Effects In HepG2 Cells Research Articles

Effect of quercetin on cell protection via erythropoietin and cell injury of HepG2 cells

Quercetin is a flavonoid that has roles in both cytoprotection and cytotoxicity. The relation of queretin's cytoprotective and cytotoxic effects are unknown. Quercetin has been shown to induce expression of hypoxia-inducible factor, a protein that is known to regulate transcription of the erythropoietin (EPO) gene, and EPO is known to have a cytoprotective effect. This study used HepG2 cells to assess whether the cell-protective and/or cytotoxic roles of quercetin are mediated by promotion of EPO production. Increases in the levels of HIF-1α protein and EPO mRNA were quercetin concentration-dependent, with significant increases observed from 10 μM quercetin. Silencing of EPO expression by si-EPO RNA attenuated quercetin-induced cytoprotection against hydrogen peroxide toxicity. Cytotoxicity, evidenced by the induction of apoptosis, was significantly increased by exposure to 50 μM quercetin. Specifically, the levels of cleaved caspase-3 and Bax and the rate of cell death increased, and the level of Bcl-2 decreased, in cells treated with 50 μM quercetin. In contrast, exposure to 10 μM quercetin attenuated cisplatin-induced apoptosis. However, quercetin's ability to protect cells from cisplatin-induced apoptosis was eliminated when EPO expression was silenced using si-EPO RNA. Together, these results suggested that quercetin's cytoprotective effects in HepG2 cells are mediated via EPO production.

Celastrol downregulates E2F1 to induce growth inhibitory effects in hepatocellular carcinoma HepG2 cells.

Celastrol, a natural compound extracted from Tripterygiumwilfordii, is known to exhibit potential anticancer activities in various types of tumor cells. E2F1 is reported to be overexpressed in several types of human tumors and its inactivation may be a valuable novel potential therapeutic strategy for cancer treatment. However, the molecular mechanism underlying the pro-apoptotic effects of celastrol on hepatocellular carcinoma (HCC) cells remains unclear, and E2F1-targeted compounds have been rarely identified. In the present study, we demonstrated that celastrol inhibited the proliferation of human HCC cells and triggered apoptosis of HepG2 cells in a caspase-dependent manner. E2F1 was potently downregulated by celastrol in a dose-and time-dependent manner at both the mRNA and protein levels. Moreover, siRNA-mediated E2F1 silencing enhanced celastrol-induced apoptosis and inhibition of proliferation. Our data imply that downregulation of E2F1 may be a key factor in the celastrol-mediated inhibitory effects in HepG2 cells, and celastrol can serve as a leading compound for the development of compounds designed to inactivate E2F1 for HCC therapy.

20(S)-25-methoxyl-dammarane-3β, 12β, 20-triol negatively regulates activation of STAT3 and ERK pathways and exhibits anti-cancer effects in HepG2 cells.

The pro-inflammatory cytokine interleukin 6 (IL-6), via activating its downstream JAK/STAT3 and Ras/ERK signaling pathways, is involved in cell growth, proliferation and anti-apoptotic activities in various malignancies. To screen inhibitors of IL-6 signaling, we constructed a STAT3 and ERK dual-pathway responsive luciferase reporter vector (Co.RE). Among several candidates, the natural compound 20(S)-25-methoxyl-dammarane-3β, 12β, 20-triol (25-OCH3-PPD, GS25) was identified to clearly inhibit the luciferase activity of Co.RE. GS25 was confirmed to indeed inhibit activation of both STAT3 and ERK pathways and expression of downstream target genes of IL-6, and to predominantly decrease the viability of HepG2 cells via induction of cell cycle arrest and apoptosis. Interestingly, GS25 showed preferential inhibition of HepG2 cell viability relative to normal liver L02 cells. Further investigation showed that GS25 could not induce apoptosis and block activation of STAT3 and ERK pathways in L02 cells as efficiently as in HepG2 cells, which may result in differential effects of GS25 on malignant and normal liver cells. In addition, GS25 was found to potently suppress the expression of endogenous STAT3 at a higher concentration and dramatically induce p38 phosphorylation in HepG2 cells, which could mediate its anti-cancer effects. Finally, we demonstrated that GS25 also inhibited tumor growth in HepG2 xenograft mice. Taken together, these findings indicate that GS25 elicits its anti-cancer effects on HepG2 cells through multiple mechanisms and has the potential to be used as an inhibitor of IL-6 signaling. Thus, GS25 may be developed as a treatment for hepatocarcinoma with low toxicity on normal liver tissues as well as other inflammation-associated diseases.

