Human Hepatoma HepG2 Cells Research Articles

Suppression Effect of Ulva lactuca Selenium Nanoparticles (USeNPs) on HepG2 Carcinoma Cells Resulting from Degradation of Epidermal Growth Factor Receptor (EGFR) with an Evaluation of Its Antiviral and Antioxidant Activities

The current study sought to assess the antitumor, anticancer, and antioxidant efficacy of Ulva lactuca-mediated selenium nanoparticles by using an in vitro model of human hepatocellular carcinoma (HepG-2 cells) and HAV HM175 strain of hepatitis A virus, with the evaluation of antioxidant activity conducted using DPPH assay. The study showed promising cytotoxicity at the highest concentrations of 250 and 500 µg/mL, with viability rates of 19.43 and 8.75% for cancer cells, and the lowest toxicity with the highest viability rates of 59.41 and 30.64% for normal cells, respectively. These concentrations also exhibited the highest inhibition rates (51.28 and 76.31%, respectively) against the Epidermal Growth Factor Receptor (EGFR) enzyme and provide an explanation of the mechanism of such cytotoxicity, as this enzyme is responsible for the degradation of EGFR. Additionally, U. lactuca-mediated selenium nanoparticles (USeNPs) showed promising antiviral activity (+++) (50–<75%) with EC50 = 57.41 μg/mL and 74.13% antiviral rates against HAV HM175 at 100 µg/mL maximum noncytotoxic conc (MNCC). Using MTT assay, its selectivity index was 5.78 ≥ 2, which indicates that USeNPs exhibited antiviral activity that outweighed its toxicity. Therefore, USeNPs is an active antiviral agent and warrants further study. Furthermore, the DPPH scavenging activity of such nanoparticles was moderate as the highest sample concentration (100 µg/mL) recorded 31.64 ± 0.03% DPPH scavenging activity (with IC50 = 158.02 ± 0.07 μg/mL), a percentage which does not exceed that of standard ascorbic acid.

Knockdown of the long noncoding RNA PURPL induces apoptosis and sensitizes liver cancer cells to doxorubicin

Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer with increasing incidence in western countries. Most HCC patients have advanced cancer at the time of diagnosis due to the asymptomatic nature of early-stage HCC and do not qualify for potentially curative surgical treatment, thus, highlighting the need for new therapeutic strategies. Long noncoding RNAs (lncRNAs) comprise a large and heterogeneous group of non-protein coding transcripts that play important regulatory roles in numerous biological processes in cancer. In this study, we performed RNA sequencing of liver biopsies from ten HCC, ten hepatitis C virus-associated HCC, and four normal livers to identify dysregulated lncRNAs in HCC. We show that the lncRNA p53-upregulated-regulator-of-p53-levels (PURPL) is upregulated in HCC biopsies and that its expression is p53-dependent in liver cancer cell lines. In addition, antisense oligonucleotide-mediated knockdown of PURPL inhibited cell proliferation, induced apoptosis, and sensitized HepG2 human HCC cells to treatment with the chemotherapeutic agent doxorubicin. In summary, our findings suggest that PURPL could serve as a new therapeutic target for reversing doxorubicin resistance in HCC.

Efficient Synthesis of Densely Functionalized Pyrido[2,3-d]Pyrimidines via Three-component One-pot Domino Knoevenagel aza-Diels Alder Reaction and Induces Apoptosis in Human Cancer Cell Lines via Inhibiting Aurora A and B Kinases

The identification of novel aurora kinase inhibitors is one of the most attractive directions in the field of anticancer research and development. In our ongoing efforts to pursue the class of inhibitors, a series of pyrido[2,3-d]pyrimidines were synthesized in DIPEAc/EtOH media. The advantages of the present methodology include a one-pot multicomponent environmentally friendly approach, cost effectiveness, broad substrate scope, operational simplicity, short reaction times, easy workup procedure and high yields. Synthesized compounds were screened against human lung carcinoma A549 cells, human hepatocellular liver carcinoma HepG2 cells and human cervical carcinoma epithelial HeLa cells. Compound 6 b i.e. diethyl 5-(4-methoxyphenyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazoline-6,7-dicarboxylate was found to be equipotent than the standard drug VX-680 against selected cancer cell lines. The selected compounds (6b, 6d and 6h) exhibit potent inhibition against Aurora A and B with IC50 values (9.4–25 mg/L). Molecular docking model showed that the compounds can bind well to the target by interacting with amino acid residues. It will provide some valuable information for the commercial Aurora Kinase inhibitors.

