizol is a plant-derived pharmacologically active sesquiterpenoid compound isolated from Curcuma xanthorrhiza. Previously, we have reported that xanthorrhizol inhibited the proliferation of HepG2 human hepatoma cells by inducing apoptotic cell death via caspase activation. Here, we attempt to further elucidate the mode of action of xanthorrhizol. Apoptosis in xanthorrhizol-treated HepG2 cells as observed by scanning electron microscopy was accompanied by truncation of BID; reduction of both anti-apoptotic Bcl-2 and Bcl-X{sub L} expression; cleavage of PARP and DFF45/ICAD proteins and DNA fragmentation. Taken together, these results suggest xanthorrhizol as a potent antiproliferative agent on HepG2 cells by inducing apoptosis via Bcl-2 family members. Hence we proposed that xanthorrhizol could be used as an anti-liver cancer drug for future studies.

  9. In vitro toxicity of cationic micelles and liposomes in cultured human hepatocyte (HepG2) and lung epithelial (A549) cell lines

    DEFF Research Database (Denmark)

    Roursgaard, Martin; Knudsen, Kristina Bram; Northeved, Helle

    2016-01-01

    The aim of this study was to compare the effects of cationic micelle and liposome drug delivery systems on liver and lung cells in a toxicological in vitro screening model, with observations on cytotoxicity and genotoxicity. A screening battery was established for assessment of a broad range...

  10. Ginseng (Panax quinquefolius and Licorice (Glycyrrhiza uralensis Root Extract Combinations Increase Hepatocarcinoma Cell (Hep-G2 Viability

    Directory of Open Access Journals (Sweden)

    David G. Popovich

    2011-01-01

    Full Text Available The combined cytoactive effects of American ginseng (Panax quinquefolius and licorice (Glycyrrhiza uralensis root extracts were investigated in a hepatocarcinoma cell line (Hep-G2. An isobolographic analysis was utilized to express the possibility of synergistic, additive or antagonistic interaction between the two extracts. Both ginseng and licorice roots are widely utilized in traditional Chinese medicine preparations to treat a variety of ailments. However, the effect of the herbs in combination is currently unknown in cultured Hep-G2 cells. Ginseng (GE and licorice (LE extracts were both able to reduce cell viability. The LC50 values, after 72 h, were found to be 0.64 ± 0.02 mg/mL (GE and 0.53 ± 0.02 mg/mL (LE. An isobologram was plotted, which included five theoretical LC50s calculated, based on the fixed fraction method of combination ginseng to licorice extracts to establish a line of additivity. All combinations of GE to LE (1/5, 1/3, 1/2, 2/3, 4/5 produced an effect on Hep-G2 cell viability but they were all found to be antagonistic. The LC50 of fractions 1/3, 1/2, 2/3 were 23%, 21% and 18% above the theoretical LC50. Lactate dehydrogenase release indicated that as the proportion of GE to LE increased beyond 50%, the influence on membrane permeability increased. Cell-cycle analysis showed a slight but significant arrest at the G1 phase of cell cycle for LE. Both GE and LE reduced Hep-G2 viability independently; however, the combinations of both extracts were found to have an antagonistic effect on cell viability and increased cultured Hep-G2 survival.

  11. Radiation induced bystander effect on hepatoma HepG2 cells under hypoxia condition

    International Nuclear Information System (INIS)

    Zhang Jianghong; Jin Yizun; Shao Chunlin; Prise KM

    2009-01-01

    Objective: To investigate radiation induced bystander effect and its mechanism on hepatoma HepG2 cells under hypoxia condition. Methods: Non-irradiated bystander hepatoma cells were co-cultured with irradiated cells or treated with the conditioned medium (CM) from irradiated cells, then micronuclei (MN) were measured for both irradiated cells and bystander cells. Results: The MN yield of irradiated HepG2 cells under hypoxic condition was significantly lower than that under normoxia, the oxygen enhancement ratio of HepG2 cells of MN was 1.6. For both hypoxic and normoxic condition, the MN yield of bystander cells were obviously enhanced to a similar high level after co-culturing with irradiated cells or with CM treatment, and it also correlated with the irradiation dose. When the hypoxic HepG2 cells were treated with either DMSO, a scavenger of reactive oxygen species (ROS), or aminoguanidine, an iNOS inhibitor, the yield of bystander MN was partly diminished, and the reducing rate of DMSO was 42.2%-46.7%, the reducing rate of aminoguanidine was 42% . Conclusion: ROS, NO and their downstream signal factors are involved in the radiation induced bystander effect of hypoxic HepG2 cells. (authors)

  12. Role of cytochrome P450 2J2 on cell proliferation and resistance to an anticancer agent in hepatocellular carcinoma HepG2 cells.

    Science.gov (United States)

    Hwang, Geun Hye; Park, So Mi; Han, Ho Jae; Baek, Kyoung Min; Kim, Joong Sun; Chang, Woochul; Lee, Ho Jin; Yun, Seung Pil; Ryu, Jung Min; Lee, Min Young

    2017-11-01

    The present study examined the role of human cytochrome P450 2J2 (CYP2J2) on cell proliferation and resistance to an anticancer agent using stable hepatocellular carcinoma HepG2 cells overexpressing CYP2J2. Overexpression of CYP2J2 significantly increased HepG2 cell proliferation and the expression levels of cell cycle regulatory proteins, including cyclin D1, cyclin E, cyclin-dependent kinase (Cdk)2 and Cdk4. CYP2J2-overexpressing HepG2 cells exhibited high levels of Akt phosphorylation compared with those observed in wild-type HepG2 cells. Although Akt phosphorylation in both cell lines was significantly attenuated by LY294002, a specific phosphoinositide 3-kinase/Akt signaling inhibitor, the levels of Akt phosphorylation following treatment with LY294002 were higher in CYP2J2-overexpressing HepG2 cells than in wild-type HepG2 cells. Cell counting revealed that proliferation was reduced by LY294002 in both cell lines; however, CYP2J2-overexpressing HepG2 cell numbers were higher than those of wild-type HepG2 cells following treatment with LY294002. These results indicated that increased cell proliferation by CYP2J2 overexpression is mediated by increased Akt activity. It was also demonstrated that doxorubicin, an anticancer agent, reduced cell viability, induced a significant increase in the B-cell lymphoma (Bcl)-2 associated X protein (Bax)/Bcl-2 ratio and decreased pro-caspase-3 levels in wild-type HepG2 cells. However, the doxorubicin-induced reduction in cell viability was significantly attenuated by enhanced upregulation of CYP2J2 expression. The increase in the Bax/Bcl-2 ratio and the decrease in pro-caspase-3 levels were also recovered by CYP2J2 overexpression. In conclusion, CYP2J2 serves important roles in cancer cell proliferation and resistance to the anticancer agent doxorubicin in HepG2 cells.

  13. Apoptosis Induction by Polygonum minus Is Related to Antioxidant Capacity, Alterations in Expression of Apoptotic-Related Genes, and S-Phase Cell Cycle Arrest in HepG2 Cell Line

    Directory of Open Access Journals (Sweden)

    Mohd Alfazari Mohd Ghazali

    2014-01-01

    Full Text Available Polygonum minus (Polygonaceae is a medicinal herb distributed throughout eastern Asia. The present study investigated antiproliferative effect of P. minus and its possible mechanisms. Four extracts (petroleum ether, methanol, ethyl acetate, and water were prepared by cold maceration. Extracts were subjected to phytochemical screening, antioxidant, and antiproliferative assays; the most bioactive was fractionated using vacuum liquid chromatography into seven fractions (F1–F7. Antioxidant activity was measured via total phenolic content (TPC, 2,2-diphenyl-1-picrylhydrazyl (DPPH, and ferric reducing antioxidant power (FRAP assays. Antiproliferative activity was evaluated using 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay. Most active fraction was tested for apoptosis induction and cell cycle arrest in HepG2 cells using flow cytometry and confocal microscopy. Apoptotic-related gene expression was studied by RT-PCR. Ethyl acetate extract was bioactive in initial assays. Its fraction, F7, exhibited highest antioxidant capacity (TPC; 113.16±6.2 mg GAE/g extract, DPPH; EC50: 30.5±3.2 μg/mL, FRAP; 1169±20.3 μmol Fe (II/mg extract and selective antiproliferative effect (IC50: 25.75±1.5 μg/mL. F7 induced apoptosis in concentration- and time-dependent manner and caused cell cycle arrest at S-phase. Upregulation of proapoptotic genes (Bax, p53, and caspase-3 and downregulation of antiapoptotic gene, Bcl-2, were observed. In conclusion, F7 was antiproliferative to HepG2 cells by inducing apoptosis, cell cycle arrest, and via antioxidative effects.

  14. Up-regulation of P-glycoprotein expression by catalase via JNK activation in HepG2 cells.

    Science.gov (United States)

    Li, Lin; Xu, Jianfeng; Min, Taishan; Huang, Weida

    2006-01-01

    Overexpression of the MDR1 gene is one of the reasons for multidrug resistance (MDR). Some studies suggested that antioxidants could down-regulate MDR1 expression as a possible cancer treatment. In this report, we try to determine the effects of antioxidants (catalase or N-acetylcysteine [NAC]) on the regulation of intrinsic MDR1 overexpression in HepG2 cells. Adding catalase or N-acetylcysteine to the HepG2 culture led to a significant increase of MDR1 mRNA and P-glycoprotein drug transporter activity. After catalase or NAC treatment, a reduced intracellular reactive oxygen species (ROS) was observed. The JNK inhibitor SP600125 abolished the positive effects of catalase on drug transporter activity in a dose-dependent manner. Furthermore, the up-regulation of P-glycoprotein functions by catalase was only observed in HepG2 cells but not in other cell lines tested (MCF-7, A549, A431). These data suggested that catalase can up-regulate P-glycoprotein expression in HepG2 cells via reducing intracellular ROS, and JNK may mediate this process.

  15. Expression of CAR in SW480 and HepG2 cells during G1 is associated with cell proliferation

    International Nuclear Information System (INIS)

    Osabe, Makoto; Sugatani, Junko; Takemura, Akiko; Yamazaki, Yasuhiro; Ikari, Akira; Kitamura, Naomi; Negishi, Masahiko; Miwa, Masao

    2008-01-01

    Constitutive androstane receptor (CAR) is a transcription factor to regulate the expression of several genes related to drug-metabolism. Here, we demonstrate that CAR protein accumulates during G1 in human SW480 and HepG2 cells. After the G1/S phase transition, CAR protein levels decreased, and CAR was hardly detected in cells by the late M phase. CAR expression in both cell lines was suppressed by RNA interference-mediated suppression of CDK4. Depletion of CAR by RNA interference in both cells and by hepatocyte growth factor treatment in HepG2 cells resulted in decreased MDM2 expression that led to p21 upregulation and repression of HepG2 cell growth. Thus, our results demonstrate that CAR expression is an early G1 event regulated by CDK4 that contributes to MDM2 expression; these findings suggest that CAR may influence the expression of genes involved in not only the metabolism of endogenous and exogenous substances but also in the cell proliferation

  16. Tyrosine kinase/phosphatase inhibitors decrease dengue virus production in HepG2 cells.

    Science.gov (United States)

    Limjindaporn, Thawornchai; Panaampon, Jutatip; Malakar, Shilu; Noisakran, Sansanee; Yenchitsomanus, Pa-Thai

    2017-01-29

    Dengue virus is the causative agent of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. High rates of dengue virus replication and virion production are related to disease severity. To identify anti-DENV compounds, we performed cell-based ELISA testing to detect the level of DENV E protein expression. Among a total of 83 inhibitors, eight were identified as inhibitors with antiviral activity. Epidermal growth factor receptor inhibitor II (EGFR/ErbB-2/ErbB-4 inhibitor II) and protein tyrosine phosphatase inhibitor IV (PTP inhibitor IV) significantly inhibited dengue virus production and demonstrated low toxicity in hepatocyte cell lines. Our results suggest the efficacy of tyrosine kinase/phosphatase inhibitors in decreasing dengue virus production in HepG2 cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Targeting and molecular imaging of HepG2 cells using surface-functionalized gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Rathinaraj, Pierson [Auckland University of Technology, Institute of Biomedical Technologies (New Zealand); Lee, Kyubae; Choi, Yuri; Park, Soo-Young [Kyungpook National University, School of Applied Chemical Engineering, Graduate School (Korea, Republic of); Kwon, Oh Hyeong [Kumoh National Institute of Technology, Department of Polymer Science and Engineering (Korea, Republic of); Kang, Inn-Kyu, E-mail: ikkang@knu.ac.kr [Kyungpook National University, School of Applied Chemical Engineering, Graduate School (Korea, Republic of)

    2015-07-15

    Mercaptosuccinic acid (M)-conjugated gold nanoparticles (GM) were prepared and characterized by transmission electron microscope and dynamic light scattering. M was used to improve the monodispersity and non-specific intracellular uptake of nanoparticles. Lactobionic acid (L) was subsequently conjugated to the GM to target preferentially HepG2 cells (liver cancer cells) that express asialoglycoprotein receptors (ASGPR) on their membrane surfaces and facilitate the transit of nanoparticles across the cell membrane. The mean size of lactobionic acid-conjugated gold nanoparticle (GL) was approximately 10 ± 0.2 nm. Finally, the Atto 680 dye (A6) was coupled to the nanoparticles to visualize their internalization into HepG2 cells. The interaction of surface-modified gold nanoparticles with HepG2 cells was studied after culturing cells in media containing the GM or L-conjugated GM (GL)

  18. Inhibitory effects of Cassia tora L. on benzo[a]pyrene-mediated DNA damage toward HepG2 cells.

    Science.gov (United States)

    Wu, C H; Hsieh, C L; Song, T Y; Yen, G C

    2001-05-01

    The effects of water extracts from Cassia tora L. (WECT) treated with different degrees of roasting on benzo[a]pyrene (B[a]P)-induced DNA damage in human hepatoma cell line HepG2 were investigated via the comet assay without exogenous activation mixtures, such as S9 mix. WECT alone, at concentrations of 0.1-2 mg/mL, showed neither cytotoxic nor genotoxic effect toward HepG2 cells. B[a]P-induced DNA damage in HepG2 cells could be reduced by WECT in a dose-dependent manner (P roasted at 150 degrees C (60%) > roasted at 250 degrees C (23%). Ethoxyresorufin-O-dealkylase activity of HepG2 cells was effectively inhibited by WECT, and a similar trend of inhibition was observed in the order unroasted (64%) > roasted at 150 degrees C (42%) > roasted at 250 degrees C (18%). The activity of NADPH cytochrome P-450 reductase was also decreased by unroasted and 150 degrees C-roasted samples (50% and 38%, respectively). Furthermore, glutathione S-transferase activity was increased by treatment with unroasted (1.26-fold) and 150 degrees C-roasted (1.35-fold) samples at 1 mg/mL. In addition, the contents of anthraquinones (AQs) in WECT, including chrysophanol, emodin, and rhein, were decreased with increasing roasting temperature. Each of these AQs also demonstrated significant antigenotoxic activity in the comet assay. The inhibitory effects of chrysophanol, emodin, and rhein on B[a]P-mediated DNA damage in HepG2 cells were 78, 86, and 71%, respectively, at 100 microM. These findings suggested that the decreased antigenotoxicity of the roasted samples might be due to a reduction in their AQs content.

  19. Time- and concentration-dependent effects of resveratrol in HL-60 and HepG2 cells

    DEFF Research Database (Denmark)

    Stervbo, Ulrik; Vang, Ole; Bonnesen, Christine

    2006-01-01

    Resveratrol, a phytochemical present in grapes, has been demonstrated to inhibit tumourigenesis in animal models. However, the specific mechanism by which resveratrol exerts its anticarcinogenic effect has yet to be elucidated. In the present study, the inhibitory effects of resveratrol on cell...... proliferation and apoptosis were evaluated in the human leukaemia cell line HL-60 and the human hepatoma derived cell line HepG2. We found that after a 2 h incubation period, resveratrol inhibited DNA synthesis in a concentration-dependent manner. The IC50 value was 15 μM in both HL-60 and HepG2 cells. When...... the time of treatment was extended, an increase in IC50 value was observed; for example, at 24 h the IC50 value was 30 μM for HL-60 cells and 60 μM for HepG2 cells. Flow cytometry revealed that cells accumulated in different phases of the cell cycle depending on the resveratrol concentration. Furthermore...

  20. Prooxidant and antioxidant properties of salicylaldehyde isonicotinoyl hydrazone iron chelators in HepG2 cells.

    Science.gov (United States)

    Caro, Andres A; Commissariat, Ava; Dunn, Caroline; Kim, Hyunjoo; García, Salvador Lorente; Smith, Allen; Strang, Harrison; Stuppy, Jake; Desrochers, Linda P; Goodwin, Thomas E

    2015-11-01

    Salicylaldehyde isonicotinoyl hydrazone (SIH) is an iron chelator of the aroylhydrazone class that displays antioxidant or prooxidant effects in different mammalian cell lines. Because the liver is the major site of iron storage, elucidating the effect of SIH on hepatic oxidative metabolism is critical for designing effective hepatic antioxidant therapies. Hepatocyte-like HepG2 cells were exposed to SIH or to analogs showing greater stability, such as N'-[1-(2-Hydroxyphenyl)ethyliden]isonicotinoyl hydrazide (HAPI), or devoid of iron chelating properties, such as benzaldehyde isonicotinoyl hydrazone (BIH), and toxicity, oxidative stress and antioxidant (glutathione) metabolism were evaluated. Autoxidation of Fe(2+)in vitro increased in the presence of SIH or HAPI (but not BIH), an effect partially blocked by Fe(2+) chelation. Incubation of HepG2 cells with SIH or HAPI (but not BIH) was non-toxic and increased reactive oxygen species (ROS) levels, activated the transcription factor Nrf2, induced the catalytic subunit of γ-glutamate cysteine ligase (Gclc), and increased glutathione concentration. Fe(2+) chelation decreased ROS and inhibited Nrf2 activation, and Nrf2 knock-down inhibited the induction of Gclc in the presence of HAPI. Inhibition of γ-glutamate cysteine ligase enzymatic activity inhibited the increase in glutathione caused by HAPI, and increased oxidative stress. SIH iron chelators display both prooxidant (increasing the autoxidation rate of Fe(2+)) and antioxidant (activating Nrf2 signaling) effects. Activation by SIH iron chelators of a hormetic antioxidant response contributes to their antioxidant properties and modulates the anti- and pro-oxidant balance. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. Validation of in vitro cell models used in drug metabolism and transport studies; genotyping of cytochrome P450, phase II enzymes and drug transporter polymorphisms in the human hepatoma (HepG2), ovarian carcinoma (IGROV-1) and colon carcinoma (CaCo-2, LS180) cell lines

    International Nuclear Information System (INIS)

    Brandon, Esther F.A.; Bosch, Tessa M.; Deenen, Maarten J.; Levink, Rianne; Wal, Everdina van der; Meerveld, Joyce B.M. van; Bijl, Monique; Beijnen, Jos H.; Schellens, Jan H.M.; Meijerman, Irma

    2006-01-01

    Human cell lines are often used for in vitro biotransformation and transport studies of drugs. In vivo, genetic polymorphisms have been identified in drug-metabolizing enzymes and ABC-drug transporters leading to altered enzyme activity, or a change in the inducibility of these enzymes. These genetic polymorphisms could also influence the outcome of studies using human cell lines. Therefore, the aim of our study was to pharmacogenotype four cell lines frequently used in drug metabolism and transport studies, HepG2, IGROV-1, CaCo-2 and LS180, for genetic polymorphisms in biotransformation enzymes and drug transporters. The results indicate that, despite the presence of some genetic polymorphisms, no real effects influencing the activity of metabolizing enzymes or drug transporters in the investigated cell lines are expected. However, this characterization will be an aid in the interpretation of the results of biotransformation and transport studies using these in vitro cell models

  2. Anti-hepatocarcinoma effects of resveratrol nanoethosomes against human HepG2 cells

    Science.gov (United States)

    Meng, Xiang-Ping; Zhang, Zhen; Chen, Tong-sheng; Wang, Yi-fei; Wang, Zhi-ping

    2017-02-01

    Hepatocarcinoma, a malignant cancer, threaten human life badly. It is a current issue to seek the effective natural remedy from plant to treat cancer due to the resistance of the advanced hepatocarcinoma to chemotherapy. Resveratrol (Res) has been widely investigated with its strong anti-tumor activity. However, its low oral bioavailability restricts its wide application. In this study, we prepared resveratrol nanoethosomes (ResN) via ethanol injection method. The in vitro anti-hepatocarcinoma effects of ResN relative to efficacy of bulk Res were evaluated on proliferation and apoptosis of human HepG2 cells. ResN were spherical vesicles and its particle diameter, zeta potential were (115.8 +/- 1.3) nm and (-12.8 +/- 1.9) mV, respectively. ResN exhibited significant inhibitory effects against human HepG2 cells by MTT assay, and the IC50 value was 49.2 μg/ml (105.4 μg/ml of Res bulk solution). By flow cytometry assay, there was an increase in G2/M phase cells treated with ResN. The results demonstrated ResN could effectively block the G2/M phase of HepG2 cells, which can also enhance the inhibitory effect of Res against HepG2 cells.

  3. Silver Nanoparticles Induce HePG-2 Cells Apoptosis Through ROS-Mediated Signaling Pathways

    Science.gov (United States)

    Zhu, Bing; Li, Yinghua; Lin, Zhengfang; Zhao, Mingqi; Xu, Tiantian; Wang, Changbing; Deng, Ning

    2016-04-01

    Recently, silver nanoparticles (AgNPs) have been shown to provide a novel approach to overcome tumors, especially those of hepatocarcinoma. However, the anticancer mechanism of silver nanoparticles is unclear. Thus, the purpose of this study was to estimate the effect of AgNPs on proliferation and activation of ROS-mediated signaling pathway on human hepatocellular carcinoma HePG-2 cells. A simple chemical method for preparing AgNPs with superior anticancer activity has been showed in this study. AgNPs were detected by transmission electronic microscopy (TEM) and energy dispersive X-ray (EDX). The size distribution and zeta potential of silver nanoparticles were detected by Zetasizer Nano. The average size of AgNPs (2 nm) observably increased the cellular uptake by endocytosis. AgNPs markedly inhibited the proliferation of HePG-2 cells through induction of apoptosis with caspase-3 activation and PARP cleavage. AgNPs with dose-dependent manner significantly increased the apoptotic cell population (sub-G1). Furthermore, AgNP-induced apoptosis was found dependent on the overproduction of reactive oxygen species (ROS) and affecting of MAPKs and AKT signaling and DNA damage-mediated p53 phosphorylation to advance HePG-2 cells apoptosis. Therefore, our results show that the mechanism of ROS-mediated signaling pathways may provide useful information in AgNP-induced HePG-2 cell apoptosis.

  4. Hyperglycemia and anthocyanin inhibit quercetin metabolism in HepG2 cells

    Science.gov (United States)

    A high glucose (Glu) milieu promotes generation of reactive oxygen species, which may not only cause cellular damage, but also modulate phase II enzymes that are responsible for the metabolism of flavonoids. Thus, we examined the effect of a high Glu milieu on quercetin (Q) metabolism in HepG2 cells...

  5. Differential genomic effects of six different TiO2 nanomaterials on human liver HepG2 cells

    Science.gov (United States)

    Engineered nanoparticles are reported to cause liver toxicity in vivo. To better assess the mechanism of the in vivo liver toxicity, we used the human hepatocarcinoma cells (HepG2) as a model system. Human HepG2 cells were exposed to 6 TiO2 nanomaterials (with dry primary partic...

  6. RNA interference mediated inhibition of dengue virus multiplication and entry in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Mohammed Abdelfatah Alhoot

    Full Text Available BACKGROUND: Dengue virus-host cell interaction initiates when the virus binds to the attachment receptors followed by endocytic internalization of the virus particle. Successful entry into the cell is necessary for infection initiation. Currently, there is no protective vaccine or antiviral treatment for dengue infection. Targeting the viral entry pathway has become an attractive therapeutic strategy to block infection. This study aimed to investigate the effect of silencing the GRP78 and clathrin-mediated endocytosis on dengue virus entry and multiplication into HepG2 cells. METHODOLOGY/PRINCIPAL FINDINGS: HepG2 cells were transfected using specific siRNAs to silence the cellular surface receptor (GRP78 and clathrin-mediated endocytosis pathway. Gene expression analysis showed a marked down-regulation of the targeted genes (87.2%, 90.3%, and 87.8% for GRP78, CLTC, and DNM2 respectively in transfected HepG2 cells when measured by RT-qPCR. Intracellular and extracellular viral RNA loads were quantified by RT-qPCR to investigate the effect of silencing the attachment receptor and clathrin-mediated endocytosis on dengue virus entry. Silenced cells showed a significant reduction of intracellular (92.4% and extracellular viral RNA load (71.4% compared to non-silenced cells. Flow cytometry analysis showed a marked reduction of infected cells (89.7% in silenced HepG2 cells compared to non-silenced cells. Furthermore, the ability to generate infectious virions using the plaque assay was reduced 1.07 log in silenced HepG2 cells. CONCLUSIONS/SIGNIFICANCE: Silencing the attachment receptor and clathrin-mediated endocytosis using siRNA could inhibit dengue virus entry and multiplication into HepG2 cells. This leads to reduction of infected cells as well as the viral load, which might function as a unique and promising therapeutic agent for attenuating dengue infection and prevent the development of dengue fever to the severe life-threatening DHF or DSS

  7. Pfaffosidic Fraction from Hebanthe paniculata Induces Cell Cycle Arrest and Caspase-3-Induced Apoptosis in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Tereza Cristina da Silva

    2015-01-01

    Full Text Available Hebanthe paniculata roots (formerly Pfaffia paniculata and popularly known as Brazilian ginseng show antineoplastic, chemopreventive, and antiproliferative properties. Functional properties of these roots and their extracts are usually attributed to the pfaffosidic fraction, which is composed mainly by pfaffosides A–F. However, the therapeutic potential of this fraction in cancer cells is not yet entirely understood. This study aimed to analyze the antitumoral effects of the purified pfaffosidic fraction or saponinic fraction on the human hepatocellular carcinoma HepG2 cell line. Cellular viability, proliferation, and apoptosis were evaluated, respectively, by MTT assay, BrdU incorporation, activated caspase-3 immunocytochemistry, and DNA fragmentation assay. Cell cycle was analyzed by flow cytometry and the cell cycle-related proteins were analyzed by quantitative PCR and Western blot. The cells exposed to pfaffosidic fraction had reduced viability and cellular growth, induced G2/M at 48 h or S at 72 h arrest, and increased sub-G1 cell population via cyclin E downregulation, p27KIP1 overexpression, and caspase-3-induced apoptosis, without affecting the DNA integrity. Antitumoral effects of pfaffosidic fraction from H. paniculata in HepG2 cells originated by multimechanisms of action might be associated with cell cycle arrest in the S phase, by CDK2 and cyclin E downregulation and p27KIP1 overexpression, besides induction of apoptosis through caspase-3 activation.

  8. Stimulation of LDL receptor activity in Hep-G2 cells by a serum factor(s)

    International Nuclear Information System (INIS)

    Ellsworth, J.L.; Brown, C.; Cooper, A.D.

    1988-01-01

    The regulation of low-density lipoprotein (LDL) receptor activity in the human hepatoma cell line Hep-G2 by serum components was examined. Incubation of dense monolayers of Hep-G2 cells with fresh medium containing 10% fetal calf serum (FM) produced a time-dependent increase in LDL receptor activity. Uptake and degradation of 125I-LDL was stimulated two- to four-fold, as compared with that of Hep-G2 cells cultured in the same media in which they had been grown to confluence (CM); the maximal 125I-LDL uptake plus degradation increased from 0.2 microgram/mg cell protein/4 h to 0.8 microgram/mg cell protein/4 h. In addition, a two-fold increase in cell surface binding of 125I-LDL to Hep-G2 cells was observed when binding was measured at 4 degrees C. There was no change in the apparent Kd. The stimulation of LDL receptor activity was suppressed in a concentration-dependent manner by the addition of cholesterol, as LDL, to the cell medium. In contrast to the stimulation of LDL receptor activity, FM did not affect the uptake or degradation of 125I-asialoorosomucoid. Addition of FM increased the protein content per dish, and DNA synthesis was stimulated approximately five-fold, as measured by [3H]thymidine incorporation into DNA; however, the cell number did not change. Cellular cholesterol biosynthesis was also stimulated by FM; [14C]acetate incorporation into unesterified and esterified cholesterol was increased approximately five-fold. Incubation of Hep-G2 cells with high-density lipoproteins (200 micrograms protein/ml) or albumin (8.0 mg/ml) in the absence of the serum factor did not significantly increase the total processed 125I-LDL. Stimulation of LDL receptor activity was dependent on a heat-stable, nondialyzable serum component that eluted in the inclusion volume of a Sephadex G-75 column

  9. Modulation of the pentose phosphate pathway alters phase I metabolism of testosterone and dextromethorphan in HepG2 cells.

    Science.gov (United States)

    Xiao, Wen-jing; Ma, Ting; Ge, Chun; Xia, Wen-juan; Mao, Yong; Sun, Run-bin; Yu, Xiao-yi; Aa, Ji-ye; Wang, Guang-ji

    2015-02-01

    The pentose phosphate pathway (PPP) is involved in the activity of glucose-6-phosphate dehydrogenase (G6PD) and generation of NADPH, which plays a key role in drug metabolism. The aim of this study was to investigate the effects of modulation of the PPP on drug metabolism capacity in vitro. A pair of hepatic cell lines, ie, the cancerous HepG2 cells and normal L02 cells, was used. The expression of CYP450 enzymes, p53 and G6PD in the cells were analyzed. The metabolism of testosterone (TEST, 10 μmol/L) and dextromethorphan (DEM, 1 μmol/L), the two typical substrates for CYP3A4 and CYP2D6, in the cells was examined in the presence of different agents. Both the expression and metabolic activities of CYP3A4 and CYP2D6 were considerably higher in HepG2 cells than in L02 cells. The metabolism of TEST and DEM in HepG2 cells was dose-dependently inhibited by the specific CYP3A4 inhibitor ketoconazole and CYP2D6 inhibitor quinidine. Addition of the p53 inhibitor cyclic PFT-α (5, 25 μmol/L) in HepG2 cells dose-dependently enhanced the metabolism of DEM and TEST, whereas addition of the p53 activator NSC 66811 (3, 10, 25 μmol/L) dose-dependently inhibited the metabolism. Furthermore, addition of the G6PD inhibitor 6-aminonicotinamide (5, 15 μmol/L) in HepG2 cells dose-dependently inhibited the metabolism of DEM and TEST, whereas addition of the PPP activity stimulator menadione (1, 5, 15 μmol/L) dose-dependently enhanced the metabolism. Modulation of p53 and the PPP alters the metabolism of DEM and TEST, suggesting that the metabolic flux pattern of PPP may be closely involved in drug metabolism and the individual variance.

  10. Caspase-independent cell death mediated by apoptosis-inducing factor (AIF) nuclear translocation is involved in ionizing radiation induced HepG2 cell death

    International Nuclear Information System (INIS)

    Sun, Hengwen; Yang, Shana; Li, Jianhua; Zhang, Yajie; Gao, Dongsheng; Zhao, Shenting

    2016-01-01

    Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. The aim of radiotherapy is to eradicate cancer cells with ionizing radiation. Except for the caspase-dependent mechanism, several lines of evidence demonstrated that caspase-independent mechanism is directly involved in the cell death responding to irradiation. For this reason, defining the contribution of caspase-independent molecular mechanisms represents the main goal in radiotherapy. In this study, we focused on the role of apoptosis-inducing factor (AIF), the caspase-independent molecular, in ionizing radiation induced hepatocellular carcinoma cell line (HepG2) cell death. We found that ionizing radiation has no function on AIF expression in HepG2 cells, but could induce AIF release from the mitochondria and translocate into nuclei. Inhibition of AIF could reduce ionizing radiation induced HepG2 cell death. These studies strongly support a direct relationship between AIF nuclear translocation and radiation induced cell death. What's more, AIF nuclear translocation is caspase-independent manner, but not caspase-dependent manner, in this process. These new findings add a further attractive point of investigation to better define the complex interplay between caspase-independent cell death and radiation therapy. - Highlights: • AIF nuclear translocation is involved in ionizing radiation induced hepatocellular carcinoma cell line HepG2 cell death. • AIF mediated cell death induced by ionizing radiation is caspase-independent. • Caspase-independent pathway is involved in ionzing radiation induced HepG2 cell death.

  11. Caspase-independent cell death mediated by apoptosis-inducing factor (AIF) nuclear translocation is involved in ionizing radiation induced HepG2 cell death

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Hengwen [Department of Radiation, Cancer Center of Guangdong General Hospital (Guangdong Academy of Medical Science), Guangzhou, 510080, Guangdong (China); Yang, Shana; Li, Jianhua [Department of Physiology, Guangzhou Medical University, Guangzhou, 510182, Guangdong (China); Zhang, Yajie [Department of Pathology, Guangzhou Medical University, Guangzhou, 510182, Guangdong (China); Gao, Dongsheng [Department of Oncology, Guangdong Medical College Affiliated Pengpai Memorial Hospital, Hai Feng, 516400, Gungdong (China); Zhao, Shenting, E-mail: zhaoshenting@126.com [Department of Physiology, Guangzhou Medical University, Guangzhou, 510182, Guangdong (China)

    2016-03-25

    Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. The aim of radiotherapy is to eradicate cancer cells with ionizing radiation. Except for the caspase-dependent mechanism, several lines of evidence demonstrated that caspase-independent mechanism is directly involved in the cell death responding to irradiation. For this reason, defining the contribution of caspase-independent molecular mechanisms represents the main goal in radiotherapy. In this study, we focused on the role of apoptosis-inducing factor (AIF), the caspase-independent molecular, in ionizing radiation induced hepatocellular carcinoma cell line (HepG2) cell death. We found that ionizing radiation has no function on AIF expression in HepG2 cells, but could induce AIF release from the mitochondria and translocate into nuclei. Inhibition of AIF could reduce ionizing radiation induced HepG2 cell death. These studies strongly support a direct relationship between AIF nuclear translocation and radiation induced cell death. What's more, AIF nuclear translocation is caspase-independent manner, but not caspase-dependent manner, in this process. These new findings add a further attractive point of investigation to better define the complex interplay between caspase-independent cell death and radiation therapy. - Highlights: • AIF nuclear translocation is involved in ionizing radiation induced hepatocellular carcinoma cell line HepG2 cell death. • AIF mediated cell death induced by ionizing radiation is caspase-independent. • Caspase-independent pathway is involved in ionzing radiation induced HepG2 cell death.

  12. LRD-22, a novel dual dithiocarbamatic acid ester, inhibits Aurora-A kinase and induces apoptosis and cell cycle arrest in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Huiling; Li, Ridong [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China); Li, Li [Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing (China); Ge, Zemei [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China); Zhou, Rouli, E-mail: rlzhou@bjmu.edu.cn [Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Runtao, E-mail: lirt@bjmu.edu.cn [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China)

    2015-02-27

    In this study we investigated the antitumor activity of the novel dual dithiocarbamatic acid ester LRD-22 in vitro and in vivo. Several cancer cell lines were employed to determine the effect of LRD-22 on cell growth, and the MTT assay showed there was a significant decrease in viable tumor cell numbers in the presence of LRD-22, especially in the HepG2 cell line. Colony formation assay also showed LRD-22 strongly inhibits HepG2 cell growth. Evaluation of the mechanism involved showed that inhibitory effects of LRD-22 on cell growth are due to induction of apoptosis and G2/M arrest. LRD-22 inhibited Aurora-A phosphorylation at Thr{sub 288} and subsequently impaired p53 phosphorylation at Ser{sub 315} which was associated with the proteasome degradation pathway. Tumor suppressor protein p53 is stabilized by this mechanism and accumulates through inhibition of Aurora-A kinase activity via treatment with LRD-22. In vivo study of HepG2 xenograft in nude mice also shows LRD-22 suppresses tumor growth at a concentration of 5 mg/kg without animals suffering loss of body weight. In conclusion, our results demonstrate LRD-22 acts as an Aurora-A kinase inhibitor to induce apoptosis and inhibit proliferation in HepG2 cells, and should be considered as a promising targeting agent for HCC therapy. - Highlights: • LRD-22 significantly inhibits cancer cell growth, especially in the HepG2 cell line. • The inhibitory effect of LRD-22 is due to induction of apoptosis and cell cycle arrest. • LRD-22 inhibits Aurora-A phosphorylation which results in subsequent impairment of the p53 pathway. • LRD-22 suppresses tumor growth in xenograft mice without body weight loss.

  13. Oridonin increases anticancer effects of lentinan in HepG2 human hepatoblastoma cells.

    Science.gov (United States)

    Sun, Zhiqiang; Han, Qinghe; Duan, Liwei; Yuan, Qinghai; Wang, Hui

    2018-02-01

    The aim of the present study was to investigate whether oridonin is able to increase the effects of lentinan (LNT) in HepG2 human hepatoblastoma cells by MTT, flow cytometry, reverse transcription-quantitative polymerase chain reaction and western blot analysis. The in vitro results demonstrated that 20 µg/ml of oridonin was a nontoxic concentration for L02 normal liver cells and HepG2 liver cancer cells. Furthermore, treatment with 0-200 µg/ml LNT was only able to decrease the viability of HepG2 liver cancer cells. The growth inhibitory rate of the LNT-L (100 µg/ml) treatment group was 20.7% and the rate of the LNT-H (200 µg/ml) treatment group was 54.8%. Notably, the growth inhibitory rate of the oridonin + LNT-H group was 84.3%. The highest percentage of apoptotic cells was observed in the oridonin + LNT-H group (20 µg/ml oridonin and 200 µg/ml LNT). The percentage of apoptotic cells in the oridonin + LNT-H group was significantly different from the percentage of apoptotic cells in the LNT-H (26.1%) and the LNT-L (16.8%) groups. Treatment with LNT produced an increase in caspase-3, caspase-9, Bcl-2-like protein 4, p53, p21, nuclear factor κB inhibitor-α mRNA and protein expression and a decrease in B-cell lymphoma 2 and nuclear factor-κB expression in HepG2 cells compared with untreated control cells. Treatment with a combination of oridonin and LNT-H induced a further increase in expression with the biggest differences in expression observed between the oridonin + LNT-H group and control. It was observed that treatment with oridonin was able to increase the anticancer effects of LNT in HepG2 cells. Therefore, oridonin may be used to sensitize cells to LNT.

  14. Aglycemia keeps mitochondrial oxidative phosphorylation under hypoxic conditions in HepG2 cells

    Czech Academy of Sciences Publication Activity Database

    Plecitá-Hlavatá, Lydie; Ježek, Jan; Ježek, Petr

    2015-01-01

    Roč. 47, č. 6 (2015), s. 467-476 ISSN 0145-479X R&D Projects: GA ČR(CZ) GAP302/10/0346; GA ČR(CZ) GA13-02033S; GA MŠk(CZ) LH11055 Institutional support: RVO:67985823 Keywords : cancer mitochondria * non-canonical response to hypoxia * hypoxia- inducible factor * glutaminolysis * HepG2 cells Subject RIV: ED - Physiology Impact factor : 2.080, year: 2015

  15. Downregulation of human paraoxonase 1 (PON1) by organophosphate pesticides in HepG2 cells.

    Science.gov (United States)

    Medina-Díaz, Irma Martha; Ponce-Ruiz, Néstor; Ramírez-Chávez, Bryana; Rojas-García, Aurora Elizabeth; Barrón-Vivanco, Briscia S; Elizondo, Guillermo; Bernal-Hernández, Yael Y

    2017-02-01

    Paraoxonase 1 (PON1) is a calcium-dependent esterase synthesized primarily in the liver and secreted into the plasma where it is associated with high-density lipoproteins (HDL). PON1 hydrolyzes and detoxifies some toxic metabolites of organophosphorus compounds (OPs) such as methyl parathion and chlorpyrifos. Thus, PON1 activity and expression levels are important for determining susceptibility against OPs poisoning. Some studies have demonstrated that OPs can modulate gene expression through interactions with nuclear receptors. In this study, we evaluated the effects of methyl parathion and chlorpyrifos on the modulation of PON1 in Human Hepatocellular Carcinoma (HepG2) cells by real-time PCR, PON1 activity assay, and western blot. The results showed that the treatments with methyl parathion and chlorpyrifos decreased PON1 mRNA and immunoreactive protein and increased inflammatory cytokines in HepG2 cells. The effects of methyl parathion and chlorpyrifos on the downregulation of PON1 gene expression in HepG2 cells may provide evidence of OPs cytotoxicity related to oxidative stress and an inflammatory response. A decrease in the expression of the PON1 gene may increase the susceptibility to OPs intoxication and the risk of diseases related to inflammation and oxidative stress. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 490-500, 2017. © 2016 Wiley Periodicals, Inc.

  16. Tripterygium regelii decreases the biosynthesis of triacylglycerol and cholesterol in HepG2 cells.

    Science.gov (United States)

    Kang, Myung-Ji; Kwon, Eun-Bin; Yuk, Heung Joo; Ryu, Hyung Won; Kim, Soo-Yeon; Lee, Mi-Kyeong; Moon, Dong-Oh; Lee, Su Ui; Oh, Sei-Ryang; Lee, Hyun-Sun; Kim, Mun-Ock

    2017-12-01

    In the course of screening to find a plant material decreasing the activity of triacylglycerol and cholesterol, we identified Tripterygium regelii (TR). The methanol extract of TR leaves (TR-LM) was shown to reduce the intracellular lipid contents consisting of triacylglycerol (TG) and cholesterol in HepG2 cells. TR-LM also downregulated the mRNA and protein expression of the lipogenic genes such as SREBP-1 and its target enzymes. Consequently, TR-LM reduced the TG biosynthesis in HepG2 cells. In addition, TR-LM decreased SREBP2 and its target enzyme HMG-CoA reductase, which is involved in cholesterol synthesis. In this study, we evaluated that TR-LM attenuated cellular lipid contents through the suppression of de novo TG and cholesterol biosynthesis in HepG2 cells. All these taken together, TR-LM could be beneficial in regulating lipid metabolism and useful preventing the hyperlipidemia and its complications, in that liver is a crucial tissue for the secretion of serum lipids.

  17. Specific binding of tubeimoside-2 with proteins in hepatocarcinoma HepG2 cells: investigation by molecular spectroscopy

    Science.gov (United States)

    Yang, Sun; Shi-Sheng, Sun; Ying-Yong, Zhao; Jun, Fan

    2012-07-01

    In this study, we compared different binding interactions of TBMS2 with proteins both in hepatocarcinoma HepG2 cells and in normal embryo hepatic L02 cells by using fluorescence, absorption, and CD spectroscopy. The fluorescence data revealed that the fluorescence intensity of proteins in the HepG2 and L02 cells decreased in the presence of TBMS2 by 30.79% and 12.01%, respectively. Binding constants and thermodynamic parameters were obtained for systems of TBMS2 with the two kinds of cell proteins. The results indicated that HepG2 cell proteins had a higher TBMS2 binding activity than those in the L02 cells. Analysis of the TBMS2 cytotoxic activities showed that TBMS2 could selectively induce apoptosis of HepG2 cells by binding to them, while its apoptotic effect on L02 cells was relatively weaker.

  18. Free radical generation from an aniline derivative in HepG2 cells: a possible captodative effect.

    Science.gov (United States)

    Horinouchi, Yuya; Summers, Fiona A; Ehrenshaft, Marilyn; Mason, Ronald P

    2015-01-01

    Xenobiotic metabolism can induce the generation of protein radicals, which are believed to play an important role in the toxicity of chemicals and drugs. It is therefore important to identify chemical structures capable of inducing macromolecular free radical formation in living cells. In this study, we evaluated the ability of four structurally related environmental chemicals, aniline, nitrosobenzene, N,N-dimethylaniline, and N,N-dimethyl-4-nitrosoaniline (DMNA), to induce free radicals and cellular damage in the hepatoma cell line HepG2. Cytotoxicity was assessed using lactate dehydrogenase assays, and morphological changes were observed using phase contrast microscopy. Protein free radicals were detected by immuno-spin trapping using in-cell western experiments and confocal microscopy to determine the subcellular locale of free radical generation. DMNA induced free radical generation, lactate dehydrogenase release, and morphological changes in HepG2 cells, whereas aniline, nitrosobenzene, N,N-dimethylaniline did not. Confocal microscopy showed that DMNA induced free radical generation mainly in the cytosol. Preincubation of HepG2 cells with N-acetylcysteine and 2,2'-dipyridyl significantly prevented free radical generation on subsequent incubation with DMNA, whereas preincubation with apocynin and dimethyl sulfoxide had no effect. These results suggest that DMNA is metabolized to reactive free radicals capable of generating protein radicals which may play a critical role in DMNA toxicity. We propose that the captodative effect, the combined action of the electron-releasing dimethylamine substituent, and the electron-withdrawing nitroso substituent, leads to a thermodynamically stabilized radical, facilitating enhanced protein radical formation by DMNA. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Investigation of quercetin-induced HepG2 cell apoptosis-associated cellular biophysical alterations by atomic force microscopy.

    Science.gov (United States)

    Pi, Jiang; Li, Baole; Tu, Lvying; Zhu, Haiyan; Jin, Hua; Yang, Fen; Bai, Haihua; Cai, Huaihong; Cai, Jiye

    2016-01-01

    Quercetin, a wildly distributed bioflavonoid, has been proved to possess excellent antitumor activity on hepatocellular carcinoma (HCC). In the present study, the biophysical properties of HepG2 cells were qualitatively and quantitatively determined using high resolution atomic force microscopy (AFM) to understand the anticancer effects of quercetin on HCC cells at nanoscale. The results showed that quercetin could induce severe apoptosis in HepG2 cells through arrest of cell cycle and disruption of mitochondria membrane potential. Additionally, the nuclei and F-actin structures of HepG2 cells were destroyed by quercetin treatment as well. AFM morphological data showed some typical apoptotic characterization of HepG2 cells with increased particle size and roughness in the ultrastructure of cell surface upon quercetin treatment. As an important biophysical property of cells, the membrane stiffness of HepG2 cells was further quantified by AFM force measurements, which indicated that HepG2 cells became much stiffer after quercetin treatment. These results collectively suggest that quercetin can be served as a potential therapeutic agent for HCC, which not only extends our understanding of the anticancer effects of quercetin against HCC cells into nanoscale, but also highlights the applications of AFM for the investigation of anticancer drugs. © Wiley Periodicals, Inc.

  20. Differential genomic effects on signaling pathways by two different CeO2 nanoparticles in HepG2 cells

    Data.gov (United States)

    U.S. Environmental Protection Agency — Differential genomic effects on signaling pathways by two different CeO2 nanoparticles in HepG2 cells. This dataset is associated with the following publication:...

  1. Toxicity evaluation of surface water treated with different disinfectants in HepG2 cells.

    Science.gov (United States)

    Marabini, Laura; Frigerio, Silvia; Chiesara, Enzo; Radice, Sonia

    2006-01-01

    It is well known that water disinfection through chlorination causes the formation of a mixture of disinfection by-products (DBPs), many of which are genotoxic and carcinogenic. To demonstrate the formation of such compounds, a pilot water plant supplied with water from Lake Trasimeno was set up at the waterworks of Castiglione del Lago (PG, Italy). The disinfectants, continuously added to pre-filtered lake water flowing into three different basins, were sodium hypochlorite, chlorine dioxide and peracetic acid, an alternative disinfectant used until now for disinfecting waste waters, but not yet studied for a possible use in drinking water treatment. The aim of this study was to evaluate the formation during the disinfection processes of some toxic compounds that could explain the genotoxic effects of drinking waters. Differently treated waters were concentrated by solid-phase adsorption on silica C(18) columns and toxicity was assessed in a line of human hepatoma cells (HepG2), a metabolically competent cellular line very useful for human risk evaluation. The seasonal variability of the physical-chemical water characteristics (AOX, UV 254 nm, potential formation of THM, pH and temperature) made indispensable experimentation with water samples taken during the various seasons. Autumn waters cause greater toxicity compared to those of other seasons, in particular dilution of the concentrate at 0.5l equivalent of disinfected waters with chlorine dioxide and peracetic acid causes a 55% reduction in cellular vitality while the cellular vitality is over 80% with the all other water concentrates. Moreover it is very interesting underline that non-cytotoxic quantities of the autumnal water concentrates cause, after 2h treatment, a decrease in GSH and a statistically significant increase in oxygen radicals, while after prolonged treatment (24h) cause a GSH increase, without variations in the oxygen radical content. This phenomenon could be interpreted as the cellular

  2. Metallomics Study of CdSe/ZnS Quantum Dots in HepG2 Cells.

    Science.gov (United States)

    Peng, Lu; He, Man; Chen, Beibei; Qiao, Yu; Hu, Bin

    2015-10-27

    Toxicity of quantum dots (QDs) has been a hot research concern in the past decade, and there is a lot of challenge in this field. The physicochemical characteristics of QDs can affect their toxicity, while little is known about the specific chemical form of QDs in living cells after incubation so far. In this work, speciation of four CdSe/ZnS QDs in HepG2 cells was carried out from the metallomics' point of view for the first time by using size exclusion chromatography (SEC) coupled with inductively coupled plasma-mass spectrometry (ICP-MS). On the basis of the signal of Cd, two kinds of chemical forms, named as QD-1 and QD-2, were observed in HepG2 cells incubated with CdSe/ZnS QDs. QD-1 was demonstrated to be a kind of QD-like nanoparticles, confirmed by chromatographic retention time, transmission electron microscopy (TEM) characterization, and fluorescence detection. QD-2 was demonstrated to be cadmium-metallothioneins complex (Cd-MTs) by reversed phase liquid chromatography (RPLC) synchronously coupled with ICP-MS and electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-Q-TOF-MS) analysis. Meanwhile, speciation of QDs in HepG2 cells incubated with different conditions was analyzed. With the variation of QDs incubation concentration/time, and elimination time, the species of QD-1 and QD-2 were also observed without other obvious species, and both the amount of QD-1 and QD-2 increased with incubation concentration and time. The obtained results provide valuable information and a strategy for the study of existing chemical form of QDs, greatly benefiting the understanding of QDs toxicity in living cells.

  3. Campomanesia adamantium (Myrtaceae fruits protect HEPG2 cells against carbon tetrachloride-induced toxicity

    Directory of Open Access Journals (Sweden)

    Thaís de Oliveira Fernandes

    2015-01-01

    Full Text Available Campomanesia adamantium (Myrtaceae is an antioxidant compounds-rich Brazilian fruit popularly known as gabiroba. In view of this, it was evaluated the hepatoprotective effects of pulp (GPE or peel/seed (GPSE hydroalcoholic extracts of gabiroba on injured liver-derived HepG2 cells by CCl4 (4 mM. The results showed the presence of total phenolic in GPSE was (60% higher when compared to GPE, associated with interesting antioxidant activity using DPPH·− assay. Additionally, HPLC chromatograms and thin layer chromatography of GPE and GPSE showed the presence of flavonoids. Pretreatment of HepG2 cells with GPE or GPSE (both at 800–1000 μg/mL significantly (p < 0.0001 protected against cytotoxicity induced by CCl4. Additionally, the cells treated with both extracts (both at 1000 μg/mL showed normal morphology (general and nuclear contrasting with apoptotic characteristics in the cells only exposed to CCl4. In these experiments, GPSE also was more effective than GPE. In addition, CCl4 induced a marked increase in AST (p < 0.05 and ALT (p < 0.0001 levels, while GPE or GPSE significantly (p < 0.0001 reduced these levels, reaching values found in the control group. In conclusion, the results suggest that gabiroba fruits exert hepatoprotective effects on HepG2 cells against the CCl4-induced toxicity, probably, at least in part, associated with the presence of antioxidant compounds, especially flavonoids.

  4. Differential Cytotoxicity of Acetaminophen in Mouse Macrophage J774.2 and Human Hepatoma HepG2 Cells: Protection by Diallyl Sulfide.

    Directory of Open Access Journals (Sweden)

    Haider Raza

    Full Text Available Non-steroidal anti-inflammatory drugs (NSAIDs, including acetaminophen (APAP, have been reported to induce cytotoxicity in cancer and non-cancerous cells. Overdose of acetaminophen (APAP causes liver injury in humans and animals. Hepatic glutathione (GSH depletion followed by oxidative stress and mitochondrial dysfunction are believed to be the main causes of APAP toxicity. The precise molecular mechanism of APAP toxicity in different cellular systems is, however, not clearly understood. Our previous studies on mouse macrophage J774.2 cells treated with APAP strongly suggest induction of apoptosis associated with mitochondrial dysfunction and oxidative stress. In the present study, using human hepatoma HepG2 cells, we have further demonstrated that macrophages are a more sensitive target for APAP-induced toxicity than HepG2 cells. Using similar dose- and time-point studies, a marked increase in apoptosis and DNA fragmentation were seen in macrophages compared to HepG2 cells. Differential effects of APAP on mitochondrial respiratory functions and oxidative stress were observed in the two cell lines which are presumably dependent on the varying degree of drug metabolism by the different cytochrome P450s and detoxification by glutathione S-transferase enzyme systems. Our results demonstrate a marked increase in the activity and expression of glutathione transferase (GST and multidrug resistance (MDR1 proteins in APAP-treated HepG2 cells compared to macrophages. This may explain the apparent resistance of HepG2 cells to APAP toxicity. However, treatment of these cells with diallyl sulfide (DAS, 200 μM, a known chemopreventive agent from garlic extract, 24 h prior to APAP (10 μmol/ml for 18h exhibited comparable cytoprotective effects in the two cell lines. These results may help in better understanding the mechanism of cytotoxicity caused by APAP and cytoprotection by chemopreventive agents in cancer and non-cancerous cellular systems.

  5. Effects of hydrogen peroxide and apolipoprotein E isoforms on apolipoprotein E trafficking in HepG2 cells.

    Science.gov (United States)

    Sabaretnam, Tharani; Harris, Matthew J; Kockx, Maaike; Witting, Paul K; Le Couteur, David G; Kritharides, Leonard

    2009-12-01

    1. The major source of apolipoprotein E (apoE) is the liver. In the present study, the effects of oxidative stress and apoE isoforms on apoE distribution and trafficking were established using the HepG2 liver tumour cell line. 2. Hydrogen peroxide (0, 25, 250 and 1000 micromol/L) was associated with rapid and concentration-dependent redistribution of apoE into the early endosomal compartment. This redistribution was achieved with a much lower concentration (25 micromol/L) than that needed to induce changes in intracellular apoE mRNA expression, apoE protein levels and markers of oxidative stress (250-1000 micromol/L). 3. Live cell imaging of apoE3-green fluorescent protein revealed a significant decrease in traffic velocity in response to oxidative stress. 4. The E4 isoform was associated with reduced trafficking velocity compared with the E3 isoform under basal conditions. 5. The results indicate that oxidative stress and apoE isoforms influence apoE trafficking and distribution within HepG2 cells. Altered apoE hepatocyte trafficking may provide a mechanistic link between oxidative stress, ageing and some diseases in older people.

  6. Crambescin C1 Exerts a Cytoprotective Effect on HepG2 Cells through Metallothionein Induction

    Science.gov (United States)

    Roel, María; Rubiolo, Juan A.; Ternon, Eva; Thomas, Olivier P.; Vieytes, Mercedes R.; Botana, Luis M.

    2015-01-01

    The Mediterranean marine sponge Crambe crambe is the source of two families of guanidine alkaloids known as crambescins and crambescidins. Some of the biological effects of crambescidins have been previously reported while crambescins have undergone little study. Taking this into account, we performed comparative transcriptome analysis to examine the effect of crambescin-C1 (CC1) on human tumor hepatocarcinoma cells HepG2 followed by validation experiments to confirm its predicted biological activities. We report herein that, while crambescin-A1 has a minor effect on these cells, CC1 protects them against oxidative injury by means of metallothionein induction even at low concentrations. Additionally, at high doses, CC1 arrests the HepG2 cell cycle in G0/G1 and thus inhibits tumor cell proliferation. The findings presented here provide the first detailed approach regarding the different effects of crambescins on tumor cells and provide a basis for future studies on other possible cellular mechanisms related to these bioactivities. PMID:26225985

  7. Taurine reduces the secretion of apolipoprotein B100 and lipids in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Nagao Koji

    2008-10-01

    Full Text Available Abstract Background Higher concentrations of serum lipids and apolipoprotein B100 (apoB are major individual risk factors of atherosclerosis and coronary heart disease. Therefore ameliorative effects of food components against the diseases are being paid attention in the affluent countries. The present study was undertaken to investigate the effect of taurine on apoB secretion and lipid metabolism in human liver model HepG2 cells. Results The results demonstrated that an addition of taurine to the culture media reduces triacylglycerol (TG-mass in the cells and the medium. Similarly, cellular cholesterol-mass was decreased. Taurine inhibited the incorporation of [14C] oleate into cellular and medium TG, suggesting the inhibition of TG synthesis. In addition, taurine reduced the synthesis of cellular cholesterol ester and its secretion, suggesting the inhibition of acyl-coenzyme A:cholesterol acyltransferase activity. Furthermore, taurine reduced the secretion of apoB, which is a major protein component of very low-density lipoprotein. Conclusion This is a first report to demonstrate that taurine inhibits the secretion of apoB from HepG2 cells.

  8. Proanthocyanidins modulate microRNA expression in human HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Anna Arola-Arnal

    Full Text Available Mi(croRNAs are small non-coding RNAs of 18-25 nucleotides in length that modulate gene expression at the post-transcriptional level. These RNAs have been shown to be involved in a several biological processes, human diseases and metabolic disorders. Proanthocyanidins, which are the most abundant polyphenol class in the human diet, have positive health effects on a variety of metabolic disorders such as inflammation, obesity, diabetes and insulin resistance. The present study aimed to evaluate whether proanthocyanidin-rich natural extracts modulate miRNA expression. Using microarray analysis and Q-PCR, we investigated miRNA expression in HepG2 cells treated with proanthocyanidins. Our results showed that when HepG2 cells were treated with grape seed proanthocyanidin extract (GSPE, cocoa proanthocyanidin extract (CPE or pure epigallocatechin gallate isolated from green tea (EGCG, fifteen, six and five differentially expressed miRNAs, respectively, were identified out of 904 mRNAs. Specifically, miR-30b* was downregulated by the three treatments, and treatment with GSPE or CPE upregulated miR-1224-3p, miR-197 and miR-532-3p. Therefore, these results provide evidence of the capacity of dietary proanthocyanidins to influence microRNA expression, suggesting a new mechanism of action of proanthocyanidins.

  9. Metabolic Flux Distribution during Defatting of Steatotic Human Hepatoma (HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Gabriel Yarmush

    2016-01-01

    Full Text Available Methods that rapidly decrease fat in steatotic hepatocytes may be helpful to recover severely fatty livers for transplantation. Defatting kinetics are highly dependent upon the extracellular medium composition; however, the pathways involved are poorly understood. Steatosis was induced in human hepatoma cells (HepG2 by exposure to high levels of free fatty acids, followed by defatting using plain medium containing no fatty acids, or medium supplemented with a cocktail of defatting agents previously described before. We measured the levels of 28 extracellular metabolites and intracellular triglyceride, and fed the data into a steady-state mass balance model to estimate strictly intracellular fluxes. We found that during defatting, triglyceride content decreased, while beta-oxidation, the tricarboxylic acid cycle, and the urea cycle increased. These fluxes were augmented by defatting agents, and even more so by hyperoxic conditions. In all defatting conditions, the rate of extracellular glucose uptake/release was very small compared to the internal supply from glycogenolysis, and glycolysis remained highly active. Thus, in steatotic HepG2 cells, glycolysis and fatty acid oxidation may co-exist. Together, these pathways generate reducing equivalents that are supplied to mitochondrial oxidative phosphorylation.

  10. Pine needle oil induces G2/M arrest of HepG2 cells by activating the ATM pathway.

    Science.gov (United States)

    Qiu, Bing; Jiang, Wei; Qiu, Wenliang; Mu, Wenling; Qin, Yujing; Zhu, Yongcui; Zhang, Jianying; Wang, Qingyi; Liu, Dongjie; Qu, Zhangyi

    2018-02-01

    Over the last two decades, inducing DNA damage of cancer cells by natural medicines has become a research hotspot in the field of cancer treatment. Although various natural medicines have anticancer effects, very few studies have been conducted to explore the anti-cancer effect of pine needle oil. In the present study, the role of pine needle oil in inducing G2/M arrest in HepG2 cells was investigated. The data revealed that pine needle oil could induce DNA damage in a dose-dependent manner. In the pine needle oil-treated HepG2 cells, the protein levels of phosphorylated (p)-ataxia-telangiectasia mutated (ATM), γ-H2A histone family, member X, p-p53, p-checkpoint kinase 2 and p-cell division cycle 25C were evidently increased, indicating that pine needle oil facilitated G2/M arrest in HepG2 cells through the ATM pathway. In response to the treatment with pine needle oil, ATM was activated in HepG2 cells, which subsequently phosphorylated downstream targets and induced G2/M arrest. In summary, the data of the present study indicated that pine needle oil induces G2/M arrest in HepG2 cells by facilitating ATM activation.

  11. Regulation of human CYP27A1 by estrogens and androgens in HepG2 and prostate cells.

    Science.gov (United States)

    Tang, Wanjin; Norlin, Maria; Wikvall, Kjell

    2007-06-01

    The regulation of the human CYP27A1 gene by estrogens and androgens was studied in human liver-derived HepG2 and prostate cells. Our results show that the promoter activity, enzymatic activity and mRNA levels of CYP27A1 in HepG2 cells are downregulated by estrogen in presence of ERalpha or ERbeta. Similar effects by estrogen were found in RWPE-1 prostate cells. In contrast, estrogen markedly upregulated the transcriptional activity of CYP27A1 in LNCaP prostate cancer cells. 5alpha-Dihydrotestosterone and androgen receptor upregulated the transcriptional activity of CYP27A1 in HepG2 cells. Progressive deletion experiments indicate that the ERbeta-mediated effects in HepG2 and LNCaP cells are conferred to the same region (-451/+42) whereas ERalpha-mediated effects on this promoter are more complex. The results indicate that the stimulating effect of androgen in HepG2 cells is conferred to a region upstream from -792 in the CYP27A1 promoter. In summary, we have identified the human CYP27A1 gene as a target for estrogens and androgens. The results imply that expression of CYP27A1 may be affected by endogenous sex hormones and pharmacological compounds with estrogenic or androgenic effects.

  12. Effects of elaidic acid in a HepG2-SF liver cell model

    DEFF Research Database (Denmark)

    Hansen, Toke Peter Krogager

    Det primære mål for dette Ph.D-studie var at identificere potentielle proteinbiomarkører for menneskelig indtagelse af elaidinsyre, hvilken er den mest almindelige transfedtsyre i fødevarer. En serum fri HepG2 celle model (HepG-SF) blev inkuberet i syv dage med elaidinsyre eller med andre...... human blodplasma. Det sekundære mål for Ph.D-studiet var at undersøge årsagerne til den specifikke cellulære respons for elaidinsyre. Det blev observeret at inkubation med elaidinsyre resulterede i at 28 % af de esterficerede fedtsyrerne i fosfolipid-fraktionen var elaidinsyre, hvilket indikerer en...... lipidmetabolismen når HepG2-SF celler blev inkuberet med elaidinsyre sammenlignet med oleinsyre eller stearinsyre. Den mest fremtrædende ændring var en opregulering af enzymer som syntetiserer kolesterol og fedtsyrer, hvilken indikerede aktivering af sterol regulatory element-binding proteins (SREBPs). Dog blev...

  13. Characterization of dengue virus entry into HepG2 cells

    Directory of Open Access Journals (Sweden)

    Suksanpaisan Lukkana

    2009-02-01

    Full Text Available Abstract Background Despite infections by the dengue virus being a significant problem in tropical and sub-tropical countries, the mechanism by which the dengue virus enters into mammalian cells remains poorly described. Methods A combination of biochemical inhibition, dominant negative transfection of Eps15 and siRNA mediated gene silencing was used to explore the entry mechanism of dengue into HepG2 cells. Results Results were consistent with entry via multiple pathways, specifically via clathrin coated pit mediated endocytosis and macropinocytosis, with clathrin mediated endocytosis being the predominant pathway. Conclusion We propose that entry of the dengue virus to mammalian cells can occur by multiple pathways, and this opens the possibility of the virus being directed to multiple cellular compartments. This would have significant implications in understanding the interaction of the dengue virus with the host cell machinery.

  14. Cytotoxicity and apoptotic effects of tea polyphenol-loaded chitosan nanoparticles on human hepatoma HepG2 cells

    International Nuclear Information System (INIS)

    Liang, Jin; Li, Feng; Fang, Yong; Yang, Wenjian; An, Xinxin; Zhao, Liyan; Xin, Zhihong; Cao, Lin; Hu, Qiuhui

    2014-01-01

    Tea polyphenols have strong antioxidant and antitumor activities. However, these health benefits are limited due to their poor in vivo stability and low bioavailability. Chitosan nanoparticles as delivery systems may provide an alternative approach for enhancing bioavailability of poorly absorbed drugs. In this study, tea polyphenol-loaded chitosan nanoparticles have been prepared using two different chitosan biomaterials, and their antitumor effects were evaluated in HepG2 cells, including cell cytotoxicity comparison, cell morphology analysis, cell apoptosis and cell cycle detection. The results indicated that the tea polyphenol-loaded chitosan nanoparticles showed a branch shape and heterogeneous distribution in prepared suspension. MTT assay suggested that tea polyphenol-loaded chitosan nanoparticles could inhibit the proliferation of HepG2 cells, and the cytotoxicity rates were increased gradually and appeared an obvious dose-dependent relationship. Transmission electron microscope images showed that the HepG2 cells treated with tea polyphenol-loaded chitosan nanoparticles exhibited some typical apoptotic features, such as microvilli disappearance, margination of nuclear chromatin, intracytoplasmic vacuoles and the mitochondrial swelling. In addition, the tea polyphenol-loaded chitosan nanoparticles had relatively weak inhibitory effects on HepG2 cancer cells compared with tea polyphenols. Tea polyphenols not only induced cancer cell apoptosis, but also promoted their necrosis. However, tea polyphenol-loaded chitosan nanoparticles exhibited their antitumor effects mainly through inducing cell apoptosis. Our results revealed that the inhibition effects of tea polyphenol-loaded chitosan nanoparticles on tumor cells probably depended on their controlled drug release and effective cell delivery. The chitosan nanoparticles themselves as the delivery carrier showed limited antitumor effects compared with their encapsulated drugs. - Highlights: • Tea polyphenol

  15. Cytotoxicity and apoptotic effects of tea polyphenol-loaded chitosan nanoparticles on human hepatoma HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Jin [Key Laboratory of Tea Biochemistry and Biotechnology of Ministry of Education and Ministry of Agriculture, Anhui Agricultural University, Hefei 230036 (China); College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); Li, Feng [College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); Fang, Yong; Yang, Wenjian [College of Food Science and Engineering, Nanjing University of Finance and Economics, Nanjing 210023 (China); An, Xinxin; Zhao, Liyan; Xin, Zhihong; Cao, Lin [College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); Hu, Qiuhui, E-mail: qiuhuihu@njau.edu.cn [College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095 (China); College of Food Science and Engineering, Nanjing University of Finance and Economics, Nanjing 210023 (China)

    2014-03-01

    Tea polyphenols have strong antioxidant and antitumor activities. However, these health benefits are limited due to their poor in vivo stability and low bioavailability. Chitosan nanoparticles as delivery systems may provide an alternative approach for enhancing bioavailability of poorly absorbed drugs. In this study, tea polyphenol-loaded chitosan nanoparticles have been prepared using two different chitosan biomaterials, and their antitumor effects were evaluated in HepG2 cells, including cell cytotoxicity comparison, cell morphology analysis, cell apoptosis and cell cycle detection. The results indicated that the tea polyphenol-loaded chitosan nanoparticles showed a branch shape and heterogeneous distribution in prepared suspension. MTT assay suggested that tea polyphenol-loaded chitosan nanoparticles could inhibit the proliferation of HepG2 cells, and the cytotoxicity rates were increased gradually and appeared an obvious dose-dependent relationship. Transmission electron microscope images showed that the HepG2 cells treated with tea polyphenol-loaded chitosan nanoparticles exhibited some typical apoptotic features, such as microvilli disappearance, margination of nuclear chromatin, intracytoplasmic vacuoles and the mitochondrial swelling. In addition, the tea polyphenol-loaded chitosan nanoparticles had relatively weak inhibitory effects on HepG2 cancer cells compared with tea polyphenols. Tea polyphenols not only induced cancer cell apoptosis, but also promoted their necrosis. However, tea polyphenol-loaded chitosan nanoparticles exhibited their antitumor effects mainly through inducing cell apoptosis. Our results revealed that the inhibition effects of tea polyphenol-loaded chitosan nanoparticles on tumor cells probably depended on their controlled drug release and effective cell delivery. The chitosan nanoparticles themselves as the delivery carrier showed limited antitumor effects compared with their encapsulated drugs. - Highlights: • Tea polyphenol

  16. Borax-induced apoptosis in HepG2 cells involves p53, Bcl-2, and Bax.

    Science.gov (United States)

    Wei, Y; Yuan, F J; Zhou, W B; Wu, L; Chen, L; Wang, J J; Zhang, Y S

    2016-06-21

    Borax, a boron compound and a salt of boric acid, is known to inhibit the growth of tumor cells. HepG2 cells have been shown to be clearly susceptible to the anti-proliferative effects of borax. However, the specific mechanisms regulating this effect are poorly understood. This study aimed to investigate the pathways underlying the growth inhibition induced by borax in HepG2 cells. The effects of borax on HepG2 cell viability were characterized using MTT. Apoptosis was also verified by annexin V/propidium iodide staining. JC-1 dye and western blotting techniques were used to measure mitochondrial membrane potential and p53, Bax, and Bcl-2 protein expression, respectively. Relevant mRNA levels were measured by qRT-PCR. Borax inhibited the proliferation of HepG2 cells in a time- and dose-dependent manner in vitro. The apoptotic process triggered by borax involved the upregulation of p53 and Bax and the downregulation of Bcl-2, which was confirmed by a change in the mitochondrial membrane potential. These results elucidate a borax-induced apoptotic pathway in HepG2 cells that involves the upregulation of p53 and Bax and the downregulation of Bcl-2.

  17. Extracellular visfatin activates gluconeogenesis in HepG2 cells through the classical PKA/CREB-dependent pathway.

    Science.gov (United States)

    Choi, Y J; Choi, S-E; Ha, E S; Kang, Y; Han, S J; Kim, D J; Lee, K W; Kim, H J

    2014-04-01

    Adipokines reportedly affect hepatic gluconeogenesis, and the adipokine visfatin is known to be related to insulin resistance and type 2 diabetes. However, whether visfatin contributes to hepatic gluconeogenesis remains unclear. Visfatin, also known as nicotinamide phosphoribosyltransferase (NAMPT), modulates sirtuin1 (SIRT1) through the regulation of nicotinamide adenine dinucleotide (NAD). Therefore, we investigated the effect of extracellular visfatin on glucose production in HepG2 cells, and evaluated whether extracellular visfatin affects hepatic gluconeogenesis via an NAD+-SIRT1-dependent pathway. Treatment with visfatin significantly increased glucose production and the mRNA expression and protein levels of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) in HepG2 cells in a time- and concentration-dependent manner. Knockdown of SIRT1 had no remarkable effect on the induction of gluconeogenesis by visfatin. Subsequently, we evaluated if extracellular visfatin stimulates the production of gluconeogenic enzymes through the classical protein kinase A (PKA)/cyclic AMP-responsive element (CRE)-binding protein (CREB)-dependent process. The phosphorylation of CREB and PKA increased significantly in HepG2 cells treated with visfatin. Additionally, knockdown of CREB and PKA inhibited visfatin-induced gluconeogenesis in HepG2 cells. In summary, extracellular visfatin modulates glucose production in HepG2 cells through the PKA/CREB pathway, rather than via SIRT1 signaling. © Georg Thieme Verlag KG Stuttgart · New York.