Pentoxifylline induces apoptosis of HepG2 cells by reducing reactive oxygen species production and activating the MAPK signaling

AimsPentoxifylline (PTX) is a methylxanthine derivative and has potent anti-tumor activity. This study aimed at investigating the anti-HCC effects of PTX and associated molecular mechanisms. Main methodsThe effects of varying doses of PTX on viability, cell cycle and apoptosis of HepG2 cells were determined by MTT and flow cytometry, respectively. The effects of PTX on the production of reactive oxygen species (ROS), expression of pro- and anti-apoptotic regulators and activation of the MAPK signaling in HepG2 cells were analyzed by flow cytometry and Western blot assays. The effects of PTX on the growth of implanted HepG2 cells and their apoptosis in mice were examined. Key findingsOur results indicated that PTX inhibited proliferation of HepG2 cells and induced HepG2 cell cycle arrest at G0/G1 phase and apoptosis in a dose- and time-dependent manner. Treatment with PTX reduced levels of ROS and Bcl-XL expression, but increased caspase 3 and caspase 9 expression and JNK and ERK1/2 phosphorylation in HepG2 cells. Pre-treatment with n-acetyl-l-cysteine (NAC), a ROS scavenger, enhanced PTX-mediated cell cycle arrest, apoptosis and the JNK and ERK MAPK activation, while pre-treatment with SP600125 or PD98509 attenuated PTX-mediated effects in HepG2 cells. Treatment with PTX inhibited the growth of implanted HCC and promoted HCC apoptosis in mice. SignificanceOur data demonstrate that PTX inhibits proliferation of HepG2 cells and induces HepG2 cell apoptosis by attenuating ROS production and enhancing the MAPK activation in HepG2 cells.

Liraglutide, a glucagon-like peptide-1 analog, induce autophagy and senescence in HepG2 cells

It has been reported that glucagon-like peptide-1 (GLP-1) agents have been associated with both the increased risk of cancer and inhibition of tumor growth and metastases. The aim of this study is to evaluate the effect of liraglutide on hepatocellular carcinoma cells – HepG2. Cytometry was used to evaluate mechanism related to decreased cell proliferation. Nuclear staining and morphometric analysis were also used to verify the process that was taking place after treatment with liraglutide, and in order to better understand the mechanism, TGF-β1 was performed. HepG2 cells decreased proliferation after liraglutide treatment without altering oxidative stress levels. Liraglutide was able to induce autophagy and senescence through the increase of TGF-β1 which possibly explains the growth decrease. We have demonstrated that liraglutide has an antiproliferative effect in HepG2 cells inducing autophagy and senescence by the increase of TGF-β1.

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.

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.

Vasculoprotective Effects of Water Extracts of Black, Green and Dark Tea in Vitro

To investigate the preventive effects of tea on hyperglycemia and vascular complications of diabetes, we report the extraction and composition as well as the vasculoprotective effects of black tea extract (BTE), green tea extract (GTE), and dark tea extract (DTE). High Performance Liquid Chromatography (HPLC) and colorimetric methods were conducted to analyze for tea catechins, caffeine, polyphenols, amino acids and polysaccharides of BTE, GTE and DTE. The inhibitory effects of α-glucosidase, aldose reductase (AR), advanced glycation end-products (AGEs) and glucose uptake promotion effect in BTE, GTE and DTE were explored in vitro. Contents of six major catechin forms and total catechin as well as polyphenols are higher in GTE and DTE than BTE. BTE, GTE, and DTE showed the inhibitory effects of a-glucosidase, AR, and AGEs, but only DTE exhibited the glucose uptake promotion effect in HepG2 cells. The results suggest that regular consumption of tea can help prevent the progression of hyperglycemia and the vascular complications of diabetes.

Genotoxic and chemopreventive assessment of Cynara scolymus L. aqueous extract in a human-derived liver cell line

Cynara scolymus L., popularly known as artichoke, is consumed as food and used as tea infusions for pharmacological purposes to treat liver dysfunctions and other conditions. Scientific data on the safety and protective effect of artichoke in human-derived liver cells is missing. This study investigated the genotoxic and modulatory effect of a liophilized extract suspended in water of C. scolymus L. leaves. Four extract concentrations (0.62, 1.25, 2.5 and 5.0 mg/mL) were evaluated using the comet assay on human hepatocyte cultures, HepG2 cells. Genotoxicity was assessed after two treatment periods, 1 and 24 h. Antigenotoxicity was evaluated against oxidative lesions induced by hydrogen peroxide in pre-, simultaneous and post-treatment protocols. Artichoke leaves aqueous extract induced genotoxic effects in HepG2 cells after 1- and 24-h treatments. In turn, extract concentrations of 0.62, 1.25 and 2.5 mg/mL, exhibited a protective effect in pretreatment, compared to hydrogen peroxide alone. However, in simultaneous and post-treatment protocols, only the lowest concentration reduced the frequency of DNA damage induced by hydrogen peroxide. In addition, in the simultaneous treatment protocol, the highest artichoke extract concentration increased hydrogen peroxide genotoxicity. It can be concluded that artichoke is genotoxic, in vitro, to HepG2 cells, but can also modulate hydrogen peroxide DNA damage.