Optimization and Application of A Bionic System of Dynamic Co-Culture with Hepatocytes and Renal Cells Based on Microfluidic Chip Technique in Evaluating Materials of Health Food.

We aimed to explore the optimization and application of a bionic system of dynamic co-culture with hepatocytes and renal cells based on the microfluidic chip technique in evaluating emodin, which might replace the conventionally cytological evaluation technique of health food. After optimal experiments, the improved bionic system was composed of human hepatocellular carcinoma cells (HepG2), human renal glomerular endothelial cells (HRGECs), rat tail collagen type I, and gelatin with optimized concentrations (1.3 mg/mL + 7.5%). The applicability of the bionic system indicated that the growth stability was appropriate (CV: 7.36%), and the cell viability of that gradually decreased with the increasing of emodin concentration from 0-100 μM, which statistic significances were at 50 and 100 μM (p < 0.05), and the stained results of dead/live cells also showed the same trend. The LDH level appeared rising trend after decline between 0 μM and 100 μM emodi, and the level of that at 100 μM emodin was significantly higher than that at 25 μM and 50 μM emodin, respectively. The BUN level continuously and significantly declined with the increasing of emodin concentration (p < 0.05). Our research realized the application of this optimized bionic system in evaluating emodin, and provided a useful platform and reference for further in vitro alternative research with regard to evaluating the efficacies of health food in the future.

Proton irradiation induced reactive oxygen species promote morphological and functional changes in HepG2 cells

The increased use of proton therapy has led to the need of better understanding the cellular mechanisms involved. The aim of this study was to investigate the effects induced by the accelerated proton beam in hepatocarcinoma cells. An existing facility in IFIN-HH, a 3 MV Tandetron™ accelerator, was used to irradiate HepG2 human hepatocarcinoma cells with doses between 0 and 3 Gy. Colony formation was used to assess the influence of radiation on cell long-term replication. Also, the changes induced at the mitochondrial level were shown by increased ROS and ATP levels as well as a decrease in the mitochondrial membrane potential. An increased dose has induced DNA damages and G2/M cell cycle arrest which leads to caspase 3/7 mediated apoptosis and senescence induction. Finally, the morphological and ultrastructural changes were observed at the membrane level and the nucleus of the irradiated cells. Thus, proton irradiation induces both morphological and functional changes in HepG2 cells.

Chemical constituents from Ficus sur Forssk (Moraceae).

Phytochemical investigation of the aerial roots of Ficus sur, a Cameroonian medicinal plant, resulted in apreviously undescribed cerebroside, suroside (1), in addition to its aglycon congener suramide (2). Moreover, six known natural products including alpinumisoflavone (3), wighteone metabolite (4), oleanolic acid (5), β-sitosterol (6), β-sitosterol-3-O-β-D-glucopyranoside (7), and epi-ѱ-taraxastanolone (8) were identified. The structures of thepreviously undescribed compounds were determined by analysis of 1D and 2D-NMR (One and two dimensional nuclear magnetic resonance), mass spectrometry, chemical conversion, and by comparison of these data with those from the literature. Wighteone metabolite (4) exhibited a weak cytotoxic activity against the human HepG2 hepatocellular carcinoma cells with an IC50 value of 51.9µM.

Co-Encapsulation of Tannic Acid and Resveratrol in Zein/Pectin Nanoparticles: Stability, Antioxidant Activity, and Bioaccessibility.