  18. Comparative cytotoxic response of nickel ferrite nanoparticles in human liver HepG2 and breast MFC-7 cancer cells.

    Science.gov (United States)

    Ahamed, Maqusood; Akhtar, Mohd Javed; Alhadlaq, Hisham A; Khan, M A Majeed; Alrokayan, Salman A

    2015-09-01

    Nickel ferrite nanoparticles (NPs) have received much attention for their potential applications in biomedical fields such as magnetic resonance imaging, drug delivery and cancer hyperthermia. However, little is known about the toxicity of nickel ferrite NPs at the cellular and molecular levels. In this study, we investigated the cytotoxic responses of nickel ferrite NPs in two different types of human cells (i.e., liver HepG2 and breast MCF-7). Nickel ferrite NPs induced dose-dependent cytotoxicity in both types of cells, which was demonstrated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazoliumbromide (MTT), neutral red uptake (NRU) and lactate dehydrogenase (LDH) assays. Nickel ferrite NPs were also found to induce oxidative stress, which was evident by the depletion of glutathione and the induction of reactive oxygen species (ROS) and lipid peroxidation. The mitochondrial membrane potential due to nickel ferrite NP exposure was also observed. The mRNA levels for the tumor suppressor gene p53 and the apoptotic genes bax, CASP3 and CASP9 were up-regulated, while the anti-apoptotic gene bcl-2 was down-regulated following nickel ferrite NP exposure. Furthermore, the activities of apoptotic enzymes (caspase-3 and caspase-9) were also higher in both types of cells treated with nickel ferrite NPs. Cytotoxicity induced by nickel ferrite was efficiently prevented by N-acetyl cysteine (ROS scavenger) treatment, which suggested that oxidative stress might be one of the possible mechanisms of nickel ferrite NP toxicity. We also observed that MCF-7 cells were slightly more susceptible to nickel ferrite NP exposure than HepG2 cells. This study warrants further investigation to explore the potential mechanisms of different cytotoxic responses of nickel ferrite NPs in different cell lines. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Octyl gallate reduces ATP levels and Ki67 expression leading HepG2 cells to cell cycle arrest and mitochondria-mediated apoptosis.

    Science.gov (United States)

    Lima, Kelly Goulart; Krause, Gabriele Catyana; da Silva, Elisa Feller Gonçalves; Xavier, Léder Leal; Martins, Léo Anderson Meira; Alice, Laura Manzoli; da Luz, Luiza Bueno; Gassen, Rodrigo Benedetti; Filippi-Chiela, Eduardo Cremonese; Haute, Gabriela Viegas; Garcia, Maria Claudia Rosa; Funchal, Giselle Afonso; Pedrazza, Leonardo; Reghelin, Camille Kirinus; de Oliveira, Jarbas Rodrigues

    2018-04-01

    Octyl gallate (OG) is an antioxidant that has shown anti-tumor, anti-diabetic and anti-amyloidogenic activities. Mitochondria play an important role in hepatocellular carcinoma, mainly by maintaining accelerated cellular proliferation through the production of ATP. Thus, the mitochondria may be a target for antitumor therapies. Here, we investigated the effects of OG in the hepatocarcinoma cell line (HepG2) and the mechanisms involved. We report, for the first time, that treatment with OG for 24h inhibited HepG2 cell growth by decreasing mitochondrial activity and mass, which led to the reduction of ATP levels. This reduction in the energy supply triggered a decrease in Ki67 protein expression, leading cells to cycle arrest. In addition, treatment with two doses of OG for 48h induced loss of mitochondrial functionality, mitochondrial swelling and apoptosis. Finally, we report that HepG2 cells had no resistance to treatment after multiple doses. Collectively, our findings indicate that metabolic dysregulation and Ki67 protein reduction are key events in the initial anti-proliferative action of OG, whereas mitochondrial swelling and apoptosis induction are involved in the action mechanism of OG after prolonged exposure. This suggests that OG targets mitochondria, thus representing a candidate for further research on therapies for hepatocarcinoma. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Inflammation response at the transcriptional level of HepG2 cells induced by multi-walled carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Piret, Jean-Pascal; Vankoningsloo, Sebastien; Noel, Florence; Saout, Christelle; Toussaint, Olivier [Research Unit in Cellular Biology (URBC), Narilis, University of Namur, 5000 Namur (Belgium); Mendoza, Jorge Mejia; Lucas, Stephane, E-mail: olivier.toussaint@fundp.ac.be [Research Center for the Physics of Matter and Radiation (PMR), Narilis, University of Namur, 5000 Namur (Belgium)

    2011-07-06

    Poor information are currently available about the biological effects of multi-walled carbon nanotubes (MWCNT) on the liver. In this study, we evaluated the effects of MWCNT at the transcriptional level on the classical in vitro model of HepG2 hepatocarcinoma cells. The expression levels of 96 transcript species implicated in the inflammatory and immune responses was studied after a 24h incubation of HepG2 cells in presence of raw MWCNT dispersed in water by stirring. Among the 46 transcript species detected, only a few transcripts including mRNA coding for interleukine-7, chemokines receptor of the C-C families CCR7, as well as Endothelin-1, were statistically more abundant after treatment with MWCNT. Altogether, these data indicate that MWCNT can only induce a weak inflammatory response in HepG2 cells.

  1. Inflammation response at the transcriptional level of HepG2 cells induced by multi-walled carbon nanotubes

    Science.gov (United States)

    Piret, Jean-Pascal; Vankoningsloo, Sébastien; Noël, Florence; Mejia Mendoza, Jorge; Lucas, Stéphane; Saout, Christelle; Toussaint, Olivier

    2011-07-01

    Poor information are currently available about the biological effects of multi-walled carbon nanotubes (MWCNT) on the liver. In this study, we evaluated the effects of MWCNT at the transcriptional level on the classical in vitro model of HepG2 hepatocarcinoma cells. The expression levels of 96 transcript species implicated in the inflammatory and immune responses was studied after a 24h incubation of HepG2 cells in presence of raw MWCNT dispersed in water by stirring. Among the 46 transcript species detected, only a few transcripts including mRNA coding for interleukine-7, chemokines receptor of the C-C families CCR7, as well as Endothelin-1, were statistically more abundant after treatment with MWCNT. Altogether, these data indicate that MWCNT can only induce a weak inflammatory response in HepG2 cells.

  2. Studies on Cytotoxic Activity against HepG-2 Cells of Naphthoquinones from Green Walnut Husks of Juglans mandshurica Maxim

    Directory of Open Access Journals (Sweden)

    Yuanyuan Zhou

    2015-08-01

    Full Text Available Twenty-seven naphthoquinones and their derivatives, including four new naphthalenyl glucosides and twenty-three known compounds, were isolated from green walnut husks, which came from Juglans mandshurica Maxim. The structures of four new naphthalenyl glucosides were elucidated based on extensive spectroscopic analyses. All of these compounds were evaluated for their cytotoxic activities against the growth of human cancer cells lines HepG-2 by MTT [3-(4,5-dimethylthiazo l-2-yl-2,5 diphenyl tetrazolium bromide] assay. The results were shown that most naphthoquinones in an aglycone form exhibited better cytotoxicity in vitro than naphthalenyl glucosides with IC50 values in the range of 7.33–88.23 μM. Meanwhile, preliminary structure-activity relationships for these compounds were discussed.

  3. [Effects of conditioned media for rat bone marrow-derived mesenchymal stem cells on palmitic acid-induced insulin resistance in HepG2 cells].

    Science.gov (United States)

    Sun, Xiaoya; Hao, Haojie; Han, Weidong; Mu, Yiming

    2015-05-01

    To study the effect of conditioned media for rat bone marrow mesenchymal stem cells (BMSCs-CM) on palmitic acid (PA)-induced insulin resistance (IR) in HepG2 cells and its underlying molecular mechanisms. HepG2 cells were treated with or without BMSCs-CM and L-DMEM in the presence or absence of PA.Glucose utilization in HepG2 cells were detected with PAS, glucose and glycogen measurements. Western blotting was used to assess the expression of phospho-insulin receptor substrate (p-IRS), phosphatidylinositol 3-kinase (PI3K) and p-AKT. (1) Incubation of HepG2 cells with 0.25 mmol/L PA for 24 hours significantly increased the glucose concentration and decreased the glycogen content (Palteration in cells pretreated with PA (Pinsulin sensitivity in HepG2 cells pretreated with PA through upregulation of insulin signaling component expression.

  4. Antiproliferative effects of cinobufacini on human hepatocellular carcinoma HepG2 cells detected by atomic force microscopy.

    Science.gov (United States)

    Wu, Qing; Lin, Wei-Dong; Liao, Guan-Qun; Zhang, Li-Guo; Wen, Shun-Qian; Lin, Jia-Ying

    2015-01-21

    To investigate the antiproliferative activity of cinobufacini on human hepatocellular carcinoma HepG2 cells and the possible mechanism of its action. HepG2 cells were treated with different concentrations of cinobufacini. Cell viability was measured by methylthiazolyl tetrazolium (MTT) assay. Cell cycle distribution was analyzed by flow cytometry (FCM). Cytoskeletal and nuclear alterations were observed by fluorescein isothiocyanate-phalloidin and DAPI staining under a laser scanning confocal microscope. Changes in morphology and ultrastructure of cells were detected by atomic force microscopy (AFM) at the nanoscale level. MTT assay indicated that cinobufacini significantly inhibited the viability of HepG2 cells in a dose-dependent manner. With the concentration of cinobufacini increasing from 0 to 0.10 mg/mL, the cell viability decreased from 74.9% ± 2.7% to 49.41% ± 2.2% and 39.24% ± 2.1% (P nuclei were not damaged seriously. Additionally, high-resolution AFM imaging revealed that cell morphology and ultrastructure changed a lot after treatment with cinobufacini. It appeared as significant shrinkage and deep pores in the cell membrane, with larger particles and a rougher cell surface. Cinobufacini inhibits the viability of HepG2 cells via cytoskeletal destruction and cell membrane toxicity.

  5. Quantification of the uptake of silver nanoparticles and ions to HepG2 cells.

    Science.gov (United States)

    Yu, Su-juan; Chao, Jing-bo; Sun, Jia; Yin, Yong-guang; Liu, Jing-fu; Jiang, Gui-bin

    2013-04-02

    The toxic mechanism of silver nanoparticles (AgNPs) is still debating, partially because of the common co-occurrence and the lack of methods for separation of AgNPs and Ag(+) in biological matrices. For the first time, Triton-X 114-based cloud point extraction (CPE) was proposed to separate AgNPs and Ag(+) in the cell lysates of exposed HepG2 cells. Cell lysates were subjected to CPE after adding Na2S2O3, which facilitated the transfer of AgNPs into the nether Triton X-114-rich phase by salt effect and the preserve of Ag(+) in the upper aqueous phase through the formation of hydrophilic complex. Then the AgNP and Ag(+) contents in the exposed cells were determined by ICP-MS after microwave digestion of the two phases, respectively. Under the optimized conditions, over 67% of AgNPs in cell lysates were extracted into the Triton X-114-rich phase while 94% of Ag(+) remained in the aqueous phase, and the limits of detection for AgNPs and Ag(+) were 2.94 μg/L and 2.40 μg/L, respectively. This developed analytical method was applied to quantify the uptake of AgNPs to the HepG2 cells. After exposure to 10 mg/L AgNPs for 24 h, about 67.8 ng Ag were assimilated per 10(4) cells, in which about 10.3% silver existed as Ag(+). Compared to the pristine AgNPs (with 5.2% Ag(+)) for exposure, the higher ratio of Ag(+) to AgNPs in the exposed cells (10.3% Ag(+)) suggests the transformation of AgNPs into Ag(+) in the cells and/or the higher uptake rate of Ag(+) than that of AgNPs. Given that the toxicity of Ag(+) is much higher than that of AgNPs, the substantial content of Ag(+) in the exposed cells suggests that the contribution of Ag(+) should be taken into account in evaluating the toxicity of AgNPs to organisms, and previous results obtained by regarding the total Ag content in organisms as AgNPs should be reconsidered.

  6. Flavonoids of Korean Citrus aurantium L. Induce Apoptosis via Intrinsic Pathway in Human Hepatoblastoma HepG2 Cells.

    Science.gov (United States)

    Lee, Seung Hwan; Yumnam, Silvia; Hong, Gyeong Eun; Raha, Suchismita; Saralamma, Venu Venkatarame Gowda; Lee, Ho Jeong; Heo, Jeong Doo; Lee, Sang Joon; Lee, Won-Sup; Kim, Eun-Hee; Park, Hyeon Soo; Kim, Gon Sup

    2015-12-01

    Korean Citrus aurantium L. has long been used as a medicinal herb for its anti-inflammatory, antioxidant, and anticancer properties. The present study investigates the anticancer role of flavonoids extracted from C. aurantium on human hepatoblastoma cell, HepG2. The Citrus flavonoids inhibit the proliferation of HepG2 cells in a dose-dependent manner. This result was consistent with the in vivo xenograft results. Apoptosis was detected by cell morphology, cell cycle analysis, and immunoblot. Flavonoids decreased the level of pAkt and other downstream targets of phosphoinositide-3-kinase/Akt pathway - P-4EBP1 and P-p70S6K. The expressions of cleaved caspase 3, Bax, and Bak were increased, while those of Bcl-2 and Bcl-xL were decreased with an increase in the expression of Bax/Bcl-xL ratio in treated cells. Loss of mitochondrial membrane potential was also observed in flavonoid-treated HepG2 cells. It was also observed that the P-p38 protein level was increased both dose and time dependently in flavonoid-treated cells. Collectively, these results suggest that flavonoid extracted from Citrus inhibits HepG2 cell proliferation by inducing apoptosis via an intrinsic pathway. These findings suggest that flavonoids extracted from C. aurantium L. are potential chemotherapeutic agents against liver cancer. Copyright © 2015 John Wiley & Sons, Ltd.

  7. Data on HepG2 cells changes following exposure to cadmium sulphide quantum dots (CdS QDs

    Directory of Open Access Journals (Sweden)

    Laura Paesano

    2017-04-01

    Full Text Available The data included in this paper are associated with the research article entitled "Markers for toxicity to HepG2 exposed to cadmium sulphide quantum dots; damage to mitochondria" (Paesano et al. [1]. The article concerns the cytotoxic and genotoxic effects of CdS QDs in HepG2 cells and the mechanisms involved. In this dataset, changes in expression levels of candidate genes are reported, together with details concerning synthesis and properties of CdS QDs, additional information obtained through literature survey, measures of the mitochondrial membrane potential and the glutathione redox state.

  8. Curcumin induced nanoscale CD44 molecular redistribution and antigen-antibody interaction on HepG2 cell surface

    Energy Technology Data Exchange (ETDEWEB)

    Wang Mu [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Ruan Yuxia [Department of Ophthalmology, The First Affiliated Hospital, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Xing Xiaobo; Chen Qian; Peng, Yuan [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Cai Jiye, E-mail: tjycai@jnu.edu.cn [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China)

    2011-07-04

    Graphical abstract: Highlights: > In this study, we investigate the changes of CD44 expression and distribution on HepG2 cells after curcumin treatment. > We find curcumin is able to change the morphology and ultrastructure of HepG2 cells. > Curcumin can reduce the expression of CD44 molecules and induce the nanoscale molecular redistribution on cell surface. > The binding force between CD44-modified AFM tip and the HepG2 cell surface decreases after curcumin-treatment. - Abstract: The cell surface glycoprotein CD44 was implicated in the progression, metastasis and apoptosis of certain human tumors. In this study, we used atomic force microscope (AFM) to monitor the effect of curcumin on human hepatocellular carcinoma (HepG2) cell surface nanoscale structure. High-resolution imaging revealed that cell morphology and ultrastructure changed a lot after being treated with curcumin. The membrane average roughness increased (10.88 {+-} 4.62 nm to 129.70 {+-} 43.72 nm) and the expression of CD44 decreased (99.79 {+-} 0.16% to 75.14 {+-} 8.37%). Laser scanning confocal microscope (LSCM) imaging showed that CD44 molecules were located on the cell membrane. The florescence intensity in control group was weaker than that in curcumin treated cells. Most of the binding forces between CD44 antibodies and untreated HepG2 cell membrane were around 120-220 pN. After being incubated with curcumin, the major forces focused on 70-150 pN (10 {mu}M curcumin-treated) and 50-120 pN (20 {mu}M curcumin-treated). These results suggested that, as result of nanoscale molecular redistribution, changes of the cell surface were in response to external treatment of curcumin. The combination of AFM and LSCM could be a powerful method to detect the distribution of cell surface molecules and interactions between molecules and their ligands.

  9. Cytotoxicity assessments of Portulaca oleracea and Petroselinum sativum seed extracts on human hepatocellular carcinoma cells (HepG2).

    Science.gov (United States)

    Farshori, Nida Nayyar; Al-Sheddi, Ebtesam Saad; Al-Oqail, Mai Mohammad; Musarrat, Javed; Al-Khedhairy, Abdulaziz Ali; Siddiqui, Maqsood Ahmed

    2014-01-01

    The Pharmacological potential, such as antioxidant, anti-inflammatory, and antibacterial activities of Portulaca oleracea (PO) and Petroselinum sativum (PS) extracts are well known. However, the preventive properties against hepatocellular carcinoma cells have not been explored so far. Therefore, the present investigation was designed to study the anticancer activity of seed extracts of PO and PS on the human hepatocellular carcinoma cells (HepG2). The HepG2 cells were exposed with 5-500 μg/ml of PO and PS for 24 h. After the exposure, cell viability by 3-(4,5-dimethylthiazol-2yl)-2,5-biphenyl tetrazolium bromide (MTT) assay, neutral red uptake (NRU) assay, and cellular morphology by phase contrast inverted microscope were studied. The results showed that PO and PS extracts significantly reduced the cell viability of HepG2 in a concentration dependent manner. The cell viability was recorded to be 67%, 31%, 21%, and 17% at 50, 100, 250, and 500 μg/ml of PO, respectively by MTT assay and 91%, 62%, 27%, and 18% at 50, 100, 250, and 500 μg/ml of PO, respectively by NRU assay. PS exposed HepG2 cells with 100 μg/ml and higher concentrations were also found to be cytotoxic. The decrease in the cell viability at 100, 250, and 500 μg/ml of PS was recorded as 70%, 33%, and 15% by MTT assay and 63%, 29%, and 17%, respectively by NRU assay. Results also showed that PO and PS exposed cells reduced the normal morphology and adhesion capacity of HepG2 cells. HepG2 cells exposed with 50 μg/ml and higher concentrations of PO and PS lost their typical morphology, become smaller in size, and appeared in rounded bodies. Our results demonstrated preliminary screening of anticancer activity of Portulaca oleracea and Petroselinum sativum extracts against HepG2 cells, which can be further used for the development of a potential therapeutic anticancer agent.

  10. Effects of the radiolysis products of sennoside A on HepG2 and PC-3 cell

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Dong Ho; Jo, Min Ho [Research Division for Biotechnology, Korea Atomic Energy Research Institute, Jeongeup (Korea, Republic of)

    2016-11-15

    Radiolysis of sennoside A was carried out by gamma irradiation and the anti-cancer activities of the radiolysis product were evaluated. An aqueous solution of sennoside A was exposed to 0.5-3 kGy of gamma irradiation and the radiolysis products were analyzed by HPLC. A fraction of radiolysis product (RLF) of sennoside A was isolated and the RLF was presumed as a rhein-8-β-D-glucoside. The anticancer effect of the RLF was compared with the sennoside and rhein using a in vitro assay system of human prostate cancer cells (PC-3) and human hepatoma HepG2 cells. The cell viability of PC-3 and HepG2 cell was significantly decreased to 12.4±1.2% and 32.4±2.1%, respectively, by the treatment of 0.6 μM of RLF. The sennoside A (range from 0 to 25 μM) had no cytotoxic effect on PC-3 and HepG2 cells, while the rhein had the effect on HepG2 cells with a LD{sub 50} at 80 μM.

  11. The Growth Suppressing Effects of Girinimbine on Hepg2 Involve Induction of Apoptosis and Cell Cycle Arrest

    Directory of Open Access Journals (Sweden)

    Tang Sook Wah

    2011-08-01

    Full Text Available Murraya koenigii is an edible herb widely used in folk medicine. Here we report that girinimbine, a carbazole alkaloid isolated from this plant, inhibited the growth and induced apoptosis in human hepatocellular carcinoma, HepG2 cells. The MTT and LDH assay results showed that girinimbine decreased cell viability and increased cytotoxicity in a dose-and time-dependent manner selectively. Girinimbine-treated HepG2 cells showed typical morphological features of apoptosis, as observed from normal inverted microscopy and Hoechst 33342 assay. Furthermore, girinimbine treatment resulted in DNA fragmentation and elevated levels of caspase-3 in HepG2 cells. Girinimbine treatment also displayed a time-dependent accumulation of the Sub-G0/G1 peak (hypodiploid and caused G0/G1-phase arrest. Together, these results demonstrated for the first time that girinimbine could effectively induce programmed cell death in HepG2 cells and suggests the importance of conducting further investigations in preclinical human hepatocellular carcinoma models, especially on in vivo efficacy, to promote girinimbine for use as an anticancer agent against hepatocellular carcinoma.

  12. Necrosis of HepG2 cancer cells induced by the vibration of magnetic particles

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Biran [Laboratoire de Physique de la Matière Condensée (LPMC), CNRS UMR 7336, Université de Nice Sophia Antipolis, Parc Valrose, 06108 Nice (France); Institut de Chimie de Nice, UMR 7272, Université de Nice Sophia Antipolis, CNRS, 28 Avenue de Valrose, F-06100 Nice (France); Bienvenu, Céline [Institut de Chimie de Nice, UMR 7272, Université de Nice Sophia Antipolis, CNRS, 28 Avenue de Valrose, F-06100 Nice (France); Mendez-Garza, Juan; Lançon, Pascal; Madeira, Alexandra [Laboratoire de Physique de la Matière Condensée (LPMC), CNRS UMR 7336, Université de Nice Sophia Antipolis, Parc Valrose, 06108 Nice (France); Vierling, Pierre [Institut de Chimie de Nice, UMR 7272, Université de Nice Sophia Antipolis, CNRS, 28 Avenue de Valrose, F-06100 Nice (France); Di Giorgio, Christophe, E-mail: christophe.di-giorgio@unice.fr [Institut de Chimie de Nice, UMR 7272, Université de Nice Sophia Antipolis, CNRS, 28 Avenue de Valrose, F-06100 Nice (France); Bossis, Georges, E-mail: bossis@unice.fr [Laboratoire de Physique de la Matière Condensée (LPMC), CNRS UMR 7336, Université de Nice Sophia Antipolis, Parc Valrose, 06108 Nice (France)

    2013-10-15

    Experiments of magnetolysis, i.e., destruction of cells induced with magnetic particles (MPs) submitted to the application of a magnetic field, were conducted on HepG2 cancer cells. We herein demonstrate the usefulness of combining anisotropic MPs with an alternative magnetic field in magnetolysis. Thus, the application of an alternative magnetic field of low frequency (a few Hertz) in the presence of anisotropic, submicronic particles allowed the destruction of cancer cells “in vitro”. We also show that a constant magnetic field is far less efficient than an oscillating one. Moreover, we demonstrate that, at equal particle volume, it is much more efficient to utilize spindle shaped particles rather than spherical ones. In order to get deeper insight into the mechanism of magnetolysis experiments, we performed a study by AFM, which strongly supports that the magnetic field induces the formation of clusters of particles becoming then large enough todamage cell membranes. - Highlights: • Magnetic force was applied on cancer cells through magnetic particles. • The penetration depth was predicted, both for spherical and ellipsoidal particles. • Alternative force was shown to damage the cells contrary to static force. • The effect of indentation of magnetic particles was compared to the one of AFM tips. • The damage was attributed to the formation of clusters of particles.

  13. Proapoptotic activity of aflatoxin B1 and sterigmatocystin in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Yang Liu

    2014-01-01

    Full Text Available Aflatoxin B1 (AFB1 and sterigmatocystin (ST are two hepatocarcinogenic mycotoxins that are commonly coexisted in cereal grains, and their co-proapoptotic activity in HepG2 cells was studied. The values of IC50, which is the dosage of mycotoxin resulting in a 50% cell growth inhibition measured by a sulforhodamine B (SRB colorimetric assay, were 16.9 μM and 7.3 μM for AFB1 and ST, respectively. Additively and dose-dependently, cell apoptosis-related toxicity endpoints of double strand DNA and ATP content were decreased while the intracellular ROS and mitochondria membrane permeability (MMP were increased. Consistently, when cell cycle is arrest at G0/G1 or S phase by AFB1 and/or ST, the experimental results from flow cytometry assay demonstrated that the rate of cell apoptosis and mitochondrial membrane potential were also additively increased and decreased, respectively, in a dose-dependent manner. Thus, the integrity of mitochondria (MMP and membrane potential that is the central component of cell apoptosis is disrupted by AFB1 and ST in an additive manner. With the immunocytochemistry analysis showing increased expression of apoptosis-related proteins of Bax, Caspase-3 and p53 and decreased expression of Bcl-2 protein, an additive nature of the co-proapoptotic activity of AFB1 and ST was revealed.

  14. Dihydrotestosterone regulating apolipoprotein M expression mediates via protein kinase C in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Yi-zhou Ye

    2012-12-01

    Full Text Available Abstract Background Administration of androgens decreases plasma concentrations of high-density lipid cholesterol (HDL-C. However, the mechanisms by which androgens mediate lipid metabolism remain unknown. This present study used HepG2 cell cultures and ovariectomized C57BL/6 J mice to determine whether apolipoprotein M (ApoM, a constituent of HDL, was affected by dihydrotestosterone (DHT. Methods HepG2 cells were cultured in the presence of either DHT, agonist of protein kinase C (PKC, phorbol-12-myristate-13-acetate (PMA, blocker of androgen receptor flutamide together with different concentrations of DHT, or DHT together with staurosporine at different concentrations for 24 hrs. Ovariectomized C57BL/6 J mice were treated with DHT or vehicle for 7d or 14d and the levels of plasma ApoM and livers ApoM mRNA were measured. The mRNA levels of ApoM, ApoAI were determined by real-time RT-PCR. ApoM and ApoAI were determined by western blotting analysis. Results Addition of DHT to cell culture medium selectively down-regulated ApoM mRNA expression and ApoM secretion in a dose-dependent manner. At 10 nM DHT, the ApoM mRNA levels were about 20% lower than in untreated cells and about 40% lower at 1000 nM DHT than in the control cells. The secretion of ApoM into the medium was reduced to a similar extent. The inhibitory effect of DHT on ApoM secretion was not blocked by the classical androgen receptor blocker flutamide but by an antagonist of PKC, Staurosporine. Agonist of PKC, PMA, also reduced ApoM. At 0.5 μM PMA, the ApoM mRNA levels and the secretion of ApoM into the medium were about 30% lower than in the control cells. The mRNA expression levels and secretion of another HDL-associated apolipoprotein AI (ApoAI were not affected by DHT. The levels of plasma ApoM and liver ApoM mRNA of DHT-treated C57BL/6 J mice were lower than those of vehicle-treated mice. Conclusions DHT directly and selectively down-regulated the level of ApoM mRNA and the

  15. Galactosylated poly(ε-caprolactone) membrane promoted liver-specific functions of HepG2 cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yan, E-mail: zhang_yan@ecust.edu.cn [The Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, Shanghai 200237 (China); Zhang, Yi [The Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, Shanghai 200237 (China); Chen, Min; Zhou, Yan [The State Key Laboratory of Bioreactor Engineering, School of Bioengineering, East China University of Science and Technology, Shanghai, 200237 (China); Lang, Meidong, E-mail: mdlang@ecust.edu.cn [The Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, Shanghai 200237 (China)

    2014-08-01

    The lack of pendant functional groups on the PCL backbone has been a great challenge for surface bioactivation of poly(ε-caprolactone) (PCL). In the present study, covalently galactosylated PCL (GPCL) was developed through coupling between the amino-functionalized PCL (NPCL) and the lactobionic acid (LA) and its potential application in maintenance of physiological functions of HepG2 cells was further evaluated. The structure and properties of GPCL were explored by {sup 1}H NMR, FT-IR, GPC and DSC. Moreover, the incorporation of galactose ligands onto GPCL membranes not only promoted higher wettability, but also radically changed surface morphology in comparison with PCL and NPCL according to the contact angle measurement and atomic force microscopy. When HepG2 cells were seeded onto these membranes, the cells on GPCL membranes showed more pronounced cell adhesion and tended to form aggregates during the initial adhesion stage and then progressively grew into multi-layer structures compared to those without galactose ligands by the observation with fluorescence microscope and scanning electron microscopy. Furthermore, live–dead assay and functional tests demonstrated that HepG2 cells on GPCL membranes had superior viability and maintained better liver-specific functions. Collectively, GPCL has great potential for hepatic tissue engineering scaffolds. - Graphical abstract: The specific recognition between the galactose ligands on the galactosylated poly(ε-caprolactone) membrane and the ASGPR on the HepG2 cell surface. The galactosylated poly(ε-caprolactone) membranes improved the cell-matrix interaction. The galactosylated functionalized PCL scaffold is a potential candidate for liver tissue engineering. - Highlights: • The specific recognition between the galactose ligands on the galactosylated poly(ε-caprolactone) membrane and the ASGPR on the HepG2 cell surface. • The galactosylated poly(ε-caprolactone) membranes improved the cell-matrix interaction.

  16. Cytotoxic effect of Agaricus bisporus and Lactarius rufus β-D-glucans on HepG2 cells.

    Science.gov (United States)

    Pires, Amanda do Rocio Andrade; Ruthes, Andrea Caroline; Cadena, Silvia Maria Suter Correia; Acco, Alexandra; Gorin, Philip Albert James; Iacomini, Marcello

    2013-07-01

    The cytotoxic activity of β-D-glucans isolated from Agaricus bisporus and Lactarius rufus fruiting bodies was evaluated on human hepatocellular carcinoma cells (HepG2). NMR and methylation analysis suggest that these β-d-glucans were composed of a linear (1→6)-linked and a branched (1→3), (1→6)-linked backbone, respectively. They both decreased cell viability at concentrations of up to 100 μg mL(-1), as shown by MTT assay. The amount of LDH released and the analysis of cell morphology corroborated these values and also showed that the β-D-glucan of L. rufus was more cytotoxic to HepG2 cells than that of A. bisporus. The treatment of HepG2 cells with L. rufus and A. bisporus β-D-glucans at a dose of 200 μg mL(-1) for 24h promoted an increase of cytochrome c release and a decrease of ATP content, suggesting that these polysaccharides could promote cell death by apoptosis. Both β-D-glucans were tested against murine primary hepatocytes at a dose of 200 μg mL(-1). The results suggest that the L. rufus β-d-glucan was as cytotoxic for hepatocytes as for HepG2 cells, whereas the A. bisporus β-D-glucan, under the same conditions, was cytotoxic only for HepG2 cells, suggesting cell selectivity. These results open new possibilities for use of mushroom β-D-glucans in cancer therapy. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Chronic ecotoxic effects to Pseudomonas putida and Vibrio fischeri, and cytostatic and genotoxic effects to the hepatoma cell line (HepG2) of ofloxacin photo(cata)lytically treated solutions

    Energy Technology Data Exchange (ETDEWEB)

    Vasquez, M.I. [University of Cyprus, Department of Civil and Environmental Engineering, University of Cyprus, 75 Kallipoleos Street, 1678 Nicosia (Cyprus); Nireas International Water Research Center, University of Cyprus (Cyprus); Garcia-Käufer, M. [University Medical Centre Freiburg, Department of Environmental Health Sciences, 115 B, Breisacher Straße, 79106 Freiburg (Germany); Hapeshi, E. [University of Cyprus, Department of Civil and Environmental Engineering, University of Cyprus, 75 Kallipoleos Street, 1678 Nicosia (Cyprus); Nireas International Water Research Center, University of Cyprus (Cyprus); Menz, J. [Institute of Sustainable and Environmental Chemistry, Leuphana University Lüneburg, Scharnhorststraße 1/C13, 21335 Lüneburg (Germany); Kostarelos, K.; Fatta-Kassinos, D. [University of Cyprus, Department of Civil and Environmental Engineering, University of Cyprus, 75 Kallipoleos Street, 1678 Nicosia (Cyprus); Nireas International Water Research Center, University of Cyprus (Cyprus); Kümmerer, K., E-mail: Klaus.Kuemmerer@uni.leuphana.de [Institute of Sustainable and Environmental Chemistry, Leuphana University Lüneburg, Scharnhorststraße 1/C13, 21335 Lüneburg (Germany)

    2013-04-15

    Ofloxacin (OFL), a broad-spectrum and widespread-used photolabile fluoroquinolone, is frequently found in treated wastewaters, aquatic and terrestrial ecosystems leading to increasing concern during the past decades regarding its effects to the environment and human health. The elimination of OFL and other xenobiotics by the application of advanced oxidation processes using photolytic (PL) and photocatalytic (PC) treatments seems promising. However, an integrated assessment scheme is needed, in which, not only the removal of the parent compound, but also the effects of the photo-transformation products (PTPs) are investigated. For this purpose, in the present study, a chronic ecotoxic assessment using representative bacteria of marine and terrestrial ecosystems and a cytostatic and genotoxic evaluation using hepatoma cell line were performed. PL and PC treatments of OFL were applied using UV radiation. The photo-transformation of OFL during the treatments was monitored by DOC measurements and UPLC–MS/MS analysis. The chronic ecotoxicity of OFL and treated samples was evaluated using Pseudomonas putida and Vibrio fischeri; whereas the cytostasis and genotoxicity were estimated by the cytokinesis-block micronucleus assay (CBMN). The main results suggest that photo-transformation of OFL took place during these treatments since the concentration of OFL decreased when the irradiation time increased, as quantified by UPLC–MS/MS analysis, and this was not coupled with an analogous DOC removal. Furthermore, nine compounds were identified as probable PTPs formed through piperazinyl dealkylation and decarboxylation. The ecotoxicity of treated solutions to the bacteria studied decreased while the cytostasis to the hepatoma cell line remained at low levels during both treatments. However, the genotoxicity to the hepatoma cell line demonstrated a different pattern in which treated samples induced a greater number of MNi for the 4–16 min of irradiation (p < 0.05) during

  18. Cyclosporine A and palmitic acid treatment synergistically induce cytotoxicity in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Yi, E-mail: yi.luo@pfizer.com; Rana, Payal; Will, Yvonne

    2012-06-01

    Immunosuppressant cyclosporine A (CsA) treatment can cause severe side effects. Patients taking immunosuppressant after organ transplantation often display hyperlipidemia and obesity. Elevated levels of free fatty acids have been linked to the etiology of metabolic syndromes, nonalcoholic fatty liver and steatohepatitis. The contribution of free fatty acids to CsA-induced toxicity is not known. In this study we explored the effect of palmitic acid on CsA-induced toxicity in HepG2 cells. CsA by itself at therapeutic exposure levels did not induce detectible cytotoxicity in HepG2 cells. Co-treatment of palmitic acid and CsA resulted in a dose dependent increase in cytotoxicity, suggesting that fatty acid could sensitize cells to CsA-induced cytotoxicity at the therapeutic doses of CsA. A synergized induction of caspase-3/7 activity was also observed, indicating that apoptosis may contribute to the cytotoxicity. We demonstrated that CsA reduced cellular oxygen consumption which was further exacerbated by palmitic acid, implicating that impaired mitochondrial respiration might be an underlying mechanism for the enhanced toxicity. Inhibition of c-Jun N-terminal kinase (JNK) attenuated palmitic acid and CsA induced toxicity, suggesting that JNK activation plays an important role in mediating the enhanced palmitic acid/CsA-induced toxicity. Our data suggest that elevated FFA levels, especially saturated FFA such as palmitic acid, may be predisposing factors for CsA toxicity, and patients with underlying diseases that would elevate free fatty acids may be susceptible to CsA-induced toxicity. Furthermore, hyperlipidemia/obesity resulting from immunosuppressive therapy may aggravate CsA-induced toxicity and worsen the outcome in transplant patients. -- Highlights: ► Palmitic acid and cyclosporine (CsA) synergistically increased cytotoxicity. ► The impairment of mitochondrial functions may contribute to the enhanced toxicity. ► Inhibition of JNK activity attenuated

  19. Biochemical Effects of six Ti02 and four Ce02 Nanomaterials in HepG2 cells

    Science.gov (United States)

    Abstract The potential mammalian hepatotoxicity of nanomaterials were explored in dose-response and structure-activity studies with human hepatic HepG2 cells exposed to between 10 and 1000 ug/ml of six different TiO2 and four CeO2 nanomaterials for 3 days. Var...

  20. Differential genomic effects on signaling pathways by two different CeO2 nanoparticles in HepG2 cells

    Science.gov (United States)

    To investigate genomic effects, human liver hepatocellular carcinoma (HepG2) cells were exposed for three days to two different forms of nanoparticles both composed of Ce02 (0.3, 3 and 30 µg/mL). The two Ce02 nanopartices had dry primary particle sizes of 8 nanometers {(M) made b...

  1. The induction of apoptosis in HepG-2 cells by ruthenium(II) complexes through an intrinsic ROS-mediated mitochondrial dysfunction pathway.

    Science.gov (United States)

    Zeng, Chuan-Chuan; Lai, Shang-Hai; Yao, Jun-Hua; Zhang, Cheng; Yin, Hui; Li, Wei; Han, Bing-Jie; Liu, Yun-Jun

    2016-10-21

    Four new ruthenium(II) polypyridyl complexes [Ru(N-N)2(dhbn)](ClO4)2 (N-N = dmb: 4,4'-dimethyl-2,2'-bipyridine 1; bpy = 2,2'-bipyridine 2; phen = 1,10-phenanthroline 3; dmp = 2,9-dimethyl-1,10-phenanthroline 4) were synthesized and characterized. The cytotoxicity in vitro of the ligand and complexes toward HepG-2, HeLa, MG-63 and A549 were assayed by MTT method. The IC50 values of the complexes against the above cells range from 17.7 ± 1.1 to 45.1 ± 2.8 μM. The cytotoxic activity of the complexes against HepG-2 cells follows the order of 4 > 2 > 3 > 1. Ligand shows no cytotoxic activity against the selected cell lines. Cellular uptake, apoptosis, comet assay, reactive oxygen species, mitochondrial membrane potential, cell cycle arrest, and the expression of proteins involved in apoptosis pathway induced by the complexes were investigated. The results indicate that complexes 1-4 induce apoptosis in HepG-2 cells through an intrinsic ROS-mediated mitochondrial dysfunction pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  2. Surface ligand dependent toxicity of zinc oxide nanoparticles in HepG2 cell model

    International Nuclear Information System (INIS)

    Bartczak, D; Baradez, M-O; Merson, S; Goenaga-Infante, H; Marshall, D

    2013-01-01

    Physicochemical properties of nanoparticles (NP) strongly affect their influence on cell behaviour, but can be significantly distorted by interactions with the proteins present in biological solutions. In this study we show how different surface functionalities of zinc oxide (ZnO) NP lead to changes in the size distribution and dissolution of the NP in serum containing cell culture media and how this impacts on NP toxicity. NPs capped with weakly bound large proteins undergo substantial transformations due to the exchange of the original surface ligands to the components of the cell culture media. Conversely, NP capped with a tight monolayer of small organic molecules or with covalently conjugated proteins show significantly higher stability. These differences in ligand exchange also affect the toxicity of the NP to the HepG2 liver cell model, with the NP capped with small organic molecules being more toxic than those capped with large proteins. This study highlights the importance of characterising NPs in biological media and the effect the media has during in-vitro analysis.

  3. Surface ligand dependent toxicity of zinc oxide nanoparticles in HepG2 cell model

    Science.gov (United States)

    Bartczak, D.; Baradez, M.-O.; Merson, S.; Goenaga-Infante, H.; Marshall, D.

    2013-04-01

    Physicochemical properties of nanoparticles (NP) strongly affect their influence on cell behaviour, but can be significantly distorted by interactions with the proteins present in biological solutions. In this study we show how different surface functionalities of zinc oxide (ZnO) NP lead to changes in the size distribution and dissolution of the NP in serum containing cell culture media and how this impacts on NP toxicity. NPs capped with weakly bound large proteins undergo substantial transformations due to the exchange of the original surface ligands to the components of the cell culture media. Conversely, NP capped with a tight monolayer of small organic molecules or with covalently conjugated proteins show significantly higher stability. These differences in ligand exchange also affect the toxicity of the NP to the HepG2 liver cell model, with the NP capped with small organic molecules being more toxic than those capped with large proteins. This study highlights the importance of characterising NPs in biological media and the effect the media has during in-vitro analysis.

  4. Analysis of changes in energy and redox states in HepG2 hepatoma and C6 glioma cells upon exposure to cadmium

    International Nuclear Information System (INIS)

    Yang, M.S.; Yu, L.C.; Gupta, R.C.

    2004-01-01

    The energy and redox states of the HepG2 hepatoma and the C6 glioma cells were studied by quantifying the levels of ATP, ADP, AMP, GSH, and GSSG. These values were used to calculate the energy charge potential (ECP = [ATP + 0.5ADP]/TAN), total adenosine nucleotides (TAN = ATP + ADP + AMP), total glutathione (TG = [GSH + GSSG]/TAN), and the redox state (GSH/GSSG ratio). For comparison between cell types, the level of each energy metabolite (ATP, ADP, and AMP) was normalized against TAN of the respective cell. The results showed that ATP:ADP:AMP ratio was 0.76:0.11:0.13 for the HepG2 cells and 0.80:0.11:0.09 for the C6 glioma cells. ECP was 0.81 ± 0.01 and 0.85 ± 0.01 for the HepG2 and the C6 glioma cells, respectively. GSH/GSSG ratio was 2.66 ± 0.16 and 3.63 ± 0.48 for HepG2 and C6 glioma cells, respectively. TG was 3.2 ± 0.54 for the HepG2 cells and 2.43 ± 0.18 for the C6 glioma cells, indicating that the level of total glutathione is more than two to three times higher than the total energy metabolites in these cell lines. Following a 3-h incubation in medium containing different concentrations of Cd, there was a dose-dependent decrease in cell viability. The 3-h LC 50 for the HepG2 cells was 0.5 mM and that for the C6 glioma cells was 0.4 mM. Cellular TAN decreased with a decrease in cell viability. Upon careful analysis of the energy state, there was a significant increase in relative amount of ATP and decrease in ADP and AMP in both cells as Cd concentration increased from 0 to 0.1, 0.2, and 0.6 mM. However, ECP in both cell lines increased, which indicated that the level of high energy phosphate was adequate. There was also a significant increase in TG and a significant decrease in GSH/GSSG in the C6 glioma cells when cells were exposed to as low as 0.1 mM Cd, which suggested that the cellular redox state was compromised. The HepG2 cells, on the other hand, showed no significant change in both TG and GSH/GSSG level until Cd concentration reached 0.6 m

  5. An untargeted multi-technique metabolomics approach to studying intracellular metabolites of HepG2 cells exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin

    Directory of Open Access Journals (Sweden)

    van Delft Joost

    2011-05-01

    Full Text Available Abstract Background In vitro cell systems together with omics methods represent promising alternatives to conventional animal models for toxicity testing. Transcriptomic and proteomic approaches have been widely applied in vitro but relatively few studies have used metabolomics. Therefore, the goal of the present study was to develop an untargeted methodology for performing reproducible metabolomics on in vitro systems. The human liver cell line HepG2, and the well-known hepatotoxic and non-genotoxic carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, were used as the in vitro model system and model toxicant, respectively. Results The study focused on the analysis of intracellular metabolites using NMR, LC-MS and GC-MS, with emphasis on the reproducibility and repeatability of the data. State of the art pre-processing and alignment tools and multivariate statistics were used to detect significantly altered levels of metabolites after exposing HepG2 cells to TCDD. Several metabolites identified using databases, literature and LC-nanomate-Orbitrap analysis were affected by the treatment. The observed changes in metabolite levels are discussed in relation to the reported effects of TCDD. Conclusions Untargeted profiling of the polar and apolar metabolites of in vitro cultured HepG2 cells is a valid approach to studying the effects of TCDD on the cell metabolome. The approach described in this research demonstrates that highly reproducible experiments and correct normalization of the datasets are essential for obtaining reliable results. The effects of TCDD on HepG2 cells reported herein are in agreement with previous studies and serve to validate the procedures used in the present work.

  6. Regulation of human gamma-glutamylcysteine synthetase: co-ordinate induction of the catalytic and regulatory subunits in HepG2 cells.

    Science.gov (United States)

    Galloway, D C; Blake, D G; Shepherd, A G; McLellan, L I

    1997-11-15

    We have shown that in HepG2 cells treatment with 75 microM t-butylhydroquinone (tBHQ) results in a 2.5-fold increase in glutathione concentration, as part of an adaptive response to chemical stress. In these cells the elevation in intracellular glutathione level was found to be accompanied by an increase of between 2-fold and 3-fold in the level of the 73 kDa catalytic subunit of gamma-glutamylcysteine synthetase (heavy subunit, GCSh) and the 31 kDa regulatory subunit (light subunit, GCSl). Levels of GCSh and GCSl mRNA were increased by up to 5-fold in HepG2 cells in response to tBHQ. To study the transcriptional regulation of GCSl, we subcloned 6.7 kb of the upstream region of the human GCSl gene (GLCLR) from a genomic clone isolated from a P1 lymphoblastoid cell line genomic library. HepG2 cells were transfected with GLCLR promoter reporter constructs and treated with tBHQ. This resulted in an induction of between 1.5-fold and 3.5-fold in reporter activity, indicating that transcriptional regulation of GLCLR is likely to contribute to the induction of GCSl by tBHQ in HepG2 cells. Sequence analysis of the promoter region demonstrated the presence of putative enhancer elements including AP-1 sites and an antioxidant-responsive element, which might be involved in the observed induction of the GLCLR promoter.

  7. Synthetic peptides from two Pf sporozoite invasion-associated proteins specifically interact with HeLa and HepG2 cells.

    Science.gov (United States)

    Arévalo-Pinzón, Gabriela; Curtidor, Hernando; Muñoz, Marina; Patarroyo, Manuel A; Patarroyo, Manuel E

    2011-09-01

    Two recently described molecules have been associated with sporozoite traversal ability and hepatocyte entry: sporozoite invasion-associated proteins (SIAP)-1 and -2. The HeLa and HepG2 cell binding ability of synthetic peptides spanning the whole SIAP-1 and -2 sequences has been studied in the search for identifying these proteins' functionally active specific regions. Twelve HepG-2 and seventeen HeLa cell high-activity binding peptides (HABPs) have been identified in SIAP-1, 8 of them having high specific binding affinity for both cell lines. Four HepG2 HABPs and two HeLa HABPs have been identified in SIAP-2, one of them interacting with both HeLa and HepG2 cells. SIAP-1 and SIAP-2 HABPs bound specifically and saturably to heparin sulfate and chondroitin sulfate-type membrane receptors on host cells. Circular dichroism assays have shown high α-helix content in SIAP-1 and SIAP-2 HABP secondary structure. Immunofluorescence analysis has revealed that specific peptides against SIAP proteins are highly immunogenic in mice and that anti-SIAP-1 and -2 antibodies recognize the native protein in Plasmodium falciparum sporozoites. Polymorphism studies have shown that a most SIAP-1 and -2 HABPs are conserved among P. falciparum strains. Our results have suggested that SIAP-1 and -2 participate in host-pathogen interactions during cell-traversal and hepatocyte invasion and highlighted the relevance of the ongoing identification and study of potentially new molecules when designing a fully protective antimalarial vaccine. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Kaempferol induces apoptosis in HepG2 cells via activation of the endoplasmic reticulum stress pathway.

    Science.gov (United States)

    Guo, Haiqing; Ren, Feng; Zhang, Li; Zhang, Xiangying; Yang, Rongrong; Xie, Bangxiang; Li, Zhuo; Hu, Zhongjie; Duan, Zhongping; Zhang, Jing

    2016-03-01

    Kaempferol is a flavonoid compound that has gained importance due to its antitumor properties; however, the underlying mechanisms remain to be fully understood. The present study aimed to investigate the molecular mechanisms of the antitumor function of kaempferol in HepG2 hepatocellular carcinoma cells. Kaempferol was determined to reduce cell viability, increase lactate dehydrogenase activity and induce apoptosis in a concentration‑ and time‑dependent manner in HepG2 cells. Additionally, kaempferol‑induced apoptosis possibly acts via the endoplasmic reticulum (ER) stress pathway, due to the significant increase in the protein expression levels of glucose‑regulated protein 78, glucose‑regulated protein 94, protein kinase R‑like ER kinase, inositol‑requiring enzyme 1α, partial activating transcription factor 6 cleavage, caspase‑4, C/EBP homologous protein (CHOP) and cleaved caspase‑3. The pro‑apoptotic activity of kaempferol was determined to be due to induction of the ER stress‑CHOP pathway, as: i) ER stress was blocked by 4‑phenyl butyric acid (4‑PBA) pretreatment and knockdown of CHOP with small interfering RNA, which resulted in alleviation of kaempferol‑induced HepG2 cell apoptosis; and ii) transfection with plasmid overexpressing CHOP reversed the protective effect of 4‑PBA in kaempferol‑induced HepG2 cells and increased the apoptotic rate. Thus, kaempferol promoted HepG2 cell apoptosis via induction of the ER stress‑CHOP signaling pathway. These observations indicate that kaempferol may be used as a potential chemopreventive treatment strategy for patients with hepatocellular carcinoma.

  9. Autophagy suppresses proliferation of HepG2 cells via inhibiting glypican-3/wnt/β-catenin signaling

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    Hu P

    2018-01-01

    Full Text Available Pei Hu,1,2 Bin Cheng,3 Yulin He,3 Zhiqiang Wei,3 Dongfang Wu,1 Zhongji Meng3,4 1Department of Pharmacy, Zhongnan Hospital of Wuhan University, Wuhan, 2Department of Clinical Laboratory Medicine, 3Institute of Biomedical Research, 4Department of Infectious Disease, Taihe Hospital, Hubei University of Medicine, Shiyan, People’s Republic of China Introduction: Autophagy plays an important role in the growth and survival of hepatocellular carcinoma (HCC cells through several target proteins or signaling pathways. Glypican-3 (GPC3 is a new reliable HCC marker, which is involved in tumor growth in HCC, primarily mediated by wnt/β-catenin signaling. Objective: The present study aimed to identify the role of autophagy in the proliferation of HepG2 cells through GPC3/wnt/β-catenin signaling. Results and discussion: Results demonstrated that induction of autophagy by nutrition starvation and rapamycin treatment led to the downregulation of GPC3 expression in HepG2 cells, accompanied by the decreased expression of wnt downstream target genes (β-catenin, c-myc and cyclin D1. On the other hand, inhibition of autophagy by 3-methyl adenine (3-MA could rescue rapamycin-directed downregulation of GPC3 and wnt/β-catenin target genes and augment the proliferation of HepG2 cells. Furthermore, interference of GPC3 by siRNA suppressed wnt/β-catenin signaling and attenuated 3-MA stimulation of HepG2 cell proliferation. More interestingly, the mRNA of GPC3 remained unchanged when the protein levels of GPC3 were decreased by autophagy activation, suggesting that induction of autophagy may accelerate the degradation of GPC3. Conclusion: These results suggest that autophagy suppresses proliferation of HepG2 cells partially by inhibition of GPC3/wnt/β-catenin signaling. Keywords: hepatocellular carcinoma, glypican-3, autophagy, proliferation, wnt/β-catenin signaling

  10. Mass spectrometric analysis of host cell proteins interacting with dengue virus nonstructural protein 1 in dengue virus-infected HepG2 cells.

    Science.gov (United States)

    Dechtawewat, Thanyaporn; Paemanee, Atchara; Roytrakul, Sittiruk; Songprakhon, Pucharee; Limjindaporn, Thawornchai; Yenchitsomanus, Pa-Thai; Saitornuang, Sawanan; Puttikhunt, Chunya; Kasinrerk, Watchara; Malasit, Prida; Noisakran, Sansanee

    2016-09-01

    Dengue virus (DENV) infection is a leading cause of the mosquito-borne infectious diseases that affect humans worldwide. Virus-host interactions appear to play significant roles in DENV replication and the pathogenesis of DENV infection. Nonstructural protein 1 (NS1) of DENV is likely involved in these processes; however, its associations with host cell proteins in DENV infection remain unclear. In this study, we used a combination of techniques (immunoprecipitation, in-solution trypsin digestion, and LC-MS/MS) to identify the host cell proteins that interact with cell-associated NS1 in an in vitro model of DENV infection in the human hepatocyte HepG2 cell line. Thirty-six novel host cell proteins were identified as potential DENV NS1-interacting partners. A large number of these proteins had characteristic binding or catalytic activities, and were involved in cellular metabolism. Coimmunoprecipitation and colocalization assays confirmed the interactions of DENV NS1 and human NIMA-related kinase 2 (NEK2), thousand and one amino acid protein kinase 1 (TAO1), and component of oligomeric Golgi complex 1 (COG1) proteins in virus-infected cells. This study reports a novel set of DENV NS1-interacting host cell proteins in the HepG2 cell line and proposes possible roles for human NEK2, TAO1, and COG1 in DENV infection. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Study of apoptotic mechanisms induced by all-trans retinoic acid and its 13-cis isomer on cellular lines of human hepato carcinoma Hep3B and HepG2

    International Nuclear Information System (INIS)

    Arce Vargas, Frederick

    2006-01-01

    Two cellular lines of liver cancer (Hep3B and HepG2) were incubated during different periods of time with some concentrations of two retinoic acid isomers (ATRA and 13-cis AR) and with 5-fu chemotherapeutic agents, cisplatin and paclitaxel. It was determined if these substances leaded cytotoxicity, apoptosis and if they modified the expression of different genes related to cellular death by apoptosis, in order to explain the hepatocellular carcinoma resistance to these drugs. HepG2 cells showed more resistance than Hep3B cells to 72 hours of treatment, as much ATRA as the 13-cis AR were toxic and produced apoptosis in two cellular lines. This type of cellular death seems to be mediated by a decrease in Bcl-xL concentration in Hep3B cells treated with both retinoids an increase in bax concentration in HepG2 cells treated with 13-cis AR. It were observed 3 and 8 proteolysis of procaspase in Hep3B cells, suggesting extrinsic via activation of the apoptosis, while cellular death in HepG2 cells seems to be independent of caspases. Cisplatin and paclitaxel leaded cytotoxicity to 48 hours of treatment, with significant differences between two cellular lines only in case of paclitaxel. Hep3B cells treated with cisplatin and HepG2 cells treated with paclytaxel suffered apoptosis. 5-FU produced toxicity only when it was used to high concentrations and the mechanism of cellular death induced by this agent seems to be primarily necrosis in Hep3B cells and apoptosis in HepG2. There was decrease in the Bcl-xL concentration in two cellular lines when it was treated with cisplatin and in HepG2 cells treated with 5-FU. Bax concentration there no was modified with no treatment. Activation of the 3 caspases seems to happen only in HepG2 cells with 5-FU and paclytaxel. These two agents, also, decreased the survivin concentration of HepG2 cells. Treatments of the three drugs produced an increase in the expression of this gen in Hep3B cells, which might explain partially the resistance

  12. Cholesterol-lowing effect of taurine in HepG2 cell.

    Science.gov (United States)

    Guo, Junxia; Gao, Ya; Cao, Xuelian; Zhang, Jing; Chen, Wen

    2017-03-16

    A number of studies indicate that taurine promotes cholesterol conversion to bile acids by upregulating CYP7A1 gene expression. Few in vitro studies are concerned the concentration change of cholesterol and its product of bile acids, and the molecular mechanism of CYP7A1 induction by taurine. The levels of intracellular total cholesterol (TC), free cholesterol (FC), cholesterol ester (EC), total bile acids (TBA) and medium TBA were determined after HepG2 cells were cultured for 24/48 h in DMEM supplemented with taurine at the final concentrations of 1/10/20 mM respectively. The protein expressions of CYP7A1, MEK1/2, c-Jun, p-c-Jun and HNF-4α were detected. Taurine significantly reduced cellular TC and FC in dose -and time-dependent ways, and obviously increased intracellular/medium TBA and CYP7A1 expressions. There was no change in c-Jun expression, but the protein expressions of MEK1/2 and p-c-Jun were increased at 24 h and inhibited at 48 h by 20 mM taurine while HNF4α was induced after both of the 24 h and 48 h treatment. Taurine could enhance CYP7A1 expression by inducing HNF4α and inhibiting MEK1/2 and p-c-Jun expressions to promote intracellular cholesterol metabolism.

  13. Evaluation of anti-hepatocarcinoma capacity of puerarin nanosuspensions against human HepG2 cells

    Science.gov (United States)

    Meng, Xiang-Ping; Zhang, Zhen; Wang, Yi-Fei; Wang, Zhi-ping; Chen, Tong-sheng

    2017-02-01

    Hepatocarcinoma, a malignant cancer, threaten human life badly. It is a current issue to seek the effective natural remedy from plant to treat cancer due to the resistance of the advanced hepatocarcinoma to chemotherapy. Puerarin (Pue), a major active ingredient in the traditional Chinese medicine Gegen, has a wide range of pharmacological properties and is considered to have anti-hepatocarcinoma effects. However its low oral bioavailability restricts its wide application. In this report, Pue nanosuspension (Pue-NS) composed of Pue and poloxamer 188 was prepared by high pressure homogenization technique. The in vitro anti-hepatocarcinoma effects of Pue-NS relative to efficacy of bulk Pue were evaluated. The particle size and zeta potential of Pue-NS were 218.5 nm and -18.8 mV, respectively. MTT assay showed that Pue-NS effectively inhibited the proliferation of HepG2 cells, and the corresponding IC50 values of Pue-NS and bulk Pue were 3.39 and 5.73 μg/ml. These results suggest that the delivery of Pue-NS is a promising approach for treating tumors.

  14. Terpenoids from Curcuma wenyujin increased glucose consumption on HepG2 cells.

    Science.gov (United States)

    Zhou, Chang-Xin; Zhang, Li-Sha; Chen, Fei-Fei; Wu, Hao-Shu; Mo, Jian-Xia; Gan, Li-She

    2017-09-01

    Thirty four terpenoids, including two new cadinane-type sesquiterpenoids containing conjugated aromatic-ketone moieties, curcujinone A (1) and curcujinone B (2), were isolated from 95% ethanol extract of the root tubers of Curcuma wenyujin. Their structures were determined by spectroscopic methods, especially 2D NMR and HRMS techniques. The relative and absolute configurations of 1 and 2 were identified by quantum chemical DFT and TDDFT calculations of the 13 C NMR chemical shifts, ECD spectra, and specific optical rotations. All compounds and extracts were evaluated for their anti-diabetic activities with a glucose consumption model on HepG2 Cells. The petroleum fraction CWP (10μg/mL) and compounds curcumenol (4), 7α,11α-epoxy-5β-hydroxy-9-guaiaen-8-one (5), curdione (17), (1S, 4S, 5S 10S)-germacrone (18), zederone (20), a mixture of curcumanolide A (25) and curcumanolide B (26), gajutsulactone B (27), and wenyujinin C (30) showed promising activities with over 45% increasing of glucose consumption at 10μM. Copyright © 2017. Published by Elsevier B.V.

  15. Cytotoxicity and apoptotic effects of tea polyphenol-loaded chitosan nanoparticles on human hepatoma HepG2 cells.

    Science.gov (United States)

    Liang, Jin; Li, Feng; Fang, Yong; Yang, Wenjian; An, Xinxin; Zhao, Liyan; Xin, Zhihong; Cao, Lin; Hu, Qiuhui

    2014-03-01

    Tea polyphenols have strong antioxidant and antitumor activities. However, these health benefits are limited due to their poor in vivo stability and low bioavailability. Chitosan nanoparticles as delivery systems may provide an alternative approach for enhancing bioavailability of poorly absorbed drugs. In this study, tea polyphenol-loaded chitosan nanoparticles have been prepared using two different chitosan biomaterials, and their antitumor effects were evaluated in HepG2 cells, including cell cytotoxicity comparison, cell morphology analysis, cell apoptosis and cell cycle detection. The results indicated that the tea polyphenol-loaded chitosan nanoparticles showed a branch shape and heterogeneous distribution in prepared suspension. MTT assay suggested that tea polyphenol-loaded chitosan nanoparticles could inhibit the proliferation of HepG2 cells, and the cytotoxicity rates were increased gradually and appeared an obvious dose-dependent relationship. Transmission electron microscope images showed that the HepG2 cells treated with tea polyphenol-loaded chitosan nanoparticles exhibited some typical apoptotic features, such as microvilli disappearance, margination of nuclear chromatin, intracytoplasmic vacuoles and the mitochondrial swelling. In addition, the tea polyphenol-loaded chitosan nanoparticles had relatively weak inhibitory effects on HepG2 cancer cells compared with tea polyphenols. Tea polyphenols not only induced cancer cell apoptosis, but also promoted their necrosis. However, tea polyphenol-loaded chitosan nanoparticles exhibited their antitumor effects mainly through inducing cell apoptosis. Our results revealed that the inhibition effects of tea polyphenol-loaded chitosan nanoparticles on tumor cells probably depended on their controlled drug release and effective cell delivery. The chitosan nanoparticles themselves as the delivery carrier showed limited antitumor effects compared with their encapsulated drugs. Copyright © 2013. Published by

  16. Structure related effects of flavonoid aglycones on cell cycle progression of HepG2 cells: Metabolic activation of fisetin and quercetin by catechol-O-methyltransferase (COMT).

    Science.gov (United States)

    Poór, Miklós; Zrínyi, Zita; Kőszegi, Tamás

    2016-10-01

    Dietary flavonoids are abundant in the Plant Kingdom and they are extensively studied because of their manifold pharmacological activities. Recent studies highlighted that cell cycle arrest plays a key role in their antiproliferative effect in different tumor cells. However, structure-activity relationship of flavonoids is poorly characterized. In our study the influence of 18 flavonoid aglycones (as well as two metabolites) on cell cycle distribution was investigated. Since flavonoids are extensively metabolized by liver cells, HepG2 tumor cell line was applied, considering the potential metabolic activation/inactivation of flavonoids. Our major observations are the followings: (1) Among the tested compounds diosmetin, fisetin, apigenin, lutelin, and quercetin provoked spectacular extent of G2/M phase cell cycle arrest. (2) Inhibition of catechol-O-methyltransferase enzyme by entacapone decreased the antiproliferative effects of fisetin and quercetin. (3) Geraldol and isorhamnetin (3'-O-methylated metabolites of fisetin and quercetin, respectively) demonstrated significantly higher antiproliferative effect on HepG2 cells compared to the parent compounds. Based on these results, O-methylated flavonoid metabolites or their chemically modified derivatives may be suitable candidates of tumor therapy in the future. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  17. Implications of altered glutathione metabolism in aspirin-induced oxidative stress and mitochondrial dysfunction in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Haider Raza

    Full Text Available We have previously reported that acetylsalicylic acid (aspirin, ASA induces cell cycle arrest, oxidative stress and mitochondrial dysfunction in HepG2 cells. In the present study, we have further elucidated that altered glutathione (GSH-redox metabolism in HepG2 cells play a critical role in ASA-induced cytotoxicity. Using selected doses and time point for ASA toxicity, we have demonstrated that when GSH synthesis is inhibited in HepG2 cells by buthionine sulfoximine (BSO, prior to ASA treatment, cytotoxicity of the drug is augmented. On the other hand, when GSH-depleted cells were treated with N-acetyl cysteine (NAC, cytotoxicity/apoptosis caused by ASA was attenuated with a significant recovery in oxidative stress, GSH homeostasis, DNA fragmentation and some of the mitochondrial functions. NAC treatment, however, had no significant effects on the drug-induced inhibition of mitochondrial aconitase activity and ATP synthesis in GSH-depleted cells. Our results have confirmed that aspirin increases apoptosis by increased reactive oxygen species production, loss of mitochondrial membrane potential and inhibition of mitochondrial respiratory functions. These effects were further amplified when GSH-depleted cells were treated with ASA. We have also shown that some of the effects of aspirin might be associated with reduced GSH homeostasis, as treatment of cells with NAC attenuated the effects of BSO and aspirin. Our results strongly suggest that GSH dependent redox homeostasis in HepG2 cells is critical in preserving mitochondrial functions and preventing oxidative stress associated complications caused by aspirin treatment.

  18. Implications of altered glutathione metabolism in aspirin-induced oxidative stress and mitochondrial dysfunction in HepG2 cells.

    Science.gov (United States)

    Raza, Haider; John, Annie

    2012-01-01

    We have previously reported that acetylsalicylic acid (aspirin, ASA) induces cell cycle arrest, oxidative stress and mitochondrial dysfunction in HepG2 cells. In the present study, we have further elucidated that altered glutathione (GSH)-redox metabolism in HepG2 cells play a critical role in ASA-induced cytotoxicity. Using selected doses and time point for ASA toxicity, we have demonstrated that when GSH synthesis is inhibited in HepG2 cells by buthionine sulfoximine (BSO), prior to ASA treatment, cytotoxicity of the drug is augmented. On the other hand, when GSH-depleted cells were treated with N-acetyl cysteine (NAC), cytotoxicity/apoptosis caused by ASA was attenuated with a significant recovery in oxidative stress, GSH homeostasis, DNA fragmentation and some of the mitochondrial functions. NAC treatment, however, had no significant effects on the drug-induced inhibition of mitochondrial aconitase activity and ATP synthesis in GSH-depleted cells. Our results have confirmed that aspirin increases apoptosis by increased reactive oxygen species production, loss of mitochondrial membrane potential and inhibition of mitochondrial respiratory functions. These effects were further amplified when GSH-depleted cells were treated with ASA. We have also shown that some of the effects of aspirin might be associated with reduced GSH homeostasis, as treatment of cells with NAC attenuated the effects of BSO and aspirin. Our results strongly suggest that GSH dependent redox homeostasis in HepG2 cells is critical in preserving mitochondrial functions and preventing oxidative stress associated complications caused by aspirin treatment.

  19. In vitro investigations of Cynara scolymus L. extract on cell physiology of HepG2 liver cells

    Directory of Open Access Journals (Sweden)

    Gesine Löhr

    2009-06-01

    Full Text Available The objective of this study was the investigation of a potential influence of artichoke leaf extract (ALE on the cell physiology and gene expression of phase I/II enzymes of human liver cells HepG2 and investigation on potential cell protective effects against ethanol-induced cell toxicity against HepG2 cells. Cell biological assays under in vitro conditions using HepG2 liver cells and investigation of mitochondrial activity (MTT test, proliferation assay (BrdU incorporation ELISA, LDH as toxicity marker, gene expression analysis by RT-PCR and enzyme activity of glutationtransferase. Artichocke extract, containing 27% caffeoylquinic acids and 7% flavonoids induced mitochondrial activity, proliferation and total protein content under in vitro conditions in human liver cells HepG2. These effects could not be correlated to the well-known artichoke secondary compounds cynarin, caffeic acid, chlorogenic acid, luteolin and luteolin-7-O-glucoside. The flavones luteolin and luteolin-7-O-glucoside had inhibitory effects at 100 µg/mL level on HepG2 cells, with luteolin being a significant stronger inhibitor compared to the respective glucoside. Artichoke leaf extract had minor stimulating effect on gene expression of CYP1A2, while CYP3A4, GGT, GPX2, GSR and GST were slightly inhibited. GST inhibition under in vitro conditions was also shown by quantification of GST enzyme activity. Induction of gene expression of CYP1A2 was shown to be supraadditive after simultaneous application of ethanol plus artichoke extract. Artichoke leaf extract exhibited cell protective effects against ethanol-induced toxicity within cotreatment under in vitro conditions. Also H2O2 damage was significantly inhibited by simultaneous artichoke incubation. Pre- and posttreatments did not exert protective effects. DMSO-induced toxicity was significantly reduced by pre-, post- and cotreatment with artichoke extract and especially with luteolin-7-O-glucoside, indicating a direct

  20. HepG2 cells develop signs of riboflavin deficiency within four days of culture in riboflavin-deficient medium*

    OpenAIRE

    Werner, Ricarda; Manthey, Karoline C.; Griffin, Jacob B.; Zempleni, Janos

    2005-01-01

    Flavin mononucleotide and flavin adenine dinucleotide are essential coenzymes in redox reactions. For example, flavin adenine dinucleotide is a coenzyme for both glutathione reductase and enzymes that mediate the oxidative folding of secretory proteins. Here we investigated short-term effects of moderately riboflavin-deficient culture medium on flavin-related responses in HepG2 hepatocarcinoma cells. Cells were cultured in riboflavin-deficient (3.1 nmol/L) medium for up to six days; controls ...

  1. Heat Killed Attenuated Leishmania Induces Apoptosis of HepG2 Cells Through ROS Mediated p53 Dependent Mitochondrial Pathway

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    Dipayan Bose

    2016-03-01

    Full Text Available Background/Aims: Cytotoxic effect of attenuated Leishmania on liver cancer cells by inducing ROS generation. Methods: Spectrophotometric study to analyze cell death and levels of different active caspases. Flow cytometric study was done to analyze apoptosis induction and ROS generation and levels of different protein. Western blot analysis was performed to study the levels of protein. Confocal microscopy was done to ascertain the expression of different apoptotic markers. Results: We have now observed that attenuated Leishmania donovani UR6 also has potentiality towards growth inhibition of HepG2 cells and investigated the mechanism of action. The effect is associated with increased DNA fragmentation, rise in number of annexinV positive cells, and cell cycle arrest at G1 phase. The detection of unregulated levels of active PARP, cleaved caspases 3 and 9, cytosolic cytochrome C, Bax, and Bad, along with the observed downregulation of Bcl-2 and loss of mitochondrial membrane potential suggested the involvement of mitochondrial pathway. Enhanced ROS and p53 levels regulate the apoptosis of HepG2 cells. NAC was found to inhibit p53 production but PFT-α has no effect on ROS generation. In conclusion, Leishmania donovani UR6 efficiently induces apoptosis in HepG2 cells through ROS mediated p53 dependent mitochondrial pathway. Conclusion: It has been reported earlier that some parasites show prominent cytotoxic effect and prevent tumor growth. From our study we found that Leishmania donovani UR6 efficiently induced apoptosis in HepG2 cells through ROS mediated p53 dependent mitochondrial pathway. This study has rejuvenated the age old idea of bio-therapy.

  2. Dryocrassin ABBA Induces Apoptosis in Human Hepatocellular Carcinoma HepG2 Cells Through a Caspase-Dependent Mitochondrial Pathway.

    Science.gov (United States)

    Jin, Zhe; Wang, Wen-Fei; Huang, Jian-Ping; Wang, He-Meng; Ju, Han-Xun; Chang, Ying

    2016-01-01

    Biological and pharmacological activities of dryocrassin ABBA, a phloroglucinol derivative extracted from Dryopteris crassirhizoma, have attracted attention. In this study, the apoptotic effect of dryocrassin ABBA on human hepatocellular carcinoma HepG2 cells was investigated. We tested the effects of dryocrassin ABBA on HepG2 in vitro by MTT, flow cytometry, real-time PCR, and Western blotting. KM male mice were used to detect the effect of dryocrassin ABBA on H22 cells in vivo. Dryocrassin ABBA inhibited the growth of HepG2 cells in a concentration-dependent manner. After treatment with 25, 50, and 75 μg/mL dryocrassin ABBA, the cell viability was 68%, 60% and 49%, respectively. Dryocrassin ABBA was able to induce apoptosis, measured by propidium iodide (PI)/annexin V-FITC double staining. The results of real-time PCR and Western ting showed that dryocrassin ABBA up-regulated p53 and Bax expression and inhibited Bcl-2 expression which led to an activation of caspase-3 and caspase-7 in the cytosol, and then induction of cell apoptosis. In vivo experiments also showed that dryocrassin ABBA treatment significantly suppressed tumor growth, without major side effects. Overall, these findings provide evidence that dryocrassin ABBA may induce apoptosis in human hepatocellular carcinoma cells through a caspase-mediated mitochondrial pathway.