Liposome-mediated RNA interference delivery against Erk1 and Erk2 does not equally promote chemosensitivity in human hepatocellular carcinoma cell line HepG2

Background: Extracellular signal-regulated kinase (Erk)1 and Erk2 are central mediators of mitogen-activated protein kinase signaling pathway, which plays a key role in proliferation and chemoresistance of cancer cells. However, the effect of Erk1 and Erk2 in these processes may not be the same. The aim of this study was to investigate differential effect of Erk1 and Erk2 down-regulation on chemoresistance in human hepatocellular carcinoma (HCC) cells. Expression level and relative expression analysis in HepG2 cells were performed using RT-PCR and qRT-PCR, respectively. Phosphorylated-Erk1/2 and apoptosis analysis was performed by flow-cytometry (FCM) technique.Results: The results showed a higher expression level of Erk2 relative to Erk1 in HepG2 cells (P < 0.01). A significant decrease in phosphorylated-Erk1/2 and a compensational response was observed after Erk1 and/or Erk2 silencing using specific small interfering ribonucleic acids (siRNAs) (P < 0.01). Furthermore, 5-fluorouracil (5-FU) chemotherapy following siRNA-mediated knockdown lead to a significant enhancement of chemosensitivity with a higher rate of early apoptosis in Erk2 silencing relative to that of Erk1) + 9%, P < 0.01). 5-FU treatment after dual knockdown of Erk1/2 showed higher rate of early apoptosis relative to single Erk1 silencing (+9.25%, P < 0.01) and also higher rate of late apoptosis compared to single Erk1 and Erk2 silencing (+4.96% and +4.66%, P < 0.01).Conclusion: Our data show that liposomal siRNA-mediated down-regulation of Erk1/2 can lead to potent chemosensitizing effects in HepG2 cells. Moreover, a higher chemosensitivity following Erk2 down-regulation than Erk1 down-regulation may be associated with the higher expression of Erk2 in human HCC.

Bioactive diarylheptanoids and stilbenes from the rhizomes of Dioscorea septemloba Thunb

The 70% ethanol extract of Dioscorea septemloba rhizomes was subjected to chromatographic columns including silica gel, ODS, D101 resin, Sephadex LH-20 and HPLC, to yield three new diarylheptanoids (1–3) and two novel stilbenes (4 and 5), together with six known ones (6–11). Their structures were elucidated on the basis of extensive 1D NMR (1H and 13C), 2D NMR (COSY, HSQC, HMBC, NOESY), IR, and MS spectroscopic data analyses. Moreover, the in vitro bioactivity screening experiment suggested that compounds 6–8 and 10 could significantly increase the glucose consumption in differentiated L6 myotubes, and that 2, 9–11 displayed triglyceride inhibitory effects in HepG2 cells.

Synthesis, Characterization and Cytotoxic Evaluation of Graphene Oxide Nanosheets: In Vitro Liver Cancer Model

Background:Graphene nanosheets have a broad spectrum of biomedical applications. Hepatocellular cancer (HCC) is a major health problem in the Egyptian population. Currently, treatment strategies are invasive and have several adverse side effects. Thus, other approaches are required for managing this aggressive type of cancer. Our objective here was to prepare and characterize graphene oxide nanosheets and evaluate cytotoxic effect at the molecular level in an in vitro human liver cancer cell model (HepG2).Methods:Graphene oxide nanosheets were generated by chemical oxidation and characterized by transmission electron microscopy and X-ray diffraction. Cytotoxic effects in HepG2 cells were monitored by sulforhodamine B (SRB) colorimetric assay followed by flow cytometric analysis. Molecular investigations of DNA fragmentation and expression of some apoptotic genes at the transcriptional RNA level were also performed.Results:Treatment of HepG2 cells with 400µg/ml graphene oxide nanosheets showed alteration in cell morphology after 24 h. Flow cytometry revealed accumulation of cells in S phase of cell cycle followed by dramatic effects on cellular DNA. Extensive evaluation of the cytotoxic effects of graphene oxide showed increased mRNA Bax apoptotic gene expression with not of P53 and caspase-3 mRNA after 24h, suggesting involvement of an intrinsic apoptotic caspase-independent pathway.Conclusion:Graphene oxide can mediate apoptotic gene signaling in human liver cancer cells opening a novel approach to cancer management. Further analyses at the molecular level are now required to confirm our results and facilitate biomedical applications in vivo.