Hydrophilic tannic acid and hydrophobic resveratrol were successfully co-encapsulated in zein nanoparticles prepared using antisolvent precipitation and then coated with pectin by electrostatic deposition. The encapsulation efficiencies of the tannic acid and resveratrol were 51.5 ± 1.9% and 77.2 ± 3.2%, respectively. The co-encapsulated nanoparticles were stable against aggregation at the investigated pH range of 2.0 to 8.0 when heated at 80 °C for 2 h and when the NaCl concentration was below 50 mM. The co-encapsulated tannic acid and resveratrol exhibited stronger in vitro antioxidant activity than ascorbic acid, as determined by 1,1-diphenyl-2-picrylhydrazyl free radical (DPPH·) and 2,2'-azinobis (3-ethylberizothiazoline-6-sulfonic acid) radical cation (ABTS+·) scavenging assays. The polyphenols-loaded nanoparticles significantly decreased the malondialdehyde (MDA) concentration and increased the superoxide dismutase (SOD) and catalase (CAT) activities in peroxide-treated human hepatoma cells (HepG2). An in vitro digestion model was used to study the gastrointestinal fate of the nanoparticles. In the stomach, encapsulation inhibited tannic acid release, but promoted resveratrol release. However, in the small intestine, it led to a relatively high bioaccessibility of 76% and 100% for resveratrol and tannic acid, respectively. These results suggest that pectin-coated zein nanoparticles have the potential for the co-encapsulation of both polar and nonpolar nutraceuticals or drugs.

Involvement of adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1 in diallyl trisulfide-induced cytotoxicity in hepatocellular carcinoma cells.

It has been demonstrated that APPL1 (adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1) is involved in the regulation of several growth-related signaling pathways and thus closely associated with the development and progression of some cancers. Diallyl trisulfide (DAT), a garlic-derived bioactive compound, exerts selective cytotoxicity to various human cancer cells through interfering with pro-survival signaling pathways. However, whether and how DAT affects survival of human hepatocellular carcinoma (HCC) cells remain unclear. Herein, we tested the hypothesis of the involvement of APPL1 in DAT-induced cytotoxicity in HCC HepG2 cells. We found that Lys 63 (K63)-linked polyubiquitination of APPL1 was significantly decreased whereas phosphorylation of APPL1 at serine residues remained unchanged in DAT-treated HepG2 cells. Compared with wild-type APPL1, overexpression of APPL1 K63R mutant dramatically increased cell apoptosis and mitigated cell survival, along with a reduction of phosphorylation of STAT3, Akt, and Erk1/2. In addition, DAT administration markedly reduced protein levels of intracellular TNF receptor-associated factor 6 (TRAF6). Genetic inhibition of TRAF6 decreased K63-linked polyubiquitination of APPL1. Moreover, the cytotoxicity impacts of DAT on HepG2 cells were greatly attenuated by overexpression of wild-type APPL1. Taken together, these results suggest that APPL1 polyubiquitination probably mediates the inhibitory effects of DAT on survival of HepG2 cells by modulating STAT3, Akt, and Erk1/2 pathways.

Gadolinium-Based MRI Contrast Agent for Targeted Imaging of Vascular Endothelial Growth Factor Receptor-2-Expressing Tumors

Development of imaging methods that can detect target molecules will be of great significance for non-invasive molecular diagnosis. Vascular endothelial growth factor receptor-2 (VEGFR-2) is a broadly expressed target, and VEGFR-2-specific agents have been used in biological therapy. In this study, amine-reactive coupling was used to label the polypeptide VEGF125–136 with a Gd(DOTA) complex to create a magnetic resonance contrast agent, VEGF125–136-Gd, which binds to VEGFR-2. Using T1-weighted magnetic resonance imaging, we explored the tumor cell-targeting ability of VEGF125–136-Gd and its enhancement of T1-weighted image intensity using human hepatoma (HepG2) cells and found that the signal-to-noise ratio (SNR) increased commensurately with the concentration of VEGF125–136-Gd. At a concentration of 2 mM, the SNR produced by VEGF125–136-Gd was 5.4-fold higher than that produced by a Gd-labelled non-targeting polypeptide (NTPP-Gd) control. We also evaluated the tumor-targeting efficiency of VEGF125–136-Gd in nude mice injected with human hepatoma (HepG2) cells. Preliminary in vivo imaging of VEGFR-2 in tumor tissue revealed a targeting peak 60 min post administration of VEGF125–136-Gd, the intensity of which was 23% higher than the SNR of NTPP-Gd at the same time point. Our findings may help lay the foundation for clinical applications of VEGF125–136-Gd in targeted imaging.