  3. PPAR{gamma} activates ABCA1 gene transcription but reduces the level of ABCA1 protein in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Mogilenko, Denis A., E-mail: denis@iem.sp.ru [Department of Biochemistry, Institute of Experimental Medicine, Russian Academy of Medical Sciences, 197376 St. Petersburg (Russian Federation); Department of Embryology, St. Petersburg State University, 199034 St. Petersburg (Russian Federation); Shavva, Vladimir S. [Department of Embryology, St. Petersburg State University, 199034 St. Petersburg (Russian Federation); Dizhe, Ella B. [Department of Biochemistry, Institute of Experimental Medicine, Russian Academy of Medical Sciences, 197376 St. Petersburg (Russian Federation); Orlov, Sergey V., E-mail: serge@iem.sp.ru [Department of Biochemistry, Institute of Experimental Medicine, Russian Academy of Medical Sciences, 197376 St. Petersburg (Russian Federation); Department of Embryology, St. Petersburg State University, 199034 St. Petersburg (Russian Federation); Perevozchikov, Andrej P., E-mail: app@iem.sp.ru [Department of Biochemistry, Institute of Experimental Medicine, Russian Academy of Medical Sciences, 197376 St. Petersburg (Russian Federation); Department of Embryology, St. Petersburg State University, 199034 St. Petersburg (Russian Federation)

    2010-11-19

    Research highlights: {yields} PPAR{gamma} activates ABCA1 gene expression but decreases ABCA1 protein content in human hepatoma cell line HepG2. {yields} Treatment of HepG2 cells with PPAR{gamma} agonist GW1929 leads to dissociation of LXR{beta} from ABCA1-LXR{beta} complex. {yields} Inhibition of protein kinases MEK1/2 abolishes PPAR{gamma}-mediated dissociation of LXR{beta} from ABCA1/LXR{beta} complex. {yields} Activation of PPAR{gamma} leads to increasing of the level of LXR{beta} associated with LXRE within ABCA1 gene promoter. -- Abstract: Synthesis of ABCA1 protein in liver is necessary for high-density lipoproteins (HDL) formation in mammals. Nuclear receptor PPAR{gamma} is known as activator of ABCA1 expression, but details of PPAR{gamma}-mediated regulation of ABCA1 at both transcriptional and post-transcriptional levels in hepatocytes have not still been well elucidated. In this study we have shown, that PPAR{gamma} activates ABCA1 gene transcription in human hepatoma cells HepG2 through increasing of LXR{beta} binding with promoter region of ABCA1 gene. Treatment of HepG2 cells with PPAR{gamma} agonist GW1929 leads to dissociation of LXR{beta} from ABCA1/LXR{beta} complex and to nuclear translocation of this nuclear receptor resulting in reduction of ABCA1 protein level 24 h after treatment. Inhibition of protein kinases MEK1/2 abolishes PPAR{gamma}-mediated dissociation of LXR{beta} from ABCA1/LXR{beta} complex, but does not block PPAR{gamma}-dependent down-regulation of ABCA1 protein in HepG2 cells. These data suggest that PPAR{gamma} may be important for regulation of the level of hepatic ABCA1 protein and indicate the new interplays between PPAR{gamma}, LXR{beta} and MEK1/2 in regulation of ABCA1 mRNA and protein expression.

  4. Protective effects of the extracts of Barringtonia racemosa shoots against oxidative damage in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Kin Weng Kong

    2016-01-01

    Full Text Available Barringtonia racemosa is a tropical plant with medicinal values. In this study, the ability of the water extracts of the leaf (BLE and stem (BSE from the shoots to protect HepG2 cells against oxidative damage was studied. Five major polyphenolic compounds consisting of gallic acid, ellagic acid, protocatechuic acid, quercetin and kaempferol were identified using HPLC-DAD and ESI-MS. Cell viability assay revealed that BLE and BSE were non-cytotoxic (cell viabilities >80% at concentration less than 250 µg/ml and 500 µg/ml, respectively. BLE and BSE improved cellular antioxidant status measured by FRAP assay and protected HepG2 cells against H2O2-induced cytotoxicity. The extracts also inhibited lipid peroxidation in HepG2 cells as well as the production of reactive oxygen species. BLE and BSE could also suppress the activities of superoxide dismutase and catalase during oxidative stress. The shoots of B. racemosa can be an alternative bioactive ingredient in the prevention of oxidative damage.

  5. In HepG2 cells, coexisting carnitine deficiency masks important indicators of marginal biotin deficiency.

    Science.gov (United States)

    Bogusiewicz, Anna; Boysen, Gunnar; Mock, Donald M

    2015-01-01

    A large number of birth defects are related to nutrient deficiencies; concern that biotin deficiency is teratogenic in humans is reasonable. Surprisingly, studies indicate that increased urinary 3-hydroxyisovalerylcarnitine (3HIAc), a previously validated marker of biotin deficiency, is not a valid biomarker in pregnancy. In this study we hypothesized that coexisting carnitine deficiency can prevent the increase in 3HIAc due to biotin deficiency. We used a 2-factor nutrient depletion design to induce isolated and combined biotin and carnitine deficiency in HepG2 cells and then repleted cells with carnitine. To elucidate the metabolic pathogenesis, we quantitated intracellular and extracellular free carnitine, acylcarnitines, and acylcarnitine ratios using liquid chromatography-tandem mass spectrometry. Relative to biotin-sufficient, carnitine-sufficient cells, intracellular acetylcarnitine increased by 90%, propionylcarnitine more than doubled, and 3HIAc increased by >10-fold in biotin-deficient, carnitine-sufficient (BDCS) cells, consistent with a defensive mechanism in which biotin-deficient cells transesterify the acyl-coenzyme A (acyl-CoA) substrates of the biotin-dependent carboxylases to the related acylcarnitines. Likewise, in BDCS cells, the ratio of acetylcarnitine to malonylcarnitine and the ratio of propionylcarnitine to methylmalonylcarnitine both more than tripled, and the ratio of 3HIAc to 3-methylglutarylcarnitine (MGc) increased by >10-fold. In biotin-deficient, carnitine-deficient (BDCD) cells, the 3 substrate-derived acylcarnitines changed little, but the substrate:product ratios were masked to a lesser extent. Moreover, carnitine repletion unmasked biotin deficiency in BDCD cells as shown by increases in acetylcarnitine, propionylcarnitine, and 3HIAc (each increased by >50-fold). Likewise, ratios of acetylcarnitine:malonylcarnitine, propionylcarnitine:methylmalonylcarnitine, and 3HIAc:MGc all increased by >8-fold. Our findings provide strong

  6. Synthesis of Functionalized Fluorescent Silver Nanoparticles and their toxicological effect in aquatic environments (Goldfish) and HEPG2 cells.

    Science.gov (United States)

    Santos, Hugo; Oliveira, Elisabete; Garcia-Pardo, Javier; Diniz, Mário; Lorenzo, Julia; Rodriguez-González, Benito; Capelo, José Luis; Lodeiro, Carlos

    2013-12-01

    Silver nanoparticles, AgNPs, are widely used in our daily life, mostly due to their antibacterial, antiviral and antifungal properties. However, their potential toxicity remains unclear. In order to unravel this issue, emissive AgNPs were first synthetized using an inexpensive photochemical method, and then their permeation was assessed in vivo in goldfish and in vitro in human hepatoma cells (HepG2). In addition, the oxidative stress caused by AgNPs was assessed in enzymes such as glutathione-S-transferase (GST), catalase (CAT) and in lipid peroxidation (LPO). This study demonstrates that the smallest sized AgNPs@3 promote the largest changes in gold fish livers, whereas AgNPs@1 were found to be toxic in HEPG2 cells depending on both the size and functionalized/stabilizer ligand.

  7. Curcumin induced nanoscale CD44 molecular redistribution and antigen-antibody interaction on HepG2 cell surface

    International Nuclear Information System (INIS)

    Wang Mu; Ruan Yuxia; Xing Xiaobo; Chen Qian; Peng, Yuan; Cai Jiye

    2011-01-01

    Graphical abstract: Highlights: → In this study, we investigate the changes of CD44 expression and distribution on HepG2 cells after curcumin treatment. → We find curcumin is able to change the morphology and ultrastructure of HepG2 cells. → Curcumin can reduce the expression of CD44 molecules and induce the nanoscale molecular redistribution on cell surface. → The binding force between CD44-modified AFM tip and the HepG2 cell surface decreases after curcumin-treatment. - Abstract: The cell surface glycoprotein CD44 was implicated in the progression, metastasis and apoptosis of certain human tumors. In this study, we used atomic force microscope (AFM) to monitor the effect of curcumin on human hepatocellular carcinoma (HepG2) cell surface nanoscale structure. High-resolution imaging revealed that cell morphology and ultrastructure changed a lot after being treated with curcumin. The membrane average roughness increased (10.88 ± 4.62 nm to 129.70 ± 43.72 nm) and the expression of CD44 decreased (99.79 ± 0.16% to 75.14 ± 8.37%). Laser scanning confocal microscope (LSCM) imaging showed that CD44 molecules were located on the cell membrane. The florescence intensity in control group was weaker than that in curcumin treated cells. Most of the binding forces between CD44 antibodies and untreated HepG2 cell membrane were around 120-220 pN. After being incubated with curcumin, the major forces focused on 70-150 pN (10 μM curcumin-treated) and 50-120 pN (20 μM curcumin-treated). These results suggested that, as result of nanoscale molecular redistribution, changes of the cell surface were in response to external treatment of curcumin. The combination of AFM and LSCM could be a powerful method to detect the distribution of cell surface molecules and interactions between molecules and their ligands.

  8. Protective role of metabolism by intestinal microflora in butyl paraben-induced toxicity in HepG2 cell cultures.

    Science.gov (United States)

    Khanal, Tilak; Kim, Hyung Gyun; Jin, Sun Woo; Shim, Eol; Han, Hwa Jeong; Noh, Keumhan; Park, Sunkyoung; Lee, Dae Hun; Kang, Wonku; Yeo, Hee Kyung; Kim, Dong Hyun; Jeong, Tae Cheon; Jeong, Hye Gwang

    2012-09-03

    Parabens are alkyl esters of p-hydroxybenzoic acid (BA), including methyl paraben (MP), ethyl paraben, propyl paraben (PP), and butyl paraben (BP). In the present study, possible role of metabolism by fecalase in BP-induced cytotoxicity was investigated in HepG2 cell cultures. As an intestinal bacterial metabolic system, a human fecalase prepared from human fecal specimen was employed. Among the parabens tested, cytotoxicity of BP was most severe. BA, the de-esterified metabolite, did not induce cytotoxicity when compared to other parabens. When BP was incubated with fecalase, it rapidly disappeared, in association with reduced cytotoxicity in HepG2 cells. In addition, BP incubated with fecalase significantly caused an increase in Bcl-2 expression together with a decrease in Bax expression and cleaved caspase-3. Moreover, anti-apoptotic effect by the incubation of BP with fecalase was also confirmed by the TUNEL assay. Furthermore, BP induced a sustained activation of the phosphorylation of JNK only when it was treated alone. Meanwhile, BP-induced cell death was reversed by the pre-incubation of BP with either fecalase or SP600125. Taken together, the findings suggested that metabolism of BP by human fecalase might have protective effects against BP-induced toxicity in HepG2 cells. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  9. Biphasic Estradiol-induced AKT Phosphorylation Is Modulated by PTEN via MAP Kinase in HepG2 Cells

    Science.gov (United States)

    Marino, Maria; Acconcia, Filippo; Trentalance, Anna

    2003-01-01

    We reported previously in HepG2 cells that estradiol induces cell cycle progression throughout the G1–S transition by the parallel stimulation of both PKC-α and ERK signaling molecules. The analysis of the cyclin D1 gene expression showed that only the MAP kinase pathway was involved. Here, the presence of rapid/nongenomic, estradiol-regulated, PI3K/AKT signal transduction pathway, its modulation by the levels of the tumor suppressor PTEN, its cross-talk with the ERK pathway, and its involvement in DNA synthesis and cyclin D1 gene promoter activity have all been studied in HepG2 cells. 17β-Estradiol induced the rapid and biphasic phosphorylation of AKT. These phosphorylations were independent of each other, being the first wave of activation independent of the estrogen receptor (ER), whereas the second was dependent on ER. Both activations were dependent on PI3K activity; furthermore, the ERK pathway modulated AKT phosphorylation by acting on the PTEN levels. The results showed that the PI3K pathway, as well as ER, were strongly involved in both G1–S progression and cyclin D1 promoter activity by acting on its proximal region (-254 base pairs). These data indicate that in HepG2 cells, different rapid/nongenomic estradiol-induced signal transduction pathways modulate the multiple steps of G1–S phase transition. PMID:12808053

  10. Determination of acute lethal and chronic lethal dose thresholds of valproic acid using 3D spheroids constructed from the immortal human hepatocyte cell line HepG2/C3A

    DEFF Research Database (Denmark)

    Fey, S. J.; Wrzesinski, K.

    2013-01-01

    Valproic acid (VPA) is a broad-spectrum antiepileptic drug that is now used commonly for several other neurological and psychiatric indications. While VPA is usually well tolerated, on rare occasions, it has been associated with severe, and sometimes, fatal liver injuries. These complications may...... also arise due to acute VPA overdose. As a branched chain carboxylic acid, VPA is readily metabolized in the liver via glucuronic acid conjugation, mitochondrial β- and cytosolic ω-oxidation to produce multiple metabolites, and it is probably some of these metabolites that are involved in its toxicity....... While the actual mechanism of VPA hepatotoxicity is still not understood, two pathways have been in focus: the first involves the formation of reactive metabolites of VPA (and their subsequent covalent binding to cellular proteins) and the second is the development of oxidative stress in the cell. We...

  11. Rosemary Extracts Upregulate Nrf2, Sestrin2, and MRP2 Protein Level in Human Hepatoma HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Xiao-pei Tong

    2017-01-01

    Full Text Available In the past few decades, the incidence of liver cancer has been rapidly rising across the world. Rosemary is known to possess antioxidant activity and is used as natural antioxidant food preservative. It is proposed to have anticancer activity in treating different tumor models. In this study, we try to explore the impact of rosemary extracts on upregulating the level of Nrf2 and Nrf2-regulatory proteins, Sestrin2 and MRP2 in HepG2 cells, and to speculate its potential mechanism. The anticancer activity of rosemary extract, including its polyphenolic diterpenes carnosic acid and carnosol, was evaluated to understand the potential effect on HepG2 cells. Rosemary extract, carnosic acid, and carnosol induced the expression of Sestrin2 and MRP2 associate with enhancement of Nrf2 protein level in HepG2 cells, in which carnosic acid showed most obvious effect. Although the activation pathway of Nrf2/ARE was not exactly assessed, it can be assumed that the enhancement of expression of Sestrin2 and MRP2 may result from upregulation of Nrf2.

  12. Bioavailability and metabolism of rosemary infusion polyphenols using Caco-2 and HepG2 cell model systems.

    Science.gov (United States)

    Achour, Mariem; Saguem, Saad; Sarriá, Beatriz; Bravo, Laura; Mateos, Raquel

    2018-01-12

    Rosmarinus officinalis is an aromatic plant used in folk medicine as a result of the therapeutic properties associated with its phenolic composition, being rich in rosmarinic acid (RA) and caffeic acid (CA). To better understand the bioactivity of these compounds, their absorption and metabolism were assessed in human Caco-2 and HepG2 cells, as small intestine and liver models, respectively, using RA and CA standards, as well as a rosemary infusion and ferulic acid (FA). Test compounds were partially up-taken and metabolized by Caco-2 and HepG2 cells, although a higher metabolization rate was observed after hepatic incubation compared to intestinal incubation. CA was the compound best absorbed followed by RA and FA, showing metabolites percentages of 30.4%, 11.8% and 4.4% in Caco-2 and 34.3%, 10.3% and 3.2% in HepG2 cells, respectively. RA in the rosemary infusion showed improved bioavailability compared to pure RA. Methyl derivatives were the main metabolites detected for CA and RA after intestinal and hepatic metabolism, followed by methyl-glucuronidates and glucuronidates. RA was also minimally hydrolyzed into CA, whereas FA only was glucuronidated. Rosemary polyphenols followed the same biotransformation pathways as the standards. In addition, phase II derivatives of luteolin were observed. Rosemary polyphenols are partially metabolized in both the intestine and liver. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

  13. Cocoa flavonoids improve insulin signalling and modulate glucose production via AKT and AMPK in HepG2 cells.

    Science.gov (United States)

    Cordero-Herrera, Isabel; Martín, María Angeles; Bravo, Laura; Goya, Luis; Ramos, Sonia

    2013-06-01

    Cocoa and (-)-epicatechin (EC), a main cocoa flavanol, have been suggested to exert beneficial effects in diabetes, but the mechanism for their insulin-like effects remains unknown. In this study, the modulation of insulin signalling by EC and a cocoa phenolic extract (CPE) on hepatic HepG2 cells was investigated by analysing key proteins of the insulin pathways, namely insulin receptor, insulin receptor substrate (IRS) 1 and 2, PI3K/AKT and 5'-AMP-activated protein kinase (AMPK), as well as the levels of the glucose transporter GLUT-2 and the hepatic glucose production. EC and CPE enhanced the tyrosine phosphorylation and total insulin receptor, IRS-1 and IRS-2 levels and activated the PI3K/AKT pathway and AMPK in HepG2 cells. CPE also enhanced the levels of GLUT-2. Interestingly, EC and CPE modulated the expression of phosphoenolpyruvate carboxykinase, a key protein involved in the gluconeogenesis, leading to a diminished glucose production. In addition, EC- and CPE-regulated hepatic gluconeogenesis was prevented by the blockage of AKT and AMPK. Our data suggest that EC and CPE strengthen the insulin signalling by activating key proteins of that pathway and regulating glucose production through AKT and AMPK modulation in HepG2 cells. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Hypertonicity-induced cation channels in HepG2 cells: architecture and role in proliferation vs. apoptosis.

    Science.gov (United States)

    Koos, Björn; Christmann, Jens; Plettenberg, Sandra; Käding, Domenic; Becker, Julia; Keteku, Melody; Klein, Christian; Imtiaz, Sarah; Janning, Petra; Bastiaens, Philippe I H; Wehner, Frank

    2018-01-25

    Na + conducting hypertonicity-induced cation channels (HICCs) are key players in the volume restoration of osmotically shrunken cells and, under isotonic conditions, considered as mediators of proliferation - thereby opposing apoptosis. In an siRNA screen of ion channels and transporters in HepG2 cells, with the regulatory volume increase (RVI) as read-out, δENaC, TRPM2 and TRPM5 were identified as HICCs. Subsequently, all permutations of these channels were tested in RVI and patch-clamp recordings and, at first sight, HICCs were found to operate in an independent mode. However, there was synergy in the siRNA perturbations of HICC currents. Accordingly, proximity ligation assays showed that δENaC was located in proximity to TRPM2 and TRPM5 suggesting a physical interaction. Furthermore, δENaC, TRPM2 and TRPM5 were identified as mediators of HepG2 proliferation - their silencing enhanced apoptosis. Our study defines the architecture of HICCs in human hepatocytes as well as their molecular functions. Hypertonicity-induced cation channels (HICCs) are a substantial element in the regulatory volume increase (RVI) of osmotically shrunken cells. Under isotonic conditions, they are key effectors in the volume gain preceding proliferation; HICC repression, in turn, significantly increases apoptosis rates. Despite these fundamental roles of HICCs in cell physiology, very little is known concerning the actual molecular architecture of these channels. Here, an siRNA screening of putative ion channels and transporters was performed, in HepG2 cells, with the velocity of RVI as the read-out; in this first run, δENaC, TRPM2 and TRPM5 could be identified as HICCs. In the second run, all permutations of these channels were tested in RVI and patch-clamp recordings, with special emphasis on the non-additivity and additivity of siRNAs - which would indicate molecular interactions or independent ways of channel functioning. At first sight, the HICCs in HepG2 cells appeared to

  15. Effects of Elaidic Acid on Lipid Metabolism in HepG2 Cells, Investigated by an Integrated Approach of Lipidomics, Transcriptomics and Proteomics

    DEFF Research Database (Denmark)

    Nielsen, L. V.; Krogager, T. P.; Young, C.

    2013-01-01

    Trans fatty acid consumption in the human diet can cause adverse health effects, such as cardiovascular disease, which is associated with higher total cholesterol, a higher low density lipoprotein-cholesterol level and a decreased high density lipoprotein-cholesterol level. The aim of the study...... was to elucidate the hepatic response to the most abundant trans fatty acid in the human diet, elaidic acid, to help explain clinical findings on the relationship between trans fatty acids and cardiovascular disease. The human HepG2 cell line was used as a model to investigate the hepatic response to elaidic acid...

  16. A high-throughput quantitative expression analysis of cancer-related genes in human HepG2 cells in response to limonene, a potential anticancer agent.

    Science.gov (United States)

    Hafidh, Rand R; Hussein, Saba Z; MalAllah, Mohammed Q; Abdulamir, Ahmed S; Abu Bakar, Fatimah

    2017-11-14

    Citrus bioactive compounds, as active anticancer agent, have been under focus by several studies worldwide. However, the underlying genes responsible for the anticancer potential have not been sufficiently highlighted. The current study investigated the gene expression profile of hepatocellular carcinoma, HepG2, cells after treatment with Limonene. The concentration that killed 50% of HepG2 cells was used to elucidate the genetic mechanisms of limonene anticancer activity. The apoptotic induction was detected by flow cytometry and confocal fluorescence microscope. Two of pro-apoptotic events, caspase-3 activation and phosphatidylserine translocation were manifested by confocal fluorescence microscopy. High-throughput real-time PCR was used to profile 1023 cancer-related genes in 16 different gene families related to the cancer development. In comparison to untreated cells, limonene increased the percentage of apoptotic cells up to 89.61%, by flow cytometry, and 48.2% by fluorescence microscopy. There was a significant limonene-driven differential gene expression of HepG2 cells in 15 different gene families. Limonene was shown to significantly (>2log) up-regulate and down-regulate 14 and 59 genes, respectively. The affected gene families, from most to least affected, were apoptosis induction, signal transduction, cancer genes augmentation, alteration in kinases expression, inflammation, DNA damage repair, and cell cycle proteins. The current study reveals that limonene could be a promising, cheap, and effective anticancer compound. The broad spectrum of limonene anticancer activity is interesting for anticancer drug development. Further research is needed to confirm the current findings and to examine the anticancer potential of limonene along with underlying mechanisms on different cell lines. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  17. Flow cytometry based micronucleus assay for evaluation of genotoxic potential of 2-ACBs in hepatic cells HepG2

    Energy Technology Data Exchange (ETDEWEB)

    Barbezan, Angélica B.; Santos, Carla J.B.; Carvalho, Luma R.; Vieira, Daniel P.; Villavicêncio, Anna L.C.H., E-mail: abarbezan@ipen.br [Instituto de Pesquisas Energéticas e Nucleares (IPEN/CNEN-SP), São Paulo, SP (Brazil); Santelli, Glaucia M.M. [Universidade de São Paulo (USP), SP (Brazil). Departamento de Biologia Celular e do Desenvolvimento

    2017-07-01

    Food irradiation is approved for use in more than 60 countries for applications and purposes in a wide variety of foods, being an effective and safe method for preservation and long-term storage. 2-Alkylcyclobutanones (2-ACBs) are the only known radiolytic products generated from foods that contain fatty acids (Triglycerides) when irradiated. The acids analyzed in this study are palmitic and stearic, which when irradiated form 2-Dodecylcyclobutanones (2-dDCB) and 2-Tetradecylcyclobutanone (2-tDCB). Part of the 2-ACBs ingested is excreted through feces and part is deposited in adipose tissues. In vitro studies so far have been only in colon cells. The work used a human hepatoma cell line (HepG2) since the accumulation of fat in this organ is quite common. Micronucleus test was selected to evaluate possible genotoxic effects of 2-dDCB and 2-tDCB compounds when exposed to high concentrations (447, 1422 and 2235 μM) for 4 and 24 hours. Tests were performed in quadriplicates using flow cytometric analysis. None detectable genotoxic damage was observed after 4 hours of exposure to the compounds, and cytotoxic effects were only significant at the highest concentration (2235 μM) of 2-dDCB. After 24 hours of exposure, slight genotoxic damage was observed at all concentrations evaluated, and cytotoxic effects were only present when exposed to compound 2-tDCB. Although there is a genotoxic and cytotoxic effect in some of the situations tested, the two compounds predominantly induced proliferation reduction effects of this hepatic tumor cell line. (author)

  18. [Preparation of a glypican-3-targeting hepatocellular carcinoma MR probe and its molecular imaging in HepG2 cells].

    Science.gov (United States)

    Gu, Yan; Zeng, Yan; Guo, Da-Jing; Yang, Jing; Zhou, Jun; Liu, Xin-Jie; Wang, Zhi-Gang

    2014-02-01

    To prepare a glypican-3 (GPC3)-targeting hepatocellular carcinoma MR molecular probe and to evaluate its targeting specificity using HepG2 cells. Poly(lactic-co-glycolic acid) (PLGA) nanoparticles were prepared by a double emulsion solvent evaporation method, and the surfaces were connected with anti-GPC3 mono-antibody and paramagnetic substance Gd3+. The physical properties of the probes were investigated using fluorescence microscopy, electron microscopy, Malvern particle size analysis, inductively coupled plasma atomic emission spectroscopy (ICP-AES) and 1.5T MR imaging. The specificity of the probes to target cultured HepG2 cells was determined by laser confocal microscopy. The signal characteristics, including signal-to-noise ratio (SNR), after co-incubation with HepG2 cells were analyzed by 1.5T MR imaging. Significance of differences between multiple groups (target group, non-target group, and control group) was assessed by one-way analysis of variance, and between two groups was assessed by the LSD-t test. A difference was considered to be statistically significant at P less than 0.05. The GPC3-targeting hepatocellular carcinoma MR molecular probes were successfully prepared. The nanoparticles had a spherical shape, size of 495 +/- 17.5 nm, uniform size distribution, good dispersibility, no obvious aggregation, and could significantly increase the T1 signal. Using the ICP-AES measurement, 1 mol PLGA carried about 12 mol Gd3+, and as the Gd3+ concentration increased, the T1 signal increased. The prepared MR molecular probes could specifically target HepG2 cells, and could increase the T1 signal. The SNR value of the target group was 3.45 +/- 0.21, of the non-target group was 1.43 +/- 0.07, and of the control group was 1.12 +/- 0.03. The SNR value of the target group was significantly higher than that of the non-target group and the control group (P less than 0.05); there was no significant difference between the non-target group and the control group (P more

  19. Basil extract inhibits the sulfotransferase mediated formation of DNA adducts of the procarcinogen 1′-hydroxyestragole by rat and human liver S9 homogenates and in HepG2 human hepatoma cells

    NARCIS (Netherlands)

    Jeurissen, S.M.F.; Punt, A.; Delatour, T.; Rietjens, I.M.C.M.

    2008-01-01

    The effects of a basil extract on the sulfation and concomitant DNA adduct formation of the proximate carcinogen 1′-hydroxyestragole were studied using rat and human liver S9 homogenates and the human hepatoma cell line HepG2. Basil was chosen since it contains the procarcinogen estragole that can

  20. Hepatitis B virus induces G1 phase arrest by regulating cell cycle genes in HepG2.2.15 cells

    Directory of Open Access Journals (Sweden)

    Zhang Chong

    2011-05-01

    Full Text Available Abstract Background To investigate the effect of HBV on the proliferative ability of host cells and explore the potential mechanism. Methods MTT, colony formation assay and tumourigenicity in nude mice were performed to investigate the effect of HBV on the proliferative capability of host cells. In order to explore the potential mechanism, cell cycle and apoptosis were analysed. The cell cycle genes controlling the G1/S phase transition were detected by immunohistochemistry, westernblot and RT-PCR. Results HepG2.2.15 cells showed decreased proliferation ability compared to HepG2 cells. G1 phase arrest was the main cause but was not associated with apoptosis. p53, p21 and total retinoblastoma (Rb were determined to be up-regulated, whereas cyclinE was down-regulated at both the protein and mRNA levels in HepG2.2.15 cells. The phosphorylated Rb in HepG2.2.15 cells was decreased. Conclusions Our results suggested that HBV inhibited the capability of proliferation of HepG2.2.15 cells by regulating cell cycle genes expression and inducing G1 arrest.

  1. Characterization and Purification of Bergamottin from Citrus grandis (L. Osbeck cv. Yongjiazaoxiangyou and Its Antiproliferative Activity and Effect on Glucose Consumption in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Yilong Liu

    2017-07-01

    Full Text Available Bergamottin is a natural furanocoumarin compound with weak polarity. Characterization and quantification of bergamottin were carried out in different fruit tissues of various citrus cultivars. Among the four citrus tissues tested, i.e., flavedo, albedo, segment membrane (SM, and juice sacs (JS in eight citrus cultivars, the highest bergamottin content was found in the flavedo of Citrus grandis (L. Osbeck cv. Yongjiazaoxiangyou (YJZXY, 666.54 μg·g−1 DW. A combination of silica gel column chromatography and high-speed counter-current chromatography (HSCCC was established to efficiently purify bergamottin from the flavedo of YJZXY. Bergamottin showed significant antiproliferative activity on three cancer cell lines, i.e., human liver cancer HepG2, promyelocytic leukemia HL-60, and gastric cancer BGC-823 cells, which showed a marked inhibition effect on these cell lines in a dose-dependent manner. In addition, bergamottin significantly increased glucose consumption in HepG2 cells also in a dose-dependent manner, which is the first report of its potential in anti-diabetes applications.

  2. BC047440 antisense eukaryotic expression vectors inhibited HepG2 cell proliferation and suppressed xenograft tumorigenicity

    International Nuclear Information System (INIS)

    Lu, Zheng; Ping, Liang; JianBo, Zhou; XiaoBing, Huang; Yu, Wen; Zheng, Wang; Jing, Li

    2012-01-01

    The biological functions of the BC047440 gene highly expressed by hepatocellular carcinoma (HCC) are unknown. The objective of this study was to reconstruct antisense eukaryotic expression vectors of the gene for inhibiting HepG 2 cell proliferation and suppressing their xenograft tumorigenicity. The full-length BC047440 cDNA was cloned from human primary HCC by RT-PCR. BC047440 gene fragments were ligated with pMD18-T simple vectors and subsequent pcDNA3.1(+) plasmids to construct the recombinant antisense eukaryotic vector pcDNA3.1(+)BC047440AS. The endogenous BC047440 mRNA abundance in target gene-transfected, vector-transfected and naive HepG 2 cells was semiquantitatively analyzed by RT-PCR and cell proliferation was measured by the MTT assay. Cell cycle distribution and apoptosis were profiled by flow cytometry. The in vivo xenograft experiment was performed on nude mice to examine the effects of antisense vector on tumorigenicity. BC047440 cDNA fragments were reversely inserted into pcDNA3.1(+) plasmids. The antisense vector significantly reduced the endogenous BC047440 mRNA abundance by 41% in HepG 2 cells and inhibited their proliferation in vitro (P < 0.01). More cells were arrested by the antisense vector at the G 1 phase in an apoptosis-independent manner (P = 0.014). Additionally, transfection with pcDNA3.1(+) BC047440AS significantly reduced the xenograft tumorigenicity in nude mice. As a novel cell cycle regulator associated with HCC, the BC047440 gene was involved in cell proliferation in vitro and xenograft tumorigenicity in vivo through apoptosis-independent mechanisms

  3. Andrographolide induces cell cycle arrest at G2/M phase and cell death in HepG2 cells via alteration of reactive oxygen species.

    Science.gov (United States)

    Li, Jieliang; Cheung, Hon-Yeung; Zhang, Zhiqiang; Chan, Gallant K L; Fong, Wang-Fun

    2007-07-30

    The cytotoxicity of andrographolide to HepG2 human hepatoma cells was investigated in the present study. Growth of HepG2 cells was affected in the presence of andrographolide with an IC(50) of 40.2 microM after 48 h treatment. Flow cytometric analysis and DNA fragmentation assay revealed that andrographolide induced cell cycle arrest at G2/M phase and a late apoptosis of the cells. The occurrence of cell cycle arrest was accompanied by the collapse of mitochondrial membrane potential (MMP) and an intracellular increase of hydrogen peroxide (H(2)O(2)) but a decrease of superoxide radicals (O(2)(-)) and reduced glutathione. In the treated cells, expression of Bax as well as the transcriptional controller of this pro-apoptotic gene, p53, was upregulated but not other apoptotic proteins such as Bad, Bcl-2 and Bcl-X(L). Although the activity of caspase-3, which has direct effect on apoptosis, was also enhanced by the presence of andrographolide, cell death of HepG2 could neither be prevented by a specific inhibitor of capsase-3 nor the pan-caspase inhibitor-zVAD (Val-Ala-Asp), indicating that it was a caspase-independent cell death. Since the overall percentage of apoptotic cells was relatively small throughout the experimental studies, we conclude that the cytotoxic effect of andrographolide on HepG2 cells is primary attributed to the induction of cell cycle arrest via the alteration of cellular redox status.

  4. Synthesis, characterization and anticancer effect of the ruthenium (II) polypyridyl complexes on HepG2 cells.

    Science.gov (United States)

    Wan, Dan; Lai, Shang-Hai; Yang, Hui-Hui; Tang, Bing; Zhang, Cheng; Yin, Hui; Zeng, Chuan-Chuan; Liu, Yun-Jun

    2016-12-01

    As one of the major cell regulated center, mitochondria are closely associated with cell proliferation, apoptosis of tumor cell. In this work, four new ruthenium (II) polypyridyl complexes [Ru(bpy) 2 (FTTP)](ClO 4 ) 2 (1) (FTTP=11-(3-fluoro-naphthalen-2-yloxy)-4,5,9,14-tetraaza-benzo[b]triphenylene, bpy=2,2'-bipyridine), [Ru(phen) 2 (FTTP)](ClO 4 ) 2 (2) (phen=1,10-phenanthroline), [Ru(bpy) 2 (PTTP)](ClO 4 ) 2 (3) (PTTP=2-phenoxy-1,4,8,9-tetraazatriphenylene) and [Ru(phen) 2 (PTTP)](ClO 4 ) 2 (4) were synthesized and characterized by elemental analysis, ESI-MS, 1 H NMR and 13 C NMR. The cytotoxic activity, ability of inhibiting cell invasion, cell cycle arrest and apoptosis-inducing mechanism of these Ru(II) complexes have been investigated in detail by MTT (3-(4,5-dimethylthiazole)-2,5-diphenyltetrazolium bromide) method, invasion assay, comet assay as well as western blotting techniques. Notably, complexes 1-4 displayed high cytotoxic activity against liver carcinoma HepG2 cells and the IC 50 values of complexes 1-4 against HepG2 cells are 10.4±1.2, 9.3±0.6, 29.1±1.5 and 5.6±1.2μM, respectively. The comet assay showed that the complexes can induce DNA damage. The acridine orange (AO) and ethidium bromide (EB) staining method indicated that the complexes can cause apoptosis in HepG2 cells. Further studies showed that complexes 1-4 caused cell cycle arrest at G0/G1 phase and induced HepG2 cells apoptosis through a ROS-mediated mitochondrial dysfunction pathway, which involved an increase in the levels of reactive oxygen species (ROS), a decrease in the mitochondrial membrane potential, activation of caspases and Bcl-2 family proteins. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Comparative cytotoxicity of nanosilver in human liver HepG2 and colon Caco2 cells in culture.

    Science.gov (United States)

    Sahu, Saura C; Zheng, Jiwen; Graham, Lesley; Chen, Lynn; Ihrie, John; Yourick, Jeffrey J; Sprando, Robert L

    2014-11-01

    The use of silver nanoparticles in food, food contact materials, dietary supplements and cosmetics has increased significantly owing to their antibacterial and antifungal properties. As a consequence, the need for validated rapid screening methods to assess their toxicity is necessary to ensure consumer safety. This study evaluated two widely used in vitro cell culture models, human liver HepG2 cells and human colon Caco2 cells, as tools for assessing the potential cytotoxicity of food- and cosmetic-related nanoparticles. The two cell culture models were utilized to compare the potential cytotoxicity of 20-nm silver. The average size of the silver nanoparticle determined by our transmission electron microscopy (TEM) analysis was 20.4 nm. The dynamic light scattering (DLS) analysis showed no large agglomeration of the silver nanoparticles. The concentration of the 20-nm silver solution determined by our inductively coupled plasma-mass spectrometry (ICP-MS) analysis was 0.962 mg ml(-1) . Our ICP-MS and TEM analysis demonstrated the uptake of 20-nm silver by both HepG2 and Caco2 cells. Cytotoxicity, determined by the Alamar Blue reduction assay, was evaluated in the nanosilver concentration range of 0.1 to 20 µg ml(-1) . Significant concentration-dependent cytotoxicity of the nanosilver in HepG2 cells was observed in the concentration range of 1 to 20 µg ml(-1) and at a higher concentration range of 10 to 20 µg ml(-1) in Caco2 cells compared with the vehicle control. A concentration-dependent decrease in dsDNA content was observed in both cell types exposed to nanosilver but not controls, suggesting an increase in DNA damage. The DNA damage was observed in the concentration range of 1 to 20 µg ml(-1) . Nanosilver-exposed HepG2 and Caco2 cells showed no cellular oxidative stress, determined by the dichlorofluorescein assay, compared with the vehicle control in the concentration range used in this study. A concentration-dependent decrease in

  6. Constitutive Effects of Lead on Aryl Hydrocarbon Receptor Gene Battery and Protection by β-carotene and Ascorbic Acid in Human HepG2 Cells.

    Science.gov (United States)

    Darwish, Wageh S; Ikenaka, Yoshinori; Nakayama, Shouta M M; Mizukawa, Hazuki; Ishizuka, Mayumi

    2016-01-01

    Lead (Pb) is an environmental pollutant that can get entry into human body through contaminated foods, drinks, and inhaled air leading to severe biological consequences, and has been responsible for many deaths worldwide. The objectives of this study were 1st to investigate the modulatory effects of environmentally relevant concentrations of Pb on AhR gene battery, which is controlling xenobiotics metabolism. 2nd, trials to reduce Pb-induced adverse effects were done using some phytochemicals like β-carotene or ascorbic acid. Human hepatoma (HepG2) cell lines were exposed to a wide range of Pb concentrations varying from physiological to toxic levels (0 to 10 mg/L) for 24 h. High Pb concentrations (1 to 10 mg/L) significantly reduced phase I (CYP1A1 and 1A2) and phase II (UGT1A6 and NQO1) xenobiotic metabolizing enzyme mRNA expression in a mechanistic manner through the AhR regulation pathway. Additionally, these Pb concentrations induced oxidative stress in HepG2 cells in terms of production of reactive oxygen species (ROS) and induced heme oxygenase-1 mRNA expression in a concentration-dependent phenomenon. Coexposure of HepG2 cells to physiological concentrations of some micronutrients, like β-carotene (10 μM) or ascorbic acid (0.1 mM), along with Pb (1 mg/L) for 24 h significantly reduced the levels of ROS production and recovered AhR mRNA expression into the normal levels. Thus, consumption of foods rich in these micronutrients may help to reduce the adverse effects of lead in areas with high levels of pollution. © 2015 Institute of Food Technologists®

  7. In vitro cytotoxicity and induction of apoptosis by silica nanoparticles in human HepG2 hepatoma cells

    Directory of Open Access Journals (Sweden)

    Lu X

    2011-09-01

    Full Text Available Xun Lu2,3, Jiangchao Qian1, Huanjun Zhou2,3, Qi Gan2,3, Wei Tang1, Jingxiong Lu3, Yuan Yuan1,2, Changsheng Liu1–31State Key Laboratory of Bioreactor Engineering, 2Key Laboratory for Ultrafine Materials of Ministry of Education, 3Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai, People’s Republic of ChinaBackground: Silica nanoparticles have been discovered to exert cytotoxicity and induce apoptosis in normal human cells. However, until now, few studies have investigated the cytotoxicity of silica nanoparticles in tumor cells.Methods: This study investigated the cytotoxicity of 7–50 nm silica nanoparticles in human HepG2 hepatoma cells, using normal human L-02 hepatocytes as a control. Cell nucleus morphology changes, cellular uptake, and expression of procaspase-9, p53, Bcl-2, and Bax, as well as the activity of caspase-3, and intracellular reactive oxygen species and glutathione levels in the silica nanoparticle-treated cells, were analyzed.Results: The antitumor activity of the silica nanoparticles was closely related to particle size, and the antiproliferation activity decreased in the order of 20 nm > 7 nm > 50 nm. The silica nanoparticles were also cytotoxic in a dose- and time-dependent manner. However, the silica nanoparticles showed only slight toxicity in the L-02 control cells, Moreover, in HepG2 cells, oxidative stress and apoptosis were induced after exposure to 7–20 nm silica nanoparticles. Expression of p53 and caspase-3 increased, and expression of Bcl-2 and procaspase-9 decreased in a dose-dependent manner, whereas the expression of Bax was not significantly changed.Conclusion: A mitochondrial-dependent pathway triggered by oxidative stress mediated by reactive oxygen species may be involved in apoptosis induced by silica nanoparticles, and hence cytotoxicity in human HepG2 hepatic cancer cells.Keywords: silica nanoparticles

  8. [Production effect comparison of SEPP and GPx between HepG2 and Hela cells with different selenocompounds].

    Science.gov (United States)

    Wang, Qin; Gao, Lina; Han, Feng; Lu, Jiaxi; Liu, Yiqun; Sun, Licui; Huang, Zhenwu

    2016-03-01

    To compare the effect of several selenocompounds on the productions of SEPP and GPx in HepG2 and Hela cells. The cultured HepG2 and Hela cells were divided into the control, Na2SeO3, SeMet and MeSeCys groups. After adding the selected selenocompounds (with the respective concentration 0.01 and 0.1 μmol/L), the experimental groups were then incubated for 48 h and 72 h. Finally, the cell culture supernatants and homogenates were collected for the SEPP and GPx concentrations detection by a double-antibody sandwich enyme-linked immuno-sorbent-assay (ELISA). The SEPP and GPx concentrations in Hela cells treated with 0.1 μmol/L SeMet and MeSeCys were significantly higher than that in the control group (P cell treated with 0.1 μmol/L selenocompounds were significantly higher than that in Hela cells (P cells are more beneficial to the production of selenoproteins than Hela cells.

  9. Increased glucose metabolism and alpha-glucosidase inhibition in Cordyceps militaris water extract-treated HepG2 cells

    Science.gov (United States)

    Kim, Dae Jung; Kang, Yun Hwan; Kim, Kyoung Kon; Kim, Tae Woo; Park, Jae Bong

    2017-01-01

    BACKGROUND/OBJECTIVES Recent living condition improvements, changes in dietary habits, and reductions in physical activity are contributing to an increase in metabolic syndrome symptoms including diabetes and obesity. Through such societal developments, humankind is continuously exposed to metabolic diseases such as diabetes, and the number of the victims is increasing. This study investigated Cordyceps militaris water extract (CMW)-induced glucose uptake in HepG2 cells and the effect of CMW treatment on glucose metabolism. MATERIALS/METHODS Colorimetric assay kits were used to determine the glucokinase (GK) and pyruvate dehydrogenase (PDH) activities, glucose uptake, and glycogen content. Either RT-PCR or western blot analysis was performed for quantitation of glucose transporter 2 (GLUT2), hepatocyte nuclear factor 1 alpha (HNF-1α), phosphatidylinositol 3-kinase (PI3k), protein kinase B (Akt), phosphorylated AMP-activated protein kinase (pAMPK), phosphoenolpyruvate carboxykinase, GK, PDH, and glycogen synthase kinase 3 beta (GSK-3β) expression levels. The α-glucosidase inhibitory activities of acarbose and CMW were evaluated by absorbance measurement. RESULTS CMW induced glucose uptake in HepG2 cells by increasing GLUT2 through HNF-1α expression stimulation. Glucose in the cells increased the CMW-induced phosphorylation of AMPK. In turn, glycolysis was stimulated, and glyconeogenesis was inhibited. Furthermore, by studying the mechanism of action of PI3k, Akt, and GSK-3β, and measuring glycogen content, the study confirmed that the glucose was stored in the liver as glycogen. Finally, CMW resulted in a higher level of α-glucosidase inhibitory activity than that from acarbose. CONCLUSION CMW induced the uptake of glucose into HepG2 cells, as well, it induced metabolism of the absorbed glucose. It is concluded that CMW is a candidate or potential use in diabetes prevention and treatment. PMID:28584574

  10. Polyethyleneimine-coated quantum dots for miRNA delivery and its enhanced suppression in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Liang G

    2016-11-01

    Full Text Available Gaofeng Liang,1 Yang Li,1 Wenpo Feng,1 Xinshuai Wang,2 Aihua Jing,1 Jinghua Li,1 Kaiwang Ma1 1Department of Biomedical Engineering, School of Medical Technology & Engineering, 2Department of Oncology, The First Affiliated Hospital, Henan University of Science & Technology, Luoyang, People’s Republic of China Abstract: Quantum dots (QDs have been intensively investigated for bioimaging, drug delivery, and labeling probes because of their unique optical properties. In this study, CdSe/ZnS QDs-based nonviral vectors with the dual functions of delivering miR-26a plasmid and bioimaging were formulated by capping the surface of CdSe/ZnS QDs with polyethyleneimine (PEI. The PEI-coated QDs were capable of condensing miR-26a expression vector into nanocomplexes that can emit strong red luminescence when loaded with CdSe/ZnS QDs. Further results showed that PEI-modified nanoparticles (NPs could transfect miR-26a plasmid into HepG2 cells in vitro. Meanwhile, imaging of living cells could be achieved based on the CdSe/ZnS QDs. Further study suggested that miR-26a transfection up-regulated miR-26a expression, induced cycle arrest, and triggered proliferation inhibition in HepG2 cells. The results indicated that PEI-coated QD NPs possess the capability of bioimaging and gene delivery and could be a promising vehicle with the engineering of QD NPs for gene therapy in the future. Keywords: miR-26a, PEI/QDs, HepG2, gene delivery, bioimaging

  11. Effects of chlorpromazine with and without UV irradiation on gene expression of HepG2 cells

    International Nuclear Information System (INIS)

    Froetschl, Roland; Weickardt, Sandra; Staszewski, Solveig; Kaufmann, Gabriele; Kasper, Peter

    2005-01-01

    Damage to DNA can trigger a variety of stress-related signals that alter the expression of genes associated with numerous biological pathways. In this study, we have used a cDNA microarray representing 1089 genes related to DNA damage and repair, cell cycle, transcription, metabolism and other toxicologically important cell functions to identify genes regulated in response to DNA damage in HepG2 cells induced by UV-activated chlorpromazine (CPZ). CPZ itself is not genotoxic but, upon UV irradiation with a non-genotoxic dose in the UVA range, it produces reactive free radical intermediates with DNA damaging properties. Genotoxicity in HepG2 cells was assessed concomitantly to gene expression profiling using the Comet assay. Kinetic studies were performed at a non-cytotoxic but clearly photogenotoxic concentration of CPZ (1.25 μg/ml) to characterize gene expression profiles at four different time points (3, 7, 15, 23 h) post short-term treatment. The results obtained from repeated experiments display a time-dependent expression pattern of up-regulated and repressed genes with distinct peaks in the number of differentially expressed genes at the 7 and 23 h time points. Most of the genes with altered expression belonged to the functional categories of cell cycle regulation and cell proliferation. A comparison with published expression profiles established in response to other genotoxic compounds showed low levels of concordance, which is most likely caused by the fact that extremely different testing conditions were used

  12. Antioxidative and cytoprotective effects of andrographolide against CCl4-induced hepatotoxicity in HepG2 cells.

    Science.gov (United States)

    Krithika, R; Verma, R J; Shrivastav, P S

    2013-05-01

    This article describes antioxidative and cytoprotective property of andrographolide, a major active component of the plant Andrographis paniculata (A. paniculata). High yields (2.7%) of andrographolide was isolated from the aerial parts of this plant via silica column chromatography. The purity of the compound was determined by high-performance thin-layer chromatography (HPTLC) and reversed phase high-performance liquid chromatography (HPLC) analysis. The structure was elucidated using techniques such as UV-visible spectrophotometry, elemental analysis, Fourier transform infrared (FT-IR), (1)H nuclear magnetic resonance ((1)H NMR), (13)C nuclear magnetic resonance ((13)C NMR) and mass spectral analysis and the data obtained were comparable with reported results. It was observed that andrographolide exhibited significant antioxidative property (IC50 = 3.2 µg/ml) by its ability to scavenge a stable free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH) as compared to known antioxidants like ascorbic acid, butylated hydroxy toluene (BHT) and the plant extract. The cytoprotective role of andrographolide against carbon tetrachloride (CCl4) toxicity in human hepatoma HepG2 cell line was assessed using trypan blue exclusion test, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, by estimation of various leakage enzymes and by measuring the glutathione levels. The recovery obtained for andrographolide treatment in the presence of CCl4 was two-fold compared to A. paniculata extract for all other related biochemical parameters investigated. The results of the study indicate that andrographolide is a potent inhibitor of CCl4-mediated lipid peroxidation.

  13. Effect of human mesenchymal stem cells on the growth of HepG2 and Hela cells.

    Science.gov (United States)

    Long, Xiaohui; Matsumoto, Rena; Yang, Pengyuan; Uemura, Toshimasa

    2013-01-01

    Human mesenchymal stem cells (hMSCs) accumulate at carcinomas and have a great impact on cancer cell's behavior. Here we demonstrated that hMSCs could display both the promotional and inhibitive effects on growth of HepG2 and Hela cells by using the conditioned media, indirect co-culture, and cell-to-cell co-culture. Cell growth was increased following the addition of lower proportion of hMSCs while decreased by treatment of higher proportion of hMSCs. We also established a novel noninvasive label way by using internalizing quantum dots (i-QDs) for study of cell-cell contact in the co-culture, which was effective and sensitive for both tracking and distinguishing different cells population without the disturbance of cells. Furthermore, we investigated the role of hMSCs in regulation of cell growth and showed that mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) signaling pathways were involved in hMSC-mediated cell inhibition and proliferation. Our findings suggested that hMSCs regulated cancer cell function by providing a suitable environment, and the discovery from the study would provide some clues for development of effective strategy for hMSC-based cancer therapies.

  14. Evaluation of N-acetyl-cysteine against tetrachlorobenzoquinone-induced genotoxicity and oxidative stress in HepG2 cells.

    Science.gov (United States)

    Dong, Hui; Xu, Demei; Hu, Lihua; Li, Lingrui; Song, Erqun; Song, Yang

    2014-02-01

    Tetrachlorobenzoquinone (TCBQ) is an active metabolite of pentachlorophenol (PCP). Although the genotoxic effect of PCP has been comprehensively investigated, there is little known about TCBQ's genotoxic effects. In the current study, TCBQ was tested for its genotoxicity using HepG2 cells as experimental model. To select the exposure concentration of interest, cell viability was measured and three concentrations were used for further investigation. In single cell gel electrophoresis (SCGE) assay, concentration-dependent increase in tail length, tail DNA percentage and tail moment were detected following TCBQ exposure. Micronucleus (MN) assay indicated TCBQ gradually increased MN frequency and decreased nuclear division index (NDI). Enzyme-linked immunosorbent assay (ELISA) and western blotting analyses both showed TCBQ caused histone H2AX phosphorylation (γ-H2AX). Furthermore, the elevation of 8-hydroxydeoxyguanosine (8-OHdG) and reactive oxygen species (ROS) level indicated TCBQ-induced genotoxicity is associated with oxidative stress. On the other hand, N-acetyl-cysteine (NAC) administration significantly protected cells from the genotoxic effect of TCBQ. Overall, our data suggested TCBQ exerted genotoxic effect possibly via an oxidative damage mechanism in HepG2 cells and this toxicity is prevented by pretreatment with NAC. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Effervescent Granules Prepared Using Eucommia ulmoides Oliv. and Moso Bamboo Leaves: Hypoglycemic Activity in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Xiang-Zhou Li

    2016-01-01

    Full Text Available Eucommia ulmoides Oliv. (E. ulmoides Oliv. and moso bamboo (Phyllostachys pubescens leaves are used as folk medicines in central-western China to treat diabetes. To investigate the hypoglycemic activity of the effervescent granules prepared using E. ulmoides Oliv. and moso bamboo leaves (EBEG in HepG2 cells, EBEG were prepared with 5% of each of polysaccharides and chlorogenic acids from moso bamboo and E. ulmoides Oliv. leaves, respectively. HepG2 cells cultured in a high-glucose medium were classified into different groups. The results displayed EBEG-treated cells showed better glucose utilization than the negative controls; thus, the hypoglycemic effect of EBEG was much greater than that of granules prepared using either component alone, thereby indicating that this effect was due to a synergistic action of the components. Further, glucose consumption levels in the cells treated with EBEG (156.35% at 200 μg/mL and the positive controls (metformin, 162.29%; insulin, 161.52% were similar. Thus, EBEG exhibited good potential for use as a natural antidiabetic agent. The hypoglycemic effect of EBEG could be due to the synergistic action of polysaccharides from the moso bamboo leaves and chlorogenic acids from E. ulmoides Oliv. leaves via the inhibition of alpha-glucosidase and glucose-6-phosphate displacement enzyme.

  16. Sagunja-Tang Improves Lipid Related Disease in a Postmenopausal Rat Model and HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Hiroe Go

    2015-01-01

    Full Text Available The present study was conducted to investigate the effect of Sagunja-tang on the lipid related disease in a rat model of menopausal hyperlipidemia and lipid accumulation in methyl-β-cyclodextrin-induced HepG2 cells. In in vivo study using menopausal hyperlipidemia rats, Sagunja-tang reduced retroperitoneal and perirenal fat, serum lipids, atherogenic index, cardiac risk factor, media thickness, and nonalcoholic steatohepatitis score, when compared to menopausal hyperlipidemia control rats. In HepG2 cells, Sagunja-tang significantly decreased the lipid accumulation, total cholesterol levels, and low-density/very-low-density lipoprotein levels. Moreover, Sagunja-tang reversed the methyl-β-cyclodextrin-induced decrease in the protein levels of critical molecule involved in cholesterol synthesis, sterol regulatory element binding protein-2, and low-density lipoprotein receptor and inhibited protein levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase as well as activity. Phosphorylation level of AMP-activated protein kinase was stimulated by Sagunja-tang. These results suggest that Sagunja-tang has effect on inhibiting hepatic lipid accumulation through regulation of cholesterol synthesis and AMPK activity in vitro. These observations support the idea that Sagunja-tang is bioavailable both in vivo and in vitro and could be developed as a preventive and therapeutic agent of hyperlipidemia in postmenopausal females.

  17. Activation of human stearoyl-coenzyme A desaturase 1 contributes to the lipogenic effect of PXR in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Jun Zhang

    Full Text Available The pregnane X receptor (PXR was previously known as a xenobiotic receptor. Several recent studies suggested that PXR also played an important role in lipid homeostasis but the underlying mechanism remains to be clearly defined. In this study, we found that rifampicin, an agonist of human PXR, induced lipid accumulation in HepG2 cells. Lipid analysis showed the total cholesterol level increased. However, the free cholesterol and triglyceride levels were not changed. Treatment of HepG2 cells with rifampicin induced the expression of the free fatty acid transporter CD36 and ABCG1, as well as several lipogenic enzymes, including stearoyl-CoA desaturase-1 (SCD1, long chain free fatty acid elongase (FAE, and lecithin-cholesterol acyltransferase (LCAT, while the expression of acyl:cholesterol acetyltransferase(ACAT1 was not affected. Moreover, in PXR over-expressing HepG2 cells (HepG2-PXR, the SCD1 expression was significantly higher than in HepG2-Vector cells, even in the absence of rifampicin. Down-regulation of PXR by shRNA abolished the rifampicin-induced SCD1 gene expression in HepG2 cells. Promoter analysis showed that the human SCD1 gene promoter is activated by PXR and a novel DR-7 type PXR response element (PXRE response element was located at -338 bp of the SCD1 gene promoter. Taken together, these results indicated that PXR activation promoted lipid synthesis in HepG2 cells and SCD1 is a novel PXR target gene.

  18. Adaptation of HepG2 cells to a steady-state reduction in the content of protein phosphatase 6 (PP6) catalytic subunit

    Energy Technology Data Exchange (ETDEWEB)

    Boylan, Joan M. [Department of Pediatrics, Brown University and Rhode Island Hospital, Providence, RI (United States); Salomon, Arthur R. [Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, RI (United States); Department of Chemistry, Brown University, Providence, RI (United States); Tantravahi, Umadevi [Division of Genetics, Department of Pathology, Brown University and Women and Infants Hospital, Providence, RI (United States); Gruppuso, Philip A., E-mail: philip_gruppuso@brown.edu [Department of Pediatrics, Brown University and Rhode Island Hospital, Providence, RI (United States); Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, RI (United States)

    2015-07-15

    Protein phosphatase 6 (PP6) is a ubiquitous Ser/Thr phosphatase involved in an array of cellular processes. To assess the potential of PP6 as a therapeutic target in liver disorders, we attenuated expression of the PP6 catalytic subunit in HepG2 cells using lentiviral-transduced shRNA. Two PP6 knock-down (PP6KD) cell lines (90% reduction of PP6-C protein content) were studied in depth. Both proliferated at a rate similar to control cells. However, flow cytometry indicated G2/M cell cycle arrest that was accounted for by a shift of the cells from a diploid to tetraploid state. PP6KD cells did not show an increase in apoptosis, nor did they exhibit reduced viability in the presence of bleomycin or taxol. Gene expression analysis by microarray showed attenuated anti-inflammatory signaling. Genes associated with DNA replication were downregulated. Mass spectrometry-based phosphoproteomic analysis yielded 80 phosphopeptides representing 56 proteins that were significantly affected by a stable reduction in PP6-C. Proteins involved in DNA replication, DNA damage repair and pre-mRNA splicing were overrepresented among these. PP6KD cells showed intact mTOR signaling. Our studies demonstrated involvement of PP6 in a diverse set of biological pathways and an adaptive response that may limit the effectiveness of targeting PP6 in liver disorders. - Highlights: • Lentiviral-transduced shRNA was used to generate a stable knockdown of PP6 in HepG2 cells. • Cells adapted to reduced PP6; cell proliferation was unaffected, and cell survival was normal. • However, PP6 knockdown was associated with a transition to a tetraploid state. • Genomic profiling showed downregulated anti-inflammatory signaling and DNA replication. • Phosphoproteomic profiling showed changes in proteins associated with DNA replication and repair.

  19. [Adenovirus-mediated killing of hepatocellular carcinoma HepG2 cells by heterogeneous fusion gene NT4p53(N15)Ant].

    Science.gov (United States)

    Li, Yue-ping; Qiu, Shu-dong; Song, Li-ping; Wang, Quan-ying; Yang, Guang-xiao

    2007-07-01

    OBJECTIVE To construct a recombinant adenovirus Ad.NT4p53(N15)Ant and explore its cytotoxic effect against hepatocellular carcinoma HepG2 cells in vitro. The recombinant adenovirus containing the fusion gene of neurotrophin 4 (NT4)signal peptide, N-terminal residues (12-26) of p53 and 17 amino acid Drosophila homeobox protein Antennapedia (Ant) was constructed by gene cloning protocol. The effect of this fusion gene on HepG2 cells was evaluated by MTT assay, PI staining and flow cytometry. The fusion gene Ad.NT4p53(N15)Ant was successfully constructed, as verified by restriction endonuclease digestion and PCR. Ad.NT4p53(N15)Ant could strongly suppress the growth of HepG2 cells (with a growth inhibition rate of 63.3% 48 h after infection) without affecting NIH-3T3 cells. Flow cytometry showed that Ad.NT4p53(N15)Ant could induce obvious apoptosis of HepG2 cells. The recombinant adenovirus containing NT4p53(N15)Ant fusion gene can inhibit the growth the of HepG2 cells in vitro partially by inducing cell apoptosis.

  20. STARD4 knockdown in HepG2 cells disrupts cholesterol trafficking associated with the plasma membrane, ER, and ERC

    DEFF Research Database (Denmark)

    Garbarino, J.; Pan, M. H.; Chin, H. F.

    2012-01-01

    STARD4, a member of the evolutionarily conserved START gene family, has been implicated in the non-vesicular intracellular transport of cholesterol. However, the direction of transport and the membranes with which this protein interacts are not clear. We present studies of STARD4 function using...... small hairpin RNA knockdown technology to reduce STARD4 expression in HepG2 cells. In a cholesterol-poor environment, we found that a reduction in STARD4 expression leads to retention of cholesterol at the plasma membrane, reduction of endoplasmic reticulum-associated cholesterol, and decreased ACAT...... synthesized cholesteryl esters. Furthermore, D4 KD cells exhibited a reduced rate of sterol transport to the endocytic recycling compartment after cholesterol repletion. Although these cells displayed normal endocytic trafficking in cholesterol-poor and replete conditions, cell surface low density lipoprotein...

  1. Comparison of gene expression profiles of HepG2 cells exposed to Crambescins C1 and A1

    Directory of Open Access Journals (Sweden)

    María R. Sánchez

    2014-06-01

    Full Text Available Crambescins are guanidine alkaloids firstly isolated in the early 90s from the encrusting Mediterranean sponge Crambe crambe (Schmidt, 1862 (Bondu et al., 2012, Laville et al., 2009, Berlinck et al., 1990. C. crambe derivatives are divided in two families named crambescins and crambescidins (Gerlinck et al., 1992. Although data on the bioactivity of these compounds is scarce, crambescidins have recognized cytotoxic, antifungal, antioxidant, antimicrobial and antiviral activities (Buscema and Van de Vyver, 1985, Jares-Erijman., 1998, Olszewski et al., 2004, Lazaro et al., 2006, Suna et al., 2007, AOKI et al., 2004. Recently we have carefully evaluated the cytotoxic activity of C816 over several human tumor cell types and characterized some of the cellular mechanisms responsible of the anti-proliferative effect of this compound on human liver-derived tumor cells (Rubiolo et al., 2013. Taking this into account, and to better understand the mechanism of action of crambescins and their potential as therapeutic agents, we made a comparative gene expression profiling of HepG2 cells after crambescin C1 (C1 and crambescin A1 (CA1 exposures. Results have shown that C1 induces genes involved in sterol and glucose metabolisms and metabolism involving growth factors. It also down regulates genes mainly involved in cell cycle control, DNA replication, recombination and repair, and drug metabolism. Flow cytometry assays revealed that C1 produces a G0/G1 arrest in HepG2 cell cycle progression. CA1 also down-regulates genes involved in cell cycle regulation, DNA recombination and pathways related to tumor cells proliferation with lower potency when compared to C1.

  2. Mercury-Induced Externalization of Phosphatidylserine and Caspase 3 Activation in Human Liver Carcinoma (HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Paul B. Tchounwou

    2006-03-01

    Full Text Available Apoptosis arises from the active initiation and propagation of a series of highly orchestrated specific biochemical events leading to the demise of the cell. It is a normal physiological process, which occurs during embryonic development as well as in the maintenance of tissue homeostasis. Diverse groups of molecules are involved in the apoptosis pathway and it functions as a mechanism to eliminate unwanted or irreparably damaged cells. However, inappropriate induction of apoptosis by environmental agents has broad ranging pathologic implications and has been associated with several diseases including cancer. The toxicity of several heavy metals such as mercury has been attributed to their high affinity to sulfhydryl groups of proteins and enzymes, and their ability to disrupt cell cycle progression and/or apoptosis in various tissues. The aim of this study was to assess the potential for mercury to induce early and late-stage apoptosis in human liver carcinoma (HepG2 cells. The Annexin-V and Caspase 3 assays were performed by flow cytometric analysis to determine the extent of phosphatidylserine externalization and Caspase 3 activation in mercury-treated HepG2 cells. Cells were exposed to mercury for 10 and 48 hours respectively at doses of 0, 1, 2, and 3 μg/mL based on previous cytotoxicity results in our laboratory indicating an LD50 of 3.5 ± 0.6 μg/mL for mercury in HepG2 cells. The study data indicated a dose response relationship between mercury exposure and the degree of early and late-stage apoptosis in HepG2 cells. The percentages of cells undergoing early apoptosis were 0.03 ± 0.03%, 5.19 ± 0.04%, 6.36 ± 0.04%, and 8.84 ± 0.02% for 0, 1, 2, and 3 μg/mL of mercury respectively, indicating a gradual increase in apoptotic cells with increasing doses of mercury. The percentages of Caspase 3 positive cells undergoing late apoptosis were 3.58 ± 0.03%, 17.06 ± 0

  3. CdTe quantum dots with daunorubicin induce apoptosis of multidrug-resistant human hepatoma HepG2/ADM cells: in vitro and in vivo evaluation

    Directory of Open Access Journals (Sweden)

    Shi Lixin

    2011-01-01

    Full Text Available Abstract Cadmium telluride quantum dots (Cdte QDs have received significant attention in biomedical research because of their potential in disease diagnosis and drug delivery. In this study, we have investigated the interaction mechanism and synergistic effect of 3-mercaptopropionic acid-capped Cdte QDs with the anti-cancer drug daunorubicin (DNR on the induction of apoptosis using drug-resistant human hepatoma HepG2/ADM cells. Electrochemical assay revealed that Cdte QDs readily facilitated the uptake of the DNR into HepG2/ADM cells. Apoptotic staining, DNA fragmentation, and flow cytometry analysis further demonstrated that compared with Cdte QDs or DNR treatment alone, the apoptosis rate increased after the treatment of Cdte QDs together with DNR in HepG2/ADM cells. We observed that Cdte QDs treatment could reduce the effect of P-glycoprotein while the treatment of Cdte QDs together with DNR can clearly activate apoptosis-related caspases protein expression in HepG2/ADM cells. Moreover, our in vivo study indicated that the treatment of Cdte QDs together with DNR effectively inhibited the human hepatoma HepG2/ADM nude mice tumor growth. The increased cell apoptosis rate was closely correlated with the enhanced inhibition of tumor growth in the studied animals. Thus, Cdte QDs combined with DNR may serve as a possible alternative for targeted therapeutic approaches for some cancer treatments.

  4. The role of the vascular endothelial growth factor/vascular endothelial growth factor receptors axis mediated angiogenesis in curcumin-loaded nanostructured lipid carriers induced human HepG2 cells apoptosis

    Directory of Open Access Journals (Sweden)

    Fengling Wang

    2015-01-01

    Full Text Available Background: Curcumin (diferuloylmethane, the active constituent of turmeric extract has potent anti-cancer properties have been demonstrated in hepatocellular carcinoma (HCC. However, its underlying molecular mechanism of therapeutic effects remains unclear. Vascular endothelial growth factor (VEGF and its receptors (VEGFRs have crucial roles in tumor angiogenesis. Purpose: The goal of this study was to investigate the role of the VEGF/VEGFRs mediated angiogenesis during the proliferation and apoptosis of human HepG2 hepatoma cell line and the effect of curcumin-loaded nanostructured lipid carriers (Cur-NLC. Materials and Methods: The proliferation of HepG2 cells was determined by methyl thiazolyl tetrazolium after exposure to Cur-NLC and native curcumin. Apoptosis was quantified by flow cytometry with annexin V-fluorescein isothiocyanate and propidium iodide staining. Cellular internalization of Cur-NLC was observed by fluorescent microscope. The level of VEGF was detected by enzyme-linked immunosorbent assay kits. The expression of VEGFRs was quantified by Western blotting. Results: Cur-NLC was more effective in inhibiting the proliferation and enhancing the apoptosis of HepG2 cells than native curcumin. Fluorescent microscope analysis showed that HepG2 cells internalized Cur-NLC more effectively than native curcumin. Furthermore, Cur-NLC down-regulated the level of VEGF and the expression of VEGFR-2, but had a slight effect on VEGFR-1. Conclusion: These results clearly demonstrated that Cur-NLC was more effective in anti-cancer activity than the free form of curcumin. These studies demonstrate for the 1 st time that Cur-NLC exerts an antitumor effect on HepG2 cells by modulating VEGF/VEGFRs signaling pathway.

  5. PLGA-based gene delivering nanoparticle enhance suppression effect of miRNA in HePG2 cells

    Directory of Open Access Journals (Sweden)

    Liang Gao

    2011-01-01

    Full Text Available Abstract The biggest challenge in the field of gene therapy is how to effectively deliver target genes to special cells. This study aimed to develop a new type of poly(D,L-lactide-co-glycolide (PLGA-based nanoparticles for gene delivery, which are capable of overcoming the disadvantages of polyethylenimine (PEI- or cationic liposome-based gene carrier, such as the cytotoxicity induced by excess positive charge, as well as the aggregation on the cell surface. The PLGA-based nanoparticles presented in this study were synthesized by emulsion evaporation method and characterized by transmission electron microscopy, dynamic light scattering, and energy dispersive spectroscopy. The size of PLGA/PEI nanoparticles in phosphate-buffered saline (PBS was about 60 nm at the optimal charge ratio. Without observable aggregation, the nanoparticles showed a better monodispersity. The PLGA-based nanoparticles were used as vector carrier for miRNA transfection in HepG2 cells. It exhibited a higher transfection efficiency and lower cytotoxicity in HepG2 cells compared to the PEI/DNA complex. The N/P ratio (ratio of the polymer nitrogen to the DNA phosphate 6 of the PLGA/PEI/DNA nanocomplex displays the best property among various N/P proportions, yielding similar transfection efficiency when compared to Lipofectamine/DNA lipoplexes. Moreover, nanocomplex shows better serum compatibility than commercial liposome. PLGA nanocomplexes obviously accumulate in tumor cells after transfection, which indicate that the complexes contribute to cellular uptake of pDNA and pronouncedly enhance the treatment effect of miR-26a by inducing cell cycle arrest. Therefore, these results demonstrate that PLGA/PEI nanoparticles are promising non-viral vectors for gene delivery.

  6. OSBP-related protein 8 (ORP8) interacts with Homo sapiens sperm associated antigen 5 (SPAG5) and mediates oxysterol interference of HepG2 cell cycle

    Energy Technology Data Exchange (ETDEWEB)

    Zhong, Wenbin [Department of Biotechnology, Jinan University, Guangzhou 510632 (China); Zhou, You [Minerva Foundation Institute for Medical Research, Helsinki (Finland); Li, Jiwei [Department of Biotechnology, Jinan University, Guangzhou 510632 (China); Mysore, Raghavendra [Minerva Foundation Institute for Medical Research, Helsinki (Finland); Luo, Wei; Li, Shiqian [Department of Biotechnology, Jinan University, Guangzhou 510632 (China); Chang, Mau-Sun [Institute of Biochemical Sciences, National Taiwan University, No. 1, Taipei, Taiwan (China); Olkkonen, Vesa M. [Minerva Foundation Institute for Medical Research, Helsinki (Finland); Yan, Daoguang, E-mail: tydg@jnu.edu.cn [Department of Biotechnology, Jinan University, Guangzhou 510632 (China)

    2014-04-01

    We earlier identified OSBP-related protein 8 (ORP8) as an endoplasmic reticulum/nuclear envelope oxysterol-binding protein implicated in cellular lipid homeostasis, migration, and organization of the microtubule cytoskeleton. Here, a yeast two-hybrid screen identified Homo sapiens sperm associated antigen 5 (SPAG5)/Astrin as interaction partner of ORP8. The putative interaction was further confirmed by pull-down and co-immunoprecipitation assays. ORP8 did not colocalize with kinetochore-associated SPAG5 in mitotic HepG2 or HuH7 cells, but overexpressed ORP8 was capable of recruiting SPAG5 onto endoplasmic reticulum membranes in interphase cells. In our experiments, 25-hydroxycholesterol (25OHC) retarded the HepG2 cell cycle, causing accumulation in G2/M phase; ORP8 overexpression resulted in the same phenotype. Importantly, ORP8 knock-down dramatically inhibited the oxysterol effect on HepG2 cell cycle, suggesting a mediating role of ORP8. Furthermore, knock-down of SPAG5 significantly reduced the effects of both ORP8 overexpression and 25OHC on the cell cycle, placing SPAG5 downstream of the two cell-cycle interfering factors. Taken together, the present results suggest that ORP8 may via SPAG5 mediate oxysterol interference of the HepG2 cell cycle. - Highlights: • The oxysterol-binding protein ORP8 was found to interact with the mitotic regulator SPAG5/Astrin. • Treatment of HepG2 cells with 25-hydroxycholesterol caused cell cycle retardation in G2/M. • ORP8 overexpression caused a similar G2/M accumulation, and ORP8 knock-down reversed the 25-hydroxycholesterol effect. • Reduction of cellular of SPAG5/Astrin reversed the cell cycle effects of both 25-hydroxycholesterol and ORP8 overexpression. • Our results suggest that ORP8 mediates via SPAG5/Astrin the oxysterol interference of HepG2 cell cycle.

  7. OSBP-related protein 8 (ORP8) interacts with Homo sapiens sperm associated antigen 5 (SPAG5) and mediates oxysterol interference of HepG2 cell cycle

    International Nuclear Information System (INIS)

    Zhong, Wenbin; Zhou, You; Li, Jiwei; Mysore, Raghavendra; Luo, Wei; Li, Shiqian; Chang, Mau-Sun; Olkkonen, Vesa M.; Yan, Daoguang

    2014-01-01

    We earlier identified OSBP-related protein 8 (ORP8) as an endoplasmic reticulum/nuclear envelope oxysterol-binding protein implicated in cellular lipid homeostasis, migration, and organization of the microtubule cytoskeleton. Here, a yeast two-hybrid screen identified Homo sapiens sperm associated antigen 5 (SPAG5)/Astrin as interaction partner of ORP8. The putative interaction was further confirmed by pull-down and co-immunoprecipitation assays. ORP8 did not colocalize with kinetochore-associated SPAG5 in mitotic HepG2 or HuH7 cells, but overexpressed ORP8 was capable of recruiting SPAG5 onto endoplasmic reticulum membranes in interphase cells. In our experiments, 25-hydroxycholesterol (25OHC) retarded the HepG2 cell cycle, causing accumulation in G2/M phase; ORP8 overexpression resulted in the same phenotype. Importantly, ORP8 knock-down dramatically inhibited the oxysterol effect on HepG2 cell cycle, suggesting a mediating role of ORP8. Furthermore, knock-down of SPAG5 significantly reduced the effects of both ORP8 overexpression and 25OHC on the cell cycle, placing SPAG5 downstream of the two cell-cycle interfering factors. Taken together, the present results suggest that ORP8 may via SPAG5 mediate oxysterol interference of the HepG2 cell cycle. - Highlights: • The oxysterol-binding protein ORP8 was found to interact with the mitotic regulator SPAG5/Astrin. • Treatment of HepG2 cells with 25-hydroxycholesterol caused cell cycle retardation in G2/M. • ORP8 overexpression caused a similar G2/M accumulation, and ORP8 knock-down reversed the 25-hydroxycholesterol effect. • Reduction of cellular of SPAG5/Astrin reversed the cell cycle effects of both 25-hydroxycholesterol and ORP8 overexpression. • Our results suggest that ORP8 mediates via SPAG5/Astrin the oxysterol interference of HepG2 cell cycle

  8. [Damage effect of Polygonum multiflorum fractions on human normal liver cells L02 and liver cancer cells HepG2].