Protective effects of Cambodian medicinal plants on tert‑butyl hydroperoxide‑induced hepatotoxicity via Nrf2‑mediated heme oxygenase‑1.

Liver diseases are considered to be primary contributors to morbidity and mortality rates in humans. Oxidative stress is critical in liver injury, and oxidant‑induced liver injury may be caused by toxins, including tert‑butyl hydroperoxide (t‑BHP). The present study investigated the hepatoprotective activities of 64 crude ethanol extracts of Cambodian medicinal plants against t‑BHP‑induced cytotoxicity in human liver‑derived HepG2 cells, and assessed their cytoprotective mechanism pertaining to the expression of heme oxygenase (HO)‑1 and nuclear factor E2‑related factor 2 (Nrf2). Protective effects in HepG2 cells were determined by MTT assay. Protein expression levels of HO‑1 and Nrf2 were determined by western blotting and mRNA expression levels were determined by reverse transcription‑quantitative polymerase chain reaction. Of the 64extracts, 19 extracts exhibited high hepatoprotective activities: Ampelocissus martini, Bauhinia bracteata, Bombax ceiba, Borassus flabellifer, Cardiospermum halicacabum, Cayratia trifolia, Cinnamomum caryophyllus, Cyperus rotundus, Dasymaschalon lomentaceum, Ficus benjamina, Mangifera duperreana, Morinda citrifolia, Pandanus humilis, Peliosanthes weberi, Phyllanthus emblica, Quisqualis indica, Smilax glabra, Tinospora crispa and Willughbeia cochinchinensis, with half maximal effective concentrations ranging between 59.23 and 157.80µg/ml. Further investigations revealed that, of these 19 extracts, HO‑1 and Nrf2 were expressed in P. weberi and T. crispa expressed in a dose‑dependent manner. In addition, the activities of reactive oxygen species were suppressed following treatment of these two extracts in t‑BHP‑induced HepG2 cells. These results indicated that, of the 64 Cambodian plants, P. weberi and T.crispa exhibited hepatoprotective effects on t‑BHP‑induced cytotoxicity in HepG2 cells, possibly by the induction of Nrf2‑mediated expression of HO‑1. Taken together, these results suggested that T.crispa or P.weberi may offer potential for therapeutic applications in liver disease characterized by oxidative stress.

Rearranged oleanane type saponins, astraisoolesaponins A1–A3 and B, from the stems of Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao

Four novel saponins, astraisoolesaponins A1–A3 and B (1–4) were isolated from the stems of Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao. Spectroscopic analysis indicated that 1–4 are the first example of saponins possessing a 28-methyl-rearranged carbon skeleton. Their absolute configurations were determined by electronic circular dichroism (ECD) calculations. Both 3 and 4 displayed triglyceride accumulation inhibitory effects in HepG2 cells and glucose uptake increase effects on L6 cells.

Role of recombinant human erythropoietin loading chitosan-tripolyphosphate nanoparticles in busulfan-induced genotoxicity: Analysis of DNA fragmentation via comet assay in cultured HepG2 cells

Busulfan is one of the most effective chemotherapeutic agents used for the treatment of chronic myeloid leukemia. Busulfan is involved in secondary malignancy due to its genotoxic potential in normal tissues. As an alkylating agent busulfan can cause DNA damage by cross-linking DNAs and DNA and proteins, induces senescence in normal cells via transient depletion of intracellular glutathione (GSH) and subsequently by a continuous increase in reactive oxygen species (ROS) production. Erythropoietin, a glycoprotein widely used against drug induced anemia in cancerous patients and regulates hematopoiesis, has been shown to exert an important cyto-protective effect in many tissues. Recombinant human erythropoietin has been demonstrated to directly limit cell injury and ROS generation during oxidative stress. Furthermore, rhEPO decreased levels of pro-apoptotic factor (Bax) and also increased expression of the anti-apoptotic factor Bcl2. According to EPO's short half-life and requirements for the frequently administration, finding the new strategies to attenuate its side effects is important. The aim of this study was to explore whether rhEPO loading chitosan-tripolyphosphate nanoparticles protects against busulfan-induced genotoxicity in HepG2 cells. For this purpose cells were incubated with busulfan alone, regular rhEPO alone and regular rhEPO and CS-TPP-EPO nanoparticles along with busulfan in pre and co-treatment condition. Our results showed that busulfan induced a noticeable genotoxic effects in HepG2 cells (p<0.0001). Both regular rhEPO and CS-TPP-EPO nanoparticles reduced the effects of busulfan significantly (p<0.0001) by reduction of the level of DNA damage via blocking ROS generation, and enhancement intracellular glutathione levels. CS-TPP-EPO nanoparticles were more effective than regular rhEPO in both pre and co-treatment conditions. In conclusion, our results show that administration of rhEPO and CS-TPP-EPO nanoparticles especially in the pre-treatment conditions, significantly decreased the level of DNA damage induced by busulfan, measured with the comet assay, in HepG2 cells compared to the regular rhEPO group.