Adiponectin Stimulates Apolipoprotein A-1 Gene Expression in HepG2 Cells via AMPK, PPARα, and LXRs Signaling Mechanisms.

Adiponectin is an adipose tissue hormone, participating in energy metabolism and involved in atherogenesis. Previously, it was found that adiponectin increases expression of the APOA1 (apolipoprotein A-1) gene in hepatocytes, but the mechanisms of this effect remained unexplored. Our aim was to investigate the role of adiponectin receptors AdipoR1/R2, AMP-activated protein kinase (AMPK), nuclear peroxisome proliferator-activated receptor alpha (PPARα) and liver X receptors (LXRs) in mediating the action of adiponectin on hepatic APOA1 expression in human hepatoma HepG2 cells. The level of APOA1 expression was determined by RT-qPCR and ELISA. We showed that the siRNA-mediated knockdown of genes coding for AdipoR1, AdipoR2, AMPK, PPARα, and LXRα and β prevented adiponectin-induced APOA1 expression in HepG2 cells and demonstrated that interaction of PPARα and LXRs with the APOA1 gene hepatic enhancer is important for the adiponectin-dependent APOA1 transcription. The results of this study point out to the involvement of both types of adiponectin receptors, AMPK, PPARα, and LXRs in the adiponectin-dependent upregulation of the APOA1 expression.

GC/TOF-MS-based phytochemical analysis, in vitro antiproliferative effects, antioxidant and antibacterial activity of Sarcophyte sanguinea subsp. piriei (Hutch.) B. Hansen

Antibiotic resistance necessitates the need for continued search for new antimicrobial compounds. In this study, we report the antibacterial activity, cytotoxic, and antioxidant activity of Sarcophyte sanguinea subsp. piriei (Hutch.) B. The extracts were evaluated for antibacterial activity using the agar well diffusion, micro-dilution, minimum bactericidal concentrations (MBC) and lactate dehydrogenase (LDH) release assay. Furthermore, the cytotoxic effects of extracts were determined using MTT assay against Human Embryonic Kidney (HEK293), Human Breast Endocrine Cells (SKBR-3), Human Colorectal Carcinoma Cells (Caco-2) and Human Hepatocellular Carcinoma (HepG2) cells. The extracts were also assessed for antioxidant activity against 2,2-diphenyl-1-picryl hydrazyl (DPPH), 2–2′-Azino-di-[3-ethylbenzthiazoline sulfonate (ABTS), nitric oxide radicals and Fe2+ chelating assays. The methanol extract (MeOH) was subjected to GC-TOF-MS analysis. MeOH extract exhibited highest zone of inhibition (ZI) of 15.67 mm against Staphylococcus aureus. The extract further exhibited 67.0% inhibition against Escherichia coli in a lactate dehydrogenase (LDH) membrane damage assay. In the cytotoxicity assay, MeOH and DCM extracts had LC50 value of 237 µg/ml and 221 µg/ml against SKBR-3 cell line respectively. Gas-chromatography time-of-flight mass spectrometry (GC/TOF-MS) analysis of methanol extract indicated the presence of 5-aminoimidazole-4-carboxamide-1-ád-ribofuranosyl 5′-monophosphate (36.827%), 3-O-methyl-d-glucose (36.827%) and phthalic acid, di(oct‑3-yl) ester (8.161%). Even though the investigated plant is suggested to have anticancer activities by traditional healers, the extracts had little inhibition on selected cancerous cell lines. Given the good antibacterial and antioxidant activity of the extracts, the plant may act as an immune booster and prevent infections in immunosuppressed cancer patients. Detected compounds may be influence the observed biological activity of the plant.