    Science.gov (United States)

    Zhang, Ruichen; Zhang, Chao; Sun, Zhenxiao; Deng, Qiaohong

    2012-06-01

    To investigate the damage effect of different fractions from Polygonum multiflorum on normal human liver and liver cancer cells, in order to seek for fractions that can obviously kill cancer cells but have less impact on normal liver cells, and make a preliminary study on different mechanism of the two kinds of cells. P. multiflorum water-eluted fraction (RW), 50% ethanol-eluted fraction (R50) and 95% ethanol-eluted fraction (R95) were successively obtained from 70% ethanol extracts of P. multiflorum, after being eluted by water, 50% ethanol and 95% ethanol and then absorbed by AB-8 macroporous resin. Normal human liver L02 cells and liver cancer HepG2 cells were incubated with cell supernatants from different fractions and cells. MTT method and inverted microscope were adopted to observe the impact of L02 on growth of HepG2 cells, screening fractions with damage effect and detect their doses and time effect. Giemsa stain showed changes in cell nucleus after administration and flow cytometry analysis was used to detect cycle and apoptosis of L02 cells. MTT method and inverted microscope showed that R50 had significant growth inhibition effects on L02 and HepG2 cells. According to giemsa stain and flow cytometry analysis, R50 showed different effect on inducing the two cells: there are much more apoptotic HepG2 cells than apoptotic L02 cells in each time phase (the proportion of the apoptosis cells in HepG2 group were 83.62%, 60.52% and 74.49%, and ID2 31.02%, 20.57% and 25.32% after treated with R50 for 24, 48, 72 h. Both cells showed less than 5% of apoptotic cells in the negative control group in each time phase). However, there is no significant impact on cycle of both cells. R50 from P. multiflorum extracts had different damage effects on human liver L02 cells and liver cancer HepG2 cells, which was caused by different degree of induction on apoptosis of the two cells in nature.

  9. Protective Effect of Curcumin against Ionizing Radiation (IR)-induced Cytotoxicity and Genotoxicity in HepG2 Cells

    International Nuclear Information System (INIS)

    Chung, Dong Min; Nasir Uddin, S. M.; Ryu, Tae Ho; Kang, Mi Young; Kim, Jin Kyu

    2013-01-01

    Ionizing radiation (IR) has many practical applications such as medicine, foods, agricultures, industries, and research laboratories. However, the increasing use of radiation is associated with radiation accidents threatening human health. It is well known that exposure to IR gives rise to genomic alterations, mutagenesis, and cell death. IR is absorbed directly by DNA, leading to various DNA damages (single or double-strand breaks, base damage, and DNA-DNA or DNA-protein cross-linkages) in many living organisms. Therefore, the development of effective and nontoxic radioprotective agents is of considerable interest. Curcumin (C 12 H 20 O 6 , structure is the major yellow component of Curcuma longa with biological activities (antioxidant, anti-proliferative and anti-inflammatory properties). It has been widely used as food and medicine for a long time. The aim of our present study is to investigate the protective effects of curcumin against IR-induced cytotoxicity and genotoxicity in cultured HepG2 cells

  10. Structure of Sphingolipids From Sea Cucumber Cucumaria frondosa and Structure-Specific Cytotoxicity Against Human HepG2 Cells.

    Science.gov (United States)

    Jia, Zicai; Song, Yu; Tao, Suyuan; Cong, Peixu; Wang, Xiaoxu; Xue, Changhu; Xu, Jie

    2016-03-01

    To investigate the relationship between structure and activity, three glucocerebroside series (CFC-1, CFC-2 and CFC-3), ceramides (CF-Cer) and long-chain bases (CF-LCB) of sea cucumber Cucumaria frondosa (C. frondosa) were isolated and evaluated in HepG2 cells. The molecular species of CFC-1, CFC-2 and CFC-3 and CF-Cer were identified using reversed-phase liquid chromatography with heated electrospray ionization coupled to high-resolution mass spectrometry (RPLC-HESI-HRMS), and determined on the basis of chemical and spectroscopic evidence: For the three glucocerebroside series, fatty acids (FA) were mainly saturated (18:0 and 22:0), monounsaturated (22:1, 23:1 and 24:1) and 2-hydroxyl FA (2-HFA) (23:1 h and 24:1 h), the structure of long-chain bases (LCB) were dihydroxy (d17:1, d18:1 and d18:2) and trihydroxy (t16:0 and t17:0), and the glycosylation was glucose; For CF-Cer, FA were primarily saturated (17:0) and monounsaturated (16:1 and 19:1), the structure of LCB were dihydroxy (d17:1 and d18:1), and trihydroxy (t16:0). The results of cell experiment indicated that all of three glucocerebroside series, CF-Cer and CF-LCB exhibited an inhibitory effects on cell proliferation. Moreover, CFC-3 was most effective in three glucocerebrosides to HepG-2 cell viability. The inhibition effect of CF-LCB was the strongest, and the inhibition effect of CF-Cer was much stronger than glucocerebrosides.

  11. Endopolysaccharides from Ganoderma resinaceum, Phlebia rufa, and Trametes versicolor affect differently the proliferation rate of HepG2 cells.

    Science.gov (United States)

    Silva, Amélia M; Miranda, Andreia; Fernandes, Elisabete; Santos, Susana; Fraga, Irene; Santos, Dario L; Dias, Albino A; Bezerra, Rui M

    2013-03-01

    Fungi have been used for medicinal purposes for long time by Asian countries, being a putative source of powerful new phytopharmaceuticals such as polysaccharides. The aim of this study was to extract endopolysaccharides (IPS) from Ganoderma resinaceum, Phlebia rufa, and Trametes versicolor, grown under submerged culture, to compare crude IPS production, total carbohydrate, and protein yield, and to study the effect of these IPS on HepG2 cells proliferation rate. Total biomass produced by G. resinaceum, P. rufa, and T. versicolor was (in gram per liter) 3.32 ± 0.80, 5.42 ± 0.58, and 4.2 ± 1.29 and the IPS yield (as the biomass percent) was 9.9 ± 0.05, 29.0 ± 6.3, and 9.1 ± 3.1 %, respectively. Characterization of IPS has shown different proportion between total sugar and protein being, on average 6.04, 10.74, and 22.62, for G. resinaceum, T. versicolor, and P. rufa, respectively. The IPS effect, at 50, 100, and 200 μg mL(-1) on HepG2 cell growth and viability was negligible for G. resinaceum and P. rufa but, in the case of T. versicolor, 200 μg mL(-1) of IPS evoked 40 % reduction on cell growth. The results suggest that the intracellular polysaccharides from T. versicolor are a potential source for bioactive molecules with anti-proliferative properties.

  12. Glucocorticoids induce mitochondrial gene transcription in HepG2 cells: role of the mitochondrial glucocorticoid receptor.

    Science.gov (United States)

    Psarra, Anna-Maria G; Sekeris, Constantine E

    2011-10-01

    Glucocorticoids are major regulators of a plethora of cellular functions, acting on target cells through glucocorticoid receptors (GR) and modulation of gene transcription, among other mechanisms. One main site of action of glucocorticoids is the hepatocyte, which responds to the hormonal stimulus with induction of several proteins among them enzymes of oxidative phosphorylation (OXPHOS), both nuclearly and mitochondrially encoded. The induction of OXPHOS is regarded as a result of a nuclear action of the receptor on the respective nuclear genes and on genes encoding mitochondrial transcription factors. The presence of GR in mitochondria and of sequences in the mitochondrial genome similar to glucocorticoid responsive elements, suggested a direct action of GR on mitochondrial transcription. We demonstrate in HepG2 hepatocarcinoma cells specific binding of GR to the regulatory D-loop region of the mitochondrial genome and show that dexamethasone induces the mitochondrial transcription factors A, B1, and B2, the mitochondrial ribosomal RNA, and several mitochondrially encoded OXPHOS genes. Applying α-amanitin, the specific inhibitor of DNA-dependent RNA polymerase II, the dexamethasone-induced transcription of the mitochondrial genes can still proceeds, whereas the DEX effect on transcription of the mitochondrial transcription factors is suppressed. Moreover, HepG2 cells overexpressing mitochondrial targeted GR showed increased RNA synthesis, cytrochrome oxidase subunit I protein expression, and mitochondrial ATP production. We conclude that glucocorticoids can stimulate directly mitochondrial transcription by the mitochondrially localized GR, affecting OXPHOS enzyme biosynthesis. This takes place in addition to their action on mitochondrial genes by way of induction of the nuclearly encoded mitochondrial transcription factors. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Regulation of HepG2 cell apolipoprotein B metabolism by the citrus flavanones hesperetin and naringenin.

    Science.gov (United States)

    Borradaile, N M; Carroll, K K; Kurowska, E M

    1999-06-01

    Our previous studies showed that replacing the drinking water of rabbits fed a casein-containing diet with either orange juice or grapefruit juice reduced serum low density lipoprotein cholesterol and hepatic cholesteryl ester concentrations. To determine whether the changes observed in rabbits were due to flavonoids present in the juices acting directly on the liver, the effects of hesperetin and naringenin on net apolipoprotein B (apoB) secretion by HepG2 cells were investigated. These flavanones dose-dependently reduced net apoB secretion by up to 81% after a 24 h incubation, while doses of 60 micrograms/mL reduced net apoB secretion by 50% after 4 h. Coincubation with the proteasome inhibitor, MG-132, did not alter the ability of the flavonoids to reduce net apoB secretion over 4 h, suggesting that the flavonoid-induced changes in apoB metabolism were not due to a direct increase in proteasomal activity. However, the flavonoids were unable to reduce net apoB secretion after 4 h in the presence of oleate, suggesting that these compounds may interfere with the availability of neutral lipids for lipoprotein assembly. Furthermore, our 14C-acetate-labeling studies showed a 50% reduction in cholesteryl ester synthesis in the presence of either flavonoid, which could account for the reduction in net apoB secretion caused by incubation with these compounds. These in vitro studies suggest that hesperetin and naringenin may, in part, reduce net apoB secretion by HepG2 cells by inhibiting cholesteryl ester synthesis and that these compounds are good candidates for further in vivo studies to determine whether they are responsible for the cholesterol-lowering properties of dietary citrus juices.

  14. Multifunctional selenium nanoparticles as carriers of HSP70 siRNA to induce apoptosis of HepG2 cells

    Directory of Open Access Journals (Sweden)

    Li Y

    2016-07-01

    Full Text Available Yinghua Li,1 Zhengfang Lin,1 Mingqi Zhao,1 Tiantian Xu,1 Changbing Wang,1 Huimin Xia,1,* Hanzhong Wang,2,* Bing Zhu1,* 1Guangzhou Women and Children’s Medical Center, Guangzhou, Guangdong, 2State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, People’s Republic of China *These authors contributed equally to this work Abstract: Small interfering RNA (siRNA as a new therapeutic modality holds promise for cancer treatment, but it is unable to cross cell membrane. To overcome this limitation, nanotechnology has been proposed for mediation of siRNA transfection. Selenium (Se is a vital dietary trace element for mammalian life and plays an essential role in the growth and functioning of humans. As a novel Se species, Se nanoparticles have attracted more and more attention for their higher anticancer efficacy. In the present study, siRNAs with polyethylenimine (PEI-modified Se nanoparticles (Se@PEI@siRNA have been demonstrated to enhance the apoptosis of HepG2 cells. Heat shock protein (HSP-70 is overexpressed in many types of human cancer and plays a significant role in several biological processes including the regulation of apoptosis. The objective of this study was to silence inducible HSP70 and promote the apoptosis of Se-induced HepG2 cells. Se@PEI@siRNA were successfully prepared and characterized by various microscopic methods. Se@PEI@siRNA showed satisfactory size distribution, high stability, and selectivity between cancer and normal cells. The cytotoxicity of Se@PEI@siRNA was lower for normal cells than tumor cells, indicating that these compounds may have fewer side effects. The gene-silencing efficiency of Se@PEI@siRNA was significantly much higher than Lipofectamine 2000@siRNA and resulted in a significantly reduced HSP70 mRNA and protein expression in cancer cells. When the expression of HSP70 was diminished, the function of cell protection was also removed and cancer cells became more

  15. Bioluminescence-based cytotoxicity assay for simultaneous evaluation of cell viability and membrane damage in human hepatoma HepG2 cells.

    Science.gov (United States)

    Uno, Katsuhiro; Murotomi, Kazutoshi; Kazuki, Yasuhiro; Oshimura, Mitsuo; Nakajima, Yoshihiro

    2018-05-01

    We have developed a bioluminescence-based non-destructive cytotoxicity assay in which cell viability and membrane damage are simultaneously evaluated using Emerald luciferase (ELuc) and endoplasmic reticulum (ER)-targeted copepod luciferase (GLuc-KDEL), respectively, by using multi-integrase mouse artificial chromosome (MI-MAC) vector. We have demonstrated that the time-dependent concentration response curves of ELuc luminescence intensity and WST-1 assay, and GLuc-KDEL luminescence intensity and lactate dehydrogenase (LDH) activity in the culture medium accompanied by cytotoxicity show good agreement in toxicant-treated ELuc- and GLuc-KDEL-expressing HepG2 stable cell lines. We have clarified that the increase of GLuc-KDEL luminescence intensity in the culture medium reflects the type of cell death, including necrosis and late apoptosis, but not early apoptosis. We have also uncovered a strong correlation between GLuc-KDEL luminescence intensity in the culture medium and the extracellular release of high mobility group box 1 (HMGB1), a representative damage-associated molecular pattern (DAMP) molecule. The bioluminescence measurement assay using ELuc and GLuc-KDEL developed in this study can simultaneously monitor cell viability and membrane damage, respectively, and the increase of GLuc-KDEL luminescence intensity in the culture medium accompanied by the increase of cytotoxicity is an index of necrosis and late apoptosis associated with the extracellular release of DAMP molecules. Copyright © 2018 John Wiley & Sons, Ltd.

  16. DIFFERENTIAL-EFFECTS OF EICOSAPENTAENOIC ACID ON GLYCEROLIPID AND APOLIPOPROTEIN-B METABOLISM IN PRIMARY HUMAN HEPATOCYTES COMPARED TO HEPG2 CELLS AND PRIMARY RAT HEPATOCYTES

    NARCIS (Netherlands)

    LIN, YG; SMIT, MJ; HAVINGA, R; VERKADE, HJ; VONK, RJ; KUIPERS, F

    1995-01-01

    We compared the effects of eicosapentaenoic acid (EPA) and oleic acid (OA) on glycerolipid and apolipoprotein B (apoB) metabolism in primary human hepatocytes, HepG2 cells and primary rat hepatocytes. Cells were incubated for 1 to 5 h with 0.25 mM bovine serum albumin in the absence (control) or

  17. Learning Boolean Networks in HepG2 cells using ToxCast High-Content Imaging Data (SOT annual meeting)

    Science.gov (United States)

    Cells adapt to their environment via homeostatic processes that are regulated by complex molecular networks. Our objective was to learn key elements of these networks in HepG2 cells using ToxCast High-content imaging (HCI) measurements taken over three time points (1, 24, and 72h...

  18. Influence of different chemical agents (H2O2, t-BHP and MMS) on the activity of antioxidant enzymes in human HepG2 and hamster V79 cells; relationship to cytotoxicity and genotoxicity.

    Science.gov (United States)

    Slamenova, D; Kozics, K; Melusova, M; Horvathova, E

    2015-01-01

    We investigated activities of antioxidant enzymes (AEs), superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) in human HepG2 and hamster V79 cells treated with a scale of concentrations of hydrogen peroxide (H2O2), tert-butyl hydroperoxide (t-BHP) and methyl methanesulfonate (MMS). Cytotoxicity and genotoxicity of these substances were evaluated simultaneously. We have found out that H2O2, t-BHP and MMS predictably induce significant concentration-dependent increase of DNA lesions in both cell lines. Cytotoxicity detected in V79 cells with help of PE test was in a good conformity with the level of DNA damage. MTT test has proved unsuitable, except for MMS-treated V79 cells. Compared with human cells HepG2, hamster cells V79 manifested approximately similar levels of SOD and CAT but ten times higher activity of GPx. Across all concentrations tested the most significant increase of activity of the enzyme CAT was found in H2O2- and t-BHP-treated HepG2 cells, of the enzyme SOD in t-BHP- and MMS-treated V79 cells, and of the enzyme GPx in H2O2-treated V79 cells. We suggest that stimulation of enzyme activity by the relevant chemical compounds may result from transcriptional or post-transcriptional regulation of the expression of the genes CAT, SOD and GPx. Several authors suggest that moderate levels of toxic reactants can induce increase of AEs activities, while very high levels of reactants can induce their decrease, as a consequence of damage of the molecular machinery required to induce AEs. Based on a great amount of experiments, which were done and described within this paper, we can say that the above mentioned principle does not apply in general. Only the reactions of t-BHP affected HepG2 cells were consistent with this idea.

  19. Cordyceps militaris induces tumor cell death via the caspase-dependent mitochondrial pathway in HepG2 and MCF-7 cells

    Science.gov (United States)

    SONG, JINGJING; WANG, YINGWU; TENG, MEIYU; ZHANG, SHIQIANG; YIN, MENGYA; LU, JIAHUI; LIU, YAN; LEE, ROBERT J; WANG, DI; TENG, LESHENG

    2016-01-01

    Cordyceps militaris (CM), an entomopathogenic fungus belonging to the class ascomycetes, possesses various pharmacological activities, including cytotoxic effects, on various types of human tumor cells. The present study investigated the anti-hepatocellular carcinoma (HCC) and anti-breast cancer effects of CM in in vitro and in vivo models. CM aqueous extract reduced cell viability, suppressed cell proliferation, inhibited cell migration ability, caused the over-release of lactate dehydrogenase, induced mitochondrial dysfunction and enhanced apoptotic rates in MCF-7 and HepG2 cells. The expression levels of cleaved poly (ADP ribose) polymerase and caspase-3, biomarkers of apoptosis, were increased following treatment with CM aqueous extract for 24 h. Furthermore, in the MCF-7 and HepG2 cells, enhanced levels of B cell-associated X protein and cleaved caspase-8 were observed in the CM-treated cells. Finally, the antitumor activities of CM in HCC and breast cancer were also confirmed in MCF-7- and HepG2-xengraft nude mice models. Collectively, the data obtained in the present study suggested that the cytotoxic effects of CM aqueous extract on HCC and breast cancer are associated with the caspase-dependent mitochondrial pathway. PMID:27109250

  20. Existence of B/E and E receptors on Hep-G2 cells: a study using colloidal gold- and 125I-labeled lipoproteins

    International Nuclear Information System (INIS)

    Hesz, A.; Ingolic, E.; Krempler, F.; Kostner, G.M.

    1987-01-01

    The presence of specific receptors for apolipoprotein B (low-density lipoproteins) and apolipoprotein E (HDL-E) on Hep-G2 cells and human skin fibroblasts was studied by chemical methods and by electron microscopy using a differential gold labeling technique. Fibroblasts bound both types of lipoproteins to one and the same receptor (B/E receptor) as deduced from competition experiments with HDL-E and LDL. Labeled HDL-E, on the other hand, was only partially displaced by cold LDL but was completely displaced by unlabeled HDL-E. Scatchard analysis of lipoprotein binding to Hep-G2 cells revealed an approx 10 times higher binding affinity of apoE-containing lipoproteins as compared to apoB-containing ones. No differences between apoE- or apoB-containing lipoproteins with respect to the morphology of cell binding and intracellular processing were observed. The results are compatible with the concept that Hep-G2 cells possess two kinds of receptors, one specific for apoB- and apoE-containing lipoproteins (B/E receptor) and another specific for apoE only. From these studies we conclude that Hep-G2 cells may serve as a suitable model for studying the lipoprotein metabolism in the liver

  1. [Over-expression of uracil DNA glycosylase 2 (UNG2) enhances the resistance to oxidative damage in HepG2 cells].

    Science.gov (United States)

    Cao, Liyan; Cheng, Shan; Du, Juan; Guo, Yanhai; Huang, Xiaofeng

    2017-04-01

    Objective To investigate the uracil glycosidic enzyme activity of uracil DNA glycosylase 2 (UNG2) and study the role of UNG2 in the resistance of antioxidant stress of HepG2 cells. Methods The UNG2-expressing vector was built. Western blotting was used to detect the expression of UNG2. Immunofluorescence staining was performed to observe the cellular location of UNG2. Oligonucleotide was used as substrate for the determination of the UNG2 glycosidic enzyme activity. H 2 O 2 toxicity assay was done to study the function of UNG2 in the antioxidant resistance of hepatocellular carcinoma HepG2 cells. Results UNG2 was successfully over-expressed in HEK293FT cells, and UNG2 was found to be mainly located in nucleus. Enzyme activity assay showed that UNG2 had significant oligonucleotide dU glycosidic enzyme activity. H 2 O 2 toxicity assay showed that over-expressed UNG2 could remarkably increase the survival of HepG2 cells after exposed to H 2 O 2 . Conclusion UNG2 possesses specific DNA glycosidic enzyme activity, and it can protect HepG2 cells against oxidative stress damage.

  2. Gelsolin negatively regulates the activity of tumor suppressor p53 through their physical interaction in hepatocarcinoma HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    An, Joo-Hee; Kim, Jung-Woong; Jang, Sang-Min; Kim, Chul-Hong; Kang, Eun-Jin; Choi, Kyung-Hee, E-mail: khchoi@cau.ac.kr

    2011-08-19

    Highlights: {yields} The actin binding protein Gelsolin (GSN) interacts with transcription factor p53. {yields} GSN interacts with transactivation- and DNA binding domains of p53. {yields} GSN represses transactivity of p53 via inhibition of nuclear translocation of p53. {yields} GSN inhibits the p53-mediated apoptosis in hepatocarcinoma HepG2 cells. -- Abstract: As a transcription factor, p53 modulates several cellular responses including cell-cycle control, apoptosis, and differentiation. In this study, we have shown that an actin regulatory protein, gelsolin (GSN), can physically interact with p53. The nuclear localization of p53 is inhibited by GSN overexpression in hepatocarcinoma HepG2 cells. Additionally, we demonstrate that GSN negatively regulates p53-dependent transcriptional activity of a reporter construct, driven by the p21-promoter. Furthermore, p53-mediated apoptosis was repressed in GSN-transfected HepG2 cells. Taken together, these results suggest that GSN binds to p53 and this interaction leads to the inhibition of p53-induced apoptosis by anchoring of p53 in the cytoplasm in HepG2 cells.

  3. Melittin restores PTEN expression by down-regulating HDAC2 in human hepatocelluar carcinoma HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Hui Zhang

    Full Text Available Melittin is a water-soluble toxic peptide derived from the venom of the bee. Although many studies show the anti-tumor activity of melittin in human cancer including glioma cells, the underlying mechanisms remain elusive. Here the effect of melittin on human hepatocelluar carcinoma HepG2 cell proliferation in vitro and further mechanisms was investigated. We found melittin could inhibit cell proliferation in vitro using Flow cytometry and MTT method. Besides, we discovered that melittin significantly downregulated the expressions of CyclinD1 and CDK4. Results of western Blot and Real-time PCR analysis indicated that melittin was capable to upregulate the expression of PTEN and attenuate histone deacetylase 2 (HDAC2 expression. Further studies demonstrated that knockdown of HDAC2 completely mimicked the effects of melittin on PTEN gene expression. Conversely, it was that the potential utility of melittin on PTEN expression was reversed in cells treated with a recombinant pEGFP-C2-HDAC2 plasmid. In addition, treatment with melittin caused a downregulation of Akt phosphorylation, while overexpression of HDAC2 promoted Akt phosphorylation. These findings suggested that the inhibitory of cell growth by melittin might be led by HDAC2-mediated PTEN upregulation, Akt inactivation, and inhibition of the PI3K/Akt signaling pathways.

  4. Alcohol dehydrogenase and cytochrome P450 2E1 can be induced by long-term exposure to ethanol in cultured liver HEP-G2 cells.

    Science.gov (United States)

    Balusikova, Kamila; Kovar, Jan

    2013-09-01

    It has been shown in previous studies that liver HEP-G2 cells (human hepatocellular carcinoma) lose their ability to express active alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). Although both are ethanol-inducible enzymes, short-term exposure to ethanol does not cause any changes in expression or activity in cultured HEP-G2 cells. Therefore, we tested the effect of long-term exposure to ethanol on the expression and activity of both ADH and CYP2E1 in these cells. The expression of ADH and CYP2E1 was assessed at the mRNA and/or protein level using real-time PCR and Western blot analysis. Specific colorimetric assays were used for the measurement of ADH and CYP2E1 enzymatic activities. Caco-2 cells (active CYP2E1 and inactive ADH) were used as control cells. Significantly increased protein expression of ADH (about 2.5-fold) as well as CYP2E1 (about 1.6-fold) was found in HEP-G2 cells after long-term (12 mo) exposure to ethanol. The activity of ADH and CYP2E1 was also significantly increased from 12 ± 3 and 6 ± 1 nmol/h/mg of total protein to 191 ± 9 and 57 ± 9 nmol/h/mg of total protein, respectively. We suggest that the loss of activity of ethanol-metabolizing enzymes in cultured HEP-G2 cells is reversible and can be induced by prolonged exposure to ethanol. We are therefore able to reactivate HEP-G2 cells metabolic functions concerning ethanol oxidation just by modification of in vitro culture conditions without necessity of transfection with its side effect - enzyme overexpression.

  5. Ashwagandha (Withania somnifera supercritical CO2 extract derived withanolides mitigates Bisphenol A induced mitochondrial toxicity in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Satyakumar Vidyashankar

    2014-01-01

    Full Text Available Bisphenol A (BPA safety aspects on human health are debated extensively for long time. In the present study, we have studied the toxicity induced by BPA at no observed adverse effect level (NOAEL using HepG2 cells. We report that BPA at 100 nM induced cytotoxicity to HepG2 cells as determined by MTT assay at 0–72 h. The toxicity was result of reduced oxygen consumption and reduced mitochondrial membrane potential associated with decreased ATP production. The BPA treatment resulted in increase of malondialdehyde (MDA content with decreased glutathione and other antioxidant enzymes. BPA derived toxicity is a concern to human health and alternative non-toxic natural products/derivatives or adjuvants that serve as antidote will be relevant. In this context, Ashwagandha (Withania somnifera a widely used herb to treat arthritis, rheumatism and to improve longevity for time immemorial is investigated for its antidote effect. Ashwagandha supercritical CO2 extract derived Withanolides (ADW at 100 μg/ml protect HepG2 cells from BPA induced toxicity by suppressing mitochondrial damage and increased ATP production. Further, cellular MDA content was significantly suppressed with increased non-enzymic and antioxidant enzyme activities. These findings derived from the present study suggest the beneficial effect of ADW in mitigating BPA induced mitochondrial toxicity in HepG2 cells.

  6. Metabolomic effects of CeO2, SiO2 and CuO metal oxide nanomaterials on HepG2 cells

    Data.gov (United States)

    U.S. Environmental Protection Agency — The data set is a matrix of cellular biochemical (metabolites) in HepG2 cells treated with various metal oxide nanomaterials composed of CeO2, SiO2 and CuO. This...

  7. Metabolomic effects of CeO2, SiO2 and CuO metal oxide nanomaterials on HepG2 cells

    Science.gov (United States)

    Abstract Background To better assess potential hepatotoxicity of nanomaterials, human liver HepG2 cells were exposed for 3 days to five different CeO2 (either 30 or 100 μg/ml), 3 SiO2 based (30 μg/ml) or 1 CuO (3 μg/ml) nanomaterials with dry primary particle sizes r...

  8. TRAF1 knockdown alleviates palmitate-induced insulin resistance in HepG2 cells through NF-κB pathway

    International Nuclear Information System (INIS)

    Zhang, Wanlu; Tang, Zhuqi; Zhu, Xiaohui; Xia, Nana; Zhao, Yun; Wang, Suxin; Cui, Shiwei; Wang, Cuifang

    2015-01-01

    High-fat diet (HFD) and inflammation are key contributors to insulin resistance (IR) and Type 2 diabetes mellitus (T2DM). With HFD, plasma free fatty acids (FFAs) can activate the nuclear factor-κB (NF-κB) in target tissues, then initiate negative crosstalk between FFAs and insulin signaling. However, the molecular link between IR and inflammation remains to be identified. We here reported that tumor necrosis factor receptor-associated factor 1 (TRAF1), an adapter in signal transduction, was involved in the onset of IR in hepatocytes. TRAF1 was significantly up-regulated in insulin-resistant liver tissues and palmitate (PA)-treated HepG2 cells. In addition, we showed that depletion of TRAF1 led to inhibition of the activity of NF-κB. Given the fact that the activation of NF-κB played a facilitating role in IR, the phosphorylation of Akt and GSK3β was also analyzed. We found that depletion of TRAF1 markedly reversed PA-induced attenuation of the phosphorylation of Akt and GSK3β in the cells. The accumulation of lipid droplets in hepatocyte and expression of two key gluconeogenic enzymes, PEPCK and G6Pase, were also determined and found to display a similar tendency with the phosphorylation of Akt and GSK3β. Glucose uptake assay indicated that knocking down TRAF1 blocked the effect of PA on the suppression of glucose uptake. These data implicated that TRAF1 knockdown might alleviate PA-induced IR in HepG2 cells through NF-κB pathway. - Highlights: • TRAF1 accelerated PA-induced IR in HepG2 cells mediated through NF-κB signaling. • Knockdown of TRAF1 alleviated PA-induced IR in HepG2 cells. • Knockdown of TRAF1 alleviated PA-induced lipid accumulation in HepG2 cells. • Knockdown of TRAF1 reversed PA-induced suppression of glucose uptake in HepG2 cells. • Knockdown of TRAF1 reversed PA-induced gluconeogenesis in HepG2 cells.

  9. TRAF1 knockdown alleviates palmitate-induced insulin resistance in HepG2 cells through NF-κB pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Wanlu [Department of Pathogen Biology, Medical College, Nantong University, 19 Qixiu Road, Nantong 226001, Jiangsu Province (China); Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qixiu Road, Nantong 226001, Jiangsu Province (China); Tang, Zhuqi; Zhu, Xiaohui [Department of Endocrinology, Affiliated Hospital of Nantong University, 20 Xisi Road, Nantong 226001, Jiangsu Province (China); Xia, Nana; Zhao, Yun; Wang, Suxin [Department of Pathogen Biology, Medical College, Nantong University, 19 Qixiu Road, Nantong 226001, Jiangsu Province (China); Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qixiu Road, Nantong 226001, Jiangsu Province (China); Cui, Shiwei, E-mail: neifenmicui@163.com [Department of Endocrinology, Affiliated Hospital of Nantong University, 20 Xisi Road, Nantong 226001, Jiangsu Province (China); Wang, Cuifang, E-mail: binghuodinghuo@163.com [Department of Endocrinology, Affiliated Hospital of Nantong University, 20 Xisi Road, Nantong 226001, Jiangsu Province (China)

    2015-11-20

    High-fat diet (HFD) and inflammation are key contributors to insulin resistance (IR) and Type 2 diabetes mellitus (T2DM). With HFD, plasma free fatty acids (FFAs) can activate the nuclear factor-κB (NF-κB) in target tissues, then initiate negative crosstalk between FFAs and insulin signaling. However, the molecular link between IR and inflammation remains to be identified. We here reported that tumor necrosis factor receptor-associated factor 1 (TRAF1), an adapter in signal transduction, was involved in the onset of IR in hepatocytes. TRAF1 was significantly up-regulated in insulin-resistant liver tissues and palmitate (PA)-treated HepG2 cells. In addition, we showed that depletion of TRAF1 led to inhibition of the activity of NF-κB. Given the fact that the activation of NF-κB played a facilitating role in IR, the phosphorylation of Akt and GSK3β was also analyzed. We found that depletion of TRAF1 markedly reversed PA-induced attenuation of the phosphorylation of Akt and GSK3β in the cells. The accumulation of lipid droplets in hepatocyte and expression of two key gluconeogenic enzymes, PEPCK and G6Pase, were also determined and found to display a similar tendency with the phosphorylation of Akt and GSK3β. Glucose uptake assay indicated that knocking down TRAF1 blocked the effect of PA on the suppression of glucose uptake. These data implicated that TRAF1 knockdown might alleviate PA-induced IR in HepG2 cells through NF-κB pathway. - Highlights: • TRAF1 accelerated PA-induced IR in HepG2 cells mediated through NF-κB signaling. • Knockdown of TRAF1 alleviated PA-induced IR in HepG2 cells. • Knockdown of TRAF1 alleviated PA-induced lipid accumulation in HepG2 cells. • Knockdown of TRAF1 reversed PA-induced suppression of glucose uptake in HepG2 cells. • Knockdown of TRAF1 reversed PA-induced gluconeogenesis in HepG2 cells.

  10. Effect of green tea and (-)-epigallocatechin gallate on ethanol-induced toxicity in HepG2 cells.

    Science.gov (United States)

    Lee, Sang Il; Kim, Hyo Jung; Boo, Yong Chool

    2008-05-01

    Despite the continuing reports supporting the hepatoprotective effects of green tea against ethanol intoxication, there remain controversies regarding the active compound(s) and molecular mechanism. These issues were addressed in the present study using cultured HepG2 cells exposed to a lethal dose of ethanol. Gamma-glutamyl transferase (GGT) was chosen as a marker of ethanol toxicity because it is widely used in clinics. When the cells were treated with ethanol at various concentrations, there was a dose-dependent increase of GGT activity in the culture media and loss of cell viability. Pretreatment of the cells with green tea extract attenuated the changes significantly. Among the green tea constituents, (-)-epigallocatechin gallate (EGCG) attenuated the ethanol cytotoxicity effectively, whereas L-theanine and caffeine had no effects. The ethanol cytotoxicity was also attenuated by alcohol dehydrogenase inhibitor 4-methyl pyrazol and GGT inhibitor acivicin as well as by thiol modulators such as S-adenosyl-L-methionine, N-acetyl-L-cysteine and glutathione. EGCG failed to prevent the intracellular glutathione loss caused by ethanol, but it appeared to be a strong GGT inhibitor. Therefore the cytoprotective effects of green tea could be attributed to the inhibition of GGT activity by EGCG. This study suggests that GGT inhibitors including EGCG may provide a novel strategy for attenuating ethanol-induced liver damage.

  11. Synergy analysis reveals association between insulin signaling and desmoplakin expression in palmitate treated HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Xuewei Wang

    Full Text Available The regulation of complex cellular activities in palmitate treated HepG2 cells, and the ensuing cytotoxic phenotype, involves cooperative interactions between genes. While previous approaches have largely focused on identifying individual target genes, elucidating interacting genes has thus far remained elusive. We applied the concept of information synergy to reconstruct a "gene-cooperativity" network for palmititate-induced cytotoxicity in liver cells. Our approach integrated gene expression data with metabolic profiles to select a subset of genes for network reconstruction. Subsequent analysis of the network revealed insulin signaling as the most significantly enriched pathway, and desmoplakin (DSP as its top neighbor. We determined that palmitate significantly reduces DSP expression, and treatment with insulin restores the lost expression of DSP. Insulin resistance is a common pathological feature of fatty liver and related ailments, whereas loss of DSP has been noted in liver carcinoma. Reduced DSP expression can lead to loss of cell-cell adhesion via desmosomes, and disrupt the keratin intermediate filament network. Our findings suggest that DSP expression may be perturbed by palmitate and, along with insulin resistance, may play a role in palmitate induced cytotoxicity, and serve as potential targets for further studies on non-alcoholic fatty liver disease (NAFLD.

  12. 3-Nitrobenzanthrone (3-NBA) induced micronucleus formation and DNA damage in human hepatoma (HepG2) cells.

    Science.gov (United States)

    Lamy, Evelyn; Kassie, Fekadu; Gminski, Richard; Schmeiser, Heinz H; Mersch-Sundermann, Volker

    2004-01-15

    3-Nitrobenzanthrone (3-NBA), identified in diesel exhaust and in airborne particulate matter, is a potent mutagen in Salmonella, induces micronuclei formation in mice and in human cells and DNA adducts in rats. In the present study, we investigated the genotoxic potency of 3-NBA in human HepG2 cells using the micronucleus (MN) assay and the single cell gel electrophoresis (SCGE). 3-NBA caused a genotoxic effect at concentrations > or =12 nM in both assays. In the micronucleus assay, we found 98.7+/-10.3 MN/1000 BNC at a concentration of 100 nM 3-NBA in comparison to 27.3+/-0.6 MN/1000 BNC with the negative control. At the same concentration, the DNA-migration (SCGE) showed an Olive tail moment (OTM) of 2.7+/-0.45 and %DNA in the tail of 8.28+/-0.76; OTM and %DNA in the tail of cells treated with the negative control were 0.73+/-0.08 and 2.81+/-0.30, respectively. The results are discussed under consideration of former studies.

  13. Release and uptake of volatile organic compounds by human hepatocellular carcinoma cells (HepG2) in vitro.

    Science.gov (United States)

    Mochalski, Paweł; Sponring, Andreas; King, Julian; Unterkofler, Karl; Troppmair, Jakob; Amann, Anton

    2013-07-17

    Volatile organic compounds (VOCs) emitted by human body offer a unique insight into biochemical processes ongoing in healthy and diseased human organisms. Unfortunately, in many cases their origin and metabolic fate have not been yet elucidated in sufficient depth, thus limiting their clinical application. The primary goal of this work was to identify and quantify volatile organic compounds being released or metabolized by HepG2 hepatocellular carcinoma cells. The hepatocellular carcinoma cells were incubated in specially designed head-space 1-L glass bottles sealed for 24 hours prior to measurements. Identification and quantification of volatiles released and consumed by cells under study were performed by gas chromatography with mass spectrometric detection (GC-MS) coupled with head-space needle trap device extraction (HS-NTD) as the pre-concentration technique. Most of the compounds were identified both by spectral library match as well as retention time comparison based on standards. A total of nine compounds were found to be metabolised and further twelve released by the cells under study (Wilcoxon signed-rank test, p3-octanone, 2-nonanone), five volatile sulphur compounds (dimethyl sulfide, ethyl methyl sulfide, 3-methyl thiophene, 2-methyl-1-(methylthio)- propane and 2-methyl-5-(methylthio) furan), n-propyl acetate, and 2-heptene. The emission and uptake of the aforementioned VOCs may reflect the activity of abundant liver enzymes and support the potential of VOC analysis for the assessment of enzymes function.

  14. Effect of annatto on micronuclei induction by direct and indirect mutagens in HepG2 cells.

    Science.gov (United States)

    Barcelos, Gustavo Rafael Mazzaron; Angeli, José Pedro Friedmann; Serpeloni, Juliana Mara; Rocha, Bruno Alves; Mantovani, Mário Sérgio; Antunes, Lusânia Maria Greggi

    2009-12-01

    Annatto (AN), a natural food colorant rich in carotenoids, has been reported as being an effective antioxidant, but little is known about its potential chemopreventive properties. In this study, we evaluated the ability of AN to protect human hepatoma cells (HepG2) from micronucleus (MN) induction against three different mutagens: benzo(a)pyrene (B(a)P), doxorubicin (DXR), and methyl methanesulfonate (MMS). In an attempt to clarify the possible mechanism of antimutagenicity of AN, three protocols of treatment were applied (pretreatment; simultaneous treatment, and post-treatment with AN following treatment with the mutagens). Also, cells exposed only to AN were assayed for cytotoxicity and mutagenicity. A dosage up to 10 microg/ml of AN was devoid of mutagenic activity. Protective effects were seen on micronuclei induced by B(a)P and DXR using pre and simultaneous treatment, but AN had no significant effect on MN induction by MMS in any of the protocols. Our results also show that exposure of cells to concentrations of AN higher than 10 microg/ml decreased cell viability. Taken together, our findings indicate that AN presents antimutagenic activity in vitro, but its protective effect is dependent on the mutagen and on type of treatment suggesting its potential use as a chemopreventive agent.

  15. STARD4 knockdown in HepG2 cells disrupts cholesterol trafficking associated with the plasma membrane, ER, and ERC.

    Science.gov (United States)

    Garbarino, Jeanne; Pan, Meihui; Chin, Harvey F; Lund, Frederik W; Maxfield, Frederick R; Breslow, Jan L

    2012-12-01

    STARD4, a member of the evolutionarily conserved START gene family, has been implicated in the nonvesicular intracellular transport of cholesterol. However, the direction of transport and the membranes with which this protein interacts are not clear. We present studies of STARD4 function using small hairpin RNA knockdown technology to reduce STARD4 expression in HepG2 cells. In a cholesterol-poor environment, we found that a reduction in STARD4 expression leads to retention of cholesterol at the plasma membrane, reduction of endoplasmic reticulum-associated cholesterol, and decreased ACAT synthesized cholesteryl esters. Furthermore, D4 KD cells exhibited a reduced rate of sterol transport to the endocytic recycling compartment after cholesterol repletion. Although these cells displayed normal endocytic trafficking in cholesterol-poor and replete conditions, cell surface low density lipoprotein receptor (LDLR) levels were increased and decreased, respectively. We also observed a decrease in NPC1 protein expression, suggesting the induction of compensatory pathways to maintain cholesterol balance. These data indicate a role for STARD4 in nonvesicular transport of cholesterol from the plasma membrane and the endocytic recycling compartment to the endoplasmic reticulum and perhaps other intracellular compartments as well.

  16. In vitro treatment of HepG2 cells with saturated fatty acids reproduces mitochondrial dysfunction found in nonalcoholic steatohepatitis

    Directory of Open Access Journals (Sweden)

    Inmaculada García-Ruiz

    2015-02-01

    Full Text Available Activity of the oxidative phosphorylation system (OXPHOS is decreased in humans and mice with nonalcoholic steatohepatitis. Nitro-oxidative stress seems to be involved in its pathogenesis. The aim of this study was to determine whether fatty acids are implicated in the pathogenesis of this mitochondrial defect. In HepG2 cells, we analyzed the effect of saturated (palmitic and stearic acids and monounsaturated (oleic acid fatty acids on: OXPHOS activity; levels of protein expression of OXPHOS complexes and their subunits; gene expression and half-life of OXPHOS complexes; nitro-oxidative stress; and NADPH oxidase gene expression and activity. We also studied the effects of inhibiting or silencing NADPH oxidase on the palmitic-acid-induced nitro-oxidative stress and subsequent OXPHOS inhibition. Exposure of cultured HepG2 cells to saturated fatty acids resulted in a significant decrease in the OXPHOS activity. This effect was prevented in the presence of a mimic of manganese superoxide dismutase. Palmitic acid reduced the amount of both fully-assembled OXPHOS complexes and of complex subunits. This reduction was due mainly to an accelerated degradation of these subunits, which was associated with a 3-tyrosine nitration of mitochondrial proteins. Pretreatment of cells with uric acid, an antiperoxynitrite agent, prevented protein degradation induced by palmitic acid. A reduced gene expression also contributed to decrease mitochondrial DNA (mtDNA-encoded subunits. Saturated fatty acids induced oxidative stress and caused mtDNA oxidative damage. This effect was prevented by inhibiting NADPH oxidase. These acids activated NADPH oxidase gene expression and increased NADPH oxidase activity. Silencing this oxidase abrogated totally the inhibitory effect of palmitic acid on OXPHOS complex activity. We conclude that saturated fatty acids caused nitro-oxidative stress, reduced OXPHOS complex half-life and activity, and decreased gene expression of mt

  17. Shifts in dietary carbohydrate-lipid exposure regulate expression of the non-alcoholic fatty liver disease-associated gene PNPLA3/adiponutrin in mouse liver and HepG2 human liver cells.

    Science.gov (United States)

    Hao, Lei; Ito, Kyoko; Huang, Kuan-Hsun; Sae-tan, Sudathip; Lambert, Joshua D; Ross, A Catharine

    2014-10-01

    Patatin-like phospholipase domain containing 3 (PNPLA3, adiponutrin) has been identified as a modifier of lipid metabolism. To better understand the physiological role of PNPLA3/adiponutrin, we have investigated its regulation in intact mice and human hepatocytes under various nutritional/metabolic conditions. PNPLA3 gene expression was determined by real-time PCR in liver of C57BL/6 mice after dietary treatments and in HepG2 cells exposed to various nutritional/metabolic stimuli. Intracellular lipid content was determined in HepG2 cells after siRNA-mediated knockdown of PNPLA3. In vivo, mice fed a high-carbohydrate (HC) liquid diet had elevated hepatic lipid content, and PNPLA3 mRNA and protein expression, compared to chow-fed mice. Elevated expression was completely abrogated by addition of unsaturated lipid emulsion to the HC diet. By contrast, in mice with high-fat diet-induced steatosis, Pnpla3 expression did not differ compared to low-fat fed mice. In HepG2 cells, Pnpla3 expression was reversibly suppressed by glucose depletion and increased by glucose refeeding, but unchanged by addition of insulin and glucagon. Several unsaturated fatty acids each significantly decreased Pnpla3 mRNA, similar to lipid emulsion in vivo. However, Pnpla3 knockdown in HepG2 cells did not alter total lipid content in high glucose- or oleic acid-treated cells. Our results provide evidence that PNPLA3 expression is an early signal/signature of carbohydrate-induced lipogenesis, but its expression is not associated with steatosis per se. Under lipogenic conditions due to high-carbohydrate feeding, certain unsaturated fatty acids can effectively suppress both lipogenesis and PNPLA3 expression, both in vivo and in a hepatocyte cell line. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. Glyoxylate is a substrate of the sulfate-oxalate exchanger, sat-1, and increases its expression in HepG2 cells.

    Science.gov (United States)

    Schnedler, Nina; Burckhardt, Gerhard; Burckhardt, Birgitta C

    2011-03-01

    Hyperoxaluria is a major problem causing nephrolithiasis. Little is known about the regulation of oxalate transport from the liver, the main organ for oxalate synthesis, into the circulation. Since the sulfate anion transporter-1(sat-1) is present in the sinusoidal membrane of hepatocytes and translocates oxalate, its impact on increased oxalate synthesis was studied. Sat-1 expressing oocytes were used for cis-inhibition, trans-stimulation, and efflux experiments with labelled sulfate and oxalate to demonstrate the interactions of oxalate, glyoxylate, and glycolate with sat-1. HepG2 cells were incubated with oxalate and its precursors (glycine, hydroxyproline, glyoxylate, and glycolate). Changes in endogenous sat-1 mRNA-expression were examined using real-time PCR. After incubation of HepG2 cells in glyoxylate, sat-1 protein-expression was analysed by Western blotting, and sulfate uptake into HepG2 cells was measured. RT-PCR was used to screen for mRNA of other transporters. While oxalate and glyoxylate inhibited sulfate uptake, glycolate did not. Sulfate and oxalate uptake were trans-stimulated by glyoxylate but not by glycolate. Glyoxylate enhanced sulfate efflux. Glyoxylate was the only oxalate precursor stimulating sat-1 mRNA-expression. After incubation of HepG2 cells in glyoxylate, both sat-1 protein-expression and sulfate uptake into the cells increased. mRNA-expression of other transporters in HepG2 cells was not affected by glyoxylate treatment. The oxalate precursor glyoxylate was identified as a substrate of sat-1. Upregulated expression of sat-1 mRNA and of a functional sat-1 protein indicates that glyoxylate may be responsible for the elevated oxalate release from hepatocytes observed in hyperoxaluria. Copyright © 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  19. Polychlorinated biphenyl quinone-induced genotoxicity, oxidative DNA damage and γ-H2AX formation in HepG2 cells.

    Science.gov (United States)

    Dong, Hui; Su, Chuanyang; Xia, Xiaomin; Li, Lingrui; Song, Erqun; Song, Yang

    2014-04-05

    Our previous study has demonstrated that PCB quinone is cytotoxic in HepG2 cells (Toxicology in Vitro 26 (2012) 841-848). However, it is not clear whether PCB quinone is also carcinogenic (or mutagenic). In the current study, we investigated the genotoxicity of PCB quinone (2,3,5-trichloro-6-phenyl-[1,4]benzoquinone, PCB29-pQ) in HepG2 cells using single cell gel electrophoresis (SCGE) assay and micronucleus (MN) assay. We found PCB29-pQ exposure significantly increased olive tail moment (OTM) and micronuclei (MN) frequencies in HepG2 cells. These data suggested that PCB29-pQ caused DNA strand breaks and chromosome breaks. We further investigated whether the genotoxicity of PCB29-pQ is associated with the generation of reactive oxygen species (ROS). Using enzyme-linked immunosorbent assay for 8-hydroxydeoxyguanosine (8-OHdG) detection, we demonstrated that the level of oxidative DNA damage was significantly evaluated with PCB29-pQ exposure concentration and time dependently. Moreover, γ-H2AX appeared after the treatment of PCB29-pQ in HepG2 cells, may indicate double strand breaks (DSBs). In addition, the pretreatment of ROS scavengers inhibited the genotoxicity of PCB29-pQ significantly. In conclusion, our data suggested that PCB29-pQ causes genotoxic effects in HepG2 cells, probably via ROS-induced oxidative DNA damage. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  20. Generation of Multilayered 3D Structures of HepG2 Cells Using a Bio-printing Technique.

    Science.gov (United States)

    Jeon, Hyeryeon; Kang, Kyojin; Park, Su A; Kim, Wan Doo; Paik, Seung Sam; Lee, Sang-Hun; Jeong, Jaemin; Choi, Dongho

    2017-01-15

    Chronic liver disease is a major widespread cause of death, and whole liver transplantation is the only definitive treatment for patients with end-stage liver diseases. However, many problems, including donor shortage, surgical complications and cost, hinder their usage. Recently, tissue-engineering technology provided a potential breakthrough for solving these problems. Three-dimensional (3D) printing technology has been used to mimic tissues and organs suitable for transplantation, but applications for the liver have been rare. A 3D bioprinting system was used to construct 3D printed hepatic structures using alginate. HepG2 cells were cultured on these 3D structures for 3 weeks and examined by fluorescence microscopy, histology and immunohistochemistry. The expression of liverspecific markers was quantified on days 1, 7, 14, and 21. The cells grew well on the alginate scaffold, and liver-specific gene expression increased. The cells grew more extensively in 3D culture than two-dimensional culture and exhibited better structural aspects of the liver, indicating that the 3D bioprinting method recapitulates the liver architecture. The 3D bioprinting of hepatic structures appears feasible. This technology may become a major tool and provide a bridge between basic science and the clinical challenges for regenerative medicine of the liver.

  1. Antagonism of Secreted PCSK9 Increases Low Density Lipoprotein Receptor Expression in HepG2 Cells

    Energy Technology Data Exchange (ETDEWEB)

    McNutt, Markey C.; Kwon, Hyock Joo; Chen, Chiyuan; Chen, Justin R.; Horton, Jay D.; Lagace, Thomas A.; (USMC); (UTSMC)

    2009-07-10

    PCSK9 is a secreted protein that degrades low density lipoprotein receptors (LDLRs) in liver by binding to the epidermal growth factor-like repeat A (EGF-A) domain of the LDLR. It is not known whether PCSK9 causes degradation of LDLRs within the secretory pathway or following secretion and reuptake via endocytosis. Here we show that a mutation in the LDLR EGF-A domain associated with familial hypercholesterolemia, H306Y, results in increased sensitivity to exogenous PCSK9-mediated cellular degradation because of enhanced PCSK9 binding affinity. The crystal structure of the PCSK9-EGF-A(H306Y) complex shows that Tyr-306 forms a hydrogen bond with Asp-374 in PCSK9 at neutral pH, which strengthens the interaction with PCSK9. To block secreted PCSK9 activity, LDLR (H306Y) subfragments were added to the medium of HepG2 cells stably overexpressing wild-type PCSK9 or gain-of-function PCSK9 mutants associated with hypercholesterolemia (D374Y or S127R). These subfragments blocked secreted PCSK9 binding to cell surface LDLRs and resulted in the recovery of LDLR levels to those of control cells. We conclude that PCSK9 acts primarily as a secreted factor to cause LDLR degradation. These studies support the concept that pharmacological inhibition of the PCSK9-LDLR interaction extracellularly will increase hepatic LDLR expression and lower plasma low density lipoprotein levels.

  2. Xanthohumol Suppresses Mylip/Idol Gene Expression and Modulates LDLR Abundance and Activity in HepG2 Cells.

    Science.gov (United States)

    Chen, Shih-Fen; Chen, Pei-Yi; Hsu, Hao-Jen; Wu, Ming-Jiuan; Yen, Jui-Hung

    2017-09-13

    Xanthohumol, a prenylated flavonoid found in hops (Humulus lupulus L.), exhibits multiple biological activities such as antiatherosclerosis and hypolipidemic activities. In this study, we aim to investigate the hypocholesterolemic effects and molecular mechanisms of xanthohumol in hepatic cells. We found that xanthohumol (10 and 20 μM) increased the amount of cell-surface low-density lipoprotein receptor (LDLR) from 100.0 ± 2.1% to 115.0 ± 1.3% and 135.2 ± 2.7%, and enhanced the LDL uptake activity from 100.0 ± 0.9% to 139.1 ± 13.2% in HepG2 cells (p Idol) mRNA and protein by approximately 45% (p Idol expression via counteracting liver X receptor (LXR) activation. The molecular docking results predicted that xanthohumol has a high binding affinity to interact with the LXRα ligand-binding domain, which may result in attenuation of LXRα-induced Mylip/Idol expression. Finally, we demonstrated that the Mylip/Idol expression and LDLR activity were synergistically changed by a combination of xanthohumol and simvastatin treatment. Our findings indicated that xanthohumol may regulate the LXR-Mylip/Idol axis to modulate hepatic LDLR abundance and activity.

  3. Zinc protects HepG2 cells against the oxidative damage and DNA damage induced by ochratoxin A

    International Nuclear Information System (INIS)

    Zheng, Juanjuan; Zhang, Yu; Xu, Wentao; Luo, YunBo; Hao, Junran; Shen, Xiao Li; Yang, Xuan; Li, Xiaohong; Huang, Kunlun

    2013-01-01

    Oxidative stress and DNA damage are the most studied mechanisms by which ochratoxin A (OTA) induces its toxic effects, which include nephrotoxicity, hepatotoxicity, immunotoxicity and genotoxicity. Zinc, which is an essential trace element, is considered a potential antioxidant. The aim of this paper was to investigate whether zinc supplement could inhibit OTA-induced oxidative damage and DNA damage in HepG2 cells and the mechanism of inhibition. The results indicated that that exposure of OTA decreased the intracellular zinc concentration; zinc supplement significantly reduced the OTA-induced production of reactive oxygen species (ROS) and decrease in superoxide dismutase (SOD) activity but did not affect the OTA-induced decrease in the mitochondrial membrane potential (Δψ m ). Meanwhile, the addition of the zinc chelator N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) strongly aggravated the OTA-induced oxidative damage. This study also demonstrated that zinc helped to maintain the integrity of DNA through the reduction of OTA-induced DNA strand breaks, 8-hydroxy-2′-deoxyguanosine (8-OHdG) formation and DNA hypomethylation. OTA increased the mRNA expression of metallothionein1-A (MT1A), metallothionein2-A (MT2A) and Cu/Zn superoxide dismutase (SOD1). Zinc supplement further enhanced the mRNA expression of MT1A and MT2A, but it had no effect on the mRNA expression of SOD1 and catalase (CAT). Zinc was for the first time proven to reduce the cytotoxicity of OTA through inhibiting the oxidative damage and DNA damage, and regulating the expression of zinc-associated genes. Thus, the addition of zinc can potentially be used to reduce the OTA toxicity of contaminated feeds. - Highlights: ► OTA decreased the intracellular zinc concentration. ► OTA induced the formation of 8-OHdG in HepG2 cells. ► It was testified for the first time that OTA induced DNA hypomethylation. ► Zinc protects against the oxidative damage and DNA damage induced by OTA in

  4. An earthworm protease cleaving serum fibronectin and decreasing HBeAg in HepG2.2.15 cells

    Directory of Open Access Journals (Sweden)

    Zhao Jing

    2008-11-01

    Full Text Available Abstract Background Virus-binding activity is one of the important functions of fibronectin (FN. It has been reported that a high concentration of FN in blood improves the transmission frequency of hepatitis viruses. Therefore, to investigate a protease that hydrolyzes FN rapidly is useful to decrease the FN concentration in blood and HBV infection. So far, however, no specific protease digesting FN in serum has been reported. Methods We employed a purified earthworm protease to digest serum proteins. The rapidly cleaved protein (FN was identified by MALDI-TOF MS and western blotting. The cleavage sites were determined by N-terminus amino acid residues sequencing. The protease was orally administrated to rats to investigate whether serum FN in vivo became decreased. The serum FN was determined by western blotting and ELISA. In cytological studies, the protease was added to the medium in the culture of HepG2.2.15 cells and then HBsAg and HBeAg were determined by ELISA. Results The protease purified from earthworm Eisenia fetida was found to function as a fibronectinase (FNase. The cleavage sites on FN by the FNase were at R and K, exhibiting a trypsin alkaline serine-like function. The earthworm fibronectinase (EFNase cleaved FN at four sites, R259, R1005, K1557 and R2039, among which the digested fragments at R259, K1557 and R2039 were related to the virus-binding activity as reported. The serum FN was significantly decreased when the earthworm fibronectinase was orally administrated to rats. The ELISA results showed that the secretion of HBeAg from HepG2.2.15 cells was significantly inhibited in the presence of the FNase. Conclusion The earthworm fibronectinase (EFNase cleaves FN much faster than the other proteins in serum, showing a potential to inhibit HBV infection through its suppressing the level of HBeAg. This suggests that EFNase is probably used as one of the candidates for the therapeutic agents to treat hepatitis virus infection.

  5. Zinc protects HepG2 cells against the oxidative damage and DNA damage induced by ochratoxin A

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Juanjuan; Zhang, Yu [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); Xu, Wentao, E-mail: xuwentaoboy@sina.com [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); The Supervision, Inspection and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083 (China); Luo, YunBo [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); The Supervision, Inspection and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083 (China); Hao, Junran [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); Shen, Xiao Li [The Supervision, Inspection and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083 (China); Yang, Xuan [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); Li, Xiaohong [The Supervision, Inspection and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083 (China); Huang, Kunlun, E-mail: hkl009@163.com [Laboratory of Food Safety and Molecular Biology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083 (China); The Supervision, Inspection and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083 (China)

    2013-04-15

    Oxidative stress and DNA damage are the most studied mechanisms by which ochratoxin A (OTA) induces its toxic effects, which include nephrotoxicity, hepatotoxicity, immunotoxicity and genotoxicity. Zinc, which is an essential trace element, is considered a potential antioxidant. The aim of this paper was to investigate whether zinc supplement could inhibit OTA-induced oxidative damage and DNA damage in HepG2 cells and the mechanism of inhibition. The results indicated that that exposure of OTA decreased the intracellular zinc concentration; zinc supplement significantly reduced the OTA-induced production of reactive oxygen species (ROS) and decrease in superoxide dismutase (SOD) activity but did not affect the OTA-induced decrease in the mitochondrial membrane potential (Δψ{sub m}). Meanwhile, the addition of the zinc chelator N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) strongly aggravated the OTA-induced oxidative damage. This study also demonstrated that zinc helped to maintain the integrity of DNA through the reduction of OTA-induced DNA strand breaks, 8-hydroxy-2′-deoxyguanosine (8-OHdG) formation and DNA hypomethylation. OTA increased the mRNA expression of metallothionein1-A (MT1A), metallothionein2-A (MT2A) and Cu/Zn superoxide dismutase (SOD1). Zinc supplement further enhanced the mRNA expression of MT1A and MT2A, but it had no effect on the mRNA expression of SOD1 and catalase (CAT). Zinc was for the first time proven to reduce the cytotoxicity of OTA through inhibiting the oxidative damage and DNA damage, and regulating the expression of zinc-associated genes. Thus, the addition of zinc can potentially be used to reduce the OTA toxicity of contaminated feeds. - Highlights: ► OTA decreased the intracellular zinc concentration. ► OTA induced the formation of 8-OHdG in HepG2 cells. ► It was testified for the first time that OTA induced DNA hypomethylation. ► Zinc protects against the oxidative damage and DNA damage induced by

  6. Cholesterol lowering effects of mono-lactose-appended β-cyclodextrin in Niemann–Pick type C disease-like HepG2 cells

    Directory of Open Access Journals (Sweden)

    Keiichi Motoyama

    2015-11-01

    Full Text Available The Niemann–Pick type C disease (NPC is one of inherited lysosomal storage disorders, emerges the accumulation of unesterified cholesterol in endolysosomes. Currently, 2-hydroxypropyl-β-cyclodextrin (HP-β-CyD has been applied for the treatment of NPC. HP-β-CyD improved hepatosplenomegaly in NPC patients, however, a high dose of HP-β-CyD was necessary. Therefore, the decrease in dose by actively targeted-β-CyD to hepatocytes is expected. In the present study, to deliver β-CyD selectively to hepatocytes, we newly fabricated mono-lactose-appended β-CyD (Lac-β-CyD and evaluated its cholesterol lowering effects in NPC-like HepG2 cells, cholesterol accumulated HepG2 cells induced by treatment with U18666A. Lac-β-CyD (degree of substitution of lactose (DSL 1 significantly decreased the intracellular cholesterol content in a concentration-dependent manner. TRITC-Lac-β-CyD was associated with NPC-like HepG2 cells higher than TRITC-β-CyD. In addition, TRITC-Lac-β-CyD was partially localized with endolysosomes after endocytosis. Thus, Lac-β-CyD entered NPC-like HepG2 cells via asialoglycoprotein receptor (ASGPR-mediated endocytosis and decreased the accumulation of intracellular cholesterol in NPC-like HepG2 cells. These results suggest that Lac-β-CyD may have the potential as a drug for the treatment of hepatosplenomegaly in NPC disease.

  7. Saponins, especially platycodin D, from Platycodon grandiflorum modulate hepatic lipogenesis in high-fat diet-fed rats and high glucose-exposed HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Yong Pil [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Department of Pharmaceutical Engineering, International University of Korea, Jinju (Korea, Republic of); Choi, Jae Ho; Kim, Hyung Gyun; Khanal, Tilak; Song, Gye Young [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Nam, Myoung Soo [College of Agriculture and Life Sciences, Chungnam National University, Daejeon (Korea, Republic of); Lee, Hyun-Sun [Molecular Cancer Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Chung, Young Chul; Lee, Young Chun [Division of Food Science, International University of Korea, Jinju (Korea, Republic of); Jeong, Hye Gwang, E-mail: hgjeong@cnu.ac.kr [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon 305-764 (Korea, Republic of)

    2013-03-01

    AMP-activated protein kinase (AMPK) plays a central role in controlling hepatic lipid metabolism through modulating the downstream acetyl CoA carboxylase (ACC) and sterol regulatory element-binding protein-1c (SREBP-1c) pathway. Saponins, particularly platycodin D, from the roots of Platycodon grandiflorum (Changkil saponins, CKS) have a variety of pharmacological properties, including antioxidant and hepatoprotective properties. The aim of this study was to investigate the effects of CKS on hepatic lipogenesis and on the expression of genes involved in lipogenesis, and the mechanisms involved. CKS attenuated fat accumulation and the induction of the lipogenic genes encoding SREBP-1c and fatty acid synthase in the livers of HFD-fed rats and in steatotic HepG2 cells. Blood biochemical analyses and histopathological examinations showed that CKS prevented liver injury. CKS and platycodin D each increased the phosphorylation of AMPK and acetyl-CoA carboxylase in HFD-fed rats and HepG2 cells. The use of specific inhibitors showed that platycodin D activated AMPK via SIRT1/CaMKKβ in HepG2 cells. This study demonstrates that CKS or platycodin D alone can regulate hepatic lipogenesis via an AMPK-dependent signalling pathway. - Highlights: ► CKS attenuated fat accumulation in HFD-fed rats and in steatotic HepG2 cells. ► CKS and its major component, platycodin D, inhibited the levels of SREBP-1 and FAS. ► CKS and platycodin D increased the phosphorylation of AMPK and ACC. ► Platycodin D activated AMPK via SIRT1/CaMKKβ in HepG2 cells.

  8. (3'R)-hydroxytabernaelegantine C: A bisindole alkaloid with potent apoptosis inducing activity in colon (HCT116, SW620) and liver (HepG2) cancer cells.

    Science.gov (United States)

    Paterna, Angela; Gomes, Sofia E; Borralho, Pedro M; Mulhovo, Silva; Rodrigues, Cecília M P; Ferreira, Maria-José U

    2016-12-24

    Tabernaemontana elegans Stapf. (Apocynaceae) is a medicinal plant traditionally used in African countries to treat cancer. To discover new apoptosis inducing lead compounds from T. elegans and provide scientific validation of the ethnopharmacological use of this plant. Through fractionation, (3'R)-hydroxytaberanelegantine C (1), a vobasinyl-iboga bisindole alkaloid, was isolated from a cytotoxic alkaloid fraction of the methanol extract of T. elegans roots. Its structure was identified by spectroscopic methods, mainly 1D and 2D NMR experiments. Compound 1 was evaluated for its ability to induce apoptosis in HCT116 and SW620 colon and HepG2 liver carcinoma cells. The cell viability of compound 1 was evaluated by the MTS and lactate dehydrogenase (LDH) assays. Induction of apoptosis was analyzed through Guava ViaCount assay, by flow cytometry, caspase-3/7 activity assays and evaluation of nuclear morphology by Hoechst staining. To determine the molecular pathways elicited by 1 exposure, immunoblot analysis was also performed. (3'R)-hydroxytaberanelegantine C (1) displayed strong apoptosis induction activity as compared to 5-fluorouracil (5-FU), the most used anticancer agent in colorectal cancer treatment. In the MTS assay, compound 1 exhibited IC 50 values similar or lower than 5-FU in the three cell lines tested. The IC 50 value of 1 was also calculated in CCD18co normal human colon fibroblasts. The lactate dehydrogenase assay showed increased LDH release by compound 1, and the Guava ViaCount assay revealed that 1 significantly increased the incidence of apoptosis to a further extent than 5-FU. Moreover, the induction of apoptosis was corroborated by evaluation of nuclear morphology by Hoechst staining and caspase-3/7 activity assays of 1 treated cells. As expected, in immunoblot analysis, compound 1 treatment led to poly(ADP-ribose) polymerase cleavage. This was accompanied by decreased anti-apoptotic proteins Bcl-2 and XIAP steady state levels in all three cancer

  9. Protective Effect of Curcumin against Ionizing Radiation (IR)-induced Cytotoxicity and Genotoxicity in HepG2 Cells

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Dong Min; Nasir Uddin, S. M.; Ryu, Tae Ho; Kang, Mi Young; Kim, Jin Kyu [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2013-10-15

    Ionizing radiation (IR) has many practical applications such as medicine, foods, agricultures, industries, and research laboratories. However, the increasing use of radiation is associated with radiation accidents threatening human health. It is well known that exposure to IR gives rise to genomic alterations, mutagenesis, and cell death. IR is absorbed directly by DNA, leading to various DNA damages (single or double-strand breaks, base damage, and DNA-DNA or DNA-protein cross-linkages) in many living organisms. Therefore, the development of effective and nontoxic radioprotective agents is of considerable interest. Curcumin (C{sub 12}H{sub 20}O{sub 6}, structure is the major yellow component of Curcuma longa with biological activities (antioxidant, anti-proliferative and anti-inflammatory properties). It has been widely used as food and medicine for a long time. The aim of our present study is to investigate the protective effects of curcumin against IR-induced cytotoxicity and genotoxicity in cultured HepG2 cells.

  10. Statins Activate Human PPAR Promoter and Increase PPAR mRNA Expression and Activation in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Makoto Seo

    2008-01-01

    Full Text Available Statins increase peroxisome proliferator-activated receptor (PPAR mRNA expression, but the mechanism of this increased PPAR production remains elusive. To examine the regulation of PPAR production, we examined the effect of 7 statins (atorvastatin, cerivastatin, fluvastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin on human PPAR promoter activity, mRNA expression, nuclear protein levels, and transcriptional activity. The main results are as follows. (1 Majority of statins enhanced PPAR promoter activity in a dose-dependent manner in HepG2 cells transfected with the human PPAR promoter. This enhancement may be mediated by statin-induced HNF-4. (2 PPAR mRNA expression was increased by statin treatment. (3 The PPAR levels in nuclear fractions were increased by statin treatment. (4 Simvastatin, pravastatin, and cerivastatin markedly enhanced transcriptional activity in 293T cells cotransfected with acyl-coenzyme A oxidase promoter and PPAR/RXR expression vectors. In summary, these data demonstrate that PPAR production and activation are upregulated through the PPAR promoter activity by statin treatment.

  11. Flow cytometric evaluation of the contribution of ionic silver to genotoxic potential of nanosilver in human liver HepG2 and colon Caco2 cells.

    Science.gov (United States)

    Sahu, Saura C; Njoroge, Joyce; Bryce, Steven M; Zheng, Jiwen; Ihrie, John

    2016-04-01

    Exposure to nanosilver found in food- and cosmetics-related consumer products is of public concern because of the lack of information about its safety. In this study, two widely used in vitro cell culture models, human liver HepG2 and colon Caco2 cells, and the flow cytometric micronucleus (FCMN) assay were evaluated as tools for rapid predictive screening of the potential genotoxicity of nanosilver. Recently, we reported the genotoxicity of 20 nm nanosilver using these systems. In the current study presented here, we tested the hypothesis that the nanoparticle size and cell types were critical determinants of its genotoxicity. To test this hypothesis, we used the FCMN assay to evaluate the genotoxic potential of 50 nm nanosilver of the same shape, composition, surface charge and obtained from the same commercial source using the same experimental conditions and in vitro models (HepG2 and Caco2) as previously tested for the 20 nm silver. Results of our study show that up to the concentrations tested in these cultured cell test systems, the smaller (20 nm) nanoparticle is genotoxic to both the cell types by inducing micronucleus (MN). However, the larger (50 nm) nanosilver induces MN only in HepG2 cells, but not in Caco2 cells. Also in this study, we evaluated the contribution of ionic silver to the genotoxic potential of nanosilver using silver acetate as the representative ionic silver. The MN frequencies in HepG2 and Caco2 cells exposed to the ionic silver in the concentration range tested are not statistically significant from the control values except at the top concentrations for both the cell types. Therefore, our results indicate that the ionic silver may not contribute to the MN-forming ability of nanosilver in HepG2 and Caco2 cells. Also our results suggest that the HepG2 and Caco2 cell cultures and the FCMN assay are useful tools for rapid predictive screening of a genotoxic potential of food- and cosmetics-related chemicals including nanosilver

  12. Static magnetic field enhances the anticancer efficacy of capsaicin on HepG2 cells via capsaicin receptor TRPV1.

    Science.gov (United States)

    Chen, Wei-Ting; Lin, Guan-Bo; Lin, Shu-Hui; Lu, Chueh-Hsuan; Hsieh, Chih-Hsiung; Ma, Bo-Lun; Chao, Chih-Yu

    2018-01-01

    Static magnetic field (SMF) has shown some possibilities for cancer therapies. In particular, the combinational effect between SMF and anti-cancer drugs has drawn scientists' attentions in recent years. However, the underlying mechanism for the drug-specific synergistic effect is far from being understood. Besides, the drugs used are all conventional chemotherapy drugs, which may cause unpleasant side effects. In this study, our results demonstrate for the first time that SMF could enhance the anti-cancer effect of natural compound, capsaicin, on HepG2 cancer cells through the mitochondria-dependent apoptosis pathway. We found that the synergistic effect could be due to that SMF increased the binding efficiency of capsaicin for the TRPV1 channel. These findings may provide a support to develop an application of SMF for cancer therapy. The present study offers the first trial in combining SMF with natural compound on anti-cancer treatment, which provides additional insight into the interaction between SMF and anti-cancer drugs and opens the door for the development of new strategies in fighting cancer with minimum cytotoxicity and side effects.