Alizarin as a New Activator of the Aryl Hydrocarbon Receptor Signaling Pathway

Alizarin (1,2-dihydroxyanthraquinone) is a natural red dye extracted from the roots of Rubia cordifolia L. (family Rubiaceae). Alizarin has been used as a biological red stain for calcium. The aryl hydrocarbon receptor (AhR) has critical roles in multiple physiological pathways. This study aimed to determine whether alizarin is an unreported ligand of AhR. In the present study, we investigated the effects on cytochrome P450 (CYP) 1A1 mRNA, protein expression, AhR nuclear translocation, aryl hydrocarbon response element (AHRE) reporter activity, and AhR-specific antagonist following alizarin treatment of cells of the human hepatoma cell line, HepG2, and murine hepatoma cell line, Hepa-1c1c7. Alizarin induced CYP1A1 mRNA and protein expression in HepG2 and Hep-1c1c7 cells. Such induction was not present in C4 (B13NBii1) cells, which are AhR signal deficient, C12 (B15ECiii2) cells, which reduce AhR protein levels. The alizarin-induced responses were blocked by CH-223191, which is an AhR antagonist. Alizarin, the same as with the AhR ligand, induced the nuclear localization of AhR, as well as stimulated the transcriptional activity of AHRE. The results of this study suggest that alizarin is an AhR agonist.

Directional screening and identification of potential cytotoxic components from Achnatherum inebrians by a combination of surface palsmon resonance and chromatography

ObjectiveTo establish a method for directional screening of the cytotoxic components from the medicinal herb of Achnatherum inebrians by a combination of surface plasmon resonance (SPR) biosensor and chromatographic isolation technology. MethodsUnder the guidance of bioactive assessment based on binding abilities between objects and the α-Mannosidase (α-Man) target, the active components from different solvents extracts, different polar extraction parts and fractions were screened orderly and directionally using SPR. Components with a high binding ability to α-Man can be precisely oriented in a narrower fractions range and are easy to isolate. Three human cancer cells were used to evaluate the cytotoxic activity of component with the highest affinity to α-Man. ResultsEight compounds were isolated and identificated from A. inebrians for the first time. Deoxyvasicinone possessed the highest affinity to α-Man among them. Moreover, deoxyvasicinone showed good effects on inhibited proliferation of human hepatoma cells HepG2 (IC50 = 5.7 μmol/L), human breast cancer cells MCF7 (IC50 = 7.21 μmol/L) and human lung cancer cells HCC827 (IC50 = 0.75 μmol/L), respectively. In particular, its inhibitory effect on HCC827 was stronger than the positive drug gefitinib (IC50 = 1.65 μmol/L). ConclusionA comprehensive strategy of directional screening potential cytotoxic components from herb based on biomolecular interaction and chromatography was established. Deoxyvasicinone as an effective anti-cancer component was initially isolated from A. inebrians. It is expected that this screening strategy could provide new perspectives for rapid screening and identification of active components from natural plants with the complex matrix.

Tumoricidal effects of unprimed and curcumin-primed adipose-derived stem cells on human hepatoma HepG2 cells under oxidative conditions

IntroductionAdipose-derived stem cells (ASCs) have been proven to have tumoricidal effects against hepatic cancer cell lines. However, it appears that exposure to oxidative microenvironment compromises the potential outcome of ASCs in real hepatoma. Herein, we aimed to examine the tumoricidal effects of ASCs under oxidative conditions and to investigate the impact of curcumin priming on ASCs’ therapeutic potential. MethodsWe used human hepatoma (HepG2) cells in a coculture system with unprimed or curcumin-primed ASCs (Cur-ASCs) under H2O2-induced oxidative conditions. To investigate HepG2 proliferation and death, MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) and annexin V staining assays were performed. To determine the HepG2 migration and invasion potential, the scratch healing and the transwell invasion assays were performed. To evaluate the expression of apoptosis-protein markers, Western blotting was performed. ResultsCur-ASCs suppressed HepG2 proliferation, migration, and invasion as well as prompted apoptosis more significantly compared to unprimed ASCs under oxidative conditions. Expressional studies also revealed an obvious decline in the BCL-2/BAX ratio in HepG2 cocultured with Cur-ASCs. In addition, we noticed a marked elevation of apoptosis and senescence in unprimed ASCs compared to Cur-ASCs after coculture experiments, which demonstrated that curcumin priming preserved the survival and growth potential of ASCs; hence, Cur-ASCs performed better tumoricidal functions under oxidative conditions. ConclusionOur findings suggest that ASCs have the intrinsic ability to induce cell death in HepG2 cells; however, their functions can be compromised under oxidative conditions. We believe that curcumin priming is an effective approach for improving the therapeutic effectiveness of ASCs in the cancerous microenvironment.

Evaluation of cytotoxicity and genotoxicity of a novel oxovanadium complex with orotate

The development of new drugs based on metal complexes requires a detailed analysis of their biological endpoints. In this study, we report the genotoxic profile and influence on cell proliferation and death of the oxovanadium(IV) complex with orotic acid ([VO(C5H4N2O4)2], VO(oro)). Human hepatocellular carcinoma cells (HepG2) were the most sensitive tumor cells to VO(oro), which interfered with the integrity of cell membranes and proliferative capacity in a dose-dependent manner, inducing cell death by apoptosis. Regarding genotoxicity, VO(oro) did not induce considerable levels of DNA damage in HepG2 cells (comet test) and gene mutations (Ames test). However, it caused a statistically significant increase in the frequency of micronuclei at the highest concentration tested (12.5 µmol.L–1), indicating aneuploidy and clastogenicity. The data presented here provide information on various biological aspects of the VO(oro) complex, which may allow the elucidation of its mechanism of action as a possible therapeutic agent.

Adenosine-Mimicking Derivatives of 3-Aminopyrazine-2-Carboxamide: Towards Inhibitors of Prolyl-tRNA Synthetase with Antimycobacterial Activity

Multidrug-resistant tuberculosis (MDR-TB) poses a significant threat to mankind and as such earned its place on the WHO list of priority pathogens. New antimycobacterials with a mechanism of action different to currently used agents are highly required. This study presents the design, synthesis, and biological evaluation of 3-acylaminopyrazine-2-carboxamides derived from a previously reported inhibitor of human prolyl-tRNA synthetase. Compounds were evaluated in vitro against various strains of mycobacteria, pathogenic bacteria, and fungi of clinical significance. In general, high activity against mycobacteria was noted, while the antibacterial and antifungal activity was minimal. The most active compounds were 4'-substituted 3-(benzamido)pyrazine-2-carboxamides, exerting MIC (Minimum Inhibitory Concentration) from 1.95 to 31.25 µg/mL. Detailed structure-activity relationships were established and rationalized in silico with regard to mycobacterial ProRS as a probable target. The active compounds preserved their activity even against multidrug-resistant strains of Mycobacterium tuberculosis. At the same time, they were non-cytotoxic against HepG2 human hepatocellular carcinoma cells. This project is the first step in the successful repurposing of inhibitors of human ProRS to inhibitors of mycobacterial ProRS with antimycobacterial activity.

The Biological Relevance of Papaverine in Cancer Cells.

Papaverine (PPV), a benzylisoquinoline alkaloid, extracted from the Papaverine somniferum plant, is currently in clinical use as a vasodilator. Research has shown that PPV inhibits phosphodiesterase 10A (PDE10A,) resulting in the accumulation of cyclic adenosine 3', 5'-monophosphate (cAMP) that affects multiple downstream pathways, including phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt), a mammalian target of rapamycin (mTOR) and vascular endothelial growth factor (VEGF). The accumulation of cAMP can further affect mitochondrial metabolism through the activation of protein kinase A (PKA), which activates the mitochondrial complex I. Literature has shown that PPV exerts anti-proliferative affects in several tumorigenic cell lines including adenocarcinoma alveolar cancer (A549) and human hepatoma (HepG-2) cell lines. Cell cycle investigations have shown varying results with the effects dependent on concentration and cell type with data suggesting an increase in cells occupying the sub-G1 phase, which is indicative of cell death. These results suggest that PPV may be a beneficial compound to explore for the use in anticancer studies. More insight into the effects of the compound on cellular and molecular mechanisms is needed. Understanding the effects PPV may exert on tumorigenic cells may better researchers' understanding of phytomedicines and the effects of PPV and PPV-derived compounds in cancer.