  13. Signaling dynamics of palmitate-induced ER stress responses mediated by ATF4 in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Cho Hyunju

    2013-01-01

    Full Text Available Abstract Background Palmitic acid, the most common saturated free fatty acid, has been implicated in ER (endoplasmic reticulum stress-mediated apoptosis. This lipoapotosis is dependent, in part, on the upregulation of the activating transcription factor-4 (ATF4. To better understand the mechanisms by which palmitate upregulates the expression level of ATF4, we integrated literature information on palmitate-induced ER stress signaling into a discrete dynamic model. The model provides an in silico framework that enables simulations and predictions. The model predictions were confirmed through further experiments in human hepatocellular carcinoma (HepG2 cells and the results were used to update the model and our current understanding of the signaling induced by palmitate. Results The three key things from the in silico simulation and experimental results are: 1 palmitate induces different signaling pathways (PKR (double-stranded RNA-activated protein kinase, PERK (PKR-like ER kinase, PKA (cyclic AMP (cAMP-dependent protein kinase A in a time dependent-manner, 2 both ATF4 and CREB1 (cAMP-responsive element-binding protein 1 interact with the Atf4 promoter to contribute to a prolonged accumulation of ATF4, and 3 CREB1 is involved in ER-stress induced apoptosis upon palmitate treatment, by regulating ATF4 expression and possibly Ca2+ dependent-CaM (calmodulin signaling pathway. Conclusion The in silico model helped to delineate the essential signaling pathways in palmitate-mediated apoptosis.

  14. Cultivation of HepG2.2.15 on Cytodex-3

    DEFF Research Database (Denmark)

    Lupberger, Joachim; Mund, Andreas; Kock, Josef

    2006-01-01

    BACKGROUND/AIMS: Several novel systems are available to study human hepatitis B virus (HBV) replication in cell culture demanding for efficient cell culture based systems for HBV production. The aim was to enhance HBV production of the HBV stably producing cell line HepG2.2.15 by cultivation...... on spherical micro substrate. METHODS: HepG2.2.15 was cultivated on microcarrier substrate Cytodex-3. HBV specific transcripts, viral protein and genome secretion, cell proliferation and MAP kinase signaling were analyzed. Infectivity of HBV particles was analyzed using primary tupaia hepatocytes. RESULTS......: Compared to stationary flask cultures, HepG2.2.15 on Cytodex-3 secreted 18-fold more HBV genomes, more HBeAg per culture volume and less HBV surface antigen per extracellular viral genome equivalent. This was reflected by a significantly higher infectivity of supernatant derived from carrier grown HepG.2...

  15. Surface Grafted Glycopolymer Brushes to Enhance Selective Adhesion of HepG2 Cells

    DEFF Research Database (Denmark)

    Chernyy, Sergey; Jensen, Bettina Elisabeth Brøgger; Shimizu, Kyoko

    2013-01-01

    This work demonstrates the application of carbohydrate based methacrylate polymer brush, poly(2-lactobionamidoethyl methacrylate), for the purpose of cell adhesion studies. The first part of the work illustrates the effects of the structure of the aminosilane based ATRP initiator layer on the pol......This work demonstrates the application of carbohydrate based methacrylate polymer brush, poly(2-lactobionamidoethyl methacrylate), for the purpose of cell adhesion studies. The first part of the work illustrates the effects of the structure of the aminosilane based ATRP initiator layer...... on the polymerization kinetics of 2-lactobionamidoethyl methacrylate) (LAMA) monomer on thermally oxidized silicon wafer. Both monolayer and multilayered aminosilane precursor layers have been prepared followed by reaction with 2-bromoisobutyrylbromide to form the ATRP initiator layer. It is inferred from the kinetic...... studies that the rate of termination is low on a multilayered initiator layer compared to a disordered monolayer structure. However both initiator types results in similar graft densities. Furthermore, it is shown that thick comb-like poly(LAMA) brushes can be constructed by initiating a second ATRP...

  16. Pro-oxidant effect of ALA is implicated in mitochondrial dysfunction of HepG2 cells.

    Science.gov (United States)

    Laafi, Jihane; Homedan, Chadi; Jacques, Caroline; Gueguen, Naig; Schmitt, Caroline; Puy, Hervé; Reynier, Pascal; Carmen Martinez, Maria; Malthièry, Yves

    2014-11-01

    Heme biosynthesis begins in the mitochondrion with the formation of delta-aminolevulinic acid (ALA). In acute intermittent porphyria, hereditary tyrosinemia type I and lead poisoning patients, ALA is accumulated in plasma and in organs, especially the liver. These diseases are also associated with neuromuscular dysfunction and increased incidence of hepatocellular carcinoma. Many studies suggest that this damage may originate from ALA-induced oxidative stress following its accumulation. Using the MnSOD as an oxidative stress marker, we showed here that ALA treatment of cultured cells induced ROS production, increasing with ALA concentration. The mitochondrial energetic function of ALA-treated HepG2 cells was further explored. Mitochondrial respiration and ATP content were reduced compared to control cells. For the 300 μM treatment, ALA induced a mitochondrial mass decrease and a mitochondrial network imbalance although neither necrosis nor apoptosis were observed. The up regulation of PGC-1, Tfam and ND5 genes was also found; these genes encode mitochondrial proteins involved in mitochondrial biogenesis activation and OXPHOS function. We propose that ALA may constitute an internal bioenergetic signal, which initiates a coordinated upregulation of respiratory genes, which ultimately drives mitochondrial metabolic adaptation within cells. The addition of an antioxidant, Manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), resulted in improvement of maximal respiratory chain capacity with 300 μM ALA. Our results suggest that mitochondria, an ALA-production site, are more sensitive to pro-oxidant effect of ALA, and may be directly involved in pathophysiology of patients with inherited or acquired porphyria. Copyright © 2014 Elsevier B.V. and Société française de biochimie et biologie Moléculaire (SFBBM). All rights reserved.

  17. Orexin A affects HepG2 human hepatocellular carcinoma cells glucose metabolism via HIF-1α-dependent and -independent mechanism.

    Science.gov (United States)

    Wan, Xing; Liu, Yuanyuan; Zhao, Yuyan; Sun, Xiaoqi; Fan, Dongxiao; Guo, Lei

    2017-01-01

    Orexins are hypothalamic neuropeptides that regulate feeding, reward, wakefulness and energy homeostasis. The present study sought to characterize the involvement of orexin A in glucose metabolism in HepG2 human hepatocellular carcinoma cells, and investigated the role of hypoxia-inducible factor-1α (HIF-1α) in the response. HepG2 cells were exposed to different concentrations of orexin A (10-9 to 10-7 M) in vitro, without or with the orexin receptor 1 (OX1R) inhibitor (SB334867), HIF-1α inhibitor (YC-1) or a combination of both inhibitors. Subsequently, OX1R, HIF-1α expression and localization, glucose uptake, glucose transporter 1 (GLUT1) expression and ATP content were measured. We further investigated the intracellular fate of glucose by measuring the gene expression of pyruvate dehydrogenase kinase 1 (PDK1), lactate dehydrogenase (LDHA) and pyruvate dehydrogenase B (PDHB), as well as metabolite levels including lactate generation and mitochondrial pyruvate dehydrogenase (PDH) activity. The activity of phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway was also assessed. Our results showed that the expression of OX1R was predominantly located in the nucleus in HepG2 cells. Orexin A oxygen-independently promoted the mRNA and protein expression of HIF-1α as well as its nuclear accumulation in HepG2 cells and the elevated HIF-1α protein was associated, at least partly, with the activation of the PI3K/Akt/mTOR pathway. Orexin A stimulated GLUT1 expression, glucose uptake as well as ATP generation in HepG2 cells via OX1R acting through the HIF-1α pathway. Moreover, orexin A inhibited LDHA, PDK1 expression and lactate production, stimulated PDHB expression and PDH enzyme activity independent of HIF-1α. Our results indicated that orexin signaling facilitated the glucose flux into mitochondrial oxidative metabolism rather than glycolysis in HepG2 cells. These findings provide new insight into the regulation of glucose metabolism

  18. Internalisation and multiple phosphorylation of γ-Conglutin, the lupin seed glycaemia-lowering protein, in HepG2 cells

    International Nuclear Information System (INIS)

    Capraro, Jessica; Magni, Chiara; Faoro, Franco; Maffi, Dario; Scarafoni, Alessio; Tedeschi, Gabriella; Maffioli, Elisa; Parolari, Anna; Manzoni, Cristina; Lovati, Maria Rosa; Duranti, Marcello

    2013-01-01

    Highlights: •A glycaemia-reducing lupin seed protein is internalized by HepG2 cells. •The protein accumulates in the cytosol in an intact form. •The internalized protein is multiply phosphorylated. -- Abstract: Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing glucose uptake by model cells. This work investigated whether γ-Conglutin is internalised into the target cells and undergoes any covalent change during the process, as a first step to understanding its mechanism of action. To this purpose, γ-Conglutin-treated and untreated HepG2 cells were submitted to confocal and transmission electron microscopy. Immune-revelation of γ-Conglutin at various intervals revealed its accumulation inside the cytosol. In parallel, 2D-electrophoresis of the cell lysates and antibody reaction of the blotted maps showed the presence of the protein intact subunits inside the treated cells, whilest no trace of the protein was found in the control cells. However, γ-Conglutin-related spots with an unexpectedly low pI were also observed in the maps. These spots were excised, trypsin-treated and submitted to MS/MS spectrometric analysis. The presence of phosphorylated amino acids was detected. These findings, by showing that γ-Conglutin is internalised by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to the understanding of the lupin γ-Conglutin insulin-mimetic activity

  19. Measurement of Hepatic Protein Fractional Synthetic Rate with Stable Isotope Labeling Technique in Thapsigargin Stressed HepG2 Cells

    Science.gov (United States)

    Song, Juquan; Zhang, Xiao-jun; Boehning, Darren; Brooks, Natasha C.; Herndon, David N.; Jeschke, Marc G.

    2012-01-01

    Severe burn-induced liver damage and dysfunction is associated with endoplasmic reticulum (ER) stress. ER stress has been shown to regulate global protein synthesis. In the current study, we induced ER stress in vitro and estimated the effect of ER stress on hepatic protein synthesis. The aim was two-fold: (1) to establish an in vitro model to isotopically measure hepatic protein synthesis and (2) to evaluate protein fractional synthetic rate (FSR) in response to ER stress. Human hepatocellular carcinoma cells (HepG2) were cultured in medium supplemented with stable isotopes 1,2-13C2-glycine and L-[ring-13C6]phenylalanine. ER stress was induced by exposing the cells to 100 nM of thapsigargin (TG). Cell content was collected from day 0 to 14. Alterations in cytosolic calcium were measured by calcium imaging and ER stress markers were confirmed by Western blotting. The precursor and product enrichments were detected by GC-MS analysis for FSR calculation. We found that the hepatic protein FSR were 0.97±0.02 and 0.99±0.05%/hr calculated from 1,2-13C2-glycine and L-[ring-13C6]phenylalanine, respectively. TG depleted ER calcium stores and induced ER stress by upregulating p-IRE-1 and Bip. FSR dramatically decreased to 0.68±0.03 and 0.60±0.06%/hr in the TG treatment group (pisotope tracer incorporation technique is a useful method for studying the effects of ER stress on hepatic protein synthesis. PMID:22298954

  20. Bystander effect in human hepatoma HepG2 cells caused by medium transfers at different times after high-LET carbon ion irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Wu Qingfeng [Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Lanzhou 730000 (China); Graduate School of Chinese Academy of Sciences, Beijing 100039 (China); Li Qiang, E-mail: liqiang@impcas.ac.c [Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Lanzhou 730000 (China); Jin Xiaodong; Liu Xinguo [Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Lanzhou 730000 (China); Dai Zhongying [Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Lanzhou 730000 (China); Graduate School of Chinese Academy of Sciences, Beijing 100039 (China)

    2011-01-15

    Although radiation-induced bystander effects have been well documented in a variety of biological systems, whether irradiated cells have the ability to generate bystander signaling persistently is still unclear and the clinical relevance of bystander effects in radiotherapy remains to be elucidated. This study examines tumor cellular bystander response to autologous medium from cell culture irradiated with high-linear energy transfer (LET) heavy ions at a therapeutically relevant dose in terms of clonogenic cell survival. In vitro experiments were performed using human hepatoma HepG2 cell line exposed to 100 keV/{mu}m carbon ions at a dose of 2 Gy. Two different periods (2 and 12 h) after irradiation, irradiated cell conditioned medium (ICCM) and replenished fresh medium were harvested and then transferred to unirradiated bystander cells. Cellular bystander responses were measured with the different medium transfer protocols. Significant higher survival fractions of unirradiated cells receiving the media from the irradiated cultures at the different times post-irradiation than those of the control were observed. Even replenishing fresh medium for unirradiated cells which had been exposed to the ICCM for 12 h could not prevent the bystander cells from the increased survival fraction. These results suggest that the irradiated cells could release unidentified signal factor(s), which induced the increase in survival fraction for the unirradiated bystander cells, into the media sustainedly and the carbon ions triggered a cascade of signaling events in the irradiated cells rather than secreting the soluble signal factor(s) just at a short period after irradiation. Based on the observations in this study, the importance of bystander effect in clinical radiotherapy was discussed and incorporating the bystander effect into the current radiobiological models, which are applicable to heavy ion radiotherapy, is needed urgently.

  1. Bystander effect in human hepatoma HepG2 cells caused by medium transfers at different times after high-LET carbon ion irradiation

    International Nuclear Information System (INIS)

    Wu Qingfeng; Li Qiang; Jin Xiaodong; Liu Xinguo; Dai Zhongying

    2011-01-01

    Although radiation-induced bystander effects have been well documented in a variety of biological systems, whether irradiated cells have the ability to generate bystander signaling persistently is still unclear and the clinical relevance of bystander effects in radiotherapy remains to be elucidated. This study examines tumor cellular bystander response to autologous medium from cell culture irradiated with high-linear energy transfer (LET) heavy ions at a therapeutically relevant dose in terms of clonogenic cell survival. In vitro experiments were performed using human hepatoma HepG2 cell line exposed to 100 keV/μm carbon ions at a dose of 2 Gy. Two different periods (2 and 12 h) after irradiation, irradiated cell conditioned medium (ICCM) and replenished fresh medium were harvested and then transferred to unirradiated bystander cells. Cellular bystander responses were measured with the different medium transfer protocols. Significant higher survival fractions of unirradiated cells receiving the media from the irradiated cultures at the different times post-irradiation than those of the control were observed. Even replenishing fresh medium for unirradiated cells which had been exposed to the ICCM for 12 h could not prevent the bystander cells from the increased survival fraction. These results suggest that the irradiated cells could release unidentified signal factor(s), which induced the increase in survival fraction for the unirradiated bystander cells, into the media sustainedly and the carbon ions triggered a cascade of signaling events in the irradiated cells rather than secreting the soluble signal factor(s) just at a short period after irradiation. Based on the observations in this study, the importance of bystander effect in clinical radiotherapy was discussed and incorporating the bystander effect into the current radiobiological models, which are applicable to heavy ion radiotherapy, is needed urgently.

  2. Antioxidant mechanism of mitochondria-targeted plastoquinone SkQ1 is suppressed in aglycemic HepG2 cells dependent on oxidative phosphorylation

    Czech Academy of Sciences Publication Activity Database

    Ježek, J.; Engstová, Hana; Ježek, Petr

    2017-01-01

    Roč. 1858, č. 9 (2017), s. 750-762 ISSN 0005-2728 R&D Projects: GA ČR(CZ) GA17-01813S Institutional support: RVO:67985823 Keywords : mitochondria-targeted antioxidant SkQ1 * mitochondrial Complex I superoxide formation * mitochondrial Complex III superoxide formation * HepG2 cells * NAD(P)H fluorescence lifetime imaging microscopy Subject RIV: ED - Physiology OBOR OECD: Cell biology Impact factor: 4.932, year: 2016

  3. Newly developed chitosan-silver hybrid nanoparticles: biosafety and apoptosis induction in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    El-Sherbiny, Ibrahim M., E-mail: ielsherbiny@Zewailcity.edu.eg; Salih, Ehab [Zewail City of Science and Technology, Center for Materials Science (Egypt); Yassin, Abdelrahman M. [Genetic Engineering and Biotechnology Research Institute, City of Scientific Research and Technology Applications, Biopharmaceutical Product Research Department (Egypt); Hafez, Elsayed E. [City of Scientific Research and Technology Applications, Plant Protection and Biomolecular Diagnosis Department (Egypt)

    2016-07-15

    The present study reports the biosafety assessment, the exact molecular effects, and apoptosis induction of newly developed chitosan-silver hybrid nanoparticles (Cs–Ag NPs) in HepG2 cells. The investigated hybrid NPs were green synthesized using Cs/grape leaves aqueous extract (Cs/GLE) or Cs/GLE NPs as reducing and stabilizing agents. The successful formation of Cs/GLE NPs and Cs–Ag hybrid NPs has been confirmed by UV–Vis spectrophotometry, FTIR spectroscopy, XRD, and HRTEM. From the TEM analysis, the prepared Cs/GLE NPs are uniform and spherical with an average size of 150 nm, and the AgNPs (5–10 nm) were formed mainly on their surface. The UV–Vis spectra of Cs–Ag NPs showed a surface plasmon resonance (SPR) peak at about 450 nm confirming their formation. The synthesized Cs–Ag NPs were found to be crystalline as shown by XRD patterns with fcc phase oriented along the (111), (200), (220), and (311) planes. The cytotoxicity patterns, the antiproliferative activities, and the possible mechanisms of anticancer activity at molecular level of the newly developed Cs–Ag hybrid NPs were investigated. Cytotoxicity patterns of all the preparations demonstrated that the nontoxic treatment concentrations are ranged from 0.39 to 50 %, and many of the newly prepared Cs–Ag hybrid NPs showed high anticancer activities against HpG2 cells, and induced cellular apoptosis by downregulating BCL2 gene and upregulating P53.Graphical Abstract.

  4. Heat-modified citrus pectin induces apoptosis-like cell death and autophagy in HepG2 and A549 cancer cells.

    Directory of Open Access Journals (Sweden)

    Lionel Leclere

    Full Text Available Cancer is still one of the leading causes of death worldwide, and finding new treatments remains a major challenge. Previous studies showed that modified forms of pectin, a complex polysaccharide present in the primary plant cell wall, possess anticancer properties. Nevertheless, the mechanism of action of modified pectin and the pathways involved are unclear. Here, we show that citrus pectin modified by heat treatment induced cell death in HepG2 and A549 cells. The induced cell death differs from classical apoptosis because no DNA cleavage was observed. In addition, Z-VAD-fmk, a pan-caspase inhibitor, did not influence the observed cell death in HepG2 cells but appeared to be partly protective in A549 cells, indicating that heat-modified citrus pectin might induce caspase-independent cell death. An increase in the abundance of the phosphatidylethanolamine-conjugated Light Chain 3 (LC3 protein and a decrease in p62 protein abundance were observed in both cell types when incubated in the presence of heat-modified citrus pectin. These results indicate the activation of autophagy. To our knowledge, this is the first time that autophagy has been revealed in cells incubated in the presence of a modified form of pectin. This autophagy activation appears to be protective, at least for A549 cells, because its inhibition with 3-methyladenine increased the observed modified pectin-induced cytotoxicity. This study confirms the potential of modified pectin to improve chemotherapeutic cancer treatments.

  5. Heat-modified citrus pectin induces apoptosis-like cell death and autophagy in HepG2 and A549 cancer cells.

    Science.gov (United States)

    Leclere, Lionel; Fransolet, Maude; Cote, Francois; Cambier, Pierre; Arnould, Thierry; Van Cutsem, Pierre; Michiels, Carine

    2015-01-01

    Cancer is still one of the leading causes of death worldwide, and finding new treatments remains a major challenge. Previous studies showed that modified forms of pectin, a complex polysaccharide present in the primary plant cell wall, possess anticancer properties. Nevertheless, the mechanism of action of modified pectin and the pathways involved are unclear. Here, we show that citrus pectin modified by heat treatment induced cell death in HepG2 and A549 cells. The induced cell death differs from classical apoptosis because no DNA cleavage was observed. In addition, Z-VAD-fmk, a pan-caspase inhibitor, did not influence the observed cell death in HepG2 cells but appeared to be partly protective in A549 cells, indicating that heat-modified citrus pectin might induce caspase-independent cell death. An increase in the abundance of the phosphatidylethanolamine-conjugated Light Chain 3 (LC3) protein and a decrease in p62 protein abundance were observed in both cell types when incubated in the presence of heat-modified citrus pectin. These results indicate the activation of autophagy. To our knowledge, this is the first time that autophagy has been revealed in cells incubated in the presence of a modified form of pectin. This autophagy activation appears to be protective, at least for A549 cells, because its inhibition with 3-methyladenine increased the observed modified pectin-induced cytotoxicity. This study confirms the potential of modified pectin to improve chemotherapeutic cancer treatments.

  6. Nanosilica induced dose-dependent cytotoxicity and cell type-dependent multinucleation in HepG2 and L-02 cells

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Yongbo [Capital Medical University, Beijing Key Laboratory for Pediatric Diseases of Otolaryngology, Head and Neck Surgery, Beijing Pediatric Research Institute, Beijing Children’s Hospital (China); Duan, Junchao; Li, Yang; Yu, Yang; Hu, Hejing; Wu, Jing; Zhang, Yannan; Li, Yanbo; CaixiaGuo; Zhou, Xianqing; Sun, Zhiwei, E-mail: zwsun@ccmu.edu.cn [Capital Medical University, School of Public Health (China)

    2016-11-15

    The prevalent exposure to nanosilica gained concerns about health effects of these particles on human beings. Although nanosilica-induced multinucleation has been confirmed previously, the underlying mechanism was still not clear; this study was to investigate the origination of multinucleated cells caused by nanosilica (62 nm) in both HepG2 and L-02 cells. Cell viability and cellular uptake was determined by MTT assay and transmission electron microscope (TEM), respectively. Giemsa staining was applied to detect multinucleation. To clarify the origination of multinucleated cells, fluorescent probes, PKH26 and PKH67, time-lapse observation were further conducted by confocal microscopy. Results indicated that nanosilica particles were internalized into cells and induced cytotoxicity in a dose-dependent manner. Quantification analysis showed that nanosilica significantly increased the rates of binucleated and multinucleated cells, which suggested mitotic catastrophe induction. Moreover, dynamic visualization verified that multinucleation resulted from cell fusion in HepG2 cells not in L-02 cells after nanosilica exposure, suggesting cell type-dependent multinucleation formation. Both multinucleation and cell fusion were involved in genetic instability, which emphasized the significance to explore the multinucleation induced by nanosilica via environmental, occupational and consumer product exposure.

  7. N-Acetyl-L-Cysteine Affords Protection against Lead-Induced Cytotoxicity and Oxidative Stress in Human Liver Carcinoma (HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Paul B. Tchounwou

    2007-06-01

    Full Text Available Although lead exposure has declined in recent years as a result of change to lead-free gasoline, several epidemiological have pointed out that it represents a medical and public health emergency, especially in young children consuming high amounts of lead-contaminated flake paints. A previous study in our laboratory indicated that lead exposure induces cytotoxicity in human liver carcinoma cells. In the present study, we evaluated the role of oxidative stress in lead-induced toxicity, and the protective effect of the anti-oxidant n-acetyl-l-cysteine (NAC. We hypothesized that oxidative stress plays a role in lead-induced cytotoxicity, and that NAC affords protection against this adverse effect. To test this hypothesis, we performed the MTT [3-(4, 5-dimethylthiazol-2-yl-2, 5-diphenyltetrazolium bromide] assay and the trypan blue exclusion test for cell viability. We also performed the thiobarbituric acid test for lipid peroxidation. Data obtained from the MTT assay indicated that NAC significantly increased the viability of HepG2 cells in a dosedependent manner upon 48 hours of exposure. Similar trend was obtained with the trypan blue exclusion test. Data generated from the thiobarbituric acid test showed a significant (p ≤ 0.05 increase of MDA levels in lead nitrate-treated HepG2 cells compared to control cells. Interestingly, the addition of NAC to lead nitrate-treated HepG2 cells significantly decreased cellular content of reactive oxygen species (ROS, as evidenced by the decrease in lipid peroxidation byproducts. Overall, findings from this study suggest that NAC inhibits lead nitrate-induced cytotoxicity and oxidative stress in HepG2 cells. Hence, NAC may be used as a salvage therapy for lead-induced toxicity in exposed persons.

  8. Copper(ii) oxide nanoparticles penetrate into HepG2 cells, exert cytotoxicity via oxidative stress and induce pro-inflammatory response

    Science.gov (United States)

    Piret, Jean-Pascal; Jacques, Diane; Audinot, Jean-Nicolas; Mejia, Jorge; Boilan, Emmanuelle; Noël, Florence; Fransolet, Maude; Demazy, Catherine; Lucas, Stéphane; Saout, Christelle; Toussaint, Olivier

    2012-10-01

    The potential toxic effects of two types of copper(ii) oxide (CuO) nanoparticles (NPs) with different specific surface areas, different shapes (rod or spheric), different sizes as raw materials and similar hydrodynamic diameter in suspension were studied on human hepatocarcinoma HepG2 cells. Both CuO NPs were shown to be able to enter into HepG2 cells and induce cellular toxicity by generating reactive oxygen species. CuO NPs increased the abundance of several transcripts coding for pro-inflammatory interleukins and chemokines. Transcriptomic data, siRNA knockdown and DNA binding activities suggested that Nrf2, NF-κB and AP-1 were implicated in the response of HepG2 cells to CuO NPs. CuO NP incubation also induced activation of MAPK pathways, ERKs and JNK/SAPK, playing a major role in the activation of AP-1. In addition, cytotoxicity, inflammatory and antioxidative responses and activation of intracellular transduction pathways induced by rod-shaped CuO NPs were more important than spherical CuO NPs. Measurement of Cu2+ released in cell culture medium suggested that Cu2+ cations released from CuO NPs were involved only to a small extent in the toxicity induced by these NPs on HepG2 cells.The potential toxic effects of two types of copper(ii) oxide (CuO) nanoparticles (NPs) with different specific surface areas, different shapes (rod or spheric), different sizes as raw materials and similar hydrodynamic diameter in suspension were studied on human hepatocarcinoma HepG2 cells. Both CuO NPs were shown to be able to enter into HepG2 cells and induce cellular toxicity by generating reactive oxygen species. CuO NPs increased the abundance of several transcripts coding for pro-inflammatory interleukins and chemokines. Transcriptomic data, siRNA knockdown and DNA binding activities suggested that Nrf2, NF-κB and AP-1 were implicated in the response of HepG2 cells to CuO NPs. CuO NP incubation also induced activation of MAPK pathways, ERKs and JNK/SAPK, playing a major

  9. Protection of HepG2 cells against acrolein toxicity by 2-cyano-3,12-dioxooleana-1,9-dien-28-imidazolide via glutathione-mediated mechanism.

    Science.gov (United States)

    Shah, Halley; Speen, Adam M; Saunders, Christina; Brooke, Elizabeth A S; Nallasamy, Palanisamy; Zhu, Hong; Li, Y Robert; Jia, Zhenquan

    2015-10-01

    Acrolein is an environmental toxicant, mainly found in smoke released from incomplete combustion of organic matter. Several studies showed that exposure to acrolein can lead to liver damage. The mechanisms involved in acrolein-induced hepatocellular toxicity, however, are not completely understood. This study examined the cytotoxic mechanisms of acrolein on HepG2 cells. Acrolein at pathophysiological concentrations was shown to cause apoptotic cell death and an increase in levels of protein carbonyl and thiobarbituric acid reactive acid substances. Acrolein also rapidly depleted intracellular glutathione (GSH), GSH-linked glutathione-S-transferases, and aldose reductase, three critical cellular defenses that detoxify reactive aldehydes. Results further showed that depletion of cellular GSH by acrolein preceded the loss of cell viability. To further determine the role of cellular GSH in acrolein-mediated cytotoxicity, buthionine sulfoximine (BSO) was used to inhibit cellular GSH biosynthesis. It was observed that depletion of cellular GSH by BSO led to a marked potentiation of acrolein-mediated cytotoxicity in HepG2 cells. To further assess the contribution of these events to acrolein-induced cytotoxicity, triterpenoid compound 2-cyano-3,12-dioxooleana-1,9-dien-28-imidazolide (CDDO-Im) was used for induction of GSH. Induction of GSH by CDDO-Im afforded cytoprotection against acrolein toxicity in HepG2 cells. Furthermore, BSO significantly inhibited CDDO-Im-mediated induction in cellular GSH levels and also reversed cytoprotective effects of CDDO-Im in HepG2 cells. These results suggest that GSH is a predominant mechanism underlying acrolein-induced cytotoxicity as well as CDDO-Im-mediated cytoprotection. This study may provide understanding on the molecular action of acrolein which may be important to develop novel strategies for the prevention of acrolein-mediated toxicity. © 2014 by the Society for Experimental Biology and Medicine.

  10. Oxidovanadium(IV) sulfate-induced glucose uptake in HepG2 cells through IR/Akt pathway and hydroxyl radicals.

    Science.gov (United States)

    Zhao, Qian; Chen, Deliang; Liu, Pingsheng; Wei, Taotao; Zhang, Fang; Ding, Wenjun

    2015-08-01

    The insulin-mimetic and anti-diabetic properties of vanadium and related compounds have been well documented both in vitro and in vivo. However, the molecular basis of the link between vanadium and the insulin signaling pathway in diabetes mellitus is not fully described. We investigated the effects of reactive oxygen species (ROS) induced by oxidovanadium(IV) sulfate (VOSO4) on glucose uptake and the insulin signaling pathway in human hepatoma cell line HepG2. Exposure of cells to VOSO4 (5-50 μM) resulted in an increase in glucose uptake, insulin receptor (IR) and protein kinase B (Akt) phosphorylation and intracellular ROS generation. Using Western blot, we found that catalase and sodium formate, but not superoxide dismutase, prevented the increase of hydroxyl radical (·OH) generation and significantly decreased VOSO4-induced IR and Akt phosphorylation. These results suggest that VOSO4-induced ·OH radical, which is a signaling species, promotes glucose uptake via the IR/Akt signaling pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. In HepG2 Cells, Coexisting Carnitine Deficiency Masks Important Indicators of Marginal Biotin Deficiency123

    Science.gov (United States)

    Bogusiewicz, Anna; Boysen, Gunnar; Mock, Donald M

    2015-01-01

    Background: A large number of birth defects are related to nutrient deficiencies; concern that biotin deficiency is teratogenic in humans is reasonable. Surprisingly, studies indicate that increased urinary 3-hydroxyisovalerylcarnitine (3HIAc), a previously validated marker of biotin deficiency, is not a valid biomarker in pregnancy. Objective: In this study we hypothesized that coexisting carnitine deficiency can prevent the increase in 3HIAc due to biotin deficiency. Methods: We used a 2-factor nutrient depletion design to induce isolated and combined biotin and carnitine deficiency in HepG2 cells and then repleted cells with carnitine. To elucidate the metabolic pathogenesis, we quantitated intracellular and extracellular free carnitine, acylcarnitines, and acylcarnitine ratios using liquid chromatography–tandem mass spectrometry. Results: Relative to biotin-sufficient, carnitine-sufficient cells, intracellular acetylcarnitine increased by 90%, propionylcarnitine more than doubled, and 3HIAc increased by >10-fold in biotin-deficient, carnitine-sufficient (BDCS) cells, consistent with a defensive mechanism in which biotin-deficient cells transesterify the acyl-coenzyme A (acyl-CoA) substrates of the biotin-dependent carboxylases to the related acylcarnitines. Likewise, in BDCS cells, the ratio of acetylcarnitine to malonylcarnitine and the ratio of propionylcarnitine to methylmalonylcarnitine both more than tripled, and the ratio of 3HIAc to 3-methylglutarylcarnitine (MGc) increased by >10-fold. In biotin-deficient, carnitine-deficient (BDCD) cells, the 3 substrate-derived acylcarnitines changed little, but the substrate:product ratios were masked to a lesser extent. Moreover, carnitine repletion unmasked biotin deficiency in BDCD cells as shown by increases in acetylcarnitine, propionylcarnitine, and 3HIAc (each increased by >50-fold). Likewise, ratios of acetylcarnitine:malonylcarnitine, propionylcarnitine:methylmalonylcarnitine, and 3HIAc:MGc all increased

  12. Saikosaponin A, an active glycoside from Radix bupleuri, reverses P-glycoprotein-mediated multidrug resistance in MCF-7/ADR cells and HepG2/ADM cells.

    Science.gov (United States)

    Ye, Rui-Ping; Chen, Zhen-Dong

    2017-02-01

    1. The expression and function of P-glycoprotein (P-gp) is associated with the phenotype of multidrug resistance (MDR). Saikosaponin A (SSA) is a triterpenoid saponin isolated from Radix Bupleuri. This study was mainly designed to understand effects of SSA on MDR in MCF-7/ADR and HepG2/ADM cells. 2. MDR reversal was examined as the alteration of cytotoxic drugs IC50 in resistant cells in the presence of SSA by MTT assay, and was compared with the non-resistant cells. Apoptosis and uptake of P-gp substrates in the tumor cells were detected by flow cytometry. Western blot was performed to assay the expression of P-gp. 3. Our results demonstrate SSA could increase the chemosensitivity of P-gp overexpressing HepG2/ADM and MCF-7/ADR cells to doxorubicin (DOX), vincristine (VCR) and paclitaxel. SSA promoted apoptosis of MCF-7/ADR cells in the presence of DOX. Moreover, it could also increase the retention of P-gp substrates DOX and rhodamine 123 in MCF-7/ADR cells, and decrease digoxin efflux ratio in Caco-2 cell monolayer. Finally, a mechanistic study showed that SSA reduced P-gp expression without affecting hydrolytic activity of P-gp. 4. In conclusion, our findings suggest that SSA could be further developed for sensitizing resistant cancer cells and used as an adjuvant therapy together with anticancer drugs to improve their therapeutic efficacies.

  13. Baicalin and its metabolites suppresses gluconeogenesis through activation of AMPK or AKT in insulin resistant HepG-2 cells.

    Science.gov (United States)

    Wang, Tao; Jiang, Hongmei; Cao, Shijie; Chen, Qian; Cui, Mingyuan; Wang, Zhijie; Li, Dandan; Zhou, Jing; Wang, Tao; Qiu, Feng; Kang, Ning

    2017-12-01

    Scutellaria baicalensis Georgi (S. baicalensis), as a traditional Chinese herbal medicine, is an important component of several famous Chinese medicinal formulas for treating patients with diabetes mellitus. Baicalin (BG), a main bioactive component of S. baicalensis, has been reported to have antidiabetic effects. However, pharmacokinetic studies have indicated that BG has poor oral bioavailability. Therefore, it is hard to explain the pharmacological effects of BG in vivo. Interestingly, several reports show that BG is extensively metabolized in rats and humans. Therefore, we speculate that the BG metabolites might be responsible for the pharmacological effects. In this study, BG and its three metabolites (M1-M3) were examined their effects on glucose consumption in insulin resistant HepG-2 cells with a commercial glucose assay kit. Real-time PCR and western blot assay were used to confirm genes and proteins of interest, respectively. The results demonstrate that BG and its metabolites (except for M3) enhanced the glucose consumption which might be associated with inhibiting the expression of the key gluconeogenic genes, including glucose-6-phosphatase (G6Pase), phosphoenolypyruvate carboxykinase (PEPCK) and glucose transporter 2 (GLUT2). Further study found that BG and M1 could suppress hepatic gluconeogenesis via activation of the AMPK pathway, while M2 could suppress hepatic gluconeogenesis via activation of the PI3K/AKT signaling pathway. Taken together, our findings suggest that both BG and its metabolites have antihyperglycemic activities, and might be the active forms of oral doses of BG in vivo. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  14. Oroxylin A reverses CAM-DR of HepG2 cells by suppressing Integrinβ1 and its related pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Binbin; Zhao, Li; Zhu, Litao; Wang, Hu; Sha, Yunying; Yao, Jing [State Key Laboratory of Natural Medicines, Jiangsu Key Laboratory of Carcinogenesis and Intervention, China Pharmaceutical University, 24 Tongjiaxiang, Nanjing 210009 (China); Li, Zhiyu [Department of Medicinal Chemistry, China Pharmaceutical University, 24 Tongjiaxiang, Nanjing 210009 (China); You, Qidong, E-mail: youqidong@gmail.com [Department of Medicinal Chemistry, China Pharmaceutical University, 24 Tongjiaxiang, Nanjing 210009 (China); Guo, Qinglong, E-mail: anticancer_drug@yahoo.com.cn [State Key Laboratory of Natural Medicines, Jiangsu Key Laboratory of Carcinogenesis and Intervention, China Pharmaceutical University, 24 Tongjiaxiang, Nanjing 210009 (China)

    2012-03-15

    Oroxylin A, a naturally occurring monoflavonoid extracted from Scutellariae radix, shows effective anticancer activities and low toxicities both in vivo and in vitro in previous studies. In this study, we investigated whether the CAM-DR model of HepG2 cells showed resistance to cytotoxic agents compared with normally cultured HepG2 cells. Furthermore, after the treatment of Paclitaxel, less inhibitory effects and decreased apoptosis rate were detected in the model. Data also revealed increased expression of Integrinβ1 might be responsible for the resistance ability. Moreover, Integrinβ1-siRNA-transfected CAM-DR HepG2 cells exhibited more inhibitory effects and higher levels of apoptosis than the non-transfected CAM-DR cells. The data corroborated that Integrinβ1 played a significant role in CAM-DR. After the treatment of weakly-toxic concentrations of Oroxylin A, the apoptosis induced by Paclitaxel in the CAM-DR model increased dramatically. Western blot assay revealed Oroxylin A markedly down-regulated the expression of Integrinβ1 and the activity of related pathway. As a conclusion, Oroxylin A can reverse the resistance of CAM-DR via inhibition of Integrinβ1 and its related pathway. Oroxylin A may be a potential candidate of a CAM-DR reversal agent. Highlights: ► Adhesion of HepG2 cells to fibronectin exhibited resistance to Paclitaxel. ► The resistance was associated with the increased expression of Integrinβ1. ► Knocking down Integrinβ1 can increase the toxicity of Paclitaxel on CAM-DR model. ► Oroxylin A reversed the resistance by suppressing Integrinβ1 and related pathway.

  15. Genotoxicity evaluation of multi-component mixtures of polyaromatic hydrocarbons (PAHs), arsenic, cadmium, and lead using flow cytometry based micronucleus test in HepG2 cells.

    Science.gov (United States)

    Muthusamy, Sasikumar; Peng, Cheng; Ng, Jack C

    2018-03-01

    Some polyaromatic hydrocarbons (PAHs) and metals are known human carcinogens and the combined toxicity data of these co-contaminants are important for assessing their health risk. In this study, we have evaluated the combined genotoxicity, AhR activity and cell cycle parameters of four PAHs (benzo[a]pyrene (Ba]P), naphthalene (Nap), phenanthrene (Phe) and pyrene (Pyr)) and three metals (arsenic (As), cadmium (Cd), and lead (Pb)) in HepG2 cells using a flow cytometry based micronucleus (MN) test CAFLUX assay and nuclear fluorescence assay, respectively. The mixtures of B[a]P and metals induced a maximum of four fold increase in the MN formation compared to B[a]P alone. The higher combination of PAHs and metals did not significantly increase the MN formation. The mixtures of metals or non-carcinogenic PAHs were found to increase or decrease the aryl hydrocarbon receptor (AhR) activation of B[a]P in HepG2 cell based CAFLUX assay. Overall, the results showed that combined genotoxicity of PAHs and metals in HepG2 cells vary depending on the concentrations and number of the chemicals that are present in the mixtures and the effects of higher order combinations appear to be largely unpredictable from binary combinations. In this study, we have demonstrated the use of flow cytometry based MN test to screen the genotoxicity of environmental chemicals and its mixtures. Copyright © 2018 Elsevier B.V. All rights reserved.

  16. Dihydrocapsaicin down-regulates apoM expression through inhibiting Foxa2 expression and enhancing LXRα expression in HepG2 cells.

    Science.gov (United States)

    Zhao, Jia-Yi; Hu, Yan-Wei; Li, Shu-Fen; Hu, Ya-Rong; Ma, Xin; Wu, Shao-Guo; Wang, Yan-Chao; Gao, Ji-Juan; Sha, Yan-Hua; Zheng, Lei; Wang, Qian

    2014-03-19

    Apolipoprotein M (apoM), as a novel apolipoprotein which is mainly expressed in liver and kidney tissues, is associated with development and progression of atherosclerosis and diabetes. Our group have recently shown that Dihydrocapsaicin(DHC)can significantly decrease atherosclerotic plaque formation in apoE-/- mice. However, the effect and possible mechanism of DHC on apoM expression remain unclear. HepG2 cells were treated with 0 μM, 25 μM, 50 μM and 100 μM DHC for 24 h or were treated with 100 μM DHC for 0, 6, 12, and 24 h, respectively. The mRNA levels and protein levels were measured by real-time quantitative PCR and western blot analysis, respectively. We found that DHC markedly decreased expression of apoM at both mRNA and protein level in HepG2 cells in a dose-dependent and time-dependent manner. Expression of Foxa2 was decreased while expression of LXRα was increased by DHC treatment in HepG2 cells. In addittion, overexpression of Foxa2 markedly compensated the inhibition effect induced by DHC on apoM expression. LXRα small interfering RNA significantly abolished the inhibition effect which induced by DHC on apoM expression. The liver of C57BL/6 mice treated with DHC had significantly lower expression of apoM. Furthermore, the liver had lower expression of Foxa2 while had higher expression of LXRα. DHC could down-regulate apoM expression through inhibiting Foxa2 expression and enhancing LXRα expression in HepG2 cells.

  17. Metabolomic effects in HepG2 cells exposed to CeO2, SiO2 and CuO nanomaterials.

    Science.gov (United States)

    To better assess potential hepatotoxicity of nanomaterials, human liver HepG2 cells were exposed for three days to 5 different CeO2 (either 30 or 100 ug/ml), 3 SiO2 based (30 ug/ml) or 1 CuO (3 ug/ml) nanomaterials with dry primary particle sizes ranging from 15 to 213 nm. Metab...

  18. Lignans from Opuntia ficus-indica seeds protect rat primary hepatocytes and HepG2 cells against ethanol-induced oxidative stress.

    Science.gov (United States)

    Kim, Jung Wha; Yang, Heejung; Kim, Hyeon Woo; Kim, Hong Pyo; Sung, Sang Hyun

    2017-01-01

    Bioactivity-guided isolation of Opuntia ficus-indica (Cactaceae) seeds against ethanol-treated primary rat hepatocytes yielded six lignan compounds. Among the isolates, furofuran lignans 4-6, significantly protected rat hepatocytes against ethanol-induced oxidative stress by reducing intracellular reactive oxygen species levels, preserving antioxidative defense enzyme activities, and maintaining the glutathione content. Moreover, 4 dose-dependently induced the heme oxygenase-1 expression in HepG2 cells.

  19. Metabolomic effects of CeO2, SiO2 and CuO metal oxide nanomaterials on HepG2 cells

    Science.gov (United States)

    To better assess potential hepatotoxicity of nanomaterials, human liver HepG2 cells were exposed for 3 days to five different CeO2 (either 30 or 100 μg/ml), 3 SiO2 based (30 μg/ml) or 1 CuO (3 μg/ml) nanomaterials with dry primary particle sizes ranging from 15 to 213 nm. Metabol...

  20. Isoorientin induces apoptosis through mitochondrial dysfunction and inhibition of PI3K/Akt signaling pathway in HepG2 cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yuan, Li; Wang, Jing; Xiao, Haifang; Xiao, Chunxia; Wang, Yutang; Liu, Xuebo, E-mail: xueboliu@yahoo.com.cn

    2012-11-15

    Isoorientin (ISO) is a flavonoid compound that can be extracted from several plant species, such as Phyllostachys pubescens, Patrinia, and Drosophyllum lusitanicum; however, its biological activity remains poorly understood. The present study investigated the effects and putative mechanism of apoptosis induced by ISO in human hepatoblastoma cancer (HepG2) cells. The results showed that ISO induced cell death in a dose-dependent manner in HepG2 cells, but no toxicity in human liver cells (HL-7702) and buffalo rat liver cells (BRL-3A) treated with ISO at the indicated concentrations. ISO-induced cell death included apoptosis which characterized by the appearance of nuclear shrinkage, the cleavage of poly (ADP-ribose) polymerase (PARP) and DNA fragmentation. ISO significantly (p < 0.01) increased the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), increased the release of cytochrome c, activated caspase-3, and enhanced intracellular levels of reactive oxygen species (ROS) and nitric oxide (NO). In addition, ISO effectively inhibited the phosphorylation of Akt and increased FoxO4 expression. The PI3K/Akt inhibitor LY294002 enhanced the apoptosis-inducing effect of ISO. However, LY294002 markedly quenched ROS and NO generation and diminished the protein expression of heme peroxidase enzyme (HO-1) and inducible nitric oxide synthase (iNOS). Furthermore, the addition of a ROS inhibitor (N-acetyl cysteine, NAC) or iNOS inhibitor (N-[3-(aminomethyl) benzyl] acetamidine, dihydrochloride, 1400W) significantly diminished the apoptosis induced by ISO and also blocked the phosphorylation of Akt. These results demonstrated for the first time that ISO induces apoptosis in HepG2 cells and indicate that this apoptosis might be mediated through mitochondrial dysfunction and PI3K/Akt signaling pathway, and has no toxicity in normal liver cells, suggesting that ISO may have good potential as a therapeutic and chemopreventive agent for liver cancer. Highlights:

  1. Isoorientin induces apoptosis through mitochondrial dysfunction and inhibition of PI3K/Akt signaling pathway in HepG2 cancer cells

    International Nuclear Information System (INIS)

    Yuan, Li; Wang, Jing; Xiao, Haifang; Xiao, Chunxia; Wang, Yutang; Liu, Xuebo

    2012-01-01

    Isoorientin (ISO) is a flavonoid compound that can be extracted from several plant species, such as Phyllostachys pubescens, Patrinia, and Drosophyllum lusitanicum; however, its biological activity remains poorly understood. The present study investigated the effects and putative mechanism of apoptosis induced by ISO in human hepatoblastoma cancer (HepG2) cells. The results showed that ISO induced cell death in a dose-dependent manner in HepG2 cells, but no toxicity in human liver cells (HL-7702) and buffalo rat liver cells (BRL-3A) treated with ISO at the indicated concentrations. ISO-induced cell death included apoptosis which characterized by the appearance of nuclear shrinkage, the cleavage of poly (ADP-ribose) polymerase (PARP) and DNA fragmentation. ISO significantly (p < 0.01) increased the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), increased the release of cytochrome c, activated caspase-3, and enhanced intracellular levels of reactive oxygen species (ROS) and nitric oxide (NO). In addition, ISO effectively inhibited the phosphorylation of Akt and increased FoxO4 expression. The PI3K/Akt inhibitor LY294002 enhanced the apoptosis-inducing effect of ISO. However, LY294002 markedly quenched ROS and NO generation and diminished the protein expression of heme peroxidase enzyme (HO-1) and inducible nitric oxide synthase (iNOS). Furthermore, the addition of a ROS inhibitor (N-acetyl cysteine, NAC) or iNOS inhibitor (N-[3-(aminomethyl) benzyl] acetamidine, dihydrochloride, 1400W) significantly diminished the apoptosis induced by ISO and also blocked the phosphorylation of Akt. These results demonstrated for the first time that ISO induces apoptosis in HepG2 cells and indicate that this apoptosis might be mediated through mitochondrial dysfunction and PI3K/Akt signaling pathway, and has no toxicity in normal liver cells, suggesting that ISO may have good potential as a therapeutic and chemopreventive agent for liver cancer. Highlights:

  2. Effects of low priming dose irradiation on cell cycle arrest of HepG2 cells caused by high dose irradiation

    International Nuclear Information System (INIS)

    Xia Jingguang; Jin Xiaodong; Chinese Academy of Sciences, Beijing; Li Wenjian; Wang Jufang; Guo Chuanling; Gao Qingxiang

    2005-01-01

    Human hepatoma cells hepG2 were irradiated twice by 60 Co γ-rays with a priming dose of 5 cGy and a higher dose of 3 Gy performed 4h or 8h after the low dose irradiation. Effects of the priming dose irradiation on cell cycle arrest caused by high dose were examined with flow cytometry. Cells in G 2 /M phase accumulated temporarily after the 5 cGy irradiation, and proliferation of tumor cells was promoted significantly by the low dose irradiation. After the 3 Gy irradiation, G 2 phase arrest occurred, and S phase delayed temporally. In comparison with 3 kGy irradiation only, the priming dose delivered 4h prior to the high dose irradiation facilitated accumulation of hepG2 cells in G 2 /M phase, whereas the priming dose delivered 8h prior to the high dose irradiation helped the cells to overcome G 2 arrest. It was concluded that effects of the priming dose treatment on cell cycle arrest caused by high dose irradiation were dependent on time interval between the two irradiations. (authors)

  3. Zanthoxylum ailanthoides Suppresses Oleic Acid-Induced Lipid Accumulation through an Activation of LKB1/AMPK Pathway in HepG2 Cells

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    Eun-Bin Kwon

    2018-01-01

    Full Text Available Zanthoxylum ailanthoides (ZA has been used as folk medicines in East Asian and recently reported to have several bioactivity; however, the studies of ZA on the regulation of triacylglycerol (TG biosynthesis have not been elucidated yet. In this study, we examined whether the methanol extract of ZA (ZA-M could reduce oleic acid- (OA- induced intracellular lipid accumulation and confirmed its mode of action in HepG2 cells. ZA-M was shown to promote the phosphorylation of AMPK and its upstream LKB1, followed by reduction of lipogenic gene expressions. As a result, treatment of ZA-M blocked de novo TG biosynthesis and subsequently mitigated intracellular neutral lipid accumulation in HepG2 cells. ZA-M also inhibited OA-induced production of reactive oxygen species (ROS and TNF-α, suggesting that ZA-M possess the anti-inflammatory feature in fatty acid over accumulated condition. Taken together, these results suggest that ZA-M attenuates OA-induced lipid accumulation and inflammation through the activation of LKB1/AMPK signaling pathway in HepG2 cells.

  4. Active Fragment of Veronica ciliata Fisch. Attenuates t-BHP-Induced Oxidative Stress Injury in HepG2 Cells through Antioxidant and Antiapoptosis Activities

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    Yiran Sun

    2017-01-01

    Full Text Available Excessive amounts of reactive oxygen species (ROS in the body are a key factor in the development of hepatopathies such as hepatitis. The aim of this study was to assess the antioxidation effect in vitro and hepatoprotective activity of the active fragment of Veronica ciliata Fisch. (VCAF. Antioxidant assays (DPPH, superoxide, and hydroxyl radicals scavenging were conducted, and hepatoprotective effects through the application of tert-butyl hydroperoxide- (t-BHP- induced oxidative stress injury in HepG2 cells were evaluated. VCAF had high phenolic and flavonoid contents and strong antioxidant activity. From the perspective of hepatoprotection, VCAF exhibited a significant protective effect on t-BHP-induced HepG2 cell injury, as indicated by reductions in cytotoxicity and the levels of ROS, 8-hydroxydeoxyguanosine (8-OHdG, and protein carbonyls. Further study demonstrated that VCAF attenuated the apoptosis of t-BHP-treated HepG2 cells by suppressing the activation of caspase-3 and caspase-8. Moreover, it significantly decreased the levels of ALT and AST, increased the activities of acetyl cholinesterase (AChE, glutathione (GSH, superoxide dismutase (SOD, and catalase (CAT, and increased total antioxidative capability (T-AOC. Collectively, we concluded that VCAF may be a considerable candidate for protecting against liver injury owing to its excellent antioxidant and antiapoptosis properties.

  5. The presence of oleate stabilized ZnO nanoparticles (NPs) and reduced the toxicity of aged NPs to Caco-2 and HepG2 cells.

    Science.gov (United States)

    Fang, Xin; Jiang, Leying; Gong, Yu; Li, Juan; Liu, Liangliang; Cao, Yi

    2017-12-25

    The presence of food components may alter the colloidal aspects and toxicity of nanoparticles (NPs). In this study, the toxicity of ZnO NPs to Caco-2 and HepG2 cells was assessed, with the emphasis on the interactions between ZnO NPs and oleate (OA). The presence of OA increased UV-Vis spectra and hydrodynamic sizes, decreased Zeta potential, and markedly reduced the release of Zn ions from the dissolution of ZnO NPs, which combined indicated that OA could coat ZnO NPs and stabilize ZnO NPs. Exposure to ZnO NPs significantly induced cytotoxicity to Caco-2 and HepG2 cells, associated with increased intracellular Zn ions but not superoxide. When OA was added to the freshly prepared ZnO NP suspensions, the cytotoxicity, intracellular Zn ions and superoxide induced by ZnO NPs were not significantly affected. However, when ZnO NPs were aged for 24 h with the presence of OA, the cytotoxicity of ZnO NPs to Caco-2 and HepG2 cells was significantly reduced, associated with a reduction of intracellular Zn ions. The results from this study suggested that the presence of OA could increase colloidal stability of ZnO NPs and consequently reduce the toxicity of ZnO NPs after aging associated with reduced accumulation of intracellular Zn ions. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. SIRT1 attenuates palmitate-induced endoplasmic reticulum stress and insulin resistance in HepG2 cells via induction of oxygen-regulated protein 150

    Science.gov (United States)

    Jung, T.W.; Lee, K.T.; Lee, M.W.; Ka, K.H.

    2012-01-01

    Endoplasmic reticulum (ER) stress has been implicated in the pathology of type 2 diabetes mellitus (T2DM). Although SIRT1 has a therapeutic effect on T2DM, the mechanisms by which SIRT1 ameliorates insulin resistance (IR) remain unclear. In this study, we investigated the impact of SIRT1 on palmitate-induced ER stress in HepG2 cells and its underlying signal pathway. Treatment with resveratrol, a SIRT1 activator significantly inhibited palmitate-induced ER stress, leading to the protection against palmitate-induced ER stress and insulin resistance. Resveratrol and SIRT1 overexpression induced the expression of oxygen-regulated protein (ORP) 150 in HepG2 cells. Forkhead box O1 (FOXO1) was involved in the regulation of ORP150 expression because suppression of FOXO1 inhibited the induction of ORP150 by SIRT1. Our results indicate a novel mechanism by which SIRT1 regulates ER stress by overexpression of ORP150, and suggest that SIRT1 ameliorates palmitate-induced insulin resistance in HepG2 cells via regulation of ER stress.

  7. Curcumin attenuates BPA-induced insulin resistance in HepG2 cells through suppression of JNK/p38 pathways.

    Science.gov (United States)

    Geng, Shanshan; Wang, Shijia; Zhu, Weiwei; Xie, Chunfeng; Li, Xiaoting; Wu, Jieshu; Zhu, Jianyun; Jiang, Ye; Yang, Xue; Li, Yuan; Chen, Yue; Wang, Xiaoqian; Meng, Yu; Zhu, Mingming; Wu, Rui; Huang, Cong; Zhong, Caiyun

    2017-04-15

    Bisphenol A (BPA) is an artificial environmental endocrine disrupting chemicals. Accumulating evidence indicates that exposure to BPA contributes to insulin resistance through diverse mechanism including inflammation and oxidative stress. Previous studies have suggested curcumin as a safe phytochemical which can improve obesity-related insulin resistance, inflammation and oxidative stress. The present study aimed to investigate the ability of curcumin to prevent BPA-induced insulin resistance in vitro and the underlying mechanism. Following the establishmet of in vitro insulin resistance via BPA treatment in human liver HepG2 cells, the protective effects of curcumin were determiend. We showed that treatment of HepG2 cells with 100nM BPA for 5days induced significantly decreased glucose consumption, impaired insulin signaling, elevation of pro-inflammatory cytokines and oxidative stress, and activation of signaling pathways; inhibition of JNK and p38 pathways, but not ERK nor NF-κB pathways, improved glucose consumption and insulin signaling in BPA-treated HepG2 cells. Moreover, we revealed that curcumin effectively attenuated the spectrum of effects of BPA-triggered insulin resistance, whereas pretreatment with JNK and p38 agonist anisomycin could significantly compensate the effects caused by curcumin. These data illustrated the role of JNK/p38 activation in BPA-induced insulin resistance and suggested curcumin as a promising candidate for the intervention of BPA-induced insulin resistance. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Activation of farnesoid X receptor increases the expression of cytokine inducible SH2-containing protein in HepG2 cells.

    Science.gov (United States)

    Xu, Zhizhen; Huang, Gang; Gong, Wei; Zhao, Yuanyin; Zhou, Peng; Zeng, Yijun; He, Fengtian

    2012-11-01

    Cytokine inducible SH2-containing protein (CISH), which negatively regulates cytokine signaling by inhibiting JAK2/STAT5 activity, is regarded as a therapeutic target for inflammatory diseases. Farnesoid X receptor (FXR), a ligand-activated transcription factor, has been proposed to play a protective function in the inflammatory responses. However, the role of FXR in modulation of CISH expression is unknown. In the present study, we for the first time identified that in human hepatoma cell line HepG2 the activation of FXR by the natural agonist chenodeoxycholic acid (CDCA) and the synthetic specific agonist GW4064 upregulated CISH at both transcriptional and translational levels, and inhibited interleukin (IL)6-induced STAT5 activation. Moreover, the in vivo experiment demonstrated that gavaging mice with CDCA increased CISH expression and reduced basal STAT5 phosphorylation in liver tissues. Reporter assay showed that FXR agonists enhanced the transcriptional activity of CISH promoter. These data suggest that FXR may serve as a novel molecular target for manipulating CISH expression in hepatocytes. FXR-mediated upregulation of CISH may play an important role in the homeostasis of cytokine signal networks and be beneficial to control cytokine-associated inflammatory diseases.

  9. Inhibition of HBV replication by delivering the dual-gene expression vector pHsa-miR16-siRNA in HepG2.2.15 cells.

    Science.gov (United States)

    Wei, Wei; Wang, Su-Fei; Yu, Bing; Ni, Ming

    2017-12-01

    This study aimed to construct the dual-gene expression vector pHsa-miR16-siRNA which can express human miR-16 and HBV X siRNA, and examine its regulatory effect on HBV gene expression in the HepG2.2.15 cell line. The expression vectors siR-1583 and pHsa-miR16-siRNA were designed and constructed. HepG2.2.15 cells were transfected with the empty vector, siR-1583, pmiR-16 and pHsa-miR16-siRNA, respectively. ELISA was performed to measure the expression of HBsAg and HBeAg in the culture supernatant 48 and72 h post transfection. Fluorescence quantitative PCR was used to measure the HBV mRNA degradation efficiency and HBV DNA copy number. The results showed that the expression of HBV genes was significantly inhibited in HepG2.2.15 cells transfected with siR-1583, pmiR-16 and pHsa-miR16-siRNA, respectively, when compared with that in cells transfected with the empty vectors, with the inhibitory effect of pHsa-miR16-siRNA being the most significant. ELISA showed that the inhibitory rates of HBsAg and HBeAg in pHsa-miR16-siRNA transfected cells were correspondingly 87.3% and 85.0% at 48 h, and 88.6% and 86.5% at 72 h post transfection (PHBV mRNA decreased by 80.2% (t=-99.22, PHBV DNA by 92.8% (t=-73.06, PHBV DNA copy number by 89.8% (t=-47.13, PHBV more efficiently than a single-gene expression vector.

  10. SphK1 inhibitor SKI II inhibits the proliferation of human hepatoma HepG2 cells via the Wnt5A/β-catenin signaling pathway.

    Science.gov (United States)

    Liu, Hong; Zhang, Cai-Xia; Ma, Yan; He, Hong-Wei; Wang, Jia-Ping; Shao, Rong-Guang

    2016-04-15

    Sphingosine 1-phosphate (S1P) promotes cell growth, proliferation and survival. Sphingosine kinase 1 (SphK1), which converts sphingosine to S1P, is a key promoter in cancer. We previously found that the SphK1 inhibitor II (SKI II), suppresses the cell growth and induces apoptosis in human hepatoma HepG2 cells. However, the precise regulatory mechanism and signaling pathway on SKI II inhibiting tumor growth remains unknown. The expressions of β-catenin and related molecules of Wnt/β-catenin signal were detected by western blot in HepG2 cells. And the mRNA expression of β-catenin was detected by RT-PCR. The Wnt5A gene was silenced by siRNA. The colony formation was determined by staining with crystal violet. And the cell growth was examined by SRB assay and BrdU assay. We found that SKI II decreased the expression of β-catenin and the downstream molecules of β-catenin signal pathway and promotes the β-catenin degradation. In addition, SKI II induced the expression of Wnt5A, and then triggered β-catenin degradation. Furthermore, silencing Wnt5A decreased the anti-tumor effects of SKI II through recovering the expressions of β-catenin and downstream molecules of β-catenin signal pathway. SKI II-induced downregulation of HepG2 cell proliferation was associated with Wnt signaling pathway through Wnt5A-mediated β-catenin degradation. Our study revealed that a novel signal pathway was involved in SKI II-inhibited cell proliferation in human hepatoma cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Alcohol depletes coenzyme-Q(10) associated with increased TNF-alpha secretion to induce cytotoxicity in HepG2 cells.

    Science.gov (United States)

    Vidyashankar, Satyakumar; Nandakumar, Krishna S; Patki, Pralhad S

    2012-12-08

    Alcohol consumption has been implicated to cause severe hepatic steatosis which is mediated by alcohol dehydrogenase (ADH) activity and CYP(450) 2E1 expression. In this context, the effect of ethanol was studied for its influence on lipogenesis in HepG2 cell which is deficient of ADH and does not express CYP(450) 2E1. The results showed that ethanol at 100mM concentration caused 40% cytotoxicity at 72h as determined by MTT assay. The incorporation of labeled [2-(14)C] acetate into triacylglycerol and phospholipid was increased by 40% and 26% respectively upon 24h incubation, whereas incorporation of labeled [2-(14)C] acetate into cholesterol was not significantly increased. Further, ethanol inhibited HMG-CoA reductase which is a rate-limiting enzyme in the cholesterol biosynthesis. It was observed that, HMG-CoA reductase inhibition was brought about by ethanol as a consequence of decreased cell viability, since incubation of HepG2 cells with mevalonate could not increase the cholesterol content and increase the cell viability. Addition of ethanol significantly increased TNF-alpha secretion and depleted mitochondrial coenzyme-Q(10) which is detrimental for cell viability. But vitamin E (10mM) could partially restore coenzyme-Q(10) and glutathione content with decreased TNF-alpha secretion in ethanol treated cells. Further, lipid peroxidation, glutathione peroxidase and superoxide dismutase enzyme activities remained unaffected. Ethanol decreased glutathione content while, GSH/GSSG ratio was significantly higher compared to other groups showing cellular pro-oxidant and antioxidant balance remained intact. Alanine amino transferase activity was increased by 4.85 folds in cells treated with ethanol confirming hepatocyte damage. Hence, it is inferred that ethanol induced cytotoxicity in HepG2 cells due to coenzyme-Q(10) depletion and increased TNF-alpha secretion. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  12. Oroxylin A regulates glucose metabolism in response to hypoxic stress with the involvement of Hypoxia-inducible factor-1 in human hepatoma HepG2 cells.

    Science.gov (United States)

    Dai, Qinsheng; Yin, Qian; Wei, Libin; Zhou, Yuxin; Qiao, Chen; Guo, Yongjian; Wang, Xiaotang; Ma, Shiping; Lu, Na

    2016-08-01

    Metabolic alteration in cancer cells is one of the most conspicuous characteristics that distinguish cancer cells from normal cells. In this study, we investigated the influence and signaling ways of oroxylin A affecting cancer cell energy metabolism under hypoxia. The data showed that oroxylin A remarkably reduced the generation of lactate and glucose uptake under hypoxia in HepG2 cells. Moreover, oroxylin A inhibited HIF-1α expression and its stability. The downstream targets (PDK1, LDHA, and HK II), as well as their mRNA levels were also suppressed by oroxylin A under hypoxia. The silencing or the overexpression of HIF-1α assays suggested that HIF-1α is required for metabolic effect of oroxylin A in HepG2 cells during hypoxia. Furthermore, oroxylin A could reduce the expression of complex III in mitochondrial respiratory chain, and then decrease the accumulation of ROS at moderate concentrations (0-50 µM) under hypoxia, which was benefit for its inhibition on glycolytic activity by decreasing ROS-mediated HIF-1 expression. Besides, oroxylin A didn't cause the loss of MMP under hypoxia and had no obvious effects on the expression of OXPHOS complexes, suggesting that oroxylin A did not affect mitochondrial mass at the moderate stress of oroxylin A. The suppressive effect of oroxylin A on glycolysis led to a significantly repress of ATP generation, for ATP generation mostly depends on glycolysis in HepG2 cells. This study revealed a new aspect of glucose metabolism regulation of oroxylin A under hypoxia, which may contribute to its new anticancer mechanism. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  13. Effect of Mesenchymal Stem Cells and a Novel Curcumin Derivative on Notch1 Signaling in Hepatoma Cell Line

    Directory of Open Access Journals (Sweden)

    Mohamed Talaat Abdel Aziz

    2013-01-01

    Full Text Available This study was conducted to evaluate the effect of mesenchymal stem cells (MSCs and a novel curcumin derivative (NCD on HepG2 cells (hepatoma cell line and to investigate their effect on Notch1 signaling pathway target genes. HepG2 cells were divided into HepG2 control group, HepG2 cells treated with MSC conditioned medium (MSCs CM, HepG2 cells treated with a NCD, HepG2 cells treated with MSCs CM and NCD, and HepG2 cells treated with MSCs CM (CM of MSCs pretreated with a NCD. Expression of Notch1, Hes1, VEGF, and cyclin D1 was assessed by real-time, reverse transcription-polymerase chain reaction (RT-PCR in HepG2 cells. In addition, HepG2 proliferation assay was performed in all groups. Notch1 and its target genes (Hes1 and cyclin D1 were downregulated in all treated groups with more suppressive effect in the groups treated with both MSCs and NCD. Also, treated HepG2 cells showed significant decrease in cell proliferation rate. These data suggest that modulation of Notch1 signaling pathway by MSCs and/or NCD can be considered as a therapeutic target in HCC.

  14. JS-K, a nitric oxide prodrug, induces DNA damage and apoptosis in HBV-positive hepatocellular carcinoma HepG2.2.15 cell.

    Science.gov (United States)

    Liu, Zhengyun; Li, Guangmin; Gou, Ying; Xiao, Dongyan; Luo, Guo; Saavedra, Joseph E; Liu, Jie; Wang, Huan

    2017-08-01

    Hepatocellular carcinoma (HCC) is the most important cause of cancer-related death, and 85% of HCC is caused by chronic HBV infection, the prognosis of patients and the reduction of HBV DNA levels remain unsatisfactory. JS-K, a nitric oxide-releasing diazeniumdiolates, is effective against various tumors, but little is known on its effects on HBV positive HCC. We found that JS-K reduced the expression of HBsAg and HBeAg in HBV-positive HepG2.2.15 cells. This study aimed to further examine anti-tumor effects of JS-K on HepG2.2.15 cells. The MTT assay and colony forming assay were used to study the cell growth inhibition of JS-K; scratch assay and transwell assay were performed to detect cell migration. The cell cycle was detected by flow cytometry. The immunofluorescence, flow cytometry analysis, and western blot were used to study DNA damage and cell apoptosis. JS-K inhibited HepG2.2.15 cell growth in a dose-dependent manner, suppressed cell colony formation and migration, arrested cells gather in the G2 phase. JS-K (1-20μM) increased the expression of DNA damage-associated protein phosphorylation H 2 AX (γH 2 AX), phosphorylation of checkpoint kinase 1 (p-Chk1), phosphorylation of checkpoint kinase 2 (p-Chk2), ataxia-telangiectasia mutated (ATM), phosphorylation of ataxia-telangiectasia mutated rad3-related (p-ATR) and apoptotic-associated proteins cleaved caspase-3, cleaved caspase-7, cleaved poly ADP-ribose polymerase (cleaved PARP). The study demonstrated JS-K is effective against HBV-positive HepG2.2.15 cells, the mechanisms are not only related to inhibition of HBsAg and HBeAg secretion, but also related with induction of DNA damage and apoptosis. JS-K is a promising anti-cancer candidate against HBV-positive HCC. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  15. Mitochondrial aquaporin-8 knockdown in human hepatoma HepG2 cells causes ROS-induced mitochondrial depolarization and loss of viability

    Energy Technology Data Exchange (ETDEWEB)

    Marchissio, Maria Julia; Francés, Daniel Eleazar Antonio; Carnovale, Cristina Ester; Marinelli, Raúl Alberto, E-mail: rmarinel@unr.edu.ar

    2012-10-15

    Human aquaporin-8 (AQP8) channels facilitate the diffusional transport of H{sub 2}O{sub 2} across membranes. Since AQP8 is expressed in hepatic inner mitochondrial membranes, we studied whether mitochondrial AQP8 (mtAQP8) knockdown in human hepatoma HepG2 cells impairs mitochondrial H{sub 2}O{sub 2} release, which may lead to organelle dysfunction and cell death. We confirmed AQP8 expression in HepG2 inner mitochondrial membranes and found that 72 h after cell transfection with siRNAs targeting two different regions of the human AQP8 molecule, mtAQP8 protein specifically decreased by around 60% (p < 0.05). Studies in isolated mtAQP8-knockdown mitochondria showed that H{sub 2}O{sub 2} release, assessed by Amplex Red, was reduced by about 45% (p < 0.05), an effect not observed in digitonin-permeabilized mitochondria. mtAQP8-knockdown cells showed an increase in mitochondrial ROS, assessed by dichlorodihydrofluorescein diacetate (+ 120%, p < 0.05) and loss of mitochondrial membrane potential (− 80%, p < 0.05), assessed by tetramethylrhodamine-coupled quantitative fluorescence microscopy. The mitochondria-targeted antioxidant MitoTempol prevented ROS accumulation and dissipation of mitochondrial membrane potential. Cyclosporin A, a mitochondrial permeability transition pore blocker, also abolished the mtAQP8 knockdown-induced mitochondrial depolarization. Besides, the loss of viability in mtAQP8 knockdown cells verified by MTT assay, LDH leakage, and trypan blue exclusion test could be prevented by cyclosporin A. Our data on human hepatoma HepG2 cells suggest that mtAQP8 facilitates mitochondrial H{sub 2}O{sub 2} release and that its defective expression causes ROS-induced mitochondrial depolarization via the mitochondrial permeability transition mechanism, and cell death. -- Highlights: ► Aquaporin-8 is expressed in mitochondria of human hepatoma HepG2 cells. ► Aquaporin-8 knockdown impairs mitochondrial H{sub 2}O{sub 2} release and increases ROS. ► Aquaporin

  16. Alcohol depletes coenzyme-Q10 associated with increased TNF-alpha secretion to induce cytotoxicity in HepG2 cells

    International Nuclear Information System (INIS)

    Vidyashankar, Satyakumar; Nandakumar, Krishna S.; Patki, Pralhad S.

    2012-01-01

    Highlights: ► Ethanol induced cytotoxicity in HepG2 cells in absence of lipogenesis. ► Ethanol inhibited HMG-CoA reductase activity. ► Ethanol induced HMG-CoA reductase inhibition is due to decreased cell viability. ► Incubation with mevalonate could not increase the cholesterol. ► Cytotoxicity brought about by CoQ10 depletion and increased TNF-alpha. -- Abstract: Alcohol consumption has been implicated to cause severe hepatic steatosis which is mediated by alcohol dehydrogenase (ADH) activity and CYP 450 2E1 expression. In this context, the effect of ethanol was studied for its influence on lipogenesis in HepG2 cell which is deficient of ADH and does not express CYP 450 2E1. The results showed that ethanol at 100 mM concentration caused 40% cytotoxicity at 72 h as determined by MTT assay. The incorporation of labeled [2- 14 C] acetate into triacylglycerol and phospholipid was increased by 40% and 26% respectively upon 24 h incubation, whereas incorporation of labeled [2- 14 C] acetate into cholesterol was not significantly increased. Further, ethanol inhibited HMG-CoA reductase which is a rate-limiting enzyme in the cholesterol biosynthesis. It was observed that, HMG-CoA reductase inhibition was brought about by ethanol as a consequence of decreased cell viability, since incubation of HepG2 cells with mevalonate could not increase the cholesterol content and increase the cell viability. Addition of ethanol significantly increased TNF-alpha secretion and depleted mitochondrial coenzyme-Q 10 which is detrimental for cell viability. But vitamin E (10 mM) could partially restore coenzyme-Q 10 and glutathione content with decreased TNF-alpha secretion in ethanol treated cells. Further, lipid peroxidation, glutathione peroxidase and superoxide dismutase enzyme activities remained unaffected. Ethanol decreased glutathione content while, GSH/GSSG ratio was significantly higher compared to other groups showing cellular pro-oxidant and antioxidant balance remained

  17. Eicosapentaenoic acid (EPA) induced apoptosis in HepG2 cells through ROS–Ca{sup 2+}–JNK mitochondrial pathways

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yuanyuan; Han, Lirong [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Qi, Wentao [Academy of State Administration of Grain, No.11 Baiwanzhuang Avenue, Xicheng District, Beijing, 100037 (China); Cheng, Dai; Ma, Xiaolei; Hou, Lihua [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Cao, Xiaohong, E-mail: caoxh@tust.edu.cn [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Wang, Chunling, E-mail: wangchunling@tust.edu.cn [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China)

    2015-01-24

    Highlights: • EPA evoked ROS formation, [Ca{sup 2+}]{sub c} accumulation, the opening of MPTP and the phosphorylation of JNK. • EPA-induced [Ca{sup 2+}]{sub c} elevation was depended on production of ROS. • EPA-induced ROS generation, [Ca{sup 2+}]{sub c} increase, and JNK activated caused MPTP opening. • The apoptosis induced by EPA was related to release of cytochrome C through the MPTP. • EPA induced HepG2 cells apoptosis through ROS–Ca{sup 2+}–JNK mitochondrial pathways. - Abstract: Eicosapentaenoic acid (EPA), a well-known dietary n−3 PUFAS, has been considered to inhibit proliferation of tumor cells. However, the molecular mechanism related to EPA-induced liver cancer cells apoptosis has not been reported. In this study, we investigated the effect of EPA on HepG2 cells proliferation and apoptosis mechanism through mitochondrial pathways. EPA inhibited proliferation of HepG2 cells in a dose-dependent manner and had no significant effect on the cell viability of humor normal liver L-02 cells. It was found that EPA initially evoked ROS formation, leading to [Ca{sup 2+}]{sub c} accumulation and the mitochondrial permeability transition pore (MPTP) opening; EPA-induced HepG2 cells apoptosis was inhibited by N-acetylcysteine (NAC, an inhibitor of ROS), 1,2-bis (2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid (BAPTA-AM, a chelator of calcium) and CsA (inhibitor of MPTP). The relationship between ROS production, the increase of cytoplasmic Ca and MPTP opening was detected. It seems that ROS may act as an upstream regulator of EPA-induced [Ca{sup 2+}]{sub c} generation, moreover, generation of ROS, overload of mitochondrial [Ca{sup 2+}]{sub c}, and JNK activated cause the opening of MPTP. Western blotting results showed that EPA elevated the phosphorylation status of JNK, processes associated with the ROS generation. Simultaneously, the apoptosis induced by EPA was related to release of cytochrome C from mitochondria to cytoplasm through the MPTP

  18. The selective target of capsaicin on FASN expression and de novo fatty acid synthesis mediated through ROS generation triggers apoptosis in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Hathaichanok Impheng

    Full Text Available The inhibition of the mammalian de novo synthesis of long-chain saturated fatty acids (LCFAs by blocking the fatty acid synthase (FASN enzyme activity in tumor cells that overexpress FASN can promote apoptosis, without apparent cytotoxic to non-tumor cells. The present study aimed to focus on the potent inhibitory effect of capsaicin on the fatty acid synthesis pathway inducing apoptosis of capsaicin in HepG2 cells. The use of capsaicin as a source for a new FASN inhibitor will provide new insight into its possible application as a selective anti-cancer therapy. The present findings showed that capsaicin promoted apoptosis as well as cell cycle arrest in the G0/G1 phase. The onset of apoptosis was correlated with a dissipation of mitochondrial membrane potential (ΔΨm. Apoptotic induction by capsaicin was mediated by inhibition of FASN protein expression which was accompanied by decreasing its activity on the de novo fatty acid synthesis. The expression of FASN was higher in HepG2 cells than in normal hepatocytes that were resistant to undergoing apoptosis following capsaicin administration. Moreover, the inhibitory effect of capsaicin on FASN expression and activity was found to be mediated by an increase of intracellular reactive oxygen species (ROS generation. Treatment of HepG2 cells with capsaicin failed to alter ACC and ACLY protein expression, suggesting ACC and ACLY might not be the specific targets of capsaicin to induce apoptosis. An accumulation of malonyl-CoA level following FASN inhibition represented a major cause of mitochondrial-dependent apoptotic induction instead of deprivation of fatty acid per se. Here, we also obtained similar results with C75 that exhibited apoptosis induction by reducing the levels of fatty acid without any change in the abundance of FASN expression along with increasing ROS production. Collectively, our results provide novel evidence that capsaicin exhibits a potent anti-cancer property by targeting

  19. Inhibition of Cholesterol Synthesis in HepG2 Cells by GINST-Decreasing HMG-CoA Reductase Expression Via AMP-Activated Protein Kinase.

    Science.gov (United States)

    Han, Joon-Seung; Sung, Jong Hwan; Lee, Seung Kwon

    2017-11-01

    GINST, a hydrolyzed ginseng extract, has been reported to have antidiabetic effects and to reduce hyperglycemia and hyperlipidemia. Hypercholesterolemia is caused by diet or genetic factors and can lead to atherosclerosis and coronary heart disease. Thus, the purpose of this study is to determine whether GINST and the ginsenoside metabolite, IH-901 (compound K), reduce cholesterol synthesis in HepG2 cells and the signal transduction pathways involved. Concentrations of cholesterol were measured by using an enzymatic method. Expression levels of sterol regulatory element-binding protein 2 (SREBP2), HMG-CoA reductase (HMGCR), peroxisome proliferators-activated receptor γ (PPARγ), CCAAT/enhancer-binding proteins α (C/EBPα), GAPDH, and phosphorylation of AMP-activated protein kinase α (AMPKα), protein kinase B (PKB, also known as Akt), and mechanistic target of rapamycin complex 1 (mTORC1) were measured using western blot. Total cholesterol concentration decreased after GINST treatment for 24 and 48 h. Expression of HMGCR decreased more with GINST than with the inhibitors, U18666A and atorvastatin, after 48 h in a dose-dependent manner. Phosphorylation of AMPKα increased 2.5x by GINST after 360 min of treatment, and phosphorylation of Akt decreased after 120 and 360 min. We separated compound K from GINST extracts flash chromatography. Compound K decreased cholesterol synthesis in HepG2 cells at 24 and 48 h. Therefore, we conclude that GINST inhibits cholesterol synthesis in HepG2 cells by decreasing HMGCR expression via AMPKα activation. GINST, a hydrolyzed ginseng extract, can inhibit cholesterol synthesis in liver cells via activation of AMPKα. IH-901 (compound K), which is the main component with bioactivity in GINST, also has anticholesterol effects. Thus, we suggest that GINST can be used to reduce hypercholesterolemia. © 2017 Institute of Food Technologists®.

  20. Transmission of HCV to a chimpanzee using virus particles produced in an RNA-transfected HepG2 cell culture.

    Science.gov (United States)

    Dash, S; Kalkeri, G; McClure, H M; Garry, R F; Clejan, S; Thung, S N; Murthy, K K

    2001-10-01

    It was demonstrated previously that HepG2 cells produce negative strand RNA and virus-like particles after transfection with RNA transcribed from a full-length hepatitis C virus (HCV) cDNA clone [Dash et al. (1997) American Journal of Pathology, 151:363-373]. To determine in vivo infectivity of these in vitro synthesized viral particles, a chimpanzee was inoculated intravenously with HCV derived from HepG2 cells. The infected chimpanzee was examined serially for elevation of liver enzymes, for the presence of HCV RNA in the serum by reverse transcription nested polymerase chain reaction (RT-PCR), anti-HCV antibodies in the serum, and inflammation in the liver. The chimpanzee developed elevated levels of liver enzymes after the second week, but the levels fluctuated over a 10-week period. HCV RNA was detected in the serum of the chimpanzee at the second, seventh and ninth weeks after inoculation, and remained positive up to 25 weeks. Liver biopsies at Weeks 18 and 19 revealed of mild inflammation. Nucleotide sequence analysis of HCV recovered from the infected chimpanzee at the second and ninth weeks showed 100% sequence homology with the clone used for transfection studies. Serum anti-HCV antibodies were not detected by EIA during the 25 weeks follow-up period. These results suggest that intravenous administration of the virus-like particles derived from RNA-transfected HepG2 cells are infectious, and therefore, the pMO9.6-T7 clone is an infectious clone. These results provide new information that in vitro synthesized HCV particles produced from full-length HCV clone can cause infection in a chimpanzee. This study will facilitate the use of innovative approaches to the study of assembly of HCV particles and mechanisms of virus infectivity in cell culture. Copyright 2001 Wiley-Liss, Inc.

  1. Chalcones suppress fatty acid-induced lipid accumulation through a LKB1/AMPK signaling pathway in HepG2 cells.

    Science.gov (United States)

    Zhang, Tianshun; Yamamoto, Norio; Ashida, Hitoshi

    2014-06-01

    Excessive lipid accumulation in the liver has been proposed to cause hyperlipidemia, diabetes and fatty liver disease. 4-Hydroxyderricin (4HD), xanthoangelol (XAG), cardamonin (CAR) and flavokawain B (FKB) are chalcones that have exhibited various biological effects against obesity, inflammation, and diabetes; however, little is known about the inhibitory effects of these chalcones on fatty liver disease. In the present study, we investigated the ability of 4HD, XAG, CAR, and FKB to reduce lipid accumulation in hepatocytes. When HepG2 cells were treated with a mixture of fatty acids (FAs; palmitic acid : oleic acid = 1 : 2 ratio), significant lipid accumulation was observed. Under the same experimental conditions, addition of chalcones at 5 μM significantly suppressed the FA-induced lipid accumulation. We found that the expression of sterol regulatory element-binding protein-1 (SREBP-1), a key molecule involved in lipogenesis, was decreased in these chalcone-treated cells. We also found that these chalcones increased the expression of peroxisome proliferator-activated receptor α (PPARα), which is involved in FA oxidation. Moreover, these chalcones increased phosphorylation of AMP-activated protein kinase (AMPK) and liver kinase B1 (LKB1), upstream regulators of SREBP-1 and PPARα. We confirmed that an AMPK inhibitor, compound C, reversed chalcone-induced changes in SREBP-1 and PPARα expression in the HepG2 cells. Collectively, we found that 4HD, XAG, CAR, and XAG attenuated lipid accumulation through activation of the LKB1/AMPK signaling pathway in HepG2 cells.

  2. Protective Effects of Maillard Reaction Products of Whey Protein Concentrate against Oxidative Stress through an Nrf2-Dependent Pathway in HepG2 Cells.

    Science.gov (United States)

    Pyo, Min Cheol; Yang, Sung-Yong; Chun, Su-Hyun; Oh, Nam Su; Lee, Kwang-Won

    2016-09-01

    Whey protein concentrate (WPC), which contains α-lactalbumin and β-lactoglobulin, is utilized widely in the food industry. The Maillard reaction is a complex reaction that produces Maillard reaction products (MRPs), which are associated with the formation of antioxidant compounds. In this study, the hepatoprotection activity of MRPs of WPC against oxidative stress through the nuclear factor-E2-related factor 2 (Nrf2)-dependent antioxidant pathway in HepG2 cells was examined. Glucose-whey protein concentrate conjugate (Glc-WPC) was obtained from Maillard reaction between WPC and glucose. The fluorescence intensity of Glc-WPC increased after 7 d compared to native WPC, and resulted in loss of 48% of the free amino groups of WPC. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of Glc-WPC showed the presence of a high-molecular-weight portion. Treatment of HepG2 cells with Glc-WPC increased cell viability in the presence of oxidative stress, inhibited the generation of intracellular reactive oxygen species by tert-butyl hydroperoxide (t-BHP), and increased the glutathione level. Nrf2 translocation and Nrf2, reduced nicotinamide adenine dinucleotide phosphate (NAD(P)H)-quinone oxidoreductase 1 (NOQ1), heme oxygenase-1 (HO-1), glutamate-L-cysteine ligase (GCL)M and GCLC mRNA levels were increased by Glc-WPC. Also, Glc-WPC increased the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK). The results of this study demonstrate that Glc-WPC activates the Nrf2-dependent pathway through the phosphorylation of ERK1/2 and JNK in HepG2 cells, and induces production of antioxidant enzymes and phase II enzymes.

  3. Up-Regulation of CYP2C19 Expression by BuChang NaoXinTong via PXR Activation in HepG2 Cells.

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    Hong Sun

    Full Text Available Cytochrome P450 2C19 (CYP2C19 is an important drug-metabolizing enzyme (DME, which is responsible for the biotransformation of several kinds of drugs such as proton pump inhibitors, platelet aggregation inhibitors and antidepressants. Previous studies showed that Buchang NaoXinTong capsules (NXT increased the CYP2C19 metabolic activity in vitro and enhanced the antiplatelet effect of clopidogrel in vivo. However, the underlying molecular mechanism remained unclear. In the present study, we examined whether Pregnane X receptor (PXR plays a role in NXT-mediated regulation of CYP2C19 expression.We applied luciferase assays, real-time quantitative PCR (qPCR, Western blotting and cell-based analysis of metabolic activity experiments to investigate the NXT regulatory effects on the CYP2C19 promoter activity, the mRNA/ protein expression and the metabolic activity.Our results demonstrated that NXT significantly increased the CYP2C19 promoter activity when co-transfected with PXR in HepG2 cells. Mutations in PXR responsive element abolished the NXT inductive effects on the CYP2C19 promoter transcription. Additionally, NXT incubation (150 and 250μg/mL also markedly up-regulated endogenous CYP2C19 mRNA and protein levels in PXR-transfected HepG2 cells. Correspondingly, NXT leaded to a significant enhancement of the CYP2C19 catalytic activity in PXR-transfected HepG2 cells.In summary, this is the first study to suggest that NXT could induce CYP2C19 expression via PXR activation.

  4. Internalisation and multiple phosphorylation of γ-Conglutin, the lupin seed glycaemia-lowering protein, in HepG2 cells.

    Science.gov (United States)

    Capraro, Jessica; Magni, Chiara; Faoro, Franco; Maffi, Dario; Scarafoni, Alessio; Tedeschi, Gabriella; Maffioli, Elisa; Parolari, Anna; Manzoni, Cristina; Lovati, Maria Rosa; Duranti, Marcello

    2013-08-09

    Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing glucose uptake by model cells. This work investigated whether γ-Conglutin is internalised into the target cells and undergoes any covalent change during the process, as a first step to understanding its mechanism of action. To this purpose, γ-Conglutin-treated and untreated HepG2 cells were submitted to confocal and transmission electron microscopy. Immune-revelation of γ-Conglutin at various intervals revealed its accumulation inside the cytosol. In parallel, 2D-electrophoresis of the cell lysates and antibody reaction of the blotted maps showed the presence of the protein intact subunits inside the treated cells, whilest no trace of the protein was found in the control cells. However, γ-Conglutin-related spots with an unexpectedly low pI were also observed in the maps. These spots were excised, trypsin-treated and submitted to MS/MS spectrometric analysis. The presence of phosphorylated amino acids was detected. These findings, by showing that γ-Conglutin is internalised by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to the understanding of the lupin γ-Conglutin insulin-mimetic activity. Copyright © 2013 Elsevier Inc. All rights reserved.

  5. Hugan Qingzhi medication ameliorates hepatic steatosis by activating AMPK and PPARα pathways in L02 cells and HepG2 cells.

    Science.gov (United States)

    Yin, JinJin; Luo, YanQin; Deng, HouLiang; Qin, ShuMin; Tang, WaiJiao; Zeng, Lu; Zhou, BenJie

    2014-05-28

    Hugan Qingzhi tablet (HQT), a lipid- lowering traditional Chinese medicine formula, has been used for the prevention and treatment of nonalcoholic fatty liver (NAFLD). This study was realized to evaluate the effects of HQT-medicated serum on hepatic steatosis using in vitro experiments with cells and explore the relevant mechanisms with method of serum pharmacology. A model of hepatic steatosis in the L02 and HepG2 cells was induced by free fatty acid (FFA). The components in the HQT-medicated serum were assayed by high-performance liquid chromatography. Intracellular lipid droplets were detected by Oil Red O staining, and their ultrastructure was examined by transmission electron microscope. The biochemical parameters, including triglyceride (TG), lactate dehydrogenase (LDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT), total antioxidant capacity (T-AOC), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione (GSH), were measured with commercial kits. Furthermore, the expression of adiponectin, AMP-activated protein kinase (AMPK) phosphorylation, sterol regulatory element-binding protein 1 (SREBP-1), peroxisome proliferator activated receptor-α (PPARα), carnitine palmitoyltransferase 1 (CPT-1), and acetyl-CoA oxidase 1 (ACOX1) was analyzed by Western blot and/or quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Moderate- and high-dose HQT-medicated serum reduced (Pcontent in L02 and HepG2 cells. They caused significant reductions (P<0.01) in LDH, AST, ALT and MDA and significant increase (P<0.05 or P<0.01) in T-AOC in the culture medium. They also caused increase (P<0.05 or P<0.01) in GSH level and SOD activity in FFA-induced steatotic L02 and HepG2 cells. Furthermore, moderate- and high-dose HQT-medicated serum enhanced (P<0.01) adiponectin expression in a concentration-dependent manner and increased (P<0.05 or P<0.01) the phosphorylation of AMPK and the expression of PPARα, CPT-1, and ACOX1, and reduced

  6. Anticancer Effects of 1,3-Dihydroxy-2-Methylanthraquinone and the Ethyl Acetate Fraction of Hedyotis Diffusa Willd against HepG2 Carcinoma Cells Mediated via Apoptosis.

    Directory of Open Access Journals (Sweden)

    Yun-Lan Li

    Full Text Available Hedyotis Diffusa Willd, used in Traditional Chinese Medicine, is a treatment for various diseases including cancer, owing to its mild effectiveness and low toxicity. The aim of this study was to identify the main anticancer components in Hedyotis Diffusa Willd, and explore mechanisms underlying their activity. Hedyotis Diffusa Willd was extracted and fractionated using ethyl acetate to obtain the H-Ethyl acetate fraction, which showed higher anticancer activity than the other fractions obtained against HepG2 cells with sulforhodamine B assays. The active component of the H-Ethyl acetate fraction was identified to be 1,3-dihydroxy-2-methylanthraquinone (DMQ with much high inhibitory rate up to 48.9 ± 3.3% and selectivity rate up to 9.4 ± 4.5 folds (p<0.01 at 125 μmol/L. HepG2 cells treated with the fraction and DMQ visualized morphologically using light and fluorescence microscopy. Annexin V--fluorescein isothiocyanate / propidium iodide staining flow cytometry, DNA ladder and cell cycle distribution assays. Mechanistic studies showed up-regulation of caspase-3, -8, and -9 proteases activities (p<0.001, indicating involvement of mitochondrial apoptotic and death receptor pathways. Further studies revealed that reactive oxygen species in DMQ and the fraction treated HepG2 cells increased (p<0.01 while mitochondrial membrane potential reduced significantly (p<0.001 compared to the control by flow cytometry assays. Western blot analysis showed that Bax, p53, Fas, FasL, p21 and cytoplasmic cytochrome C were up-regulated (p<0.01, while Bcl-2, mitochondrial cytochrome C, cyclin E and CDK 2 were down-regulated dose-dependently (p<0.01. The reverse transcriptase-polymerase chain reaction showed that mRNA expressions of p53 and Bax increased (p<0.001 while that of Bcl-2 decreased (p<0.001. Pre-treatment with caspase-8 inhibitor Z-IETD-FMK, or caspase-9 inhibitor Z-LEHD-FMK, attenuated the growth-inhibitory and apoptosis-inducing effects of DMQ and the

  7. Palmitic acid suppresses apolipoprotein M gene expression via the pathway of PPAR{sub β/δ} in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Guanghua; Shi, Yuanping; Zhang, Jun; Mu, Qinfeng; Qin, Li; Zheng, Lu; Feng, Yuehua [Comprehensive Laboratory, The Third Affiliated Hospital of Soochow University, Changzhou 213003 (China); Berggren-Söderlund, Maria; Nilsson-Ehle, Peter [Division of Clinical Chemistry and Pharmacology, Department of Laboratory Medicine, Lund University, S-221 85 Lund (Sweden); Zhang, Xiaoying, E-mail: zhangxy6689996@163.com [Department of Cardiothoracic Surgery, The Third Affiliated Hospital of Soochow University, Changzhou 213003 (China); Xu, Ning, E-mail: ning.xu@med.lu.se [Division of Clinical Chemistry and Pharmacology, Department of Laboratory Medicine, Lund University, S-221 85 Lund (Sweden)

    2014-02-28

    Highlights: • Palmitic acid significantly inhibited APOM gene expression in HepG2 cells. • Palmitic acid could obviously increase PPARB/D mRNA levels in HepG2 cells. • PPAR{sub β/δ} antagonist, GSK3787, had no effect on APOM expression. • GSK3787 could reverse the palmitic acid-induced down-regulation of APOM expression. • Palmitic acid induced suppression of APOM expression is mediated via the PPAR{sub β/δ} pathway. - Abstract: It has been demonstrated that apolipoprotein M (APOM) is a vasculoprotective constituent of high density lipoprotein (HDL), which could be related to the anti-atherosclerotic property of HDL. Investigation of regulation of APOM expression is of important for further exploring its pathophysiological function in vivo. Our previous studies indicated that expression of APOM could be regulated by platelet activating factor (PAF), transforming growth factors (TGF), insulin-like growth factor (IGF), leptin, hyperglycemia and etc., in vivo and/or in vitro. In the present study, we demonstrated that palmitic acid could significantly inhibit APOM gene expression in HepG2 cells. Further study indicated neither PI-3 kinase (PI3K) inhibitor LY294002 nor protein kinase C (PKC) inhibitor GFX could abolish palmitic acid induced down-regulation of APOM expression. In contrast, the peroxisome proliferator-activated receptor beta/delta (PPAR{sub β/δ}) antagonist GSK3787 could totally reverse the palmitic acid-induced down-regulation of APOM expression, which clearly demonstrates that down-regulation of APOM expression induced by palmitic acid is mediated via the PPAR{sub β/δ} pathway.

  8. Effect of diphenyl ether herbicides and oxadiazon on porphyrin biosynthesis in mouse liver, rat primary hepatocyte culture and HepG2 cells.

    Science.gov (United States)

    Krijt, J; van Holsteijn, I; Hassing, I; Vokurka, M; Blaauboer, B J

    1993-01-01

    The effects of the herbicides fomesafen, oxyfluorfen, oxadiazon and fluazifop-butyl on porphyrin accumulation in mouse liver, rat primary hepatocyte culture and HepG2 cells were investigated. Ten days of herbicide feeding (0.25% in the diet) increased the liver porphyrins in male C57B1/6J mice from 1.4 +/- 0.6 to 4.8 +/- 2.1 (fomesafen) 16.9 +2- 2.9 (oxyfluorfen) and 25.9 +/- 3.1 (oxadiazon) nmol/g wet weight, respectively. Fluazifop-butyl had no effect on liver porphyrin metabolism. Fomesafen, oxyfluorfen and oxadiazon increased the cellular porphyrin content of rat hepatocytes after 24 h of incubation (control, 3.2 pmol/mg protein, fomesafen, oxyfluorfen and oxadiazon at 0.125 mM concentration 51.5, 54.3 and 44.0 pmol/mg protein, respectively). The porphyrin content of HepG2 cells increased from 1.6 to 18.2, 10.6 and 9.2 pmol/mg protein after 24 h incubation with the three herbicides. Fluazifop-butyl increased hepatic cytochrome P450 levels and ethoxy- and pentoxyresorufin O-dealkylase (EROD and PROD) activity, oxyfluorfen increased PROD activity. Peroxisomal palmitoyl CoA oxidation increased after fomesafen and fluazifop treatment to about 500% of control values both in mouse liver and rat hepatocytes. Both rat hepatocytes and HepG2 cells can be used as a test system for the porphyrogenic potential of photobleaching herbicides.

  9. Anti-hepatitis B virus activity of Boehmeria nivea leaf extracts in human HepG2.2.15 cells

    Science.gov (United States)

    WEI, JINGCHEN; LIN, LIANKU; SU, XIAOJIAN; QIN, SHAOYAN; XU, QING; TANG, ZUNIAN; DENG, YAN; ZHOU, YUEHAN; HE, SONGQING

    2014-01-01

    Boehmeria nivea (Linn.) Gaudich of the Urticaceae family is a perennial ratoon herbal plant, the root of which is used in traditional Chinese medicine and possesses a variety of pharmacological properties. The 20% ethanol Boehmeria nivea root extract was shown to exert an anti-hepatitis B virus (HBV) effect in vitro and in vivo; however, whether the Boehmeria nivea leaf (BNL) extract possesses similar properties has not been determined. In this study, we aimed to investigate the anti-HBV effects of the BNL extract in HepG2.2.15 cells transfected with human HBV DNA. Our results demonstrated that the secretion of HBsAg and HBeAg was reduced in HepG2.2.15 cells treated with the BNL extract, without any recorded cytotoxic effects. In addition, the chloroform fraction (CF) and ethyl acetate fraction (EAF) of BNL were shown to be more potent compared to the other fractions: CF (100 mg/l) inhibited the secretion of HBsAg by 94.00±1.78% [inhibitory concentration 50 (IC50) = 20.92 mg/l] and that of HBeAg by 100.19±0.35% (IC50=19.67 mg/l) after 9 days of treatment. Similarly, EAF (200 mg/l) inhibited the secretion of HBsAg by 89.95±2.26% (IC50=39.90 mg/l) and that of HBeAg by 98.90±1.42% (IC50=36.45 mg/l). Furthermore, we observed that the content of HBV DNA in the medium secreted by the HepG2.2.15 cells was significantly decreased under CF (100 mg/l) or EAF (200 mg/l) treatment. Thus, we concluded that the BNL extracts exhibited anti-HBV activity, with CF and EAF being the most potent among the fractions. PMID:24649087

  10. The species origin of the serum in the culture medium influences the in vitro toxicity of silica nanoparticles to HepG2 cells.

    Science.gov (United States)

    Pisani, Cédric; Rascol, Estelle; Dorandeu, Christophe; Gaillard, Jean-Charles; Charnay, Clarence; Guari, Yannick; Chopineau, Joël; Armengaud, Jean; Devoisselle, Jean-Marie; Prat, Odette

    2017-01-01

    The formation of a protein corona around nanoparticles can influence their toxicity, triggering cellular responses that may be totally different from those elicited by pristine nanoparticles. The main objective of this study was to investigate whether the species origin of the serum proteins forming the corona influences the in vitro toxicity assessment of silica nanoparticles. Coronas were preformed around nanoparticles before cell exposures by incubation in fetal bovine (FBS) or human (HS) serum. The compositions of these protein coronas were assessed by nano-LC MS/MS. The effects of these protein-coated nanoparticles on HepG2 cells were monitored using real-time cell impedance technology. The nanoparticle coronas formed in human or fetal bovine serum comprised many homologous proteins. Using human compared with fetal bovine serum, nanoparticle toxicity in HepG2 cells decreased by 4-fold and 1.5-fold, when used at 50 and 10μg/mL, respectively. It is likely that "markers of self" are present in the serum and are recognized by human cell receptors. Preforming a corona with human serum seems to be more appropriate for in vitro toxicity testing of potential nanocarriers using human cells. In vitro cytotoxicity assays must reflect in vivo conditions as closely as possible to provide solid and useful results.

  11. Predictivity of dog co-culture model, primary human hepatocytes and HepG2 cells for the detection of hepatotoxic drugs in humans

    Energy Technology Data Exchange (ETDEWEB)

    Atienzar, Franck A., E-mail: franck.atienzar@ucb.com [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Novik, Eric I. [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Gerets, Helga H. [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Parekh, Amit [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Delatour, Claude; Cardenas, Alvaro [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); MacDonald, James [Chrysalis Pharma Consulting, LLC, 385 Route 24, Suite 1G, Chester, NJ 07930 (United States); Yarmush, Martin L. [Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854 (United States); Dhalluin, Stéphane [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium)

    2014-02-15

    Drug Induced Liver Injury (DILI) is a major cause of attrition during early and late stage drug development. Consequently, there is a need to develop better in vitro primary hepatocyte models from different species for predicting hepatotoxicity in both animals and humans early in drug development. Dog is often chosen as the non-rodent species for toxicology studies. Unfortunately, dog in vitro models allowing long term cultures are not available. The objective of the present manuscript is to describe the development of a co-culture dog model for predicting hepatotoxic drugs in humans and to compare the predictivity of the canine model along with primary human hepatocytes and HepG2 cells. After rigorous optimization, the dog co-culture model displayed metabolic capacities that were maintained up to 2 weeks which indicates that such model could be also used for long term metabolism studies. Most of the human hepatotoxic drugs were detected with a sensitivity of approximately 80% (n = 40) for the three cellular models. Nevertheless, the specificity was low approximately 40% for the HepG2 cells and hepatocytes compared to 72.7% for the canine model (n = 11). Furthermore, the dog co-culture model showed a higher superiority for the classification of 5 pairs of close structural analogs with different DILI concerns in comparison to both human cellular models. Finally, the reproducibility of the canine system was also satisfactory with a coefficient of correlation of 75.2% (n = 14). Overall, the present manuscript indicates that the dog co-culture model may represent a relevant tool to perform chronic hepatotoxicity and metabolism studies. - Highlights: • Importance of species differences in drug development. • Relevance of dog co-culture model for metabolism and toxicology studies. • Hepatotoxicity: higher predictivity of dog co-culture vs HepG2 and human hepatocytes.

  12. Predictivity of dog co-culture model, primary human hepatocytes and HepG2 cells for the detection of hepatotoxic drugs in humans

    International Nuclear Information System (INIS)

    Atienzar, Franck A.; Novik, Eric I.; Gerets, Helga H.; Parekh, Amit; Delatour, Claude; Cardenas, Alvaro; MacDonald, James; Yarmush, Martin L.; Dhalluin, Stéphane

    2014-01-01

    Drug Induced Liver Injury (DILI) is a major cause of attrition during early and late stage drug development. Consequently, there is a need to develop better in vitro primary hepatocyte models from different species for predicting hepatotoxicity in both animals and humans early in drug development. Dog is often chosen as the non-rodent species for toxicology studies. Unfortunately, dog in vitro models allowing long term cultures are not available. The objective of the present manuscript is to describe the development of a co-culture dog model for predicting hepatotoxic drugs in humans and to compare the predictivity of the canine model along with primary human hepatocytes and HepG2 cells. After rigorous optimization, the dog co-culture model displayed metabolic capacities that were maintained up to 2 weeks which indicates that such model could be also used for long term metabolism studies. Most of the human hepatotoxic drugs were detected with a sensitivity of approximately 80% (n = 40) for the three cellular models. Nevertheless, the specificity was low approximately 40% for the HepG2 cells and hepatocytes compared to 72.7% for the canine model (n = 11). Furthermore, the dog co-culture model showed a higher superiority for the classification of 5 pairs of close structural analogs with different DILI concerns in comparison to both human cellular models. Finally, the reproducibility of the canine system was also satisfactory with a coefficient of correlation of 75.2% (n = 14). Overall, the present manuscript indicates that the dog co-culture model may represent a relevant tool to perform chronic hepatotoxicity and metabolism studies. - Highlights: • Importance of species differences in drug development. • Relevance of dog co-culture model for metabolism and toxicology studies. • Hepatotoxicity: higher predictivity of dog co-culture vs HepG2 and human hepatocytes

  13. Protective Effects of Sweet Orange, Unshiu Mikan, and Mini Tomato Juice Powders on t-BHP-Induced Oxidative Stress in HepG2 Cells

    Science.gov (United States)

    Jannat, Susoma; Ali, Md Yousof; Kim, Hyeung-Rak; Jung, Hyun Ah; Choi, Jae Sue

    2016-01-01

    The aim of this study was to investigate the protective effects of juice powders from sweet orange [Citrus sinensis (L.) Osbeck], unshiu mikan (Citrus unshiu Marcow), and mini tomato (Solanum lycopersicum L.), and their major flavonoids, hesperidin, narirutin, and rutin in tert-butyl hydroperoxide (t-BHP)-induced oxidative stress in HepG2 cells. The increased reactive oxygen species and decreased glutathione levels observed in t-BHP-treated HepG2 cells were ameliorated by pretreatment with juice powders, indicating that the hepatoprotective effects of juice powders and their major flavonoids are mediated by induction of cellular defense against oxidative stress. Moreover, pretreatment with juice powders up-regulated phase-II genes such as heme oxygenase-1 (HO-1), thereby preventing cellular damage and the resultant increase in HO-1 expression. The high-performance liquid chromatography profiles of the juice powders confirmed that hesperidin, narirutin, and rutin were the key flavonoids present. Our results suggest that these fruit juice powders and their major flavonoids provide a significant cytoprotective effect against oxidative stress, which is most likely due to the flavonoid-related bioactive compounds present, leading to the normal redox status of cells. Therefore, these fruit juice powders could be advantageous as bioactive sources for the prevention of oxidative injury in hepatoma cells. PMID:27752497

  14. Antiproliferative Effect of the Isoquinoline Alkaloid Papaverine in Hepatocarcinoma HepG-2 Cells — Inhibition of Telomerase and Induction of Senescence

    Directory of Open Access Journals (Sweden)

    Sakineh Kazemi Noureini

    2014-08-01

    Full Text Available Cancer cells are often immortal through up-regulation of the hTERT gene, which encodes the catalytic subunit of a special reverse transcriptase to overcome end-replication problem of chromosomes. This study demonstrates that papaverine, an isoquinoline alkaloid from the Papaveraceae, can overcome telomerase dependent immortality of HepG-2 cells that was used as a model of hepatocarcinoma. Although this alkaloid does not directly interact with telomeric sequences, papaverine inhibits telomerase through down-regulation of hTERT, which was analysed using thermal FRET and qRT-PCR, respectively. The IC50 values for the reduction of both telomerase activity and hTERT expression was 60 µM, while IC50 for cytotoxicity was 120 µM. Repeated treatments of the cells with very low non-toxic concentrations of papaverine resulted in growth arrest and strong reduction of population doublings after 40 days. This treatment induced senescent morphology in HepG-2 cells, which was evaluated by beta-galactosidase staining. Altogether, papaverine can be regarded as a promising model compound for drug design targeting cancer development.

  15. Strawberry (cv. Romina Methanolic Extract and Anthocyanin-Enriched Fraction Improve Lipid Profile and Antioxidant Status in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Tamara Y. Forbes-Hernández

    2017-05-01

    Full Text Available Dyslipidemia and oxidation of low density lipoproteins (LDL are recognized as critical factors in the development of atherosclerosis. Healthy dietary patterns, with abundant fruit and vegetable consumption, may prevent the onset of these risk factors due to the presence of phytochemical compounds. Strawberries are known for their high content of polyphenols; among them, flavonoids are the major constituents, and it is presumed that they are responsible for the biological activity of the fruit. Nevertheless, there are only a few studies that actually evaluate the effects of different fractions isolated from strawberries. In order to assess the effects of two different strawberry extracts (whole methanolic extract/anthocyanin-enriched fraction on the lipid profile and antioxidant status in human hepatocellular carcinoma (HepG2 cells, the triglycerides and LDL-cholesterol content, lipid peroxidation, intracellular reactive oxygen species (ROS content and antioxidant enzymes’ activity on cell lysates were determined. Results demonstrated that both strawberry extracts not only improved the lipid metabolism by decreasing triglycerides and LDL-cholesterol contents, but also improved the redox state of HepG2 cells by modulating thiobarbituric acid-reactive substances production, antioxidant enzyme activity and ROS generation. The observed effects were more pronounced for the anthocyanin-enriched fraction.

  16. Comparison of the lipid-lowering effects of four different n-3 highly unsaturated fatty acids in HepG2 cells.

    Science.gov (United States)

    Nagao, Koji; Nakamitsu, Kazusa; Ishida, Hiroki; Yoshinaga, Kazuaki; Nagai, Toshiharu; Mizobe, Hoyo; Kojima, Koichi; Yanagita, Teruyoshi; Beppu, Fumiaki; Gotoh, Naohiro

    2014-01-01

    The effects on lipid metabolism of four different n-3 highly unsaturated fatty acids (n-3HUFA) including eicosapentaenoic acid (EPA, 20:5n-3), docosapentaenoic acid (DPA, 22:5n-3), docosahexaenoic acid (DHA, 22:6n-3), and tetracosahexaenoic acid (THA, 24:6n-3) were compared in the HepG2 cell model. None of the n-3HUFAs affected the viability of the cells. THA exerted the strongest suppression on the synthesis of triacylglycerol and cholesteryl ester (ChE), and the order of the strength of suppression was found to be THA > DHA > DPA > EPA. The mRNA level of fatty acid synthase was suppressed by the n-3HUFAs and the order of the strength of suppression by n-3HUFAs was the same in both triacylglycerol and ChE synthesis. These findings support previous animal test results using EPA, DPA, and DHA. In conclusion, both the number of carbon atoms and double bonds in an n-3HUFA structure has an effect on lipid metabolism in HepG2 cells.

  17. Selected cholesterol biosynthesis inhibitors produce accumulation of the intermediate FF-MAS that targets nucleus and activates LXRα in HepG2 cells.

    Science.gov (United States)

    Gatticchi, Leonardo; Cerra, Bruno; Scarpelli, Paolo; Macchioni, Lara; Sebastiani, Bartolomeo; Gioiello, Antimo; Roberti, Rita

    2017-09-01

    Sterol intermediates of the cholesterol biosynthetic pathway have drawn attention for novel biological activities. Follicular fluid meiosis activating sterol (FF-MAS) is a LXRα ligand and a potential modulator of physiologic processes regulated by nuclear receptors, such as lipid homeostasis and cell proliferation. In this work, we established a model to selectively accumulate FF-MAS in HepG2 cells, by using a combination of the inhibitors AY9944 and 17-hydroxyprogesterone to block C14-sterol reductases and the downstream C4-demethylase complex. We investigated the effects produced by altered levels of cholesterol biosynthesis intermediates, in order to dissect their influence on LXRα signaling. In particular, endogenously accumulated FF-MAS was able to modulate the expression of key genes in cholesterol metabolism, to activate LXRα nuclear signaling resulting in increased lipogenesis, and to inhibit HepG2 cells proliferation. Moreover, a fluorescent ester derivative of FF-MAS localized in nuclear lipid droplets, suggesting a role for these organelles in the storage of signaling lipids interacting with nuclear partners. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Wild Edible Mushrooms from Turkey as Possible Anticancer Agents on HepG2 Cells Together with Their Antioxidant and Antimicrobial Properties.

    Science.gov (United States)

    Sadi, Gokhan; Kaya, Abdullah; Yalcin, Hicret Asli; Emsen, Bugrahan; Kocabas, Aytac; Kartal, Deniz Irtem; Altay, Ahmet

    2016-01-01

    This study was designed to reveal cell growth inhibitory potential of six different edible mushrooms: Ramaria flava, Agrocybe molesta, Volvopluteus gloiocephalus, Lactarius deliciosus, Bovista plumbea, and Tricholoma terreum on HepG2 cells together with their antioxidant and antibacterial power. Methanolic extracts of V gloiocephalus and aqueous extracts of R. flava had the most potential cytotoxic effects over HepG2 cells. The best results for 2,2-diphenyl-1-picrylhydrazyl radical scavenging activities were obtained from both aqueous and methanolic extracts of R. flava. Methanolic extracts of T. terreum (IC50 = 1.62 mg/mL) and aqueous extracts of B. plumbea (IC50 = 0.49 mg/mL) showed maximum metal chelating activity. The highest reducing capacities were observed among the methanolic extracts of R. flava (EC50 = 1.65 mg/mL) and aqueous extracts of B. plumbea (EC50 = 1.71 mg/ mL). High-performance liquid chromatography analysis revealed the presence of many phenolic compounds in macrofungi; gallic acid and p-coumaric acid were the two main phenolics identified in all extracts. Antibacterial studies indicated that all six tested mushrooms showed antibacterial activity on at least three microorganisms. These results indicate that different extracts of the investigated mushrooms have considerable cytotoxic, antioxidant, and antibacterial properties and may be utilized as a promising source of therapeutics.

  19. Cocoa flavonoids attenuate high glucose-induced insulin signalling blockade and modulate glucose uptake and production in human HepG2 cells.

    Science.gov (United States)

    Cordero-Herrera, Isabel; Martín, María Ángeles; Goya, Luis; Ramos, Sonia

    2014-02-01

    Insulin resistance is the primary characteristic of type 2 diabetes. Cocoa and its main flavanol, (-)-epicatechin (EC), display some antidiabetic effects, but the mechanisms for their preventive activities related to glucose metabolism and insulin signalling in the liver remain largely unknown. In the present work, the preventive effect of EC and a cocoa polyphenolic extract (CPE) on insulin signalling and on both glucose production and uptake are studied in insulin-responsive human HepG2 cells treated with high glucose. Pre-treatment of cells with EC or CPE reverted decreased tyrosine-phosphorylated and total levels of IR, IRS-1 and -2 triggered by high glucose. EC and CPE pre-treatment also prevented the inactivation of the PI3K/AKT pathway and AMPK, as well as the diminution of GLUT-2 levels induced by high glucose. Furthermore, pre-treatment of cells with EC and CPE avoided the increase in PEPCK levels and the diminished glucose uptake provoked by high glucose, returning enhanced levels of glucose production and decreased glycogen content to control values. These findings suggest that EC and CPE improved insulin sensitivity of HepG2 treated with high glucose, preventing or delaying a potential hepatic dysfunction through the attenuation of the insulin signalling blockade and the modulation of glucose uptake and production. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Salmonella typhimurium strain SL7207 induces apoptosis and inhibits the growth of HepG2 hepatoma cells in vitro and in vivo

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    Baowei Li

    2012-12-01

    Full Text Available Salmonella typhimurium is probably most extensively studied tumor-targeting bacteria and SL7207 is one of its attenuated strains. SL7207 was first made for bacterial vaccine development and its therapeutic efficacy and safety for hepatocellular carcinoma has not been characterized. In this study, the inhibitory ability of SL7207-lux on human hepatoma HepG2 cells was tested in vitro and in vivo. A bacterial luminescent gene cluster (lux CDABE was transfected into SL7207 to better monitor the invasion of the bacteria. The results show that SL7207-lux can rapidly enter HepG2 cells and localize in the cytoplasm. This invasion represses cell proliferation and induces apoptosis. In vivo real-time invasion studies showed that the bacteria gradually accumulate in the tumor. This enrichment was confirmed by anatomic observation at 5 days after inoculation. About 40% of tumor growth was inhibited by SL7207-lux at 34 days post-treatment without significant loss of body weight. The area of necrosis of tumor tissue was clearly increased in the treated group. Bacterial quantification showed that the number of colony-forming units per gram of bacteria within tumor tissue was approximately 1000-fold higher than that of liver and spleen. These data suggest that attenuated S. typhimurium strain SL7207 has potential for the treatment of cancers.

  1. Antioxidative Effects of Germinated Brown Rice-Derived Extracts on H2O2-Induced Oxidative Stress in HepG2 Cells

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    Nur Diyana Md Zamri

    2014-01-01

    Full Text Available The antioxidant properties of germinated brown rice (GBR are likely mediated by multiple bioactives. To test this hypothesis, HepG2 cells pretreated with GBR extracts, rich in acylated steryl glycoside (ASG, gamma amino butyric acid GABA, phenolics or oryzanol, were incubated with hydrogen peroxide (H2O2 and their hydroxyl radical (OH• scavenging capacities and thiobarbituric acid-reactive substances (TBARS generation were evaluated. Results showed that GBR-extracts increased OH• scavenging activities in both cell-free medium and posttreatment culture media, suggesting that the extracts were both direct- and indirect-acting against OH•. The levels of TBARS in the culture medium after treatment were also reduced by all the extracts. In addition, H2O2 produced transcriptional changes in p53, JNK, p38 MAPK, AKT, BAX, and CDK4 that were inclined towards apoptosis, while GBR-extracts showed some transcriptional changes (upregulation of BAX and p53 that suggested an inclination for apoptosis although other changes (upregulation of antioxidant genes, AKT, JNK, and p38 MAPK suggested that GBR-extracts favored survival of the HepG2 cells. Our findings show that GBR bioactive-rich extracts reduce oxidative stress through improvement in antioxidant capacity, partly mediated through transcriptional regulation of antioxidant and prosurvival genes.

  2. Effect of Cudrania tricuspidata and Kaempferol in Endoplasmic Reticulum Stress-Induced Inflammation and Hepatic Insulin Resistance in HepG2 Cells.

    Science.gov (United States)

    Kim, Ok-Kyung; Jun, Woojin; Lee, Jeongmin

    2016-01-21

    In this study, we quantitated kaempferol in water extract from Cudrania tricuspidata leaves (CTL) and investigated its effects on endoplasmic reticulum (ER) stress-induced inflammation and insulin resistance in HepG2 cells. The concentration of kaempferol in the CTL was 5.07 ± 0.08 mg/g. The HepG2 cells were treated with 300 µg/mL of CTL, 500 µg/mL of CTL, 1.5 µg/mL of kaempferol or 2.5 µg/mL of kaempferol, followed immediately by stimulation with 100 nM of thapsigargin for ER stress induction for 24 h. There was a marked increase in the activation of the ER stress and inflammation response in the thapsigargin-stimulated control group. The CTL treatment interrupted the ER stress response and ER stress-induced inflammation. Kaempferol partially inhibited the ER stress response and inflammation. There was a significant increase in serine phosphorylation of insulin receptor substrate (IRS)-1 and the expression of C/EBPα and gluconeogenic genes in the thapsigargin-stimulated control group compared to the normal control. Both CTL and kaempferol suppressed serine phosphorylation of IRS-1, and the treatments did not interrupt the C/EBPα/gluconeogenic gene pathway. These results suggest that kaempferol might be the active compound of CTL and that it might protect against ER stress-induced inflammation and hyperglycemia.

  3. Protective effect of polysaccharide from maca (Lepidium meyenii) on Hep-G2 cells and alcoholic liver oxidative injury in mice.

    Science.gov (United States)

    Zhang, Lijun; Zhao, Qingsheng; Wang, Liwei; Zhao, Mingxia; Zhao, Bing

    2017-06-01

    To study the characterization and hepatoprotective activity of polysaccharide from maca (Lepidium meyenii), the main polysaccharide from maca (MP-1) was obtained by DEAE-52 cellulose column. The average molecular weight of MP-1 was 1067.3kDa and the polysaccharide purity was 91.63%. In order to assess the antioxidant activities of MP-1, four kinds of methods were used, including scavenging hydroxyl radical, DPPH, superoxide anion radical, and FRAP, and the results indicated high antioxidant activities. Furthermore, hepatoprotective activity of MP-1 was studied both in vitro and vivo. In vitro, the alcohol induced Hep-G2 cells model was established to evaluate the protective effect of MP-1, which demonstrated MP-1 can alleviate alcohol damage in Hep-G2 cells. In vivo, the Institute of Cancer Researcch (ICR) mice were used to evaluate hepatoprotecive effects of MP-1 on alcoholic liver disease (ALD). Supplement with MP-1 supressed the triglyceride level both in serum and in hepatic tissue. In addition, MP-1 ameliorated serous transaminases increase induced by alcohol, including aspartate transaminase, alanine aminotransferase, and γ-glutamyl transpeptidase. Moreover, MP-1 also dramatically increased the superoxide dismutase, glutathione peroxidase, and glutathione s-transferase levels in alcoholic mice. Meantime, histopathologic results MP-1 lighten inflammation induced by alcohol. These results indicate that MP-1 possesses hepatoprotective activity against hepatic injury induced by alcohol. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Fucoidan from Fucus vesiculosus Protects against Alcohol-Induced Liver Damage by Modulating Inflammatory Mediators in Mice and HepG2 Cells

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    Jung Dae Lim

    2015-02-01

    Full Text Available Fucoidan is an l-fucose-enriched sulfated polysaccharide isolated from brown algae and marine invertebrates. In this study, we investigated the protective effect of fucoidan from Fucus vesiculosus on alcohol-induced murine liver damage. Liver injury was induced by oral administration of 25% alcohol with or without fucoidan (30 mg/kg or 60 mg/kg for seven days. Alcohol administration increased serum aspartate aminotransferase and alanine aminotransferase levels, but these increases were suppressed by the treatment of fucoidan. Transforming growth factor beta 1 (TGF-β1, a liver fibrosis-inducing factor, was highly expressed in the alcohol-fed group and human hepatoma HepG2 cell; however, the increase in TGF-β1 expression was reduced following fucoidan administration. Treatment with fucoidan was also found to significantly reduce the production of inflammation-promoting cyclooygenase-2 and nitric oxide, while markedly increasing the expression of the hepatoprotective enzyme, hemeoxygenase-1, on murine liver and HepG2 cells. Taken together, the antifibrotic and anti-inflammatory effects of fucoidan on alcohol-induced liver damage may provide valuable insights into developing new therapeutics or interventions.

  5. Polysaccharides extraction from Opuntia stricta and their protective effect against HepG2 cell death and hypolipidaemic effects on hyperlipidaemia rats induced by high-fat diet.

    Science.gov (United States)

    Harrabi, Bahira; Athmouni, Khaled; Hamdaoui, Latifa; Ben Mahmoud, Lobna; Hakim, Ahmed; El Feki, Abdelfattah; Zeghal, Khaled; Ghozzi, Hanéne

    2017-10-01

    The aim of this study was to analyse cytoprotective effect of polysaccharides compound from Opuntia stricta (O. stricta) cladode (POS) in vitro including its radical scavenging activities and protective effects against hypercholesterolaemia. Our results showed that glucose was the dominant monosaccharides (30.35%). Arabinose, pyranose, fructose, galactose, glucose, sorbitol, S-inositol, M-inositol, trehalose and saccharose found in this species. O. stricta polysaccharides did not cause any cytotoxic effect on HepG2 cells within the range of concentrations tested (0-400 μgml -1 ). Pre-treatment of HepG2 cells with POS (100 μgml -1 ) significantly (p < .05) protected against cytotoxicity induced by DPPH and ABTS radicals. The POS showed strong antioxidant potential in vitro. The results indicated also that POS significantly prevented hypercholesterolaemia-induced elevation of serum biomarkers and induced increase in serum lipid profile. Moreover, the hypercholesterolaemia characterised by elevated lipid peroxidation (MDA) and reduced antioxidant enzyme defences (SOD, CAT and GPx) was restored by POS treatment.

  6. Diosgenin and 5-Methoxypsoralen Ameliorate Insulin Resistance through ER-α/PI3K/Akt-Signaling Pathways in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Ke Fang

    2016-01-01

    Full Text Available To determine the effects and the underlying mechanism of diosgenin (DSG and 5-methoxypsoralen (5-MOP, two main active components in the classical Chinese prescription Hu-Lu-Ba-Wan (HLBW, on insulin resistance, HepG2 cells were incubated in medium containing insulin. Treatments with DSG, 5-MOP, and their combination were performed, respectively. The result showed that the incubation of HepG2 cells with high concentration insulin markedly decreased glucose consumption and glycogen synthesis. However, treatment with DSG, 5-MOP, or their combination significantly reversed the condition and increased the phosphorylated expression of estrogen receptor-α (ERα, sarcoma (Src, Akt/protein kinase B, glycogen synthase kinase-3β (GSK-3β, and the p85 regulatory subunit of phosphatidylinositol 3-kinase p85 (PI3Kp85. At the transcriptional level, expression of the genes mentioned above also increased except for the negative regulation of GSK-3β mRNA. The increased expression of glucose transport-4 (GLUT-4 was meanwhile observed through immunofluorescence. Nevertheless, the synergistic effect of DSG and 5-MOP on improving glycometabolism was not obvious in the present study. These results suggested that DSG and 5-MOP may improve insulin resistance through an ER-mediated PI3K/Akt activation pathway which may be a new strategy for type 2 diabetes mellitus, especially for women in an estrogen-deficient condition.

  7. Usefulness of monitoring γ-H2AX and cell cycle arrest in HepG2 cells for estimating genotoxicity using a high-content analysis system.

    Science.gov (United States)

    Ando, Masamitsu; Yoshikawa, Keisuke; Iwase, Yumiko; Ishiura, Shoichi

    2014-10-01

    Formation of the phosphorylated protein γ-H2AX is a well-established marker of DNA strand breakage induced by DNA-damaging compounds. Many of these genotoxic compounds also inhibit cell division, leading to arrest at specific points in the cell cycle. Detection of γ-H2AX in combination with cell cycle arrest may therefore be useful for estimating the genotoxicity of experimental compounds. In this study, we examined γ-H2AX formation and cell cycle arrest using high-content screening (HCS) as a method for determining genotoxicity. HepG2 cells were treated with a panel of compounds and then stained with Hoechst 33342 and anti-γ-H2AX, anti-phospho-histone H3, and anti-tubulin antibodies. In total, 19 genotoxic and 7 nongenotoxic compounds were tested in this study. γ-H2AX production was observed within 1 h posttreatment for the majority of Ames-positive compounds, topoisomerase inhibitors, and DNA polymerase inhibitors. Cell cycle arrest in either the S or G2 phase was detected for all DNA-damaging compounds 24 h posttreatment, whereas tubulin-targeting compounds were shown to induce cell cycle arrest in the mitotic phase. Together, these results show that HCS is a simple, rapid, and effective tool for estimating the genotoxicity of compounds through detection of γ-H2AX production and cell cycle arrest. © 2014 Society for Laboratory Automation and Screening.

  8. Differences in TCDD-elicited gene expression profiles in human HepG2, mouse Hepa1c1c7 and rat H4IIE hepatoma cells

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    Burgoon Lyle D

    2011-04-01

    Full Text Available Abstract Background 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD is an environmental contaminant that elicits a broad spectrum of toxic effects in a species-specific manner. Current risk assessment practices routinely extrapolate results from in vivo and in vitro rodent models to assess human risk. In order to further investigate the species-specific responses elicited by TCDD, temporal gene expression responses in human HepG2, mouse Hepa1c1c7 and rat H4IIE cells were compared. Results Microarray analysis identified a core set of conserved gene expression responses across species consistent with the role of AhR in mediating adaptive metabolic responses. However, significant species-specific as well as species-divergent responses were identified. Computational analysis of the regulatory regions of species-specific and -divergent responses suggests that dioxin response elements (DREs are involved. These results are consistent with in vivo rat vs. mouse species-specific differential gene expression, and more comprehensive comparative DRE searches. Conclusions Comparative analysis of human HepG2, mouse Hepa1c1c7 and rat H4IIE TCDD-elicited gene expression responses is consistent with in vivo rat-mouse comparative gene expression studies, and more comprehensive comparative DRE searches, suggesting that AhR-mediated gene expression is species-specific.

  9. Lower concentrations of blueberry polyphenolic-rich extract differentially alter HepG2 cell proliferation and expression of genes related to cell-cycle, oxidation and epigenetic machinery

    Science.gov (United States)

    In vitro cancer models have been used to study the effect of relatively high concentrations (>200 ug/ml) of phenolic plant extracts upon cell proliferation. In this study we report that the treatment of human hepatocarcinoma HepG2 cells with lower concentrations of blueberry phenolic extract (6.5-10...

  10. Potentiation of LPS-Induced Apoptotic Cell Death in Human Hepatoma HepG2 Cells by Aspirin via ROS and Mitochondrial Dysfunction: Protection by N-Acetyl Cysteine

    Science.gov (United States)

    Raza, Haider; John, Annie; Shafarin, Jasmin

    2016-01-01

    Cytotoxicity and inflammation-associated toxic responses have been observed to be induced by bacterial lipopolysaccharides (LPS) in vitro and in vivo respectively. Use of nonsteroidal anti-inflammatory drugs (NSAIDs), such as aspirin, has been reported to be beneficial in inflammation-associated diseases like cancer, diabetes and cardiovascular disorders. Their precise molecular mechanisms, however, are not clearly understood. Our previous studies on aspirin treated HepG2 cells strongly suggest cell cycle arrest and induction of apoptosis associated with mitochondrial dysfunction. In the present study, we have further demonstrated that HepG2 cells treated with LPS alone or in combination with aspirin induces subcellular toxic responses which are accompanied by increase in reactive oxygen species (ROS) production, oxidative stress, mitochondrial respiratory dysfunction and apoptosis. The LPS/Aspirin induced toxicity was attenuated by pre-treatment of cells with N-acetyl cysteine (NAC). Alterations in oxidative stress and glutathione-dependent redox-homeostasis were more pronounced in mitochondria compared to extra- mitochondrial cellular compartments. Pre-treatment of HepG2 cells with NAC exhibited a selective protection in redox homeostasis and mitochondrial dysfunction. Our results suggest that the altered redox metabolism, oxidative stress and mitochondrial function in HepG2 cells play a critical role in LPS/aspirin-induced cytotoxicity. These results may help in better understanding the pharmacological, toxicological and therapeutic properties of NSAIDs in cancer cells exposed to bacterial endotoxins. PMID:27441638

  11. Upregulation of heat shock protein 70 and the differential protein expression induced by tumor necrosis factor-alpha enhances migration and inhibits apoptosis of hepatocellular carcinoma cell HepG2.

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    Huang, Bee-Piao; Lin, Chun-Shiang; Wang, Chau-Jong; Kao, Shao-Hsuan

    2017-01-01

    Tumor necrosis factor alpha (TNFα) plays diverse roles in liver damage and hepatocarcinogenesis with its multipotent bioactivity. However, the influence of TNFα on protein expression of hepatocellular carcinoma (HCC) is incompletely understood. Therefore, we aimed to investigate the differential protein expression of HCC in response to TNFα stimulus. We observed that HepG2 cell revealed a higher resistance to TNFα-induced apoptosis as compared to the non-tumorigenic hepatocyte THLE-2. By using a label-free quantitative proteomic analysis, we found that 520 proteins were differentially expressed in the HepG2 cells exposed to TNFα, including 211 up-regulated and 309 down-regulated proteins. We further confirmed several proteins with significant expression change (TNFα/control ratio>2.0 or expressed proteins using Gene ontology and KEGG annotations, and the results implicated that TNFα might regulate ribosome, spliceosome, antigen processing and presentation, and energy metabolism in HepG2 cells. Moreover, we demonstrated that upregulation of heat shock protein 70 (HSP70) was involved in both the promoted migration and the inhibited apoptosis of HepG2 cells in response to TNFα. Collectively, these findings indicate that TNFα alters protein expression such as HSP70, which triggering specific molecular processes and signaling cascades that promote migration and inhibit apoptosis of HepG2 cells.

  12. Acoustic and hybrid 3D-printed electrochemical biosensors for the real-time immunodetection of liver cancer cells (HepG2).

    Science.gov (United States)

    Damiati, Samar; Küpcü, Seta; Peacock, Martin; Eilenberger, Christoph; Zamzami, Mazin; Qadri, Ishtiaq; Choudhry, Hani; Sleytr, Uwe B; Schuster, Bernhard

    2017-08-15

    This study presents an efficient acoustic and hybrid three-dimensional (3D)-printed electrochemical biosensors for the detection of liver cancer cells. The biosensors function by recognizing the highly expressed tumor marker CD133, which is located on the surface of liver cancer cells. Detection was achieved by recrystallizing a recombinant S-layer fusion protein (rSbpA/ZZ) on the surface of the sensors. The fused ZZ-domain enables immobilization of the anti-CD133 antibody in a defined manner. These highly accessible anti-CD133 antibodies were employed as a sensing layer, thereby enabling the efficient detection of liver cancer cells (HepG2). The recognition of HepG2 cells was investigated in situ using a quartz crystal microbalance with dissipation monitoring (QCM-D), which enabled the label-free, real-time detection of living cells on the modified sensor surface under controlled conditions. Furthermore, the hybrid 3D additive printing strategy for biosensors facilitates both rapid development and small-scale manufacturing. The hybrid strategy of combining 3D-printed parts and more traditionally fabricated parts enables the use of optimal materials: a ceramic substrate with noble metals for the sensing element and 3D-printed capillary channels to guide and constrain the clinical sample. Cyclic voltammetry (CV) measurements confirmed the efficiency of the fabricated sensors. Most importantly, these sensors offer low-cost and disposable detection platforms for real-world applications. Thus, as demonstrated in this study, both fabricated acoustic and electrochemical sensing platforms can detect cancer cells and therefore may have further potential in other clinical applications and drug-screening studies. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. The effects of herbal composition Gambigyeongsinhwan (4) on hepatic steatosis and inflammation in Otsuka Long-Evans Tokushima fatty rats and HepG2 cells.

    Science.gov (United States)

    Yoon, Seolah; Kim, Jeongjun; Lee, Hyunghee; Lee, Haerim; Lim, Jonghoon; Yang, Heejeong; Shin, Soon Shik; Yoon, Michung

    2017-01-04

    Hepatic steatosis has risen rapidly in parallel with a dramatic increase in obesity. The aim of this study was to determine whether the herbal composition Gambigyeongsinhwan (4) (GGH(4)), composed of Curcuma longa L. (Zingiberaceae), Alnus japonica (Thunb.) Steud. (Betulaceae), and the fermented traditional Korean medicine Massa Medicata Fermentata, regulates hepatic steatosis and inflammation. The effects of GGH(4) on hepatic steatosis and inflammation in Otsuka Long-Evans Tokushima fatty (OLETF) rats and HepG2 cells were examined using Oil red O, hematoxylin and eosin, and toluidine blue staining, immunohistochemistry, quantitative real-time polymerase chain reaction, and peroxisome proliferator-activated receptor α (PPARα) transactivation assay. Administration of GGH(4) to OLETF rats improved hepatic steatosis and lowered serum levels of alanine transaminase, total cholesterol, triglycerides, and free fatty acids. GGH(4) increased mRNA levels of fatty acid oxidation enzymes (ACOX, HD, CPT-1, and MCAD) and decreased mRNA levels of lipogenesis genes (FAS, ACC1, C/EBPα, and SREBP-1c) in the liver of OLETF rats. In addition, infiltration of inflammatory cells and expression of inflammatory cytokines (CD68, TNFα, and MCP-1) in liver tissue were reduced by GGH(4). Treatment of HepG2 cells with a mixture of oleic acid and palmitoleic acid induced significant lipid accumulation, but GGH(4) inhibited lipid accumulation by regulating the expression of hepatic fatty acid oxidation and lipogenic genes. GGH(4) also increased PPARα reporter gene expression. These effects of GGH(4) were similar to those of the PPARα activator fenofibrate, whereas the PPARα antagonist GW6471 reversed the inhibitory effects of GGH(4) on lipid accumulation in HepG2 cells. These results suggest that GGH(4) inhibits obesity-induced hepatic steatosis and that this process may be mediated by regulation of the expression of PPARα target genes and lipogenic genes. GGH(4) also suppressed obesity

  14. Isolation of major phenolics from Launaea spinosa and their protective effect on HepG2 cells damaged with t-BHP.

    Science.gov (United States)

    Abdallah, Hossam; Farag, Mohamed; Osman, Samir; Kim, Da hye; Kang, Kyungsu; Pan, Cheol-Ho; Abdel-Sattar, Essam

    2016-01-01

    Some Launaea species (Asteraceae) are used traditionally to treat liver oxidative stress. The present study investigates the protective effects of isolated compounds from Launaea spinosa Sch. Bip. (Asteraceae) against oxidative stress on t-BHP-induced HepG2 cells. Major phenolic content from flowering aerial parts of L. spinosa was isolated and identified. The protective effects of isolated compounds (10 and 20 μM) against oxidative stress induced by tert-butyl hydroperoxide (t-BHP) in HepG2 cells were investigated through the measurement of aspartate aminotransferase (AST), alanine transaminase (ALT), and superoxide dismutase (SOD) levels. A new phenolic compound identified as 2,3-diferulyl R,R-(+) methyl tartrate (6), in addition to five known metabolites, esculetin (1), esculetin-7-O-d-glucoside (cichoriin) (2), fertaric acid (3), acacetin-7-O-d-glucoside (4), and acacetin-7-O-d-glucuronic acid (5), were isolated. Oxidant-induced damage by 200 μM t-BHP in HepG2 cells was inhibited by compounds 1, 4, and 5 (10 and 20 μM), or quercetin (10 μM; positive control). The protective effects of compounds 1, 4, and 5 were associated with decreasing in AST, ALT, and SOD levels. Compound 4 (20 μM) decreased the AST level from 128.5 ± 13.9 to 7.9 ±1.8 U/mL. Meanwhile, compound 1 (20 μM) decreased ALT activity from 20.3 ± 7.0 to 7.6 ± 2.4 U/mL, while compound 5 decreased SOD levels from 41.6 ± 9.0 to 28.3 ± 3.4 mU/mg. The major phenolic compounds isolated from L. spinosa displayed a significant cytoprotective effect against oxidative stress, leading to maintenance of the normal redox status of the cell.

  15. Morin impedes Yap nuclear translocation and fosters apoptosis through suppression of Wnt/β-catenin and NF-κB signaling in Mst1 overexpressed HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Perumal, NaveenKumar [Department of Biochemistry, University of Madras, Guindy Campus, Chennai 600025, Tamil Nadu (India); Perumal, MadanKumar [Department of Biochemistry, University of Madras, Guindy Campus, Chennai 600025, Tamil Nadu (India); Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, Texas 75390 (United States); Kannan, Anbarasu [Department of Cellular and Molecular Biology, The University of Texas Health Science Center, Tyler, Texas (United States); Subramani, Kumar [Centre for Biotechnology, Anna University, Chennai 600025, Tamil Nadu (India); Halagowder, Devaraj [Department of Zoology, University of Madras, Guindy Campus, Chennai 600025, Tamil Nadu (India); Sivasithamparam, NiranjaliDevaraj, E-mail: profniranjali@gmail.com [Department of Biochemistry, University of Madras, Guindy Campus, Chennai 600025, Tamil Nadu (India)

    2017-06-15

    Recent clinical and experimental evidences strongly acclaim Yes-associated protein (Yap), a key oncogenic driver in liver carcinogenesis, as a therapeutic target. Of the known multiple schemes to inhibit Yap activity, activation of Mammalian Sterile 20-like Kinase 1 (Mst1), an upstream regulator of Yap, appears to be a promising one. In this study, we hypothesize that morin, a bioflavonoid, mediates its anti-cancer effect through the activation of Mst1/hippo signaling in liver cancer cells. To test this hypothesis, both full length Mst1 (F-Mst1) and kinase active N-terminal Mst1 (N-Mst1)-overexpressed HepG2 cells were used. Exposure of F-Mst1 overexpressed HepG2 cells to morin activated Mst1 by caspase-3 cleavage and thereby inhibited Yap nuclear translocation and fostered apoptosis. Morin suppressed NF-κB p65 and Wnt/β-catenin signaling through Mst1 activation via cleavage and phosphorylation, leading to cell death. Annexin-V/PI staining further confirmed the induction of apoptosis in morin treated F-Mst1 overexpressed cells. The present study shows that morin targets cell survival molecules such as NF-κB p65 and β-catenin through activation of hippo signaling. Therefore, morin could be considered as a potential anti-cancer agent against liver cancer. - Highlights: • Morin induced cytotoxicity in cultured HepG2 cells. • Morin activated hippo pathway via Mst1 activation in transfected HepG2 cells. • Morin suppressed Wnt/β-catenin signaling and induced G0/G1 cell cycle arrest. • Morin inhibited NF-κB signaling through Mst1 activation in transfected HepG2 cells. • Morin potentiates apoptosis through Mst1-JNK-caspase mediated mechanism in HepG2 cells.

  16. Inhibition of Tumor Growth of Human Hepatocellular Carcinoma HepG2 Cells in a Nude Mouse Xenograft Model by the Total Flavonoids from Arachniodes exilis

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    Huimin Li

    2017-01-01

    Full Text Available A tumor growth model of human hepatocellular carcinoma HepG2 cells in nude mice was employed to investigate the antitumor activity of the total flavonoids extracted from Arachniodes exilis (TFAE in vivo. Several biochemical assays including hematoxylin-eosin (HE staining, immunohistochemistry, and Western blot were performed to elucidate the mechanism of action of total flavonoids extracted from Arachniodes exilis (TFAE. The results showed that TFAE effectively inhibited the tumor growth of hepatocellular carcinoma in nude mice and had no significant effect on body weight, blood system, and functions of liver and kidney. Expression levels of proapoptotic proteins Bax and cleaved caspase-3 remarkably increased while the expressions of Bcl-2, HIF-1α, and VEGF were suppressed by TFAE. These results suggested that the antitumor potential of TFEA was implied by the apoptosis of tumor cells and the inhibition of angiogenesis in tumor tissue.

  17. Regulation of the Lactobacillus Strains on HMGCoA Reductase Gene Transcription in Human HepG2 Cells via Nuclear Factor-κB.

    Science.gov (United States)

    Chen, Kun; Li, Shaocong; Chen, Fang; Li, Jun; Luo, Xuegang

    2016-02-01

    Lactic acid bacteria have been identified to be effective in reducing cholesterol levels. Most of the mechanistic studies were focused on the bile salt deconjugation ability of bile salt hydrolase in lactic acid bacteria. However, the mechanism by which Lactobacillus decreases cholesterol levels has not been thoroughly studied in intact primate cells. 3-Hydroxy-3- methyl-glutaryl-coenzyme A reductase (HMGCR) is the vital enzyme in cholesterol synthesis. To confirm the effect of probiotic Lactobacillus strains on HMGCR level, in the present study, human hepatoma HepG2 cells were treated with Lactobacillus strains, and then the HMGCR level was illustrated by luciferase reporter assay and RT-PCR. The results showed that the level of HMGCR was suppressed after being treated with the live Lactobacillus strains. These works might set a foundation for the following study of the antihyperlipidemic effects of L. acidophilus, and contribute to the development of functional foods or drugs that benefit patients suffering from hyperlipidemia diseases.

  18. Cadmium telluride quantum dots cause oxidative stress leading to extrinsic and intrinsic apoptosis in hepatocellular carcinoma HepG2 cells

    International Nuclear Information System (INIS)

    Nguyen, Kathy C.; Willmore, William G.; Tayabali, Azam F.

    2013-01-01

    The mechanisms of toxicity related to human hepatocellular carcinoma HepG2 cell exposures to cadmium telluride quantum dots (CdTe-QDs) were investigated. CdTe-QDs caused cytotoxicity in HepG2 cells in a dose- and time-dependent manner. Treated cells showed an increase in reactive oxygen species (ROS). Altered antioxidant levels were demonstrated by depletion of reduced glutathione (GSH), a decreased ratio of reduced glutathione to oxidized glutathione (GSH/GSSG) and an increased NF-E2-related Factor 2 (Nrf2) activation. Enzyme assays showed that superoxide dismutase (SOD) activity was elevated whereas catalase (CAT) and glutathione-S-transferase (GST) activities were depressed. Further analyses revealed that CdTe-QD exposure resulted in apoptosis, indicated by changes in levels of caspase-3 activity, poly ADP-ribose polymerase (PARP) cleavage and phosphatidylserine externalization. Extrinsic apoptotic pathway markers such as Fas levels and caspase-8 activity increased as a result of CdTe-QD exposure. Involvement of the intrinsic/mitochondrial apoptotic pathway was indicated by decreased levels of B-cell lymphoma 2 (Bcl2) protein and mitochondrial cytochrome c, and by increased levels of mitochondrial Bcl-2-associated X protein (Bax) and cytosolic cytochrome c. Further, mitogen-activated protein kinases (MAPKs) such as c-Jun N-terminal kinases (JNK), extracellular signal-regulated kinases (Erk1/2), and p38 were all activated. Our findings reveal that CdTe-QDs cause oxidative stress, interfere with antioxidant defenses and activate protein kinases, leading to apoptosis via both extrinsic and intrinsic pathways. Since the effects of CdTe-QDs on selected biomarkers were similar or greater compared to those of CdCl 2 at equivalent concentrations of cadmium, the study suggests that the toxicity of CdTe-QDs arises from a combination of the effects of cadmium and ROS generated from the NPs

  19. Acerinol, a cyclolanstane triterpenoid from Cimicifuga acerina, reverses ABCB1-mediated multidrug resistance in HepG2/ADM and MCF-7/ADR cells.