Genotoxicity of pyrrolizidine alkaloids in metabolically inactive human cervical cancer HeLa cells co-cultured with human hepatoma HepG2 cells

Pyrrolizidine alkaloids (PAs) are secondary plant metabolites, which can be found as contaminant in various foods and herbal products. Several PAs can cause hepatotoxicity and liver cancer via damaging hepatic sinusoidal endothelial cells (HSECs) after hepatic metabolization. HSECs themselves do not express the required metabolic enzymes for activation of PAs. Here we applied a co-culture model to mimic the in vivo hepatic environment and to study PA-induced effects on not metabolically active neighbour cells. In this co-culture model, bioactivation of PA was enabled by metabolically capable human hepatoma cells HepG2, which excrete the toxic and mutagenic pyrrole metabolites. The human cervical epithelial HeLa cells tagged with H2B-GFP were utilized as non-metabolically active neighbours because they can be identified easily based on their green fluorescence in the co-culture. The PAs europine, riddelliine and lasiocarpine induced micronuclei in HepG2 cells, and in HeLa H2B-GFP cells co-cultured with HepG2 cells, but not in HeLa H2B-GFP cells cultured alone. Metabolic inhibition of cytochrome P450 enzymes with ketoconazole abrogated micronucleus formation. The efflux transporter inhibitors verapamil and benzbromarone reduced micronucleus formation in the co-culture model. Furthermore, mitotic disturbances as an additional genotoxic mechanism of action were observed in HepG2 cells and in HeLa H2B-GFP cells co-cultured with HepG2 cells, but not in HeLa H2B-GFP cells cultured alone. Overall, we were able to show that PAs were activated by HepG2 cells and the metabolites induced genomic damage in co-cultured HeLa cells.

Chemical Composition and Cytotoxic Activity of Extracts from Carpesium divaricatum: In Vitro- versus Field-Grown Plants.

Carpesium divaricatum Sieb. & Zucc. is a plant species rich in terpenoids of anti-inflammatory and cytotoxic activity, especially germacranolides of potential medicinal value. The present study describes in vitro multiplication of C. divaricatum, analysis of active constituents in the multiple shoots, and assessment of cytotoxic activities of extracts prepared from in vitro- and field-grown plants. The plant extracts were evaluated for cytotoxicity using two melanoma cell lines (HTB140 and A375); human keratinocytes (HaCaT); two colon cancer cell lines (Caco2 and HT29); human hepatocellular carcinoma cells (HepG2); two lines of prostate cancer cells (DU145 and PC3) and prostate epithelial cells (PNT2). Chemical compositions of the assayed extracts were analyzed by HPLC/DAD, in reference to isolated compounds. Maximum of 4.07 ± 1.61 shoots regenerated from a nodal explant of C. divaricatum, cultivated in a liquid MS medium supplemented with thidiazuron (1 μM). In vitro grown shoots and plantlets of C. divaricatum accumulated terpenoids that are known as active constituents of the intact plant. Cytotoxic activity of the extracts prepared from the in vitro cultured plants was like that demonstrated by the extracts prepared from field-grown plants and seemed to be more selective than cytotoxicities of the individual germacranolides.

Effects of the emerging contaminant 1,3,6,8-tetrabromocarbazole on the NF-κB and correlated mechanism in human hepatocellular carcinoma cells

1,3,6,8-Tetrabromocarbazole (1368-BCZ) is identified as an emerging contaminant that exerts angiogenic effects. Multiple studies indicated there was a positive correlation between angiogenesis and nuclear factor kappa B (NF-κB) activation. While the role of NF-κB in inflammation and apoptosis has been well known, the potential biological effects of 1368-BCZ on NF-κB signaling and related mechanism remain unclear. We, therefore, explored the possible effects of 1368-BCZ on the NF-κB pathway at the gene and protein levels and confirmed that NF-κB activation by 1368-BCZ exposure caused an augmented phosphorylated protein level, induction of NF-κB response element (κBRE)-driven luciferase activity and upregulation of transcriptional level of downstream responsive genes. Although 1368-BCZ did not produce detectable changes in hepatic fibrosis in vivo, it obviously altered the apoptosis in human hepatocellular carcinoma (HepG2) cells. Furthermore, the induction of apoptosis was confirmed by the increased cleaved caspase-3 level. These data revealed the activating effects of 1368-BCZ on NF-κB and its involvement in the underlying mechanisms, providing additional information for toxicology studies of emerging contaminants and introducing a mechanism-based toxicological evaluation of emerging pollutants.