    Science.gov (United States)

    Liu, Dao-Lu; Li, Ying-Jie; Yao, Nan; Xu, Jun; Chen, Zhe-Sheng; Yiu, Anita; Zhang, Cui-Xian; Ye, Wen-Cai; Zhang, Dong-Mei

    2014-06-15

    Persistent cancer chemotherapy can lead to multidrug resistance which is one of the most common reasons for failure of chemotherapy. The ABCB1 transporter is a member of the ATP-binding cassette superfamily and it is frequently over-expressed in multidrug resistant cancer cells. Active ingredients derived from traditional Chinese medicinal herbs have been reported to reverse multidrug resistance mediated by ATP-binding cassette transporters. In this study, acerinol, isolated from Cimicifuga acerina, was tested for its potential to modulate the ABCB1 transporter. Our results demonstrated that acerinol could increase the chemosensitivity of ABCB1-overexpressing HepG2/ADM and MCF-7/ADR cells to chemotherapeutic drugs, doxorubicin, vincristine and paclitaxel. Furthermore, it could also increase the retention of ABCB1 substrates doxorubicin and rhodamine 123 in HepG2/ADM and MCF-7/ADR cells. A mechanistic study showed that acerinol significantly stimulated the activity of ABCB1 ATPase without affecting the expression of ABCB1 on neither mRNA nor protein level. Acerinol was also found to reverse the resistance of MCF-7/ADR cells to vincristine, dependent partly on ABCB1. In addition, acerinol׳s action was reversible, suggesting that acerinol may act as a competitive inhibitor of ABCB1 by competing with other drug substrates like doxorubicin. Indeed, docking analysis indicated that acerinol would most likely bind to the sites on ABCB1 that partly overlapped with that of verapamil. In conclusion, the present study is the first to show that acerinol from C. acerina significantly enhances the cytotoxicity of chemotherapeutic drugs by modulating the function of ABCB1. It is hopeful to develop acerinol as a new multidrug resistance reversal agent. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Phytochemical study and evaluation of cytotoxicity, mutagenicity, cell cycle kinetics and gene expression of Bauhinia holophylla (Bong.) Steud. in HepG2 cells in vitro.

    Science.gov (United States)

    Ribeiro, Diego Luis; Cilião, Heloísa Lizotti; Specian, Ana Flávia Leal; Serpeloni, Juliana Mara; De Oliveira, Marcelo Tempesta; Varanda, Eliana Aparecida; Vilegas, Wagner; Saldanha, Luiz Leonardo; Martínez-López, Wilner; Dokkedal, Anne Lígia; Cólus, Ilce Mara Syllos

    2018-04-01

    Bauhinia holophylla (Bong.) Steud. (Fabaceae) is a plant used in Brazilian folk medicine to treat diabetes and inflammation. This study evaluated the phytochemical properties, cytotoxic, apoptotic, mutagenic/antimutagenic effects and alterations in gene expression (RNAm) in HepG2 cells treated with the B. holophylla extract. The phytochemical profile highlight the presence of flavonoids isorhamentin and quercetin derivates. The MTT assay was used to evaluate the cytotoxicity of different concentrations for different treatment times. Three concentrations (7.5, 15, 30 µg/mL) were chosen for assessment of apoptosis (AO/EB), mutagenicity (micronucleus), and cell cycle kinetics (flow cytometry). Thereafter, the concentration of 7.5 µg/mL was chosen to evaluate the protective effects against DNA damage induced by benzo[a]pyrene (B[a]P). At concentrations higher than 7.5 µg/mL (between 10 and 50 µg/mL), the extract was cytotoxic, induced apoptosis, and caused antiproliferative effects. However, it did not induce micronucleus and a reduction of apoptotic and micronucleated cells was observed in treatments that included the extract and B[a]P. The protective effect is attributable to the presence of flavonoids, described as antioxidants, inhibitors of DNA adduct and activators of detoxifying enzymes. The results of the present study such as absence of cytotoxic and mutagenic effects and protective effects against known carcinogens suggest that B. holophylla has potential for use soon as herbal medicine.

  1. Live Cell Imaging Confocal Microscopy Analysis of HBV Myr-PreS1 Peptide Binding and Uptake in NTCP-GFP Expressing HepG2 Cells.

    Science.gov (United States)

    König, Alexander; Glebe, Dieter

    2017-01-01

    To obtain basic knowledge about specific molecular mechanisms involved in the entry of pathogens into cells is the basis for establishing pharmacologic substances blocking initial viral binding, infection, and subsequent viral spread. Lack of information about key cellular factors involved in the initial steps of HBV infection has hampered the characterization of HBV binding and entry for decades. However, recently, the liver-specific sodium-dependent taurocholate cotransporting polypeptide (NTCP) has been discovered as a functional receptor for HBV and HDV, thus opening the field for new concepts of basic binding and entry of HBV and HDV. Here, we describe practical issues of a basic in vitro assay system to examine kinetics and mechanisms of receptor-dependent HBV binding, uptake, and intracellular trafficking by live-cell imaging confocal microscopy. The assay system is comprised of HepG2 cells expressing a NTCP-GFP fusion-protein and chemically synthesized, fluorophore-labeled part of HBV surface protein, spanning the first N-terminal 48 amino acids of preS1 of the large hepatitis B virus surface protein.

  2. Anticancer Effects of 1,3-Dihydroxy-2-Methylanthraquinone and the Ethyl Acetate Fraction of Hedyotis Diffusa Willd against HepG2 Carcinoma Cells Mediated via Apoptosis.

    Science.gov (United States)

    Li, Yun-Lan; Zhang, Jiali; Min, Dong; Hongyan, Zhou; Lin, Niu; Li, Qing-Shan

    2016-01-01

    Hedyotis Diffusa Willd, used in Traditional Chinese Medicine, is a treatment for various diseases including cancer, owing to its mild effectiveness and low toxicity. The aim of this study was to identify the main anticancer components in Hedyotis Diffusa Willd, and explore mechanisms underlying their activity. Hedyotis Diffusa Willd was extracted and fractionated using ethyl acetate to obtain the H-Ethyl acetate fraction, which showed higher anticancer activity than the other fractions obtained against HepG2 cells with sulforhodamine B assays. The active component of the H-Ethyl acetate fraction was identified to be 1,3-dihydroxy-2-methylanthraquinone (DMQ) with much high inhibitory rate up to 48.9 ± 3.3% and selectivity rate up to 9.4 ± 4.5 folds (pEthyl acetate fraction of Hedyotis Diffusa Willd showed potential anticancer effects. Furthermore, the mechanisms of action may involve mitochondrial apoptotic and death receptor pathways.

  3. GEP100/Arf6 is required for epidermal growth factor-induced ERK/Rac1 signaling and cell migration in human hepatoma HepG2 cells.

    Directory of Open Access Journals (Sweden)

    ZhenZhen Hu

    Full Text Available BACKGROUND: Epidermal growth factor (EGF signaling is implicated in the invasion and metastasis of hepatoma cells. However, the signaling pathways for EGF-induced motility of hepatoma cells remain undefined. METHODOLOGY/PRINCIPAL FINDINGS: We found that EGF dose-dependently stimulated the migration of human hepatoma cells HepG2, with the maximal effect at 10 ng/mL. Additionally, EGF increased Arf6 activity, and ectopic expression of Arf6 T27N, a dominant negative Arf6 mutant, largely abolish EGF-induced cell migration. Blocking GEP100 with GEP100 siRNA or GEP100-△PH, a pleckstrin homology (PH domain deletion mutant of GEP100, blocked EGF-induced Arf6 activity and cell migration. EGF also increased ERK and Rac1 activity. Ectopic expression GEP100 siRNA, GEP100-△PH, or Arf6-T27N suppressed EGF-induced ERK and Rac1 activity. Furthermore, blocking ERK signaling with its inhibitor U0126 remarkably inhibited both EGF-induced Rac1 activation as well as cell migration, and ectopic expression of inactive mutant form of Rac1 (Rac1-T17N also largely abolished EGF-induced cell migration. CONCLUSIONS/SIGNIFICANCE: Taken together, this study highlights the function of the PH domain of GEP100 and its regulated Arf6/ERK/Rac1 signaling cascade in EGF-induced hepatoma cell migration. These findings could provide a rationale for designing new therapy based on inhibition of hepatoma metastasis.

  4. A Novel Polysaccharide Conjugate from Bullacta exarata Induces G1-Phase Arrest and Apoptosis in Human Hepatocellular Carcinoma HepG2 Cells.

    Science.gov (United States)

    Liao, Ningbo; Sun, Liang; Chen, Jiang; Zhong, Jianjun; Zhang, Yanjun; Zhang, Ronghua

    2017-03-01

    Bullacta exarata has been consumed in Asia, not only as a part of the normal diet, but also as a traditional Chinese medicine with liver- and kidney-benefitting functions. Several scientific investigations involving extraction of biomolecules from this mollusk and pharmacological studies on their biological activities have been carried out. However, little is known regarding the antitumor properties of polysaccharides from B. exarata , hence the polysaccharides from B. exarata have been investigated here. One polysaccharide conjugate BEPS-IA was isolated and purified from B. exarata . It mainly consisted of mannose and glucose in a molar ratio of 1:2, with an average molecular weight of 127 kDa. Thirteen general amino acids were identified to be components of the protein-bound polysaccharide. Methylation and NMR studies revealed that BEPS-IA is a heteropolysaccharide consisting of 1,4-linked-α-d-Glc, 1,6-linked-α-d-Man, 1,3,6-linked-α-d-Man, and 1-linked-α-d-Man residue, in a molar ratio of 6:1:1:1. In order to test the antitumor activity of BEPS-IA, we investigated its effect against the growth of human hepatocellular carcinoma cells HepG2 in vitro. The result showed that BEPS-IA dose-dependently exhibited an effective HepG2 cells growth inhibition with an IC 50 of 112.4 μg/mL. Flow cytometry analysis showed that BEPS-IA increased the populations of both apoptotic sub-G1 and G1 phase. The result obtained from TUNEL assay corroborated apoptosis which was shown in flow cytometry. Western blot analysis suggested that BEPS-IA induced apoptosis and growth inhibition were associated with up-regulation of p53, p21 and Bax, down-regulation of Bcl-2. These findings suggest that BEPS-IA may serve as a potential novel dietary agent for hepatocellular carcinoma.

  5. Ruthenium Complexes Induce HepG2 Human Hepatocellular Carcinoma Cell Apoptosis and Inhibit Cell Migration and Invasion through Regulation of the Nrf2 Pathway

    Directory of Open Access Journals (Sweden)

    Yiyu Lu

    2016-05-01

    Full Text Available Ruthenium (Ru complexes are currently the focus of substantial interest because of their potential application as chemotherapeutic agents with broad anticancer activities. This study investigated the in vitro and in vivo anticancer activities and mechanisms of two Ru complexes—2,3,7,8,12,13,17,18-Octaethyl-21H,23H-porphine Ru(II carbonyl (Ru1 and 5,10,15,20-Tetraphenyl-21H,23H-porphine Ru(II carbonyl (Ru2—against human hepatocellular carcinoma (HCC cells. These Ru complexes effectively inhibited the cellular growth of three human hepatocellular carcinoma (HCC cells, with IC50 values ranging from 2.7–7.3 μM. In contrast, the complexes exhibited lower toxicity towards L02 human liver normal cells with IC50 values of 20.4 and 24.8 μM, respectively. Moreover, Ru2 significantly inhibited HepG2 cell migration and invasion, and these effects were dose-dependent. The mechanistic studies demonstrated that Ru2 induced HCC cell apoptosis, as evidenced by DNA fragmentation and nuclear condensation, which was predominately triggered via caspase family member activation. Furthermore, HCC cell treatment significantly decreased the expression levels of Nrf2 and its downstream effectors, NAD(PH: quinone oxidoreductase 1 (NQO1 and heme oxygenase 1 (HO1. Ru2 also exhibited potent in vivo anticancer efficacy in a tumor-bearing nude mouse model, as demonstrated by a time- and dose-dependent inhibition on tumor growth. The results demonstrate the therapeutic potential of Ru complexes against HCC via Nrf2 pathway regulation.

  6. Ruthenium Complexes Induce HepG2 Human Hepatocellular Carcinoma Cell Apoptosis and Inhibit Cell Migration and Invasion through Regulation of the Nrf2 Pathway

    Science.gov (United States)

    Lu, Yiyu; Shen, Ting; Yang, Hua; Gu, Weiguang

    2016-01-01

    Ruthenium (Ru) complexes are currently the focus of substantial interest because of their potential application as chemotherapeutic agents with broad anticancer activities. This study investigated the in vitro and in vivo anticancer activities and mechanisms of two Ru complexes—2,3,7,8,12,13,17,18-Octaethyl-21H,23H-porphine Ru(II) carbonyl (Ru1) and 5,10,15,20-Tetraphenyl-21H,23H-porphine Ru(II) carbonyl (Ru2)—against human hepatocellular carcinoma (HCC) cells. These Ru complexes effectively inhibited the cellular growth of three human hepatocellular carcinoma (HCC) cells, with IC50 values ranging from 2.7–7.3 μM. In contrast, the complexes exhibited lower toxicity towards L02 human liver normal cells with IC50 values of 20.4 and 24.8 μM, respectively. Moreover, Ru2 significantly inhibited HepG2 cell migration and invasion, and these effects were dose-dependent. The mechanistic studies demonstrated that Ru2 induced HCC cell apoptosis, as evidenced by DNA fragmentation and nuclear condensation, which was predominately triggered via caspase family member activation. Furthermore, HCC cell treatment significantly decreased the expression levels of Nrf2 and its downstream effectors, NAD(P)H: quinone oxidoreductase 1 (NQO1) and heme oxygenase 1 (HO1). Ru2 also exhibited potent in vivo anticancer efficacy in a tumor-bearing nude mouse model, as demonstrated by a time- and dose-dependent inhibition on tumor growth. The results demonstrate the therapeutic potential of Ru complexes against HCC via Nrf2 pathway regulation. PMID:27213353

  7. NMR-based metabolomics reveals that conjugated double bond content and lipid storage efficiency in HepG2 cells are affected by fatty acid cis/trans configuration and chain length

    DEFF Research Database (Denmark)

    Najbjerg, Heidi; Young, Jette F; Bertram, Hanne Christine S.

    2011-01-01

    In the present study the metabolic response to various fatty acids was investigated in HepG2 cells by using a 1HNMRbased approach. To elucidate the effect of cis/trans configuration, the cells were exposed to either oleic acid (C18:1 cis-9), elaidic acid (C18:1 trans-9), vaccenic acid (C18:1 tran...

  8. A comparative assessment of antiproliferative properties of resveratrol and ethanol leaf extract of Anogeissus leiocarpus (DC) Guill and Perr against HepG2 hepatocarcinoma cells.

    Science.gov (United States)

    Olugbami, Jeremiah Olorunjuwon; Damoiseaux, Robert; France, Bryan; Onibiyo, Esther Modupe; Gbadegesin, Michael Adedapo; Sharma, Shivani; Gimzewski, James Kazimierz; Odunola, Oyeronke Adunni

    2017-08-02

    Epidemiological and experimental evidences have shown cancer as a leading cause of death worldwide. Although the folklore use of plants as a reliable source of health-restoring principles is well-documented, the search for more of such plants that are active against diseases, such as cancer, continues. We report here a laboratory-based evidence of the relevance of an ethanol leaf extract of Anogeissus leiocarpus (A2L) in comparison with resveratrol, a natural polyphenol, in cancer therapy. The quantitative assessment of flavonoid and phenolic contents involved quercetin and gallic acid as standards, respectively were determined using spectrophotometry. Cytotoxicity was determined fluorometrically using propidium-iodide-staining method. Antioxidant status, adenosine triphosphate (ATP) levels, caspase activities and mitochondrial integrity were assessed using fluorometry/luminometry. The antioxidant assay demonstrated that A2L possesses a strong antioxidant capacity as compared with the reference compounds, ascorbic acid and butylated hydroxytoluene. This is further buttressed by the significantly high level of phenolics obtained in the quantitative assessment of the extract. A 72-h post-treatment examination indicated that both A2L and resveratrol modulate the proliferation of HepG2 liver carcinoma cells in a time- and concentration-dependent manner. Determination of the total nuclei area, propidium-iodide negative and positive nuclei areas all further buttress the modulation of cell proliferation by A2L and resveratrol with the indication that the observed cell death is due to apoptosis and necrosis at lower and higher concentrations of treatments respectively. At lower concentrations (0.39-3.13 μg/mL), resveratrol possesses higher tendencies to activate caspases 3 and 7. Bioenergetically, both resveratrol and A2L do not adversely affect the cells at lower concentrations (0.39-6.25 μg/mL for resveratrol and 12.5-100.0 μg/mL for A2L) except at higher

  9. Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.

    Science.gov (United States)

    Plecitá-Hlavatá, Lydie; Engstová, Hana; Alán, Lukáš; Špaček, Tomáš; Dlasková, Andrea; Smolková, Katarína; Špačková, Jitka; Tauber, Jan; Strádalová, Vendula; Malínský, Jan; Lessard, Mark; Bewersdorf, Joerg; Ježek, Petr

    2016-05-01

    The relationship of the inner mitochondrial membrane (IMM) cristae structure and intracristal space (ICS) to oxidative phosphorylation (oxphos) is not well understood. Mitofilin (subunit Mic60) of the mitochondrial contact site and cristae organizing system (MICOS) IMM complex is attached to the outer membrane (OMM) via the sorting and assembly machinery/topogenesis of mitochondrial outer membrane β-barrel proteins (SAM/TOB) complex and controls the shape of the cristae. ATP synthase dimers determine sharp cristae edges, whereas trimeric OPA1 tightens ICS outlets. Metabolism is altered during hypoxia, and we therefore studied cristae morphology in HepG2 cells adapted to 5% oxygen for 72 h. Three dimensional (3D), super-resolution biplane fluorescence photoactivation localization microscopy with Eos-conjugated, ICS-located lactamase-β indicated hypoxic ICS expansion with an unchanged OMM (visualized by Eos-mitochondrial fission protein-1). 3D direct stochastic optical reconstruction microscopy immunocytochemistry revealed foci of clustered mitofilin (but not MICOS subunit Mic19) in contrast to its even normoxic distribution. Mitofilin mRNA and protein decreased by ∼20%. ATP synthase dimers vs monomers and state-3/state-4 respiration ratios were lower during hypoxia. Electron microscopy confirmed ICS expansion (maximum in glycolytic cells), which was absent in reduced or OMM-detached cristae of OPA1- and mitofilin-silenced cells, respectively. Hypoxic adaptation is reported as rounding sharp cristae edges and expanding cristae width (ICS) by partial mitofilin/Mic60 down-regulation. Mitofilin-depleted MICOS detaches from SAM while remaining MICOS with mitofilin redistributes toward higher interdistances. This phenomenon causes partial oxphos dormancy in glycolytic cells via disruption of ATP synthase dimers.-Plecitá-Hlavatá, L., Engstová, H., Alán, L., Špaček, T., Dlasková, A., Smolková, K., Špačková, J., Tauber, J., Strádalová, V., Malínský, J

  10. Caffeine Increases Apolipoprotein A-1 and Paraoxonase-1 but not Paraoxonase-3 Protein Levels in Human-Derived Liver (HepG2) Cells.

    Science.gov (United States)

    Sayılan Özgün, Gülben; Özgün, Eray; Tabakçıoğlu, Kıymet; Süer Gökmen, Selma; Eskiocak, Sevgi; Çakır, Erol

    2017-12-01

    Apolipoprotein A-1, paraoxonase-1 and paraoxonase-3 are antioxidant and anti-atherosclerotic structural high-density lipoprotein proteins that are mainly synthesized by the liver. No study has ever been performed to specifically examine the effects of caffeine on paraoxonase enzymes and on liver apolipoprotein A-1 protein levels. To investigate the dose-dependent effects of caffeine on liver apolipoprotein A-1, paraoxonase-1 and paraoxonase-3 protein levels. In vitro experimental study. HepG2 cells were incubated with 0 (control), 10, 50 and 200 μM of caffeine for 24 hours. Cell viability was evaluated by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. Apolipoprotein A-1, paraoxonase-1 and paraoxonase-3 protein levels were measured by western blotting. We observed a significant increase on apolipoprotein A-1 and paraoxonase-1 protein levels in the cells incubated with 50 µM of caffeine and a significant increase on paraoxonase-1 protein level in the cells incubated with 200 µM of caffeine. Our study showed that caffeine does not change paraoxonase-3 protein level, but the higher doses used in our study do cause an increase in both apolipoprotein A-1 and paraoxonase-1 protein levels in liver cells.

  11. The synergistic radiosensitizing effect of tirapazamine-conjugated gold nanoparticles on human hepatoma HepG2 cells under X-ray irradiation

    Directory of Open Access Journals (Sweden)

    Liu X

    2016-07-01

    Full Text Available Xi Liu,1–4 Yan Liu,1–4 Pengcheng Zhang,1–4 Xiaodong Jin,1–3 Xiaogang Zheng,1–4 Fei Ye,1–4 Weiqiang Chen,1–3 Qiang Li1–3 1Institute of Modern Physics, 2Key Laboratory of Heavy Ion Radiation Biology and Medicine, Chinese Academy of Sciences, 3Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou, 4School of Life Science, University of Chinese Academy of Sciences, Beijing, People’s Republic of China Abstract: Reductive drug-functionalized gold nanoparticles (AuNPs have been proposed to enhance the damage of X-rays to cells through improving hydroxyl radical production by secondary electrons. In this work, polyethylene glycol-capped AuNPs were conjugated with tirapazamine (TPZ moiety, and then thioctyl TPZ (TPZs-modified AuNPs (TPZs-AuNPs were synthesized. The TPZs-AuNPs were characterized by transmission electron microscopy, ultraviolet-visible spectra, dynamic light scattering, and inductively coupled plasma mass spectrometry to have a size of 16.6±2.1 nm in diameter and a TPZs/AuNPs ratio of ~700:1. In contrast with PEGylated AuNPs, the as-synthesized TPZs-AuNPs exhibited 20% increment in hydroxyl radical production in water at 2.0 Gy, and 19% increase in sensitizer enhancement ratio at 10% survival fraction for human hepatoma HepG2 cells under X-ray irradiation. The production of reactive oxygen species in HepG2 cells exposed to X-rays in vitro demonstrated a synergistic radiosensitizing effect of AuNPs and TPZ moiety. Thus, the reductive drug-conjugated TPZs-AuNPs as a kind of AuNP radiosensitizer with low gold loading provide a new strategy for enhancing the efficacy of radiation therapy. Keywords: AuNPs, radiation enhancement, synergistic effect, human hepatoma cells, hydroxyl radical production

  12. EFFECTS OF THE ANTIMUTAGENS VANILLIN AND CINNAMALDEHYDE ON SPONTANEOUS MUTATION IN E. COLI LACL STRAINS AND ON GLOBAL GENE EXPRESSION IN SALMONELLA TA104 AND HUMAN HEPG2 CELLS

    Science.gov (United States)

    Effects of the Antimutagens Vanillin and Cinnamaldehyde on Spontaneous Mutation in E. coli lacI Strains and on Global Gene Epression in Salmonella TAlO4 and Human HepG2 Cells In previous work we have shown that vanillin (VAN) and cinnamaldehyde (CIN) are dietary antimutag...

  13. The Interactions between ZnO Nanoparticles (NPs) and α-Linolenic Acid (LNA) Complexed to BSA Did Not Influence the Toxicity of ZnO NPs on HepG2 Cells.

    Science.gov (United States)

    Zhou, Yiwei; Fang, Xin; Gong, Yu; Xiao, Aiping; Xie, Yixi; Liu, Liangliang; Cao, Yi

    2017-04-24

    Nanoparticles (NPs) entering the biological environment could interact with biomolecules, but little is known about the interaction between unsaturated fatty acids (UFA) and NPs. This study used α-linolenic acid (LNA) complexed to bovine serum albumin (BSA) for UFA and HepG2 cells for hepatocytes. The interactions between BSA or LNA and ZnO NPs were studied. The presence of BSA or LNA affected the hydrodynamic size, zeta potential, UV-Vis, fluorescence, and synchronous fluorescence spectra of ZnO NPs, which indicated an interaction between BSA or LNA and NPs. Exposure to ZnO NPs with the presence of BSA significantly induced the damage to mitochondria and lysosomes in HepG2 cells, associated with an increase of intracellular Zn ions, but not intracellular superoxide. Paradoxically, the release of inflammatory cytokine interleukin-6 (IL-6) was decreased, which indicated the anti-inflammatory effects of ZnO NPs when BSA was present. The presence of LNA did not significantly affect all of these endpoints in HepG2 cells exposed to ZnO NPs and BSA. the results from the present study indicated that BSA-complexed LNA might modestly interact with ZnO NPs, but did not significantly affect ZnO NPs and BSA-induced biological effects in HepG2 cells.

  14. The Interactions between ZnO Nanoparticles (NPs and α-Linolenic Acid (LNA Complexed to BSA Did Not Influence the Toxicity of ZnO NPs on HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Yiwei Zhou

    2017-04-01

    Full Text Available Background: Nanoparticles (NPs entering the biological environment could interact with biomolecules, but little is known about the interaction between unsaturated fatty acids (UFA and NPs. Methods: This study used α-linolenic acid (LNA complexed to bovine serum albumin (BSA for UFA and HepG2 cells for hepatocytes. The interactions between BSA or LNA and ZnO NPs were studied. Results: The presence of BSA or LNA affected the hydrodynamic size, zeta potential, UV-Vis, fluorescence, and synchronous fluorescence spectra of ZnO NPs, which indicated an interaction between BSA or LNA and NPs. Exposure to ZnO NPs with the presence of BSA significantly induced the damage to mitochondria and lysosomes in HepG2 cells, associated with an increase of intracellular Zn ions, but not intracellular superoxide. Paradoxically, the release of inflammatory cytokine interleukin-6 (IL-6 was decreased, which indicated the anti-inflammatory effects of ZnO NPs when BSA was present. The presence of LNA did not significantly affect all of these endpoints in HepG2 cells exposed to ZnO NPs and BSA. Conclusions: the results from the present study indicated that BSA-complexed LNA might modestly interact with ZnO NPs, but did not significantly affect ZnO NPs and BSA-induced biological effects in HepG2 cells.

  15. Fucoxanthin Enhances Cisplatin-Induced Cytotoxicity via NFκB-Mediated Pathway and Downregulates DNA Repair Gene Expression in Human Hepatoma HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Cheng-Ling Liu

    2013-01-01

    Full Text Available Cisplain, a platinum-containing anticancer drug, has been shown to enhance DNA repair and to inhibit cell apoptosis, leading to drug resistance. Thus, the combination of anticancer drugs with nutritional factors is a potential strategy for improving the efficacy of cisplatin chemotherapy. In this study, we investigated the anti-proliferative effects of a combination of fucoxanthin, the major non-provitamin A carotenoid found in Undaria Pinnatifida, and cisplatin in human hepatoma HepG2 cells. We found that fucoxanthin (1–10 μΜ pretreatment for 24 h followed by cisplatin (10 μΜ for 24 h significantly decreased cell proliferation, as compared with cisplatin treatment alone. Mechanistically, we showed that fucoxanthin attenuated cisplatin-induced NFκB expression and enhanced the NFκB-regulated Bax/Bcl-2 mRNA ratio. Cisplatin alone induced mRNA expression of excision repair cross complementation 1 (ERCC1 and thymidine phosphorylase (TP through phosphorylation of ERK, p38 and PI3K/AKT pathways. However, fucoxanthin pretreatment significantly attenuated cisplatin-induced ERCC1 and TP mRNA expression, leading to improvement of chemotherapeutic efficacy of cisplatin. The results suggest that a combined treatment with fucoxanthin and cisplatin could lead to a potentially important new therapeutic strategy against human hepatoma cells.

  16. Berberine Attenuates Development of the Hepatic Gluconeogenesis and Lipid Metabolism Disorder in Type 2 Diabetic Mice and in Palmitate-Incubated HepG2 Cells through Suppression of the HNF-4α miR122 Pathway.

    Science.gov (United States)

    Wei, Shengnan; Zhang, Ming; Yu, Yang; Lan, Xiaoxin; Yao, Fan; Yan, Xin; Chen, Li; Hatch, Grant M

    2016-01-01

    Berberine (BBR) has been shown to exhibit protective effects against diabetes and dyslipidemia. Previous studies have indicated that BBR modulates lipid metabolism and inhibits hepatic gluconeogensis by decreasing expression of Hepatocyte Nuclear Factor-4α (HNF-4α). However, the mechanism involved in this process was unknown. In the current study, we examined the mechanism of how BBR attenuates hepatic gluconeogenesis and the lipid metabolism alterations observed in type 2 diabetic (T2D) mice and in palmitate (PA)-incubated HepG2 cells. Treatment with BBR for 4 weeks improve all biochemical parameters compared to T2D mice. Treatment of T2D mice for 4 weeks or treatment of PA-incubated HepG2 cells for 24 h with BBR decreased expression of HNF-4α and the microRNA miR122, the key gluconeogenesis enzymes Phosphoenolpyruvate carboxykinase (PEPCK) and Glucose-6-phosphatase (G6Pase) and the key lipid metabolism proteins Sterol response element binding protein-1 (SREBP-1), Fatty acid synthase-1 (FAS-1) and Acetyl-Coenzyme A carboxylase (ACCα) and increased Carnitine palmitoyltransferase-1(CPT-1) compared to T2D mice or PA-incubated HepG2 cells. Expression of HNF-4α in HepG2 cells increased expression of gluconeogenic and lipid metabolism enzymes and BBR treatment or knock down of miR122 attenuated the effect of HNF-4α expression. In contrast, BBR treatment did not alter expression of gluconeogenic and lipid metabolism enzymes in HepG2 cells with knockdown of HNF-4α. In addition, miR122 mimic increased expression of gluconeogenic and lipid metabolism enzymes in HepG2 cells with knockdown of HNF-4α. These data indicate that miR122 is a critical regulator in the downstream pathway of HNF-4α in the regulation of hepatic gluconeogenesis and lipid metabolism in HepG2 cells. The effect of BBR on hepatic gluconeogenesis and lipid metabolism is mediated through HNF-4α and is regulated downstream of miR122. Our data provide new evidence to support HNF-4α and miR122

  17. Effects of Salvia officinalis and Thymus vulgaris on oxidant-induced DNA damage and antioxidant status in HepG2 cells.

    Science.gov (United States)

    Kozics, Katarína; Klusová, Veronika; Srančíková, Annamária; Mučaji, Pavol; Slameňová, Darina; Hunáková, Lubica; Kusznierewicz, Barbara; Horváthová, Eva

    2013-12-01

    Salvia officinalis (SO) and Thymus vulgaris (TV) are medicinal plants well known for their curative powers. However, the molecular mechanisms responsible for these abilities of sage and thyme have not been fully understood yet. In this study we investigated the composition and the quantitative estimation of plant extracts, the protective effects of plant extracts against hydrogen peroxide- and 2,3-dimethoxy-1,4-naphthoquinone-induced DNA damage, and levels of enzymatic and non-enzymatic antioxidants (superoxide dismutase, glutathione peroxidase, glutathione) in human HepG2 cells. To measure antioxidative activity of plant extracts we used three assays: 1,1-diphenyl-2-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). The results showed that the oxidant-induced DNA lesions were significantly reduced in cells pre-treated with the plant extracts studied. The observed DNA-protective activity could be explained by both elevation of GPx activity in cells pre-treated with SO and TV and antioxidant activity of SO and TV. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Anthocyanin-Rich Grape Pomace Extract (Vitis vinifera L. from Wine Industry Affects Mitochondrial Bioenergetics and Glucose Metabolism in Human Hepatocarcinoma HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Nathalia F. F. de Sales

    2018-03-01

    Full Text Available Cancer cells demand high ATP provisions to support proliferation, and targeting of energy metabolism is a good strategy to increase their sensitivity to treatments. In Brazil, wine manufacture is expanding, increasing the amount of pomace that is produced. We determined the phenolic composition and antioxidant properties of a dark skin Grape Pomace Extract and its effects on metabolism and redox state in human hepatocarcinoma HepG2 cells. The material and the methods used represented the industrial process since pomace derived from white wine production and the extract concentrated by pilot plant scale reverse osmosis. Grape pomace extract was rich in polyphenols, mainly anthocyanins, and presented high antioxidant capacity. Short-term metabolic effects, irrespective of any cytotoxicity, involved increased mitochondrial respiration and antioxidant capacity and decreased glycolytic metabolism. Long-term incubation was cytotoxic and cells died by necrosis and GPE was not toxic to non-cancer human fibroblasts. To the best of our knowledge, this is the first report to characterize pomace extract from white wine production from Brazilian winemaking regarding its effects on energy metabolism, suggesting its potential use for pharmaceutical and nutraceutical purposes.

  19. Anthocyanin-Rich Grape Pomace Extract (Vitis vinifera L.) from Wine Industry Affects Mitochondrial Bioenergetics and Glucose Metabolism in Human Hepatocarcinoma HepG2 Cells.

    Science.gov (United States)

    de Sales, Nathalia F F; Silva da Costa, Leandro; Carneiro, Talita I A; Minuzzo, Daniela A; Oliveira, Felipe L; Cabral, Lourdes M C; Torres, Alexandre G; El-Bacha, Tatiana

    2018-03-08

    Cancer cells demand high ATP provisions to support proliferation, and targeting of energy metabolism is a good strategy to increase their sensitivity to treatments. In Brazil, wine manufacture is expanding, increasing the amount of pomace that is produced. We determined the phenolic composition and antioxidant properties of a dark skin Grape Pomace Extract and its effects on metabolism and redox state in human hepatocarcinoma HepG2 cells. The material and the methods used represented the industrial process since pomace derived from white wine production and the extract concentrated by pilot plant scale reverse osmosis. Grape pomace extract was rich in polyphenols, mainly anthocyanins, and presented high antioxidant capacity. Short-term metabolic effects, irrespective of any cytotoxicity, involved increased mitochondrial respiration and antioxidant capacity and decreased glycolytic metabolism. Long-term incubation was cytotoxic and cells died by necrosis and GPE was not toxic to non-cancer human fibroblasts. To the best of our knowledge, this is the first report to characterize pomace extract from white wine production from Brazilian winemaking regarding its effects on energy metabolism, suggesting its potential use for pharmaceutical and nutraceutical purposes.

  20. Schisandra chinensis peptidoglycan-assisted transmembrane transport of lignans uniquely altered the pharmacokinetic and pharmacodynamic mechanisms in human HepG2 cell model.

    Directory of Open Access Journals (Sweden)

    Charng-Cherng Chyau

    Full Text Available Schisandra chinensis (Turz Baill (S. chinensis (SC fruit is a hepatoprotective herb containing many lignans and a large amount of polysaccharides. A novel polysaccharide (called SC-2 was isolated from SC of MW 841 kDa, which exhibited a protein-to-polysaccharide ratio of 0.4089, and showed a characteristic FTIR spectrum of a peptidoglycan. Powder X-ray diffraction revealed microcrystalline structures within SC-2. SC-2 contained 10 monosaccharides and 15 amino acids (essential amino acids of 78.12%w/w. In a HepG2 cell model, SC-2 was shown by MTT and TUNEL assay to be completely non-cytotoxic. A kinetic analysis and fluorescence-labeling technique revealed no intracellular disposition of SC-2. Combined treatment of lignans with SC-2 enhanced the intracellular transport of schisandrin B and deoxyschisandrin but decreased that of gomisin C, resulting in alteration of cell-killing bioactivity. The Second Law of Thermodynamics allows this type of unidirectional transport. Conclusively, SC-2 alters the transport and cell killing capability by a "Catcher-Pitcher Unidirectional Transport Mechanism".

  1. A Novel Polysaccharide Conjugate from Bullacta exarata Induces G1-Phase Arrest and Apoptosis in Human Hepatocellular Carcinoma HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Ningbo Liao

    2017-03-01

    Full Text Available Bullacta exarata has been consumed in Asia, not only as a part of the normal diet, but also as a traditional Chinese medicine with liver- and kidney-benefitting functions. Several scientific investigations involving extraction of biomolecules from this mollusk and pharmacological studies on their biological activities have been carried out. However, little is known regarding the antitumor properties of polysaccharides from B. exarata, hence the polysaccharides from B. exarata have been investigated here. One polysaccharide conjugate BEPS-IA was isolated and purified from B. exarata. It mainly consisted of mannose and glucose in a molar ratio of 1:2, with an average molecular weight of 127 kDa. Thirteen general amino acids were identified to be components of the protein-bound polysaccharide. Methylation and NMR studies revealed that BEPS-IA is a heteropolysaccharide consisting of 1,4-linked-α-d-Glc, 1,6-linked-α-d-Man, 1,3,6-linked-α-d-Man, and 1-linked-α-d-Man residue, in a molar ratio of 6:1:1:1. In order to test the antitumor activity of BEPS-IA, we investigated its effect against the growth of human hepatocellular carcinoma cells HepG2 in vitro. The result showed that BEPS-IA dose-dependently exhibited an effective HepG2 cells growth inhibition with an IC50 of 112.4 μg/mL. Flow cytometry analysis showed that BEPS-IA increased the populations of both apoptotic sub-G1 and G1 phase. The result obtained from TUNEL assay corroborated apoptosis which was shown in flow cytometry. Western blot analysis suggested that BEPS-IA induced apoptosis and growth inhibition were associated with up-regulation of p53, p21 and Bax, down-regulation of Bcl-2. These findings suggest that BEPS-IA may serve as a potential novel dietary agent for hepatocellular carcinoma.

  2. Added value of stress related gene inductions in HepG2 cells as effect measurement in monitoring of air pollution

    Science.gov (United States)

    Nobels, Ingrid; Vanparys, Caroline; Van den Heuvel, Rosette; Vercauteren, Jordy; Blust, Ronny

    2012-08-01

    In this study we studied the effects of particulate matter samples (PM) through gene expression analysis in a routine air quality monitoring campaign by the Flemish Environment Agency (VMM, Belgium). We selected a human hepatoma (HepG2) multiple endpoint reporter assay for targeted stress related endpoint screening. Organic extracts of air samples (total suspended particles, TSP) were collected during one year in an industrial, urban and background location in Flanders, Belgium. Simultaneously, meteorological conditions (temperature, wind speed and precipitation) and particulate matter size ≤ 10 μM (PM10), organic (OC), elemental (EC) and total (TC) carbon were monitored and air samples were collected for chemical analysis (11 PAHs). Correlations between the induction of the different stress genes and the chemical pollutants were analysed. Exposure of HepG2 cells to daily air equivalents (20 m3) of organic TSP extracts revealed the dominant induction of the xenobiotic response element (Xre) and phase I (Cyp1A1) and phase II (GstYa) biotransformation enzymes. Additional effects were the induction of c-Fos, a proto-oncogen and Gadd45, a marker for cell cycle disturbance and responsive to genotoxic compounds. Inductions of other relevant pathways, such as sequestration of heavy metals, retinoids response, protein misfolding and increased cAMP levels were measured occasionally. A significant correlation was found between the genes Cyp1A1 (a typical marker for presence of PAHs and dioxin like compounds), c-Fos, Gadd45, (responsive to DNA damaging compounds) and the amount of PM10 and elemental carbon (EC) whereas no correlation was found between these genes and total PAHs content. This may suggest that the observed induction of Cyp1A1 and DNA damage related genes was provoked (partially) by other particle bound compounds (e.g. pesticides, PCBs, brominated flame retardants, dioxins, …), than PAHs. The contribution of particle bound compounds, other than PAHs might

  3. (Asteraceae) Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    Purpose: To investigate the anti-proliferative and apoptotic activity of crude and dichloromethane fraction of A. sieberi against seven cancer cell lines (Colo20, HCT116, DLD, MCF7, Jurkat, HepG2 and L929). Methods: A. sieberi was extracted with methanol and further purification was carried out using liquidliquid extraction ...

  4. Purification of naringin and neohesperidin from Huyou (Citrus changshanensis) fruit and their effects on glucose consumption in human HepG2 cells.

    Science.gov (United States)

    Zhang, Jiukai; Sun, Chongde; Yan, Youyou; Chen, Qingjun; Luo, Fenglei; Zhu, Xiaoyan; Li, Xian; Chen, Kunsong

    2012-12-01

    Huyou (Citrus changshanensis) is rich in naringin and neohesperidin, which are natural flavanone glycosides with a range of biological activities. Among the different fruit parts, i.e. flavedo, albedo, segment membrane (SM), and juice sacs (JS), albedo showed the highest contents of both compounds, with 27.00 and 19.09mg/g DW for naringin and neohesperidin, respectively. Efficient simultaneous purification of naringin and neohesperidin from Huyou albedo was established by the combination of macroporous D101 resin chromatography and high-speed counter-current chromatography (HSCCC). Purified naringin and neohesperidin were identified by both HPLC and LC-MS, and their effects on glucose consumption were investigated in HepG2 cells. Cells treated with naringin and neohesperidin showed increased consumption of glucose, and this was associated with increased phosphorylation of AMP-activated protein kinase (AMPK). Therefore, naringin and neohesperidin from Huyou may act as potential hypoglycaemic agents through regulation of glucose metabolism. Copyright © 2012 Elsevier Ltd. All rights reserved.

  5. Synergistic Hepatoprotective and Antioxidant Effect of Artichoke, Fig, Blackberry Herbal Mixture on HepG2 Cells and Their Metabolic Profiling Using NMR Coupled with Chemometrics.

    Science.gov (United States)

    Youssef, Fadia S; Labib, Rola M; Eldahshan, Omayma A; Singab, Abdel Nasser B

    2017-12-01

    The edible plants have long been reported to possess a lot of biological activities. Herein, the hepatoprotective and the antioxidant activities of the aqueous infusion of the edible parts of Cynara cardunculus, Ficus carica, and Morus nigra and their herbal mixture (CFM) was investigated in vitro using CCl 4 induced damage in HepG2 cells. The highest amelioration was observed via the consumption of CFM at 1 mg/ml showing 47.00% and 37.09% decline in aspartate transaminase and alanine transaminase and 77.32% and 101.02% increase in reduced glutathione and superoxide dismutase comparable to CCl 4 treated cells. Metabolic profiling of their aqueous infusions was done using nuclear magnetic resonance spectroscopic experiments coupled with chemometrics particularly hierarchical cluster analysis (HCA) and principal component analysis (PCA). The structural closeness of the various metabolites existing in black berry and the mixture as reflected in the PCA score plot and HCA processed from the 1 H-NMR spectral data could eventually explained the close values in their biological behavior. For fig and artichoke, the existence of different phenolic metabolites that act synergistically could greatly interpret their potent biological behavior. Thus, it can be concluded that a herbal mixture composed of black berry, artichoke, and fig could afford an excellent natural candidate to combat oxidative stress and counteract hepatic toxins owing to its phenolic compounds. © 2017 Wiley-VHCA AG, Zurich, Switzerland.

  6. Hyaluronic acid co-functionalized gold nanoparticle complex for the targeted delivery of metformin in the treatment of liver cancer (HepG2 cells).

    Science.gov (United States)

    Kumar, C Senthil; Raja, M D; Sundar, D Sathish; Gover Antoniraj, M; Ruckmani, K

    2015-09-05

    In this study, green synthesis of gold nanoparticles (AuNPs) was achieved using the extract of eggplant as a reducing agent. Hyaluronic acid (HA) serves as a capping and targeting agent. Metformin (MET) was successfully loaded on HA capped AuNPs (H-AuNPs) and this formulation binds easily on the surface of the liver cancer cells. The synthesized nanoparticles were characterized by UV-Vis spectrophotometer, HR-TEM, particle size analyser and zeta potential measurement. Toxicity studies of H-AuNPs in zebra fish confirmed the in vivo safety of the AuNPs. The in vitro cytotoxicity results showed that the amount of MET-H-AuNPs enough to achieve 50% inhibition (IC50) was much lower than free MET. Flow cytometry analysis showed the significant reduction in G2/M phase after treatment with MET-H-AuNPs, and molecular level apoptosis were studied using western blotting. The novelty of this study is the successful synthesis of AuNPs with a higher MET loading and this formulation exhibited better targeted delivery as well as increased regression activity than free MET in HepG2 cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Effective delivery of hydrophobic drugs to breast (MCF-7) and Liver (HepG2) cancer cells: A detailed investigation using Cytotoxicity assays, fluorescence imaging and flow cytometry.

    Science.gov (United States)

    Manatunga, Danushika C; de Silva, Rohini M; Nalin de Silva, K M; Neelika Malavige, Gathsaurie; Wijeratne, Dulharie T; Williams, Gareth R; Jayasinghe, Chanika D; Udagama, Preethi V

    2018-04-03

    This study aimed to develop a drug carrier system consisting of a polymer containing hydroxyapatite (HAp) shell and a magnetic core of iron oxide nanoparticles. Doxorubicin and/or curcumin were loaded into the carrier via a simple diffusion deposition approach, with encapsulation efficiencies (EE) for curcumin and doxorubicin of 93.03 ± 0.3% and 97.37 ± 0.12% respectively. The co-loading of curcumin and doxorubicin led to a total EE of 76.02 ± 0.48%. Release studies were carried out at pH 7.4 and 5.3, and revealed higher release was at pH 5.3 expressing the potential application in tumor microenvironments. Cytotoxicity assays, fluorescence imaging and flow cytometry showed the formulations could effectively inhibit the growth of MCF-7 and HEpG2 cancer cells, being more potent than the free drug molecules both in dose and time dependent manner. Additionally, hemolysis tests and cytotoxicity evaluations determined the drug-loaded carriers to be non-toxic towards non-cancerous cells. These formulations thus have great potential in the development of new cancer therapeutics. Copyright © 2018. Published by Elsevier B.V.

  8. [Transmembrane transport behavior of in vitro HepG2 cells of ananas and its effect on lipids and glucose distribution].

    Science.gov (United States)

    Pang, Yu-Nong; Chai, Yu-Shuang; Jiang, Jing-Fei; Wang, Xin-Pei; Yu, Xuan; Lei, Fan; Xing, Dong-Ming; Du, Li-Jun

    2014-08-01

    Pineapple (Ananas comosus) leaves contain mainly phenolic components with antioxidant and hypolipidemic effects. One of the principle components is p-coumaric acid. In this study, the transport behavior of p-coumaric acid, was observed after the administration of pineapple leaf phenols in vitro. Simultaneously, the effect of the phenols on glucose, total cholesterol and triglycerides transportation and metabolism in HepG2 cells was also observed. The results showed that the phenols had good transport characteristics. 5 min after the administration, p-coumaric acid of the phenols could be detected, and the content of p-coumaric acid reached the peak concentration after 60 min of the administration. p-coumaric acid of phenols have time-and dose-dependent manner. While promoting glucose transporter (GLUT4) and low density lipoprotein receptor (LDLR) expression, the phenols decreased intracellular lipid content. This reduction of intracellular lipid content was highly correlated with the promotion of lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) expression, while the reduction of intracellular glucose levels was correlated with glycogen synthesis in the cells.

  9. Effects of individual and combined toxicity of bisphenol A, dibutyl phthalate and cadmium on oxidative stress and genotoxicity in HepG 2 cells.

    Science.gov (United States)

    Li, Xiaohui; Yin, Pinghe; Zhao, Ling

    2017-07-01

    Bisphenol A, dibutyl phthalate and cadmium can be found in environment simultaneously. Several studies suggested that they had genotoxic effect. In this study, mono-exposure and co-exposure treatments, designed by 3 × 3 full factorial, were established to determine the individual toxicity and binary mixtures' combined effects on the oxidative stress and genotoxicity in HepG 2 cells. The highest oxidative damage was observed in the Cd treatments groups. Compared with control groups, the maximum level of reactive oxygen species and malondialdehyde were ∼1.4 fold and ∼2.22 fold respectively. And a minimum level of superoxide dismutase activity was found with the decrease of 43%. The mechanism that excessive oxidative stress led to the DNA damage was inferred. However, cells treated with BPA showed the worst DNA damage rather than Cd, which may because Cd mainly damages DNA repairing mechanism. For the joint effect, different interactions can be found in different biological endpoints for different combinations since different mechanisms have been clarified in mixture toxicity studies. It is sure that the co-exposure groups enhanced cytotoxicity, oxidative stress and genotoxicity compared to the mono-exposures. Synergistic and additive interactions were considered, which means greater threat to organisms when exposed to multiple estrogenic endocrine disruptors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Sensitivity to anti-Fas is independent of increased cathepsin D activity and adrenodoxin reductase expression occurring in NOS-3 overexpressing HepG2 cells.

    Science.gov (United States)

    Linares, Clara I; Ferrín, Gustavo; Aguilar-Melero, Patricia; González-Rubio, Sandra; Rodríguez-Perálvarez, Manuel; Sánchez-Aragó, María; Chicano-Gálvez, Eduardo; Cuezva, José M; Montero-Álvarez, José L; Muntané, Jordi; de la Mata, Manuel

    2015-05-01

    Stable overexpression of endothelial nitric oxide synthase (NOS-3) in HepG2 cells (4TO-NOS) leads to increased nitro-oxidative stress and upregulation of the cell death mediators p53 and Fas. Thus, NOS-3 overexpression has been suggested as a useful antiproliferative mechanism in hepatocarcinoma cells. We aimed to identify the underlying mechanism of cell death induced by NOS-3 overexpression at basal conditions and with anti-Fas treatment. The intracellular localization of NOS-3, the nitro-oxidative stress and the mitochondrial activity were analysed. In addition, the protein expression profile in 4TO-NOS was screened for differentially expressed proteins potentially involved in the induction of apoptosis. NOS-3 localization in the mitochondrial outer membrane was not associated with changes in the respiratory cellular capacity, but was related to the mitochondrial biogenesis increase and with a higher protein expression of mitochondrial complex IV. Nitro-oxidative stress and cell death in NOS-3 overexpressing cells occurred with the expression increase of pro-apoptotic genes and a higher expression/activity of the enzymes adrenodoxin reductase mitochondrial (AR) and cathepsin D (CatD). CatD overexpression in 4TO-NOS was related to the apoptosis induction independently of its catalytic activity. In addition, CatD activity inhibition by pepstatin A was not effective in blocking apoptosis induced by anti-Fas. In summary, NOS-3 overexpression resulted in an increased sensitivity to anti-Fas induced cell death, independently of AR expression and CatD activity. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Egr2 enhances insulin resistance via JAK2/STAT3/SOCS-1 pathway in HepG2 cells treated with palmitate.

    Science.gov (United States)

    Lu, Lin; Ye, Xinhua; Yao, Qing; Lu, Aijiao; Zhao, Zhen; Ding, Yang; Meng, Chuchen; Yu, Wenlong; Du, Yunfeng; Cheng, JinLuo

    2018-05-01

    Insulin resistance is generally responsible for the pathogenesis of type 2 diabetes mellitus (T2DM). Early growth response proteins-2 (Egr2) has been reported to be able to increase the expression of the suppressors of cytokine signaling-1 (SOCS-1), and impair insulin signaling pathway through suppression of insulin receptor substrates (IRS), including IRS-1 and IRS-2. However, whether Egr2 is directly involved in the development of insulin resistance, and how its potential contributions to insulin resistance still remain unknown. Here, our present investigation found that the expression levels of Egr2 were up-regulated when insulin resistance occurs, and knockdown of Egr2 abolished the effect of insulin resistance in HepG2 cells induced with palmitate (PA). Importantly, inhibition of Egr2 decreased the expression of SOCS-1 as well as reduced phosphorylation of JAK2 and STAT3. And, our data indicated that silencing of Egr2 accelerated hepatic glucose uptake and reversed the impaired lipid metabolism upon insulin resistance. In summary, the present study confirms that Egr2 could deteriorate insulin resistance via the pathway of JAK2/STAT3/SOCS-1 and may shed light on resolving insulin resistance and further the pathogenesis of T2DM. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Body weight management effect of burdock (Arctium lappa L.) root is associated with the activation of AMP-activated protein kinase in human HepG2 cells.

    Science.gov (United States)

    Kuo, Daih-Huang; Hung, Ming-Chi; Hung, Chao-Ming; Liu, Li-Min; Chen, Fu-An; Shieh, Po-Chuen; Ho, Chi-Tang; Way, Tzong-Der

    2012-10-01

    Burdock (Arcticum lappa L.) root is used in folk medicine and also as a vegetable in Asian countries. In the present study, burdock root treatment significantly reduced body weight in rats. To evaluate the bioactive compounds, we successively extracted the burdock root with ethanol (AL-1), and fractionated it with n-hexane (AL-2), ethyl acetate (AL-3), n-butanol (AL-4), and water (AL-5). Among these fractions, AL-2 contained components with the most effective hypolipidemic potential in human hepatoma HepG2 cells. AL-2 decreased the expression of fatty acid synthase (FASN) and inhibited the activity of acetyl-coenzyme A carboxylase (ACC) by stimulating AMP-activated protein kinase (AMPK) through the LKB1 pathway. Three active compounds were identified from the AL-2, namely α-linolenic acid, methyl α-linolenate, and methyl oleate. These results suggest that burdock root is expected to be useful for body weight management. Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Cytoprotective effect exerted by geraniin in HepG2 cells is through microRNA mediated regulation of BACH-1 and HO-1.

    Science.gov (United States)

    Aayadi, Hoda; Mittal, Smriti P K; Deshpande, Anjali; Gore, Makarand; Ghaskadbi, Saroj S

    2017-11-01

    Geraniin, a hydrolysable tannin, used in traditional medicine in Southeast Asia, is known to exhibit various biological activities. As an antioxidant it is known to up-regulate phase II enzyme Heme oxygenase-1 (HO-1). However its mechanism is not clearly understood. Nuclear factor erythroid-derived 2 related factor 2 (Nrf-2) is transcriptionally up-regulated by Extracellular signal-regulated kinase (ERK) 1/2 and retained in nucleus due to inactivated Glycogen synthase kinase 3 beta (GSK-3β). Geraniin additionally down-regulates expression of microRNA 217 and 377 (miR-217 and miR-377) which target HO-1 mRNA. Expression of BTB and CNC homolog 1 (BACH-1), another regulator of HO-1, is also down-regulated by up-regulating microRNA 98 (miR-98), a negative regulator of BACH-1. Thus, geraniin up-regulates HO-1 expression both through activating its positive regulator Nrf-2 and by down-regulating its negative regulator BACH-1. Up-regulation of HO-1 also confers protection to HepG2 cells from tertiary butyl hydroperoxide (TBH) induced cytotoxicity. [BMB Reports 2017; 50(11): 560-565].

  14. Anti-hepatocarcinoma effects of berberine nanosuspension against human HepG2 and Huh7 cells as well as H22 tumor bearing mice

    Science.gov (United States)

    Wang, Zhi-ping; Wu, Jun-biao; Zhou, Qun; Wang, Yi-fei; Chen, Tongsheng

    2014-09-01

    Hepatocarcinoma, a malignant cancer, threaten human life badly. It is a current issue to seek the effective natural remedy from plant to treat cancer due to the resistance of the advanced hepatocarcinoma to chemotherapy. Berberine (Ber), an isoquinoline derivative alkaloid, has a wide range of pharmacological properties and is considered to have anti-hepatocarcinoma effects. However its low oral bioavailability restricts its wide application. In this report, Ber nanosuspension (Ber-NS) composed of Ber and D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) was prepared by high pressure homogenization technique. Both in vitro and in vivo anti-hepatocarcinoma effects of Ber-NS relative to effcacy of bulk Ber were evaluated. The particle size and zeta potential of Ber-NS were 73.1 +/- 3.7 nm and 6.99 +/- 0.17 mV, respectively. Ber-NS exhibited significant inhibitory effects against human HepG2 and Huh7 cells, and the corresponding IC50 values were 8.1 and 4.7 μg/ml (18.3 and 6.5 μg/ml of Ber solution). In vivo studies also showed higher antitumor efficacy, and inhibition rates was 63.7% (41.4 % of Ber solution) at 100 mg/kg intragastric administration in the H22 solid tumor bearing mice. These results suggest that the delivery of Ber as a nanosuspension is a promising approach for treating hepatocarcinoma.

  15. Gdf11/Smad signalling and Cdx proteins cooperate to activate the Hoxc8 early enhancer in HepG2 cells.

    Science.gov (United States)

    Gaunt, Stephen J

    2017-01-01

    Developing anatomy along the head-tail axis of bilaterian embryos is specified, to a large extent, by the overlapping patterns of expression of the Hox genes. Hox gene enhancers respond to a variety of signals in order to regulate these discreet domains of expression. For mouse Hoxc8, the 399bp "early enhancer" plays a major role. Activation of this enhancer is now examined using luciferase expression constructs transfected into HepG2 cells. Constructs are activated by the combined actions of Gdf11/Smad and Cdx protein signalling pathways, both of which are functional in early embryos. Each of these pathways alone has little stimulatory effect. Stimulation by the two pathways together exceeds the sum of the effects of each pathway alone, indicating synergistic activity. By mutation analysis, two Smad binding motifs are identified as mediators of the Gdf11 effect and two Cdx binding motifs mediate the Cdx effect. The two Smad motifs and one of the Cdx sites are conserved from fish to mammals. Gdf11 stimulation is partially inhibited by Specific Inhibitor of Smad3, suggesting that Smad3 plays a part in signal transduction. Fgf2 increases luciferase activation by the Hoxc8 enhancer, but not, apparently, by specific interactions with either Gdf11 or Cdx effects.

  16. The polyphenol extract from Sechium edule shoots inhibits lipogenesis and stimulates lipolysis via activation of AMPK signals in HepG2 cells.

    Science.gov (United States)

    Wu, Cheng-Hsun; Ou, Ting-Tsz; Chang, Chun-Hua; Chang, Xiao-Zong; Yang, Mon-Yuan; Wang, Chau-Jong

    2014-01-22

    Fatty liver may have implications for metabolic syndrome, such as obesity, hypertension, and diabetes. Therefore, the development of pharmacological or natural agents to reduce fat accumulation in the liver is an important effort. The Sechium edule shoots have already been verified to decrease serum lipids and cholesterol and prevent atherosclerosis. However, how Sechium edule shoots modulate hepatic lipid metabolism is unclear. This study was designed to investigate the effects and mechanisms of polyphenol extracts (SPE) of Sechium edule shoots in reducing lipid accumulation in oleic acid-treated HepG2 cells. We found that water extracts (SWE) of Sechium edule shoots could decrease serum and hepatic lipid contents (e.g., triacylglycerol and cholesterol). Furthermore, SWE and SPE through the AMPK (AMP-activating protein kinase) signaling pathway could decrease lipogenic relative enzymes, such as FAS (fatty acid synthase), HMGCoR (HMG-CoA reductase), and SREBPs (sterol regulatory element binding proteins), and increase the expression of CPT-I (carnitine palmitoyltransferase I) and PPARα (peroxisome proliferators activated receptor α), which are critical regulators of hepatic lipid metabolism. These observations suggested that Sechium edule shoots have potential for developing health foods for preventing and remedying fatty liver.

  17. Docosahexanoic acid modifies low-density lipoprotein receptor abundance in HepG2 cells via suppression of the LXRα-Idol pathway.

    Science.gov (United States)

    Zhou, Ying; Guo, Yue; Zhuang, Xiaodong; Du, Zhimin

    2015-03-01

    As a daily supplement, omega‑3 fatty acid is confirmed to be of benefit in hypertriglyceridemia. However, the effect of omega‑3 fatty acids on the low‑density lipoprotein cholesterol (LDL‑C) metabolism remains a controversial issue. In this study, we focused on the regulatory effect of docosahexanoic acid (DHA), one type of omega‑3 fatty acid, exerted on the LDL receptor (LDLR), a determinant regulator of the LDL‑C metabolism, and explored the potential mechanism. We observed that DHA increased hepatic LDLR protein in the presence of 25‑hydroxycholesterol in HepG2 cells but did not alter the mRNA level. Previous studies have identified inducible degrader of the LDLR (Idol) as a novel negative post‑translational modulator of LDLR and a direct transcriptional target of liver X receptor α (LXRα). Since DHA had no effect on the transcriptional level of LDLR, we speculated that the post‑transcriptional pathway LXRα‑Idol participated in this regulation. The results reveal that DHA downregulated the expression of LXRα and Idol in coordination with the upregulation of LDLR expression. Multiple mechanisms are involved in the regulation of LDLR by DHA, and the suppression of the LXRα‑Idol pathway is one of these mechanisms.

  18. Selection of scFvs specific for the HepG2 cell line using ribosome ...

    Indian Academy of Sciences (India)

    Madhsudhan

    USA) was used for in vitro transcription and translation. The reaction volume was 25 μl and the reaction mixture .... translation mixture during panning. The results showed that no false band had been .... the targeted treatment of liver cancer. However, further studies are needed to investigate the binding specificity of the.

  19. Contribution of a 300 kHz alternating magnetic field on magnetic hyperthermia treatment of HepG2 cells.

    Science.gov (United States)

    Wang, Xiaowen; Chen, Youping; Huang, Changshuo; Wang, Xufei; Zhao, Linyun; Zhang, Xiaodong; Tang, Jintian

    2013-02-01

    We investigated the relative contributions of temperature and a 300 kHz alternating magnetic field (AMF) on magnetic hyperthermia treatment (MHT). Our system consisted of an induction coil, which generated AMF by electric current flow, and a newly developed, temperature-controlled circulating water-jacketed glass bottle placed inside the coil. The AMF generator operated at a frequency of 300 kHz with variable field strength ranging from 0 to 11 mT. Four treatment conditions were employed: (A) control (37 °C, 0 mT), (B) AMF exposure (37 °C, 11 mT), (C) hyperthermia (46 °C, 0 mT), and (D) hyperthermia plus AMF exposure (46 °C, 11 mT) for 30 min. Cell viability and apoptotic death rate were estimated. The relative contributions or interactions of hyperthermia (46 °C) and AMF (11 mT) on MHT were evaluated using 2 × 2 factorial experiment analysis. Group A was statistically different (P magnetic hyperthermia. Copyright © 2012 Wiley Periodicals, Inc.

  20. Identification of interacting proteins of retinoid-related orphan nuclear receptor gamma in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Ze-Min Huang1,#, Jun Wu2,#, Zheng-Cai Jia1, Yi Tian1, Jun Tang3, Yan Tang1, Ying Wang2, Yu-Zhang Wu1,* & Bing Ni1,*

    2012-06-01

    Full Text Available The retinoid-related orphan nuclear receptor gamma (RORγplays critical roles in regulation of development, immunity andmetabolism. As transcription factor usually forms a proteincomplex to function, thus capturing and dissecting of theRORγ protein complex will be helpful for exploring themechanisms underlying those functions. After construction ofthe recombinant tandem affinity purification (TAP plasmid,pMSCVpuro RORγ-CTAP(SG, the nuclear localization ofRORγ-CTAP(SG fusion protein was verified. Followingisolation of RORγ protein complex by TAP strategy, sevencandidate interacting proteins were identified. Finally, the heatshock protein 90 (HSP90 and receptor-interacting protein 140(RIP140 were confirmed to interplay with RORγ byco-immunoprecipitation. Interference of HSP90 or/and RIP140genes resulted in dramatically decreased expression ofCYP2C8 gene, the RORγ target gene. Data from this studydemonstrate that HSP90 and RIP140 proteins interact withRORγ protein in a complex format and function asco-activators in the RORγ-mediated regulatory processes ofHepG2 cells.

  1. Regulatory role of kinases and phosphatases on the internalisation of caveolae in HepG2 cells.

    Science.gov (United States)

    Botos, Erzsébet; Turi, Agnes; Müllner, Nándor; Kovalszky, Ilona; Tátrai, Péter; Kiss, Anna L

    2007-01-01

    The caveolar cycle is thought to be regulated by synchronised function of kinases and phosphatases. Using ocadaic acid--a serine/threonine protein phosphatase inhibitor--and an inhibitor of tyrosine phosphatase (sodium orthovanadate) we have followed the internalisation of caveolae. Since albumin binding to its receptor (gp60) can induce pinching off of caveolae from the plasma membrane, we also used this physiological ligand to induce the internalisation. Our confocal microscopic results show that both ocadaic acid and vanadate treatments have significantly decreased caveolin (caveolin-1 and -2) labelling on the cell surface, while the cytoplasmic labelling became much stronger. Quite often large, strongly labelled "granules" appear at the perinuclear region. Very strong caveolin labelling was detected along the actin-cytoskeleton suggesting that caveolae might move along these filaments. Our electron microscopic results also show an intensive caveolae pinching off from the plasma membrane. After ocadaic acid and vanadate treatments the number of surface connected vesicles (caveolae) decreases. At the same time, large multivesicular bodies (termed caveosomes) appear in the perinuclear area of the cytoplasm. By immunoprecipitation and Western blot analysis we detect an increased tyrosine phosphorylation of a approximately 29kDa protein in ocadaic acid and vanadate treated samples. This protein was identified as caveolin-2. No significant change in the tyrosine phosphorylation of caveolin-1 was found. From these data we can conclude that caveolae internalisation is regulated by phosphorylation of caveolin-2.

  2. HNF-4α regulated miR-122 contributes to development of gluconeogenesis and lipid metabolism disorders in Type 2 diabetic mice and in palmitate-treated HepG2 cells.

    Science.gov (United States)

    Wei, Shengnan; Zhang, Ming; Yu, Yang; Xue, Huan; Lan, Xiaoxin; Liu, Shuping; Hatch, Grant; Chen, Li

    2016-11-15

    Hepatocyte Nuclear Factor-4α (HNF-4α) is a key nuclear receptor protein required for liver development. miR-122 is a predominant microRNA expressed in liver and is involved in the regulation of cholesterol and fatty acid metabolism. HNF-4α is know to regulate expression of miR-122 in liver. We examined how HNF-4α regulated gluconeogenesis and lipid metabolism through miR-122 in vivo and in vitro. Expression of miR-122, HNF-4α, phosphoenolpyruvate carboxykinase (PEPCK), glucose-6-phosphatase (G6Pase), sterol response elementary binding protein-1 (SREBP-1), fatty acid synthase-1 (FAS-1), carnitine palmitoyltransferase-1 (CPT-1) and acetyl Coenzyme A carboxylase alpha (ACCα) were determined in livers of Type 2 diabetic mice and in insulin resistant palmitate-treated HepG2 cells. CPT-1 and phosphorylated ACCα expression were significantly decreased in livers of Type 2 diabetic mice and in palmitate-treated HepG2 cells compared to controls. In contrast, expression of miR-122, HNF-4α, PEPCK, G6Pase, SREBP-1, FAS-1 and ACCα were significantly elevated in liver of Type 2 diabetic mice and in palmitate-treated HepG2 cells compared to controls. Expression of HNF-4α increased whereas siRNA knockdown of HNF-4α decreased miR-122 levels in HepG2 cells compared to controls. In addition, expression of HNF-4α in HepG2 cells increased PEPCK, G6Pase, SREBP-1, FAS-1, ACCα mRNA and protein expression and decreased CPT-1 and p-ACCα mRNA and protein expression compared to controls. Addition of miR-122 inhibitors attenuated the HNF-4α mediated effect on expression of these gluconeogenic and lipid metabolism proteins. The results indicate that HNF-4α regulated miR-122 contributes to development of the gluconeogenic and lipid metabolism alterations observed in Type 2 diabetic mice and in palmitate-treated HepG2 cells. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Microarray analysis revealed different gene expression patterns in HepG2 cells treated with low and high concentrations of the extracts of Anacardium occidentale shoots.

    Science.gov (United States)

    Khaleghi, Shaghayegh; Aziz, Azlina Abdul; Razali, Nurhanani; Junit, Sarni Mat

    2011-11-01

    In this study, the effects of low and high concentrations of the Anacardium occidentale shoot extracts on gene expression in liver HepG2 cells were investigated. From MTT assays, the concentration of the shoot extracts that maintained 50% cell viability (IC(50)) was 1.7 mg/ml. Cell viability was kept above 90% at both 0.4 mg/ml and 0.6 mg/ml of the extracts. The three concentrations were subsequently used for the gene expression analysis using Affymetrix Human Genome 1.0 S.T arrays. The microarray data were validated using real-time qRT-PCR. A total of 246, 696 and 4503 genes were significantly regulated (P < 0.01) by at least 1.5-fold in response to 0.4, 0.6 and 1.7 mg/ml of the extracts, respectively. Mutually regulated genes in response to the three concentrations included CDKN3, LOC100289612, DHFR, VRK1, CDC6, AURKB and GABRE. Genes like CYP24A1, BRCA1, AURKA, CDC2, CDK2, CDK4 and INSR were significantly regulated at 0.6 mg/ml and 1.7 mg but not at 0.4 mg/ml. However, the expression of genes including LGR5, IGFBP3, RB1, IDE, LDLR, MTTP, APOB, MTIX, SOD2 and SOD3 were exclusively regulated at the IC(50) concentration. In conclusion, low concentrations of the extracts were able to significantly regulate a sizable number of genes. The type of genes that were expressed was highly dependent on the concentration of the extracts used.

  4. Inhibition of SH2-domain-containing inositol 5-phosphatase (SHIP2) ameliorates palmitate induced-apoptosis through regulating Akt/FOXO1 pathway and ROS production in HepG2 cells

    International Nuclear Information System (INIS)

    Gorgani-Firuzjaee, Sattar; Adeli, Khosrow; Meshkani, Reza

    2015-01-01

    The serine–threonine kinase Akt regulates proliferation and survival by phosphorylating a network of protein substrates; however, the role of a negative regulator of the Akt pathway, the SH2-domain-containing inositol 5-phosphatase (SHIP2) in apoptosis of the hepatocytes, remains unknown. In the present study, we studied the molecular mechanisms linking SHIP2 expression to apoptosis using overexpression or suppression of SHIP2 gene in HepG2 cells exposed to palmitate (0.5 mM). Overexpression of the dominant negative mutant SHIP2 (SHIP2-DN) significantly reduced palmitate-induced apoptosis in HepG2 cells, as these cells had increased cell viability, decreased apoptotic cell death and reduced the activity of caspase-3, cytochrome c and poly (ADP-ribose) polymerase. Overexpression of the wild-type SHIP2 gene led to a massive apoptosis in HepG2 cells. The protection from palmitate-induced apoptosis by SHIP2 inhibition was accompanied by a decrease in the generation of reactive oxygen species (ROS). In addition, SHIP2 inhibition was accompanied by an increased Akt and FOXO-1 phosphorylation, whereas overexpression of the wild-type SHIP2 gene had the opposite effects. Taken together, these findings suggest that SHIP2 expression level is an important determinant of hepatic lipoapotosis and its inhibition can potentially be a target in treatment of hepatic lipoapoptosis in diabetic patients. - Highlights: • Lipoapoptosis is the major contributor to the development of NAFLD. • The PI3-K/Akt pathway regulates apoptosis in different cells. • The role of negative regulator of this pathway, SHIP2 in lipoapoptosis is unknown. • SHIP2 inhibition significantly reduces palmitate-induced apoptosis in HepG2 cells. • SHIP2 inhibition prevents palmitate induced-apoptosis by regulating Akt/FOXO1 pathway

  5. Inhibition of SH2-domain-containing inositol 5-phosphatase (SHIP2) ameliorates palmitate induced-apoptosis through regulating Akt/FOXO1 pathway and ROS production in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Gorgani-Firuzjaee, Sattar [Department of Biochemistry, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Islamic Republic of Iran (Iran, Islamic Republic of); Adeli, Khosrow [Division of Clinical Biochemistry, The Hospital for Sick Children, University of Toronto, Toronto (Canada); Meshkani, Reza, E-mail: rmeshkani@tums.ac.ir [Department of Biochemistry, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Islamic Republic of Iran (Iran, Islamic Republic of)

    2015-08-21

    The serine–threonine kinase Akt regulates proliferation and survival by phosphorylating a network of protein substrates; however, the role of a negative regulator of the Akt pathway, the SH2-domain-containing inositol 5-phosphatase (SHIP2) in apoptosis of the hepatocytes, remains unknown. In the present study, we studied the molecular mechanisms linking SHIP2 expression to apoptosis using overexpression or suppression of SHIP2 gene in HepG2 cells exposed to palmitate (0.5 mM). Overexpression of the dominant negative mutant SHIP2 (SHIP2-DN) significantly reduced palmitate-induced apoptosis in HepG2 cells, as these cells had increased cell viability, decreased apoptotic cell death and reduced the activity of caspase-3, cytochrome c and poly (ADP-ribose) polymerase. Overexpression of the wild-type SHIP2 gene led to a massive apoptosis in HepG2 cells. The protection from palmitate-induced apoptosis by SHIP2 inhibition was accompanied by a decrease in the generation of reactive oxygen species (ROS). In addition, SHIP2 inhibition was accompanied by an increased Akt and FOXO-1 phosphorylation, whereas overexpression of the wild-type SHIP2 gene had the opposite effects. Taken together, these findings suggest that SHIP2 expression level is an important determinant of hepatic lipoapotosis and its inhibition can potentially be a target in treatment of hepatic lipoapoptosis in diabetic patients. - Highlights: • Lipoapoptosis is the major contributor to the development of NAFLD. • The PI3-K/Akt pathway regulates apoptosis in different cells. • The role of negative regulator of this pathway, SHIP2 in lipoapoptosis is unknown. • SHIP2 inhibition significantly reduces palmitate-induced apoptosis in HepG2 cells. • SHIP2 inhibition prevents palmitate induced-apoptosis by regulating Akt/FOXO1 pathway.

  6. Exploiting antidiabetic activity of silver nanoparticles synthesized using Punica granatum leaves and anticancer potential against human liver cancer cells (HepG2).

    Science.gov (United States)

    Saratale, Rijuta G; Shin, Han Seung; Kumar, Gopalakrishnan; Benelli, Giovanni; Kim, Dong-Su; Saratale, Ganesh D

    2018-02-01

    This study first time reports the novel synthesis of silver nanoparticles (AgNPs) using a Punica granatum leaf extract (PGE). The synthesized AgNPs were characterized by various analytical techniques including UV-Vis, Fourier transform infrared (FTIR), X-ray powder diffraction (XRD), X-ray photoelectron spectroscopy (XPS), field emission scanning electron microscopy and energy-dispersive spectra (FESEM-EDS) and high-resolution transmission electron microscopy (HRTEM). FTIR analysis revealed that the involvement of biological macromolecules of P. granatum leaf extract were distributed and involved in the synthesis and stabilization of AgNPs. A surface-sensitive technique of XPS was used to analyse the composition and oxidation state of synthesized AgNPs. The analytical results confirmed that the AgNPs were crystalline in nature with spherical shape. The zeta potential study revealed that the surface charge of synthesized AgNPs was highly negative (-26.6 mV) and particle size distribution was ranging from ∼35 to 60 nm and the average particle size was about 48 nm determined by dynamic light scattering (DLS). The PGE-AgNPs antidiabetic potential exhibited effective inhibition against α-amylase and α-glucosidase (IC 50 ; 65.2 and 53.8 μg/mL, respectively). The PGE-AgNPs showed a dose-dependent response against human liver cancer cells (HepG2) (IC 50 ; 70 μg/mL) indicating its greater efficacy in killing cancer cells. They also possessed in vitro free radical-scavenging activity in terms of ABTS (IC 50 ; 52.2 μg/mL) and DPPH (IC 50 ; 67.1 μg/mL) antioxidant activity. PGE-AgNPs displayed strong antibacterial activity and potent synergy with standard antibiotics against pathogenic bacteria. Thus, synthesized PGE-AgNPs show potential biomedical and industrial applications.

  7. Quantification of homocysteine-related metabolites and the role of betaine-homocysteine S-methyltransferase in HepG2 cells

    Czech Academy of Sciences Publication Activity Database

    Kořínek, M.; Šístek, V.; Mládková, Jana; Mikeš, P.; Jiráček, Jiří; Selicharová, Irena

    2013-01-01

    Roč. 27, č. 1 (2013), s. 111-121 ISSN 0269-3879 R&D Projects: GA ČR(CZ) GAP207/10/1277 Institutional support: RVO:61388963 Keywords : homocysteine * BHMT * LC-MS/MS * HepG2 * metabolites Subject RIV: CE - Biochemistry Impact factor: 1.662, year: 2013

  8. Comparative Cytotoxicity of Glycyrrhiza glabra Roots from Different Geographical Origins Against Immortal Human Keratinocyte (HaCaT), Lung Adenocarcinoma (A549) and Liver Carcinoma (HepG2) Cells.

    Science.gov (United States)

    Basar, Norazah; Oridupa, Olayinka Ayotunde; Ritchie, Kenneth J; Nahar, Lutfun; Osman, Nashwa Mostafa M; Stafford, Angela; Kushiev, Habibjon; Kan, Asuman; Sarker, Satyajit D

    2015-06-01

    Glycyrrhiza glabra L. (Fabaceae), commonly known as 'liquorice', is a well-known medicinal plant. Roots of this plant have long been used as a sweetening and flavouring agent in food and pharmaceutical products, and also as a traditional remedy for cough, upper and lower respiratory ailments, kidney stones, hepatitis C, skin disorder, cardiovascular diseases, diabetes, gastrointestinal ulcers and stomach ache. Previous pharmacological and clinical studies have revealed its antitussive, antiinflammatory, antiviral, antimicrobial, antioxidant, immunomodulatory, hepatoprotective and cardioprotective properties. While glycyrrhizin, a sweet-tasting triterpene saponin, is the principal bioactive compound, several bioactive flavonoids and isoflavonoids are also present in the roots of this plant. In the present study, the cytotoxicity of the methanol extracts of nine samples of the roots of G. glabra, collected from various geographical origins, was assessed against immortal human keratinocyte (HaCaT), lung adenocarcinoma (A549) and liver carcinoma (HepG2) cell lines using the in vitro 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazoliumbromide cell toxicity/viability assay. Considerable variations in levels of cytotoxicity were observed among various samples of G. glabra. Copyright © 2015 John Wiley & Sons, Ltd.

  9. The Inhibitory Effect of 3β-Hydroxy-12-oleanen-27-oic Acid on Growth and Motility of Human Hepatoma HepG2 Cells through JNK and Akt Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Juanjuan Wang

    2013-01-01

    Full Text Available 3β-Hydroxy-12-oleanen-27-oic acid (ATA was a main antitumor active triterpene from the rhizomes of Astilbe chinensis. In this study, we investigated its effects on growth, apoptosis, cell cycle, motility/invasion, and metatasis in human hepatoma HepG2 cells in vitro and antimetastasis of B16-F10 melanoma in mice in vivo, as well as its molecular mechanisms of action using a high-throughput Cancer Pathway Finder PCR Array. ATA could not only induce tumor cells into apoptosis through the activation of both extrinsic and intrinsic pathways, arrest HepG2 cells in G2/M phase, but also suppress the invasion and metastasis abilities of HepG2 cells and the lung metastasis of B16-F10 melanoma in mice. PCR array assay revealed that ATA upregulated 9 genes including CDKN1A, MDM2, CFLAR (CASPER, TNFRSF10B (DR5, c-Jun, IL-8, THBS1, SERPINB5 (maspin, and TNF and downregulated 8 genes such as CCNE1, AKT, ANGPT1, TEK, TGFBR1, MMP9, U-PA, and S100A4. These results indicate that ATA could exert antitumor effects through activating JNK/MAPK and suppressing AKT signal transduction pathways and that ATA might be a potent anticancer agent.

  10. Apolipoprotein M regulates the orphan nuclear receptor LRH-1 gene expression through binding to its promoter region in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Pan Y

    2015-04-01

    Full Text Available Yi Pan,1,2,* Hou-gang Zhou,1,* Hui Zhou,3 Min Hu,1 Li-jun Tang2 1Clinical Laboratory, The Second Xiangya Hospital of Central South University, Changsha, Hunan, People’s Republic of China; 2Molecular Biology Research Center, School of Life Sciences, Central South University, Changsha, Hunan, People’s Republic of China; 3Tumor Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, People’s Republic of China *These authors contributed equally to this work Abstract: Apolipoprotein M (ApoM is predominantly located in the high-density lipoprotein in human plasma. It has been demonstrated that ApoM expression could be regulated by several crucial nuclear receptors that are involved in the bile acid metabolism. In the present study, by combining gene-silencing experiments, overexpression studies, and chromatin immunoprecipitation assays, we showed that ApoM positively regulated liver receptor homolog-1 (LRH-1 gene expression via direct binding to an LRH-1 promoter region (nucleotides -406/-197. In addition, we investigated the effects of farnesoid X receptor agonist GW4064 on hepatic ApoM expression in vitro. In HepG2 cell cultures, both mRNA and protein levels of ApoM and LRH-1 were decreased in a time-dependent manner in the presence of 1 µM GW4064, and the inhibition effect was gradually attenuated after 24 hours. In conclusion, our findings present supportive evidence that ApoM is a regulator of human LRH-1 transcription, and further reveal the importance of ApoM as a critical regulator of bile acids metabolism. Keywords: bile acids, chromatin immunoprecipitation assay, farnesoid X receptor, GW4064, high-density lipoprotein 

  11. The Maillard reaction of a shrimp by-product protein hydrolysate: chemical changes and inhibiting effects of reactive oxygen species in human HepG2 cells.

    Science.gov (United States)

    Zha, Fengchao; Wei, Binbin; Chen, Shengjun; Dong, Shiyuan; Zeng, Mingyong; Liu, Zunying

    2015-06-01

    Recently, much attention has been given to improving the antioxidant activity of protein hydrolysates via the Maillard reaction, but little is known about the cellular antioxidant activity of Maillard reaction products (MRPs) from protein hydrolysates. We first investigated chemical characterization and the cellular antioxidant activity of MRPs in a shrimp (Litopenaeus vannamei) by-product protein hydrolysate (SBH)-glucose system at 110 °C for up to 10 h of heating. Solutions of SBH and glucose were also heated alone as controls. The Maillard reaction greatly resulted in the increase of hydroxymethylfurfural (HMF) and browning intensity, high molecular weight fraction, and reduction of the total amino acid in SBH with the heating time, which correlated well with the free radical scavenging activity of MRPs. MRPs had stronger inhibiting effects on oxidative stress of human HepG2 cells than the original SBH, and its cellular antioxidant activity strongly correlated with free radical scavenging activity, but less affected by the browning intensity and HMF level. The caramelization of glucose partially affected the HMF level and free radical scavenging activity of MRPs, but it was not related to the cellular antioxidant activity. The cellular antioxidant activity of MRPs for 5 h of heating time appeared to reach a maximum level, which was mainly due to carbonyl ammonia condensation reaction. In conclusion, the Maillard reaction is a potential method to increase the cellular antioxidant activity of a shrimp by-product protein hydrolysate, but the higher HMF levels and the lower amino acid content in MRPs should also be considered.

  12. Linalool induces cell cycle arrest and apoptosis in HepG2 cells through oxidative stress generation and modulation of Ras/MAPK and Akt/mTOR pathways.

    Science.gov (United States)

    Rodenak-Kladniew, Boris; Castro, Agustina; Stärkel, Peter; De Saeger, Christine; García de Bravo, Margarita; Crespo, Rosana

    2018-04-15

    Linalool is a plant-derived monoterpene with anticancer activity, however its mechanisms of action remain poorly understood. The aim of this work was to elucidate the anticancer mechanisms of action of linalool in hepatocellular carcinoma (HCC) HepG2 cells. Cell viability and proliferation were determined by WST-1 assay and BrdU incorporation, respectively. Cell cycle analysis was assessed through flow cytometry (FC) and western blot (WB). Apoptosis was determined by caspase-3 activity, TUNEL assay and WB. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were analyzed by FC and fluorescence microscopy. Expression of Ras, MAPKs (ERK, JNK and p38) and Akt/mTOR pathways were evaluated by WB. Linalool (0-2.5 mM) dose-dependently inhibited cell proliferation by inducing G0/G1 cell cycle arrest, through Cdk4 and cyclin A downregulation, p21 and p27 upregulation, and apoptosis, characterized by MMP loss, caspase-3 activation, PARP cleavage and DNA fragmentation. Low concentrations of linalool (1.0 mM) reduced membrane-bound Ras and Akt activity whereas higher amounts (2.0 mM) triggered mTOR inhibition and ROS generation, in correlation with MAPKs activation and Akt phosphorylation. ROS scavenger N-acetyl-L-cysteine partially rescued HepG2 cell growth and prevented MPP depolarization, ERK and JNK activation. Moreover, specific ERK and Akt phosphorylation inhibitors potentiated linalool anti-cancer activity, pointing Akt and ERK activation as pro-survival mechanisms in response to higher concentrations of linalool. This report reveals that linalool induces G0/G1 arrest and apoptosis in HepG2 cells involving Ras, MAPKs and Akt/mTOR pathways and suggests that linalool is a promising anticancer agent for HCC therapy. Copyright © 2018 Elsevier Inc. All rights reserved.

  13. HCV core protein-induced down-regulation of microRNA-152 promoted aberrant proliferation by regulating Wnt1 in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Shifeng Huang

    Full Text Available Hepatitis C virus (HCV has been reported to regulate cellular microRNAs (miRNAs. The HCV core protein is considered to be a potential oncoprotein in HCV-related hepatocellular carcinoma (HCV-HCC, but HCV core-regulated miRNAs are largely unknown. Our preliminary experiments revealed significant down-regulation of microRNA-152 (miR-152 by HCV core protein in HepG2 cells. Through target gene prediction softwares, Wnt1 was predicted to be a potential target of miR-152. The present study was initiated to investigate whether miR-152 is aberrantly regulated by the HCV core protein, and involved in the regulation of the aberrant proliferation of HCV-HCC cells.MiR-152 levels were examined by stem-loop real-time RT-PCR (SLqRT-PCR. Cell proliferation was analyzed by MTT and colony formation assay. Cell cycle analysis was performed by flow cytometry. Luciferase reporter assay was conducted to confirm miRNA-target association. Wnt1 expression was determined by real-time qPCR and Western blotting.HCV core protein significantly suppressed miR-152 expression, and led to significant Wnt1 up-regulation with a concomitant aberrantly promoted proliferation. Moreover, we validated that miR-152 inhibition promoted, while miR-152 mimics inhibited cell proliferation. Using, qRT-PCR and western blot, Wnt1 was demonstrated to be regulated by miR-152. Luciferase activity assay showed that while miR-152 mimics significantly reduced the luciferase activity by 83.76% (P<0.0001, miR-152 inhibitor showed no effect on luciferase reporter. Most notably, salvage expression of miR-152 after Ad-HCV core infection for 24 h almost totally reversed the proliferation-promoting effect of the HCV core protein, and meanwhile, reduced the expression of both Wnt1 mRNA and protein to basal levels.These findings provide important evidence that the reduced miR-152 expression by HCV core protein can indirectly lose an inhibitory effect on Wnt1, which might, at least partially lead to cell

  14. Investigation of testosterone, androstenone, and estradiol metabolism in HepG2 cells and primary culture pig hepatocytes and their effects on 17βHSD7 gene expression.

    Directory of Open Access Journals (Sweden)

    Gang Chen

    Full Text Available Steroid metabolism is important in various species. The accumulation of androgen metabolite, androstenone, in pig adipose tissue is negatively associated with pork flavor, odour and makes the meat unfit for human consumption. The 17β-hydroxysteroid dehydrogenase type 7 (17βHSD7 expressed abundantly in porcine liver, and it was previously suggested to be associated with androstenone levels. Understanding the enzymes and metabolic pathways responsible for androstenone as well as other steroids metabolism is important for improving the meat quality. At the same time, metabolism of steroids is known to be species- and tissue-specific. Therefore it is important to investigate between-species variations in the hepatic steroid metabolism and to elucidate the role of 17βHSD7 in this process. Here we used an effective methodological approach, liquid chromatography coupled with mass spectrometry, to investigate species-specific metabolism of androstenone, testosterone and beta-estradiol in HepG2 cell line, and pig cultured hepatocytes. Species- and concentration-depended effect of steroids on 17βHSD7 gene expression was also investigated. It was demonstrated that the investigated steroids can regulate the 17βHSD7 gene expression in HepG2 and primary cultured porcine hepatocytes in a concentration-dependent and species-dependent pattern. Investigation of steroid metabolites demonstrated that androstenone formed a 3'-hydroxy compound 3β-hydroxy-5α-androst-16-ene. Testosterone was metabolized to 4-androstene-3,17-dione. Estrone was found as the metabolite for β-estradiol. Inhibition study with 17βHSD inhibitor apigenin showed that apigenin didn't affect androstenone metabolism. Apigenin at high concentration (50 µM tends to inhibit testosterone metabolism but this inhibition effect was negligible. Beta-estradiol metabolism was notably inhibited with apigenin at high concentration. The study also established that the level of testosterone and

  15. Investigation of testosterone, androstenone, and estradiol metabolism in HepG2 cells and primary culture pig hepatocytes and their effects on 17βHSD7 gene expression.

    Science.gov (United States)

    Chen, Gang; Li, Sicong; Dong, Xinxing; Bai, Ying; Chen, Ailiang; Yang, Shuming; Fang, Meiying; Zamaratskaia, Galia; Doran, Olena

    2012-01-01

    Steroid metabolism is important in various species. The accumulation of androgen metabolite, androstenone, in pig adipose tissue is negatively associated with pork flavor, odour and makes the meat unfit for human consumption. The 17β-hydroxysteroid dehydrogenase type 7 (17βHSD7) expressed abundantly in porcine liver, and it was previously suggested to be associated with androstenone levels. Understanding the enzymes and metabolic pathways responsible for androstenone as well as other steroids metabolism is important for improving the meat quality. At the same time, metabolism of steroids is known to be species- and tissue-specific. Therefore it is important to investigate between-species variations in the hepatic steroid metabolism and to elucidate the role of 17βHSD7 in this process. Here we used an effective methodological approach, liquid chromatography coupled with mass spectrometry, to investigate species-specific metabolism of androstenone, testosterone and beta-estradiol in HepG2 cell line, and pig cultured hepatocytes. Species- and concentration-depended effect of steroids on 17βHSD7 gene expression was also investigated. It was demonstrated that the investigated steroids can regulate the 17βHSD7 gene expression in HepG2 and primary cultured porcine hepatocytes in a concentration-dependent and species-dependent pattern. Investigation of steroid metabolites demonstrated that androstenone formed a 3'-hydroxy compound 3β-hydroxy-5α-androst-16-ene. Testosterone was metabolized to 4-androstene-3,17-dione. Estrone was found as the metabolite for β-estradiol. Inhibition study with 17βHSD inhibitor apigenin showed that apigenin didn't affect androstenone metabolism. Apigenin at high concentration (50 µM) tends to inhibit testosterone metabolism but this inhibition effect was negligible. Beta-estradiol metabolism was notably inhibited with apigenin at high concentration. The study also established that the level of testosterone and β-estradiol metabolites

  16. Upregulation of heat shock protein 70 and the differential protein expression induced by tumor necrosis factor-alpha enhances migration and inhibits apoptosis of hepatocellular carcinoma cell HepG2

    OpenAIRE

    Huang, Bee-Piao; Lin, Chun-Shiang; Wang, Chau-Jong; Kao, Shao-Hsuan

    2017-01-01

    Tumor necrosis factor alpha (TNF?) plays diverse roles in liver damage and hepatocarcinogenesis with its multipotent bioactivity. However, the influence of TNF? on protein expression of hepatocellular carcinoma (HCC) is incompletely understood. Therefore, we aimed to investigate the differential protein expression of HCC in response to TNF? stimulus. We observed that HepG2 cell revealed a higher resistance to TNF?-induced apoptosis as compared to the non-tumorigenic hepatocyte THLE-2. By usin...

  17. Epigallocatechin gallate induces an up-regulation of LDL receptor accompanied by a reduction of PCSK9 via the annexin A2-independent pathway in HepG2 cells.

    Science.gov (United States)

    Kitamura, Kohei; Okada, Yudai; Okada, Kenji; Kawaguchi, Yuya; Nagaoka, Satoshi

    2017-08-01

    In animal studies, epigallocatechin gallate (EGCG), the dominant catechin in green tea, has been shown to improve cholesterol metabolism. However, the molecular mechanisms of EGCG underlying these functions have not been fully understood. In this study, we aimed to clarify the molecular mechanisms of the effect of EGCG on cholesterol metabolism mainly in HepG2 cells. We found that EGCG induced a reduction of the extracellular proprotein convertase subtilisin/kexin 9 (PCSK9) level accompanied by an up-regulation of the LDL receptor (LDLR) in HepG2 cells. The EGCG-induced up-regulation of LDLR occurred via the extracellular signal-regulated kinase (ERK) signaling pathway. Moreover, we showed that EGCG induced a significant early reduction of the extracellular PCSK9 protein level. However, there were no significant changes in the PCSK9 mRNA and the intracellular PCSK9 protein levels induced by EGCG. Annexin A2 knockdown affected the basal LDLR expression and did not affect the EGCG-induced reduction of the extracellular PCSK9 protein level or the up-regulation of LDLR. Annexin A2 possesses an essential function for the basal LDLR expression in HepG2 cells. But, EGCG induces the suppression of PCSK9 accompanied by an up-regulation of LDLR in an annexin A2-independent manner. EGCG attenuates the statin-induced an increase in PCSK9 level. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. MicroRNA-21 promotes hepatocellular carcinoma HepG2 cell proliferation through repression of mitogen-activated protein kinase-kinase 3

    International Nuclear Information System (INIS)

    Xu, Guangxian; Zhang, Yilin; Wei, Jun; Jia, Wei; Ge, Zhaohui; Zhang, Zhaobo; Liu, Xiaoming

    2013-01-01

    microRNA 21 (miR-21) has been demonstrated to be significantly elevated in many types of cancers, including the hepatocellular carcinoma (HCC). In the present study, we investigated the role of miR-21 in HCC by identifying its novel targets, as well as its underlying molecular mechanism. The expression of mitogen-activated protein kinase-kinase 3 (MAP2K3) in human HCC tumor tissues and adjacent non-tumor tissues was determined by immunohistochemistry staining (IHC) analysis. The 3’-untranslated region (3’-UTR) of MAP2K3 combined with miR-21 was experimentally verified by a miRNA luciferase reporter approach. Moreover, the role of miR-21 in regulating HCC cell proliferation was analyzed by an MTT assay infected with miR-21mimics/sponge inhibitor Adenoviral viral vectors. By immunohistochemistry staining analysis, we found that mitogen-activated protein kinase-kinase 3 (MAP2K3) was strikingly repressed in the human HCC tumor tissues, in comparison with the adjacent non-tumor tissues in clinical settings. More importantly, the repression of MAP2K3 was inversely correlated with the expression of miR-21 in HCC. Further study demonstrated that the MAP2K3 was a novel direct target of miR-21, which was experimentally validated by a miRNA luciferase reporter approach. In HepG2 cells, inhibition of miR-21 expression with an adenoviral miR-21 sponge vector profoundly suppressed cell proliferation by up-regulating MAP2K3 expression at both mRNA and protein levels. These results provide a clinical evidence that MAP2K3 may be a tumor repressor gene, and it is a direct target of miR-21 in HCC, indicating an underlying mechanism by which miR-21 is able to directly target MAP2K3 and inhibit its expression during the carcinogenesis of HCC, at both transcriptional and post-translational levels. This study also suggests that targeting miR-21-MAP2K3 pathway may be a promising strategy in the prevention and treatment of HCC

  19. Cytotoxicity and Expression of c-fos, HSP70, and GADD45/153 Proteins in Human Liver Carcinoma (HepG2 Cells Exposed to Dinitrotoluenes

    Directory of Open Access Journals (Sweden)

    Paul B. Tchounwou

    2005-08-01

    Full Text Available Dinitrotoluenes (DNTs are byproducts of the explosive trinitrotoluene (TNT, and exist as a mixture of 2 to 6 isomers, with 2,4-DNT and 2,6-DNT being the most significant. The main route of human exposure at ammunition facilities is inhalation. The primary targets of DNTs toxicity are the hematopoietic system, cardiovascular system, nervous system and reproductive system. In factory workers, exposure to DNTs has been linked to many adverse health effects, including: cyanosis, vertigo, headache, metallic taste, dyspnea, weakness and lassitude, loss of appetite, nausea, and vomiting. Other symptoms including pain or parasthesia in extremities, abdominal discomfort, tremors, paralysis, chest pain, and unconsciousness have been documented. An association between DNTs exposure and increased risk of hepatocellular carcinomas and subcutaneous tumors in rats, as well as renal tumors in mice, has been established. This research was therefore designed targeting the liver to assess the cellular and molecular responses of human liver carcinoma cells following exposure to 2,4-DNT and 2,6-DNT. Cytotoxicity was evaluated using the MTT assay. Upon 48 hrs of exposure, LC50 values of 245 + 14.72μg/mL, and 300 + 5.92μg/mL were recorded for 2,6-DNT and 2,4-DNT respectively, indicating that both DNTs are moderately toxic, and 2,6-DNT is slightly more toxic to HepG2 cells than 2,4-DNT. A dose response relationship was recorded with respect to the cytotoxicity of both DNTs. Western blot analysis resulted in a significant expression (p<0.05 of the 70-kDa heat shock protein in 2,6-DNT-treated cells compared to the control cells and at the 200 μg/mL dose for 2,4-DNT. A statistically significant expression in c-fos was also observed at the 200 and 250 μg/mL treatment level for 2,4- and 2,6-DNT, respectively. However, no statistically significant expression of this protooncogene-related protein was observed at the doses of 0, 100, or 300

  20. 15-deoxy-Δ12,14-prostaglandin J2 Down-Regulates Activin-Induced Activin Receptor, Smad, and Cytokines Expression via Suppression of NF-κB and MAPK Signaling in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Seung-Won Park

    2013-01-01

    Full Text Available 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2 and activin are implicated in the control of apoptosis, cell proliferation, and inflammation in cells. We examined both the mechanism by which 15d-PGJ2 regulates the transcription of activin-induced activin receptors (ActR and Smads in HepG2 cells and the involvement of the nuclear factor-κB (NF-κB and mitogen-activated protein kinase (MAPK pathways in this regulation. Activin A (25 ng/mL inhibited HepG2 cell proliferation, whereas 15d-PGJ2 (2 μM and 5 μM had no effect. Activin A and 15d-PGJ2 showed different regulatory effects on ActR and Smad expression, NF-κB p65 activity and MEK/ERK phosphorylation, whereas they both decreased IL-6 production and increased IL-8 production. When co-stimulated with 15d-PGJ2 and activin, 15d-PGJ2 inhibited the activin-induced increases in ActR and Smad expression, and decreased activin-induced IL-6 production. However, it increased activin-induced IL-8 production. In addition, 15d-PGJ2 inhibited activin-induced NF-κB p65 activity and activin-induced MEK/ERK phosphorylation. These results suggest that 15d-PGJ2 suppresses activin-induced ActR and Smad expression, down-regulates IL-6 production, and up-regulates IL-8 production via suppression of NF-κB and MAPK signaling pathway in HepG2 cells. Regulation of ActR and Smad transcript expression and cytokine production involves NF-κB and the MAPK pathway via interaction with 15d-PGJ2/activin/Smad signaling.

  1. SM22{alpha}-induced activation of p16{sup INK4a}/retinoblastoma pathway promotes cellular senescence caused by a subclinical dose of {gamma}-radiation and doxorubicin in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Tae Rim; Lee, Hee Min; Lee, So Yong; Kim, Eun Jin; Kim, Kug Chan [Department of Radiation Biology, Environmental Radiation Research Group, Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Paik, Sang Gi [Department of Biology, School of Biosciences and Biotechnology, Chungnam National University, Daejeon (Korea, Republic of); Cho, Eun Wie, E-mail: ewcho@kribb.re.kr [Daejeon-KRIBB-FHCRC Cooperation Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Kim, In Gyu, E-mail: igkim@kaeri.re.kr [Department of Radiation Biology, Environmental Radiation Research Group, Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2010-09-10

    Research highlights: {yields} SM22{alpha} overexpression in HepG2 cells leads cells to a growth arrest state, and the treatment of a subclinical dose of {gamma}-radiation or doxorubicin promotes cellular senescence. {yields} SM22{alpha} overexpression elevates p16{sup INK4a} followed by pRB activation, but there are no effects on p53/p21{sup WAF1/Cip1} pathway. {yields} SM22{alpha}-induced MT-1G activates p16{sup INK4a}/pRB pathway, which promotes cellular senescence by damaging agents. -- Abstract: Smooth muscle protein 22-alpha (SM22{alpha}) is known as a transformation- and shape change-sensitive actin cross-linking protein found in smooth muscle tissue and fibroblasts; however, its functional role remains uncertain. We reported previously that SM22{alpha} overexpression confers resistance against anti-cancer drugs or radiation via induction of metallothionein (MT) isozymes in HepG2 cells. In this study, we demonstrate that SM22{alpha} overexpression leads cells to a growth arrest state and promotes cellular senescence caused by treatment with a subclinical dose of {gamma}-radiation (0.05 and 0.1 Gy) or doxorubicin (0.01 and 0.05 {mu}g/ml), compared to control cells. Senescence growth arrest is known to be controlled by p53 phosphorylation/p21{sup WAF1/Cip1} induction or p16{sup INK4a}/retinoblastoma protein (pRB) activation. SM22{alpha} overexpression in HepG2 cells elevated p16{sup INK4a} followed by pRB activation, but did not activate the p53/p21{sup WAF1/Cip1} pathway. Moreover, MT-1G, which is induced by SM22{alpha} overexpression, was involved in the activation of the p16{sup INK4a}/pRB pathway, which led to a growth arrest state and promoted cellular senescence caused by damaging agents. Our findings provide the first demonstration that SM22{alpha} modulates cellular senescence caused by damaging agents via regulation of the p16{sup INK4a}/pRB pathway in HepG2 cells and that these effects of SM22{alpha} are partially mediated by MT-1G.

  2. SM22α-induced activation of p16INK4a/retinoblastoma pathway promotes cellular senescence caused by a subclinical dose of γ-radiation and doxorubicin in HepG2 cells

    International Nuclear Information System (INIS)

    Kim, Tae Rim; Lee, Hee Min; Lee, So Yong; Kim, Eun Jin; Kim, Kug Chan; Paik, Sang Gi; Cho, Eun Wie; Kim, In Gyu

    2010-01-01

    Research highlights: → SM22α overexpression in HepG2 cells leads cells to a growth arrest state, and the treatment of a subclinical dose of γ-radiation or doxorubicin promotes cellular senescence. → SM22α overexpression elevates p16 INK4a followed by pRB activation, but there are no effects on p53/p21 WAF1/Cip1 pathway. → SM22α-induced MT-1G activates p16 INK4a /pRB pathway, which promotes cellular senescence by damaging agents. -- Abstract: Smooth muscle protein 22-alpha (SM22α) is known as a transformation- and shape change-sensitive actin cross-linking protein found in smooth muscle tissue and fibroblasts; however, its functional role remains uncertain. We reported previously that SM22α overexpression confers resistance against anti-cancer drugs or radiation via induction of metallothionein (MT) isozymes in HepG2 cells. In this study, we demonstrate that SM22α overexpression leads cells to a growth arrest state and promotes cellular senescence caused by treatment with a subclinical dose of γ-radiation (0.05 and 0.1 Gy) or doxorubicin (0.01 and 0.05 μg/ml), compared to control cells. Senescence growth arrest is known to be controlled by p53 phosphorylation/p21 WAF1/Cip1 induction or p16 INK4a /retinoblastoma protein (pRB) activation. SM22α overexpression in HepG2 cells elevated p16 INK4a followed by pRB activation, but did not activate the p53/p21 WAF1/Cip1 pathway. Moreover, MT-1G, which is induced by SM22α overexpression, was involved in the activation of the p16 INK4a /pRB pathway, which led to a growth arrest state and promoted cellular senescence caused by damaging agents. Our findings provide the first demonstration that SM22α modulates cellular senescence caused by damaging agents via regulation of the p16 INK4a /pRB pathway in HepG2 cells and that these effects of SM22α are partially mediated by MT-1G.

  3. Induction of Apoptosis by Ethanolic Extract of Corchorus olitorius Leaf in Human Hepatocellular Carcinoma (HepG2 Cells via a Mitochondria-Dependent Pathway

    Directory of Open Access Journals (Sweden)

    Shih-Fang Tsang

    2012-08-01

    Full Text Available Corchorus olitorius L., is a culinary and medicinal herb, widely used as a vegetable in several countries in Asia. Many studies have shown that C. olitorius contains several antioxidants and exhibits anti-inflammatory and anti-proliferative activities in various in vitro and in vivo settings. Recently, C. olitorius has been approved for its antitumor activity; however, the underlying molecular mechanisms remain unclear. The goal of this study was to investigate the effects of ethanol extract of C. olitorius (ECO on the growth of human hepatocellular carcinoma (HepG2 cells and gain some insights into the underlying mechanisms of its action. We found that HepG2 cells, treated with ECO for 24 h at a concentration higher than 12.5 μg/mL, displayed a strong reduction in cell viability, whereas normal FL83B hepatocytes were not affected. DNA fragmentation and nuclear condensation were evidenced by the increased subG1 population of ECO-treated HepG2 cells. ECO triggered the activation of procaspases-3 and -9 and caused the cleavage of downstream substrate, poly ADP-ribose polymerase (PARP, followed by down-regulation of the inhibitor of caspase-activated DNase (ICAD signaling. Moreover, the increased release of cytochrome c from mitochondria with decreased membrane potential demonstrated the apoptosis induced through the caspases cascade. Our findings indicated that ECO might be effective against hepatocellular carcinoma through induction of apoptosis via mitochondria-dependent pathway.

  4. Fumosorinone, a novel PTP1B inhibitor, activates insulin signaling in insulin-resistance HepG2 cells and shows anti-diabetic effect in diabetic KKAy mice

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhi-Qin [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China); College of Pharmaceutical Sciences, key laboratory of pharmaceutical quality control of Hebei province, Hebei University, Baoding 071002 (China); Liu, Ting; Chen, Chuan [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China); Li, Ming-Yan; Wang, Zi-Yu; Chen, Ruo-song; Wei, Gui-xiang; Wang, Xiao-yi [College of Pharmaceutical Sciences, key laboratory of pharmaceutical quality control of Hebei province, Hebei University, Baoding 071002 (China); Luo, Du-Qiang, E-mail: duqiangluo999@126.com [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China)

    2015-05-15

    Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of the insulin signaling pathways, and its increased activity and expression are implicated in the pathogenesis of insulin resistance. Therefore, the inhibition of PTP1B is anticipated to become a potential therapeutic strategy to treat T2DM. Fumosorinone (FU), a new natural product isolated from insect fungi Isaria fumosorosea, was found to inhibit PTP1B activity in our previous study. Herein, the effects of FU on insulin resistance and mechanism in vitro and in vivo were investigated. FU increased the insulin-provoked glucose uptake in insulin-resistant HepG2 cells, and also reduced blood glucose and lipid levels of type 2 diabetic KKAy mice. FU decreased the expression of PTP1B both in insulin-resistant HepG2 cells and in liver tissues of diabetic KKAy mice. Furthermore, FU increased the phosphorylation of IRβ, IRS-2, Akt, GSK3β and Erk1/2 in insulin-resistant HepG2 cells, as well as the phosphorylation of IRβ, IRS-2, Akt in liver tissues of diabetic KKAy mice. These results showed that FU increased glucose uptake and improved insulin resistance by down-regulating the expression of PTP1B and activating the insulin signaling pathway, suggesting that it may possess antidiabetic properties. - Highlights: • Fumosorinone is a new PTP1B inhibitor isolated from insect pathogenic fungi. • Fumosorinone attenuated the insulin resistance both in vitro and in vivo. • Fumosorinone decreased the expression of PTP1B both in vitro and in vivo. • Fumosorinone activated the insulin signaling pathway both in vitro and in vivo.

  5. Fumosorinone, a novel PTP1B inhibitor, activates insulin signaling in insulin-resistance HepG2 cells and shows anti-diabetic effect in diabetic KKAy mice

    International Nuclear Information System (INIS)

    Liu, Zhi-Qin; Liu, Ting; Chen, Chuan; Li, Ming-Yan; Wang, Zi-Yu; Chen, Ruo-song; Wei, Gui-xiang; Wang, Xiao-yi; Luo, Du-Qiang

    2015-01-01

    Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of the insulin signaling pathways, and its increased activity and expression are implicated in the pathogenesis of insulin resistance. Therefore, the inhibition of PTP1B is anticipated to become a potential therapeutic strategy to treat T2DM. Fumosorinone (FU), a new natural product isolated from insect fungi Isaria fumosorosea, was found to inhibit PTP1B activity in our previous study. Herein, the effects of FU on insulin resistance and mechanism in vitro and in vivo were investigated. FU increased the insulin-provoked glucose uptake in insulin-resistant HepG2 cells, and also reduced blood glucose and lipid levels of type 2 diabetic KKAy mice. FU decreased the expression of PTP1B both in insulin-resistant HepG2 cells and in liver tissues of diabetic KKAy mice. Furthermore, FU increased the phosphorylation of IRβ, IRS-2, Akt, GSK3β and Erk1/2 in insulin-resistant HepG2 cells, as well as the phosphorylation of IRβ, IRS-2, Akt in liver tissues of diabetic KKAy mice. These results showed that FU increased glucose uptake and improved insulin resistance by down-regulating the expression of PTP1B and activating the insulin signaling pathway, suggesting that it may possess antidiabetic properties. - Highlights: • Fumosorinone is a new PTP1B inhibitor isolated from insect pathogenic fungi. • Fumosorinone attenuated the insulin resistance both in vitro and in vivo. • Fumosorinone decreased the expression of PTP1B both in vitro and in vivo. • Fumosorinone activated the insulin signaling pathway both in vitro and in vivo

  6. Bitter melon extract ameliorates palmitate-induced apoptosis via inhibition of endoplasmic reticulum stress in HepG2 cells and high-fat/high-fructose-diet-induced fatty liver

    Directory of Open Access Journals (Sweden)

    Hwa Joung Lee

    2018-03-01

    Full Text Available Background: Bitter melon (BM improves glucose level, lipid homeostasis, and insulin resistance in vivo. However, the preventive mechanism of BM in nonalcoholic fatty liver disease (NAFLD has not been elucidated yet. Aim & Design: To determine the protective mechanism of bitter melon extract (BME, we performed experiments in vitro and in vivo. BME were treated palmitate (PA-administrated HepG2 cells. C57BL/6J mice were divided into two groups: high-fat/high-fructose (HF/HFr without or with BME supplementation (100 mg/kg body weight. Endoplasmic reticulum (ER stress, apoptosis, and biochemical markers were then examined by western blot and real-time PCR analyses. Results: BME significantly decreased expression levels of ER-stress markers (including phospho-eIF2α, CHOP, and phospho-JNK [Jun N-terminal kinases] in PA-treated HepG2 cells. BME also significantly decreased the activity of cleaved caspase-3 (a well known apoptotic-induced molecule and DNA fragmentation. The effect of BME on ER stress–mediated apoptosis in vitro was similarly observed in HF/HFr-fed mice in vivo. BME significantly reduced HF/HFr-induced hepatic triglyceride (TG and serum alanine aminotransferase (ALT as markers of hepatic damage in mice. In addition, BME ameliorated HF/HFr-induced serum TG and serum-free fatty acids. Conclusion: These data indicate that BME has protective effects against ER stress mediated apoptosis in HepG2 cells as well as in HF/HFr-induced fatty liver of mouse. Therefore, BME might be useful for preventing and treating NAFLD.

  7. Protective effect of fucosterol isolated from the edible brown algae, Ecklonia stolonifera and Eisenia bicyclis, on tert-butyl hydroperoxide- and tacrine-induced HepG2 cell injury.

    Science.gov (United States)

    Choi, Jae Sue; Han, Yu Ran; Byeon, Jeong Su; Choung, Se-Young; Sohn, Hee Sook; Jung, Hyun Ah

    2015-08-01

    Fucosterol is the primary sterol found in brown algae. Recently, considerable interest has been generated regarding fucosterol due to its potential antioxidant, anti-inflammatory and antidiabetic effects. The aim of this study was to investigate the protective effects of fucosterol on tert-butyl hydroperoxide (t-BHP)- and tacrine-induced oxidative stress in HepG2 cells. Fucosterol by itself exhibited no cytotoxicity at concentrations below 100 μm by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The increased intracellular reactive oxygen species (ROS) and decreased glutathione levels observed in t-BHP- and tacrine-treated HepG2 cells were ameliorated by fucosterol pretreatment, indicating that the protective effects of fucosterol are mediated by the induction of cellular defence mechanisms against oxidative stress. Moreover, elevated alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in tacrine-treated mice were significantly reduced after oral administration of fucosterol. The hepatoprotective effects of fucosterol may occur via an increase in the hepatic level of glutathione and a decrease in ROS production, thereby preventing hepatic damage and the resultant increases in ALT and AST activity. These results suggest that fucosterol may be an effective hepatoprotective agent that could be useful for preventive therapies against oxidative stress-related hepatotoxicity. © 2015 Royal Pharmaceutical Society.

  8. Concomitant activation of caspase-9 and down-regulation of IAP proteins as a mechanism of apoptotic death in HepG2, T47D and HCT-116 cells upon exposure to a derivative from 4-aryl-4H-chromenes family.

    Science.gov (United States)

    Mahdavi, Majid; Davoodi, Jamshid; Zali, Mohammad Reza; Foroumadi, Alireza

    2011-06-01

    Apoptosis inducing activity of 2-amino-4-(3-nitrophenyl)-3-cyano-7-(dimethylamino)-4H-chromene (3-NC) was investigated in three human cancer cell lines of HepG2, liver cancer, T47D, breast cancer, and HCT116, colon carcinoma cancer. This compound was found to be highly active cell proliferation inhibitor with IC50 values of 55, 60 and 50 nM, respectively as determined by thiazolyl blue tetrazolium bromide (MTT) cell proliferation assays. Proliferation of HepG2, T47D and HCT116 cells was diminished by more than 70% and viability was decreased by about 50% upon 72 h of treatment at IC50 concentration of the compound. Apoptosis as the mechanism of cell death was confirmed morphologically by Hoechst 33258 staining, caspase-3 activation assay, as well as DNA ladder formation. In addition to increased caspase-3 like activity as assessed by the cleavage of DEVD-pNA, caspase-9 was also cleaved indicating the activation of the intrinsic apoptosis pathway. Furthermore, Western Blot analysis revealed that treatment with 3-NC down-regulated the expression of inhibitor of apoptosis proteins, cIAP2, XIAP and survivin in cell line dependent manner. The effectiveness of the compound was the highest when all of the above-mentioned IAPs were down-regulated simultaneously, suggesting that for efficient cancer therapy, multiple IAPs rather than an individual IAP like XIAP should be targeted. It further suggests that 3-NC may be useful for the treatment of cancer types prone to down-regulation of these IAPs. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  9. Exposure to the common food additive carrageenan leads to glucose intolerance, insulin resistance and inhibition of insulin signalling in HepG2 cells and C57BL/6J mice.

    Science.gov (United States)

    Bhattacharyya, S; O-Sullivan, I; Katyal, S; Unterman, T; Tobacman, J K

    2012-01-01

    The aim of this study was to determine the impact of the common food additive carrageenan (E407) on glucose tolerance, insulin sensitivity and insulin signalling in a mouse model and human hepatic cells, since carrageenan is known to cause inflammation through interaction with toll-like receptor (TLR)4, which is associated with inflammation in diabetes. Male C57BL/6J mice were given carrageenan (10 mg/l) in their drinking water, and underwent a glucose tolerance test (GTT), an insulin tolerance test (ITT) and an ante-mortem intraperitoneal insulin injection. HepG2 cells were exposed to carrageenan (1 mg/l × 24 h) and insulin. Levels of phospho(Ser473)-protein kinase B (Akt), phospho(Ser307)-IRS1, phosphoinositide 3-kinase (PI3K) activity and phospho(Ser32)-inhibitor of κB (IκBα) were determined by western blotting and ELISA. Glucose tolerance was significantly impaired in carrageenan-treated 12-week-old mice compared with untreated controls at all time points (n = 12; p < 0.0001). Baseline insulin and insulin levels at 30 min after taking glucose during the GTT were significantly higher following carrageenan treatment. During the ITT, glucose levels declined by more than 80% in controls, but not in carrageenan-treated mice. Carrageenan exposure completely inhibited insulin-induced increases in phospho-(Ser473)-Akt and PI3K activity in vivo in mouse liver and in human HepG2 cells. Carrageenan increased phospho(Ser307)-IRS1 levels, and this was blocked when carrageenan-induced inflammation was inhibited. This is the first report of the impact of carrageenan on glucose tolerance and indicates that carrageenan impairs glucose tolerance, increases insulin resistance and inhibits insulin signalling in vivo in mouse liver and human HepG2 cells. These effects may result from carrageenan-induced inflammation. The results demonstrate extra-colonic manifestations of ingested carrageenan and suggest that carrageenan in the human diet may contribute to the

  10. Contribution of ionic silver to genotoxic potential of nanosilver in human liver HepG2 and colon Caco2 cells evaluated by the cytokinesis-block micronucleus assay.

    Science.gov (United States)

    Sahu, Saura C; Roy, Shambhu; Zheng, Jiwen; Ihrie, John

    2016-04-01

    Extensive human exposure to food- and cosmetics-related consumer products containing nanosilver is of public concern because of the lack of information about their safety. Genotoxicity is an important endpoint for the safety and health hazard assessment of regulated products including nanomaterials. The in vitro cytokinesis-block micronucleus (CBMN) assay is a very useful test for predictive genotoxicity testing. Recently, we have reported the genotoxicity of 20 nm nanosilver in human liver HepG2 and colon Caco2 cells evaluated using the CBMN assay. The objective of our present study was three-fold: (i) to evaluate if HepG2 and Caco2 cells are valuable in vitro models for rapid genotoxicity screening of nanosilver; (ii) to test the hypothesis that the nanoparticle size and cell types are critical determinants of its genotoxicity; and (iii) to determine if ionic silver contributes to the nanosilver genotoxicity. With these objectives in mind, we evaluated the genotoxic potential of 50 nm nanosilver of the same shape, composition, surface charge, obtained from the same commercial source, under the same experimental conditions and the same genotoxic CBMN endpoint used for the previously tested 20 nm silver. The ionic silver (silver acetate) was also evaluated under the same conditions. Results of our study show that up to the concentrations tested in these cell types, the smaller (20 nm) nanosilver induces micronucleus formation in both the cell types but the larger (50 nm) nanosilver and the ionic silver provide a much weaker response compared with controls under the same conditions. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  11. Modulation of apoptosis in human hepatocellular carcinoma (HepG2 cells) by a standardized herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes with anti- hepatocarcinogenic effects.

    Science.gov (United States)

    Samarakoon, Sameera R; Thabrew, Ira; Galhena, Prasanna B; Tennekoon, Kamani H

    2012-03-29

    A standardized poly-herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes used traditionally in Sri Lanka for cancer therapy has been demonstrated previously, to have anti-hepatocarcinogenic potential. Cytotoxicity, antioxidant activity, anti-inflammatory activity, and up regulation of p53 and p21 activities are considered to be some of the possible mechanisms through which the above decoction may mediate its anti-hepatocarcinogenic action. The main aim of the present study was to determine whether apoptosis is also a major mechanism by which the decoction mediates its anti-hepatocarcinogenic action. Evaluation of apoptosis in HepG2 cells was carried out by (a) microscopic observations of cell morphology, (b) DNA fragmentation analysis, (c) activities of caspase 3 and 9, as well as by (d) analysis of the expression of pro-apoptotic (Bax) and anti-apoptotic (Bcl-2) proteins associated with cell death. The results demonstrated that in HepG2 cells, the decoction can induce (a) DNA fragmentation and (b) characteristic morphological changes associated with apoptosis (nuclear condensation, membrane blebbing, nuclear fragmentation and apoptotic bodies). The decoction could also, in a time and dose dependent manner, up regulate the expression of the pro-apoptotic gene Bax and down regulate expression of anti-apoptotic Bcl-2 gene (as evident from RT-PCR analysis, immunohistochemistry and western blotting). Further, the decoction significantly (p < .001) enhanced the activities of caspase-3 and caspase-9 in a time and dose dependent manner. Overall findings provide confirmatory evidence to demonstrate that the decoction may mediate its reported anti-hepatocarcinogenic effect, at least in part, through modulation of apoptosis.

  12. Solid lipid nanoparticles for hydrophilic biotech drugs: optimization and cell viability studies (Caco-2 & HEPG-2 cell lines)

    Czech Academy of Sciences Publication Activity Database

    Severino, P.; Andreani, T.; Jäger, Alessandro; Chaud, M. V.; Santana, M. H. A.; Silva, A. M.; Souto, E. B.

    2014-01-01

    Roč. 81, 23 June (2014), s. 28-34 ISSN 0223-5234 R&D Projects: GA ČR GAP208/10/1600 Institutional support: RVO:61389013 Keywords : lipid nanoparticles * double emulsion * hydrophilic biotech drugs Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.447, year: 2014

  13. Trivalent dimethylarsenic compound induces histone H3 phosphorylation and abnormal localization of Aurora B kinase in HepG2 cells

    International Nuclear Information System (INIS)

    Suzuki, Toshihide; Miyazaki, Koichi; Kita, Kayoko; Ochi, Takafumi

    2009-01-01

    Trivalent dimethylarsinous acid [DMA(III)] has been shown to induce mitotic abnormalities, such as centrosome abnormality, multipolar spindles, multipolar division, and aneuploidy, in several cell lines. In order to elucidate the mechanisms underlying these mitotic abnormalities, we investigated DMA(III)-mediated changes in histone H3 phosphorylation and localization of Aurora B kinase, which is a key molecule in cell mitosis. DMA(III) caused the phosphorylation of histone H3 (ser10) and was distributed predominantly in mitotic cells, especially in prometaphase cells. By contrast, most of the phospho-histone H3 was found to be localized in interphase cells after treatment with inorganic arsenite [iAs(III)], suggesting the involvement of a different pathway in phosphorylation. DMA(III) activated Aurora B kinase and slightly activated ERK MAP kinase. Phosphorylation of histone H3 by DMA(III) was effectively reduced by ZM447439 (Aurora kinase inhibitor) and slightly reduced by U0126 (MEK inhibitor). By contrast, iAs(III)-dependent histone H3 phosphorylation was markedly reduced by U0126. Aurora B kinase is generally localized in the midbody during telophase and plays an important role in cytokinesis. However, in some cells treated with DMA(III), Aurora B was not localized in the midbody of telophase cells. These findings suggested that DMA(III) induced a spindle abnormality, thereby activating the spindle assembly checkpoint (SAC) through the Aurora B kinase pathway. In addition, cytokinesis was not completed because of the abnormal localization of Aurora B kinase by DMA(III), thereby resulting in the generation of multinucleated cells. These results provide insight into the mechanism of arsenic tumorigenesis.

  14. Cellular Interactions and Biological Responses to Titanium Dioxide Nanoparticles in HepG2 and BEAS-2B Cells: Role of Cell Culture Media

    Science.gov (United States)

    ABSTRACT We have shown previously that the composition of the biological medium used in vitro can affect the cellular interaction and biological response of titanium dioxide nanoparticles (nano-TiO2) in human lung epithelial cells. However, it is unclear if these effects are co...

  15. Radiosensitivity evaluation of Human tumor cell lines by single cell gel electrophoresis

    International Nuclear Information System (INIS)

    Zhang Yipei; Cao Jia; Wang Yan; Du Liqing; Li Jin; Wang Qin; Fan Feiyue; Liu Qiang

    2011-01-01

    Objective: To explore the feasibility of determining radiosensitivity of human tumor cell lines in vitro using single cell gel electrophoresis (SCGE). Methods: Three human tumor cell lines were selected in this study, HepG 2 , EC-9706 and MCF-7. The surviving fraction (SF) and DNA damage were detected by MTT assay, nested PCR technique and comet assay respectively. Results: MTT assay: The SF of HepG 2 and EC-9706 after irradiated by 2, 4 and 8 Gy was lower significantly than that of MCF-7, which showed that the radiosensitivity of HepG 2 and EC-9706 was higher than that of MCF-7. But there was no statistical difference of SF between HepG 2 and EC-9706. SCGE: The difference of radiosensitivity among these three tumor cell lines was significant after 8 Gy γ-ray irradiation. Conclusion: The multi-utilization of many biological parameter is hopeful to evaluate the radiosensitivity of tumor cells more objectively and exactly. (authors)

  16. Sodium alginate-polyvinyl alcohol-bovin serum albumin coated Fe3O4 nanoparticles as anticancer drug delivery vehicle: Doxorubicin loading and in vitro release study and cytotoxicity to HepG2 and L02 cells.

    Science.gov (United States)

    Prabha, G; Raj, V

    2017-10-01

    The challenging part of this work was to research the potential aspects of sodium alginate (SA)-polyvinyl alcohol (PVA)-bovin serum albumin (BSA) coated Fe 3 O 4 nanoparticles (Fe 3 O 4 -SA-PVA-BSA) as a drug delivery system for doxorubicin (DOX). The anticancer drug doxorubicin was selected as a model drug which is powerful for numerous cancer treatments. Superparamagnetic Fe 3 O 4 nanoparticles were prepared by co-precipitation method. The mixture solution of Fe 3 O 4 -sodium alginate (SA) - doxorubicin (DOX) was crosslinked with Ca 2+ to form (Fe 3 O 4 -SA-DOX) nanoparticles and addition of PVA and BSA with (Fe 3 O 4 -SA-DOX) nanoparticles was prepared by coating procedure. Doxorubicin drug loaded NPs were prepared by a simple crosslinking method by calcium chloride solution. The prepared polymer coated magnetic nanoparticles (Fe 3 O 4 -SA-PVA-BSA) were characterized by using SEM, AFM, FT-IR, XRD and VSM. The mean sizes of the obtained drug loaded nanoparticles (Fe 3 O 4 -SA-DOX, Fe 3 O 4 -SA-DOX-PVA and Fe 3 O 4 -SA-DOX-PVA-BSA) were between 240±8.3 and 460±8.7nm and zeta potential of the particles also was evaluated using Malvern Zetasizer which ranged between -48.1±2.3 and -22.4±4.1mV. The encapsulation efficiency, was between 36.2±0.01 and 96.45±2.12. Moreover drug loading and drug release properties of the polymer coated magnetic nanoparticles loaded with doxorubicin (Fe 3 O 4 -SA-DOX-PVA-BSA) were also studied. In addition, the cytotoxicity of the created nanoparticles was performed by using MTT assay analysis which showed that DOX loaded nanoparticles (Fe 3 O 4 -SA-DOX-PVA-BSA) were toxic to HepG2 cell lines and non-toxic to L02 cell lines. The in-vitro drug release was studied by using UV-Visible spectrophotometer at acidic environment (pH5.0) and basic environment (pH7.4) as well as at different temperatures (37°C and 42°C). It was found that DOX drug is released much faster in acidic environment (pH5.0) than in the basic environment (pH7

  17. Intrinsic anticarcinogenic effects of Piper sarmentosum ethanolic extract on a human hepatoma cell line

    Directory of Open Access Journals (Sweden)

    Senafi Sahidan

    2009-03-01

    Full Text Available Abstract Background Piper sarmentosum, locally known as kaduk is belonging to the family of Piperaceae. It is our interest to evaluate their effect on human hepatoma cell line (HepG2 for the potential of anticarcinogenic activity. Results The anticarcinogenic activity of an ethanolic extract from Piper sarmentosum in HepG2 and non-malignant Chang's liver cell lines has been previously determined using (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT assays, where the IC50 value was used as a parameter for cytotoxicity. The ethanolic extract that showed anticarcinogenic properties in HepG2 cells had an IC50 of 12.5 μg mL-1, while IC50 values in the non-malignant Chang's liver cell line were greater than 30 μg mL-1. Apoptotic morphological changes in HepG2 cells were observed using an inverted microscope and showed chromatin condensation, cell shrinkage and apoptotic bodies following May-Grunwald-Giemsa's staining. The percentage of apoptotic cells in the overall population (apoptotic index showed a continuously significant increase (p -1 ethanolic extract-treated cells at 24, 48 and 72 hours compared to controls (untreated cells. Following acridine orange and ethidium bromide staining, treatment with 10, 12 and 14 μg mL-1 of ethanolic extracts caused typical apoptotic morphological changes in HepG2 cells. Molecular analysis of DNA fragmentation was used to examine intrinsic apoptosis induced by the ethanolic extracts. These results showed a typical intrinsic apoptotic characterisation, which included fragmentation of nuclear DNA in ethanolic extract-treated HepG2 cells. However, the non-malignant Chang's liver cell line produced no DNA fragmentation. In addition, the DNA genome was similarly intact for both the untreated non-malignant Chang's liver and HepG2 cell lines. Conclusion Therefore, our results suggest that the ethanolic extract from P. sarmentosum induced anticarcinogenic activity through an intrinsic apoptosis

  18. Ku70 acetylation and modulation of c-Myc/ATF4/CHOP signaling axis by SIRT1 inhibition lead to sensitization of HepG2 cells to TRAIL through induction of DR5 and down-regulation of c-FLIP

    DEFF Research Database (Denmark)

    Kim, Mi-Ju; Hong, Kyung-Soo; Kim, Hak-Bong

    2013-01-01

    In this study, we investigated the role of c-Myc/ATF4/CHOP signaling pathway in sensitization of human hepatoma HepG2 cells to TRAIL. Knockdown of SIRT1 or treatment with SIRT1 inhibitor caused the up-regulation of DR5 and down-regulation of c-FLIP through modulation of c-Myc/ATF4/CHOP pathway......-FLIP. Moreover, Ku70(-/-) MEF and Ku70-knockdown HepG2 cells showed the increased levels of c-Myc, ATF4, CHOP, and DR5 and decreased level of c-FLIP. These results were followed by increased sensitivity of Ku70(-/-) MEF cells and Ku70-knockdown HepG2 cells to TRAIL compared with their control cells....... These findings reveal for the first time that SIRT1 inhibition increases Ku70 acetylation, and the acetylated Ku70 with a decreased function mediates the induction of DR5 and the down-regulation of c-FLIP by up-regulating c-Myc/ATF4/CHOP pathway, and consequently promotes the TRAIL-induced apoptosis of HepG2...

  19. Concurrent acetylation of FoxO1/3a and p53 due to sirtuins inhibition elicit Bim/PUMA mediated mitochondrial dysfunction and apoptosis in berberine-treated HepG2 cells

    International Nuclear Information System (INIS)

    Shukla, Shatrunajay; Sharma, Ankita; Pandey, Vivek Kumar; Raisuddin, Sheikh; Kakkar, Poonam

    2016-01-01

    Post-translational modifications i.e. phosphorylation and acetylation are pivotal requirements for proper functioning of eukaryotic proteins. The current study aimed to decode the impact of acetylation/deacetylation of non-histone targets i.e. FoxO1/3a and p53 of sirtuins (NAD + dependent enzymes with lysine deacetylase activity) in berberine treated human hepatoma cells. Berberine (100 μM) inhibited sirtuins significantly (P < 0.05) at transcriptional level as well as at translational level. Combination of nicotinamide (sirtuin inhibitor) with berberine potentiated sirtuins inhibition and increased the expression of FoxO1/3a and phosphorylation of p53 tumor suppressor protein. As sirtuins deacetylate non-histone targets including FoxO1/3a and p53, berberine increased the acetylation load of FoxO1/3a and p53 proteins. Acetylated FoxO and p53 proteins transcriptionally activate BH3-only proteins Bim and PUMA (3.89 and 3.87 fold respectively, P<0.001), which are known as direct activator of pro-apoptotic Bcl-2 family protein Bax that culminated into mitochondria mediated activation of apoptotic cascade. Bim/PUMA knock-down showed no changes in sirtuins' expression while cytotoxicity induced by berberine and nicotinamide was curtailed up to 28.3% (P < 0.001) and it restored pro/anti apoptotic protein ratio in HepG2 cells. Sirtuins inhibition was accompanied by decline in NAD + /NADH ratio, ATP generation, enhanced ROS production and decreased mitochondrial membrane potential. TEM analysis confirmed mitochondrial deterioration and cell damage. SRT-1720 (1–10 μM), a SIRT-1 activator, when pre-treated with berberine (25 μM), reversed sirtuins expression comparable to control and significantly restored the cell viability (P < 0.05). Thus, our findings suggest that berberine mediated sirtuins inhibition resulting into FoxO1/3a and p53 acetylation followed by BH3-only protein Bim/PUMA activation may in part be responsible for mitochondria-mediated apoptosis

  20. Prediction of radiosensitivity of human tumor cell lines in vitro by determining 4977bp deletion in mitochondrial DNA

    International Nuclear Information System (INIS)

    Rong Qinglin; Cao Yongzhen; Zhang Yaowen; Zhao Xinran; Wang Qin; Li Jin; Liu Qiang

    2008-01-01

    Objective: To evaluate the possibility of predicting the radiosensitivity of tumor cell lines using the assay of the mtDNA4977bp deletion. Methods: The mtDNA4977bp deletion of HepG 2 cells and PC-3 cells were detected by nested PCR after irradiated by various doses of x-ray. Results: The radiation-induced mtDNA4977bp deletion of the tumor cell lines of HepG 2 and PC-3 were detected after irradiated. There was a dose dependent in the mtDNA4977bp deletion of two tumor cell lines. The deletion rate of HepG 2 was higher significantly than that of PC-3 at each point of radiation dose (P 2 was higher than that of PC-3. Conclusion: The assay of the mtDNA4977bp deletion may be an approach to predict the radiosensitivity of tumor cells. (authors)

  1. Flavone polyphenols dominate in Thymus schimperi Ronniger: LC-ESI-MS/MS characterization and study of anti-proliferative effects of plant extract on AGS and HepG2 cancer cells.

    Science.gov (United States)

    Desta, Kebede Taye; Kim, Gon Sup; Abd El-Aty, A M; Raha, Suchismita; Kim, Man-Bae; Jeong, Ji Hoon; Warda, Mohamad; Hacımüftüoğlu, Ahmet; Shin, Ho-Chul; Shim, Jae-Han; Shin, Sung Chul

    2017-05-15

    Thymus schimperi is a highly localized and a rare plant endemic to Ethiopia. An optimized and validated high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) method was applied to characterize 23 polyphenolic compounds found in ethyl acetate extracts of the plant. From those, flavones dominated and luteolin was the major component contributing 21.83% of the total composition (or 46.05±0.59g/kg of fresh sample weight). Validation data showed a determination coefficient (R 2 )≥0.997. Limits of detection (LOD) and quantification (LOQ) were 0.03-0.97 and 0.11-3.23mg/L, while recovery values spiked at 5 and 50mg/L were between 70.89-115.39 and 67.65-120.19%, respectively. Except for caffeic acid and epicatechin gallate, the relative standard deviations (%RSDs) were far below 15%, showing acceptable precision values. The plant extracts inhibited cell proliferation and induced cell death in human gastric adenocarcinoma (AGS) and liver hepatocellular carcinoma (HepG2) cancer cells. This is the first report of polyphenolic components from T. schimperi being characterized using HPLC-ESI-MS/MS. Being components of many edible vegetables, fruits, and spices, the identified polyphenols suggest that T. schimperi could be a potential food with promising health benefits. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Caffeic Acid Phenethyl Ester (Propolis Extract) Ameliorates Insulin Resistance by Inhibiting JNK and NF-κB Inflammatory Pathways in Diabetic Mice and HepG2 Cell Models.

    Science.gov (United States)

    Nie, Jiarui; Chang, Yaning; Li, Yujia; Zhou, Yingjun; Qin, Jiawen; Sun, Zhen; Li, Haibin

    2017-10-18

    Caffeic acid phenethyl ester (CAPE), extracted from propolis, was evaluated for the ameliorative effects on insulin resistance and the mechanisms were identified, using non-insulin-dependent diabetes mellitus (NIDDM) model mice and insulin resistance (IR) model cells. After 5 weeks of CAPE supplementation, insulin sensitivity, hyperlipidemia, and peroxisome proliferator-activated receptor-α (PPAR-α) levels were improved in mice. Proinflammatory cytokines in serum and the expressions of tumor necrosis factor-alpha (TNF-α) mRNA in tissues were markedly downregulated from CAPE-treated mice. In vitro, CAPE supplement significantly improved glucose consumption, glucose uptake, glycogen content, and oxidative stress and decreased expression of glucose-6-phosphatase (G6Pase) mRNA in cells. Both in vivo and in vitro, CAPE enhanced p-Akt (Ser473) and p-insulin receptor substrate (IRS)-1 (Tyr612), but inhibited p-JNK (Thr183/Tyr185), p-NF-κB p65 (Ser536), and nuclear translocation of p-NF-κB p65 (Ser536). In summary, CAPE can ameliorate insulin resistance through modulation of JNK and NF-κB signaling pathway in mice and HepG2 cells.

  3. Inclusion Complex of Zerumbone with Hydroxypropyl-β-Cyclodextrin Induces Apoptosis in Liver Hepatocellular HepG2 Cells via Caspase 8/BID Cleavage Switch and Modulating Bcl2/Bax Ratio

    Directory of Open Access Journals (Sweden)

    Nabilah Muhammad Nadzri

    2013-01-01

    Full Text Available Zerumbone (ZER isolated from Zingiber zerumbet was previously encapsulated with hydroxypropyl-β-cyclodextrin (HPβCD to enhance ZER’s solubility in water, thus making it highly tolerable in the human body. The anticancer effects of this new ZER-HPβCD inclusion complex via apoptosis cell death were assessed in this study for the first time in liver hepatocellular cells, HepG2. Apoptosis was ascertained by morphological study, nuclear stain, and sub-G1 cell population accumulation with G2/M arrest. Further investigations showed the release of cytochrome c and loss of mitochondrial membrane potential, proving mitochondrial dysfunction upon the ZER-HPβCD treatment as well as modulating proapoptotic and anti-apototic Bcl-2 family members. A significant increase in caspase 3/7, caspase 9, and caspase 8 was detected with the depletion of BID cleaved by caspase 8. Collectively, these results prove that a highly soluble inclusion complex of ZER-HPβCD could be a promising anticancer agent for the treatment of hepatocellular carcinoma in humans.

  4. Comparison on hypoglycemic and antioxidant activities of the fresh and dried Portulaca oleracea L. in insulin-resistant HepG2 cells and streptozotocin-induced C57BL/6J diabetic mice.

    Science.gov (United States)

    Gu, Jun-Fei; Zheng, Zhi-Yin; Yuan, Jia-Rui; Zhao, Bing-Jie; Wang, Chun-Fei; Zhang, Li; Xu, Qing-Yu; Yin, Guo-Wen; Feng, Liang; Jia, Xiao-Bin

    2015-02-23

    Fresh Portulaca oleracea L. (family: Portulacaceae; POL) has been used as a folk medicine for the treatment of diabetes mellitus for a long time. More bioactive components with higher activity could be retained in fresh medicinal herbs compared to the dried ones. The present study was conducted to compare different antidiabetic activity between fresh and dried POL, including hypoglycemic and antioxidant activities both in vivo and in vitro. Furthermore, in order to explore which components were responsible for the antidiabetic activity, the difference on chemical components between fresh and dried POL was analyzed and compared. Insulin-resistant HepG2 cells induced by insulin were used to evaluate the promoting effect of the fresh and dried POL on glucose utilization in vitro. Streptozotocin (STZ)-induced C57BL/6J diabetic mice were used to compare the differences on hypoglycemic and antioxidant activities of fresh and dried POL, including the fasting blood glucose, glucose tolerance, serum insulin level, malondialdehyde (MDA) level and superoxide dismutase (SOD) activity in vivo. UPLC/Q-TOF-MS method was performed to analyze the difference of antidiabetic components between fresh and dried POL. Compared with the dried POL extract, the fresh POL extract significantly increased the consumption of extracellular glucose in insulin-resistant HepG2 cells (P<0.05). In STZ-induced C57BL/6J diabetic mice, both fresh and dried extracts decreased markedly the fasting blood glucose (FBG) levels, and improved significantly oral glucose tolerance test (OGTT), as well as enhanced significantly insulin secretion and antioxidative activities (P<0.05; P<0.01). Furthermore, the fresh extract showed stronger antidiabetic activity (P<0.05). The UPLC/Q-TOF-MS analysis results also revealed that the relative contents of polyphenols and alkaloids in the fresh herbs were more abundant than those in the dried POL. Our results indicated that both fresh and dried POL possessed antidiabetic

  5. Effect of New Water-Soluble Dendritic Phthalocyanines on Human Colorectal and Liver Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Ebru YABAŞ

    2017-08-01

    Full Text Available Human hepatocellular carcinoma (HepG2 cells and colorectal adenocarcinoma (DLD-1 cells were treated with the synthesized water soluble phthalocyanine derivatives to understand the effect of the compounds both on colorectal and liver cancer cells. The compounds inhibited cell proliferation and displayed cytotoxic effect on these cancer cell lines however; the effect of the compounds on healthy control fibroblast cell line was comparatively lower. The compounds can be employed for cancer treatment as anticancer agents.

  6. Protective effects of rambutan (Nephelium lappaceum) peel phenolics on H2O2-induced oxidative damages in HepG2 cells and d-galactose-induced aging mice.

    Science.gov (United States)

    Zhuang, Yongliang; Ma, Qingyu; Guo, Yan; Sun, Liping

    2017-10-01

    Rambutan peel phenolic (RPP) extracts were prepared via dynamic separation with macroporous resin. The total phenolic content and individual phenolics in RPP were determined. Results showed that the total phenolic content of RPP was 877.11 mg gallic acid equivalents (GAE)/g extract. The content of geranin (122.18 mg/g extract) was the highest among those of the 39 identified phenolic compounds. RPP protected against oxidative stress in H 2 O 2 -induced HepG2 cells in a dose-response manner. The inhibitory effects of RPP on cell apoptosis might be related to its inhibitory effects on the generation of intracellular reactive oxygen species and increased effects on superoxide dismutase activity. The in vivo anti-aging activity of RPP was evaluated using an aging mice model that was induced by d-galactose (d-gal). The results showed that RPP enhanced the antioxidative status of experimental mice. Moreover, histological analysis indicated that RPP effectively reduced d-gal-induced liver and kidney tissue damage in a dose-dependent manner. Therefore, RPP can be used as a natural antioxidant and anti-aging agent in the pharmaceutical and food industries. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. [Effect of transcription activity regulated by VNTR-ZNF and -14C/T variants in the promoter region of ATP-binding cassette transporter 1 in HepG2 cells].

    Science.gov (United States)

    Gao, Shenxia; Zhao, Lili; Zhang, Ying; Mao, Yongmin

    2016-10-01

    To explore the effect of VNTR-ZNF and -14C/T variants of the promoter region of the ABCA1 gene on the transcription activity of genes in vitro. The recombinants were constructed by ligating DNA fragment containing VNTR-ZNF ACCCC inserted/deleted allele with or without -14C/T substitution fragments with a PGL2-basic vector containing luciferase reporter gene. The recombinants were then transfected into HepG2 cells using the cationic lipid method. After 48 h, transfected cells were collected and used to detect the luciferase activity. Luciferase activity of PGL2-ZNF-ACCCCDel was greater than that of PGL2-ZNF-ACCCCIns. Luciferase activity of PGL2-ZNFDel-14C was greater than that of PGL2-ZNFDel-14T, PGL2-ZNFIns-14C, PGL2-ZNFIns-14T. Compared with the insertion type, the ACCCC-deleted type of VNTR-ZNF can significantly enhance the transcription activity of ABCA1. And co-transfection of -14 C allele can further enhance this activity.

  8. HDMCP uncouples yeast mitochondrial respiration and alleviates steatosis in L02 and hepG2 cells by decreasing ATP and H2O2 levels: a novel mechanism for NAFLD.

    Science.gov (United States)

    Jin, Xi; Yang, Yi-da; Chen, Kun; Lv, Zhi-yuan; Zheng, Lin; Liu, Ya-ping; Chen, Shao-hua; Yu, Chao-hui; Jiang, Xue-yuan; Zhang, Cheng-yu; Li, You-ming

    2009-05-01

    To explore the uncoupling activity of hepatocelluar downregulated mitochondrial carrier protein (HDMCP) in a yeast expression system and its function in non-alcoholic fatty liver disease (NAFLD). Molecular cloning and RT-PCR were used for yeast protein expression and uncoupling activity was assessed. Western blot analysis was used to determine HDMCP level in rat NAFLD and steatotic L02 and hepG2 cell models where their presence was confirmed by pathologic (Nile red and H-E staining) and biochemical changes. RNA interference was used to knock down HDMCP level and mitochondrial ATP and hydroperoxide levels were measured for potential mechanism exploration. We found a significant GDP insensitive uncoupling activity of HDMCP in yeast mitochondria and its increased expression in animal and cell models. HDMCP was significantly increased with culture time and steatosis was aggravated when HDMCP level was knocked down. Furthermore, we found that HDMCP might function through promoting ATP depletion and decreasing H(2)O(2) production. This study adds supportive data to the hypothesis that HDMCP might be a long postulated liver-specific uncoupling protein and broadens our understanding of the pathogenesis of NAFLD. More importantly, HDMCP might become a novel drug target for its ability in alleviating hepatic steatosis.

  9. Carboxymethyl chitin-glucan (CM-CG) protects human HepG2 and HeLa cells against oxidative DNA lesions and stimulates DNA repair of lesions induced by alkylating agents.

    Science.gov (United States)

    Slamenová, Darina; Kováciková, Ines; Horváthová, Eva; Wsólová, Ladislava; Navarová, Jana

    2010-10-01

    A large number of functional foods, including those that contain β-d-glucans, have been shown to prevent human DNA against genotoxic effects and associated development of cancer and other chronic diseases. In this paper, carboxymethyl chitin-glucan (CM-CG) isolated from Aspergillus niger was investigated from two standpoints: (1) DNA-protective effects against oxidative DNA damage induced by H(2)O(2) and alkylating DNA damage induced by MMS and MNNG, and (2) a potential effect on rejoining of MMS- and MNNG-induced single strand DNA breaks. The results obtained by the comet assay in human cells cultured in vitro showed that CM-CG reduced significantly the level of oxidative DNA lesions induced by H(2)O(2) but did not change the level of alkylating DNA lesions induced by MMS or MNNG. On the other side, the efficiency of DNA-rejoining of single strand DNA breaks induced by MMS and MNNG was significantly h