Effects of Epothilone A in Combination with the Antidiabetic Drugs Metformin and Sitagliptin in HepG2 Human Hepatocellular Cancer Cells: Role of Transcriptional Factors NF-κB and p53.

Type 2 diabetes mellitus patients are at increased risk of many forms of malignancies, especially of the pancreas, colon and hepatocellular cancer. Unfortunately, little is known of the possible interaction between antidiabetic drugs and anticancer agents. The present study investigates the influence of metformin (MET) and sitagliptin (SITA) on the in vitro anticancer activity of the microtubule depolymerization inhibitor agent epothilone A (EpoA). Hepatocellular liver carcinoma cell line (HepG2) viability and apoptosis were determined by the MTT test and by double staining with PO-PRO-1 and 7-aminoactinomycin D, respectively, after treatment with EpoA, metformin or sitagliptin. The levels of nuclear factor NF-κB and p53 were evaluated in the presence and absence of inhibitors. While EpoA and MET inhibited HepG2 cell proliferation, SITA did not. EpoA and SITA induced higher p53 levels than MET. All tested drugs increased the level of NF-κB. Only MET enhanced the proapoptotic effect of EpoA. The EpoA+MET combination evoked the highest cytotoxic effect on HepG2 cells and led to apoptosis independent of p53, decreasing the level of NF-κB. These findings support the link between NF-κB and p53 in the modulation of apoptotic effects in HepG2 cells treated by EpoA. Our studies indicate that the combination of EpoA and MET applied in subtoxic doses has a stronger cytotoxic effect on liver cancer cells than each of the compounds alone. The therapeutic advantages of the combination of EpoA with MET may be valuable in the treatment of patients with diabetes mellitus type 2 (T2DM) and liver cancer.

Preparation and Sonodynamic Antitumor Effect of Protohemin-Conjugated Multiwalled Carbon Nanotubes Functionalized with Carboxylic Group.

Preclinical Research Sonodynamic therapy (SDT) is a cutting edge approach to treating cancer that involves necrosis and/or apoptosis. Multiwalled carbon nanotubes functionalized with carboxylic groups (MWCNTs-COOH) due their physicochemical structure represent a novel drug delivery system in the field of nanomedicine. The purpose of the research reported in this paper was to increase the antitumor potency and reduce the potential side effects of protohemin (Ph), a sonosensitizer for SDT, which was noncovalently encapsulated into MWCNTs-COOH (MWCNTs-Ph). The Ph loading efficiency in MWCNTs-COOH carrier was determined as approximately 68.8% (w/w). The growth inhibition rate of MWCNTs-Ph (Ph: 180 μg/mL) was approximately 95 ± 8.5%, whereas Ph-F (Ph: 180 μg/mL) inhibited 58 ± 4.5% of tumor cell. Ph (Ph: 180 μg/mL) alone had no antitumor effect in HepG-2 cells using ultrasound treatment at 1.0 MHz and 0.5 W/cm(2) for 100 s. Assessment of the antitumor effects of MWCNTs-Ph and Ph-F at day 11 after SDT showed that he tumor inhibition ratio for MWCNTs-Ph (6.18 × 10(-2) g·kg(-1) ·d(-1) ) was 82.8%, twice that of Ph-F (6.18 × 10(-2) g·kg(-1) ·d(-1) ) ay 41.8%. In conclusion, MWCNTs-Ph had increased antitumor efficiency and also decreased potential side effects. Drug Dev Res 77 : 152-158, 2016. © 2016 Wiley Periodicals, Inc.

In vitro protective effects of Hoveniae Semen cum Fructus extracts against oxidative stress

We evaluated the in vitro cytoprotective effects of Hoveniae Semen cum Fructus (HSCF) extracts against oxidative stress-mediated cell damage using HepG2 cells. Cytotoxic effects of HSCF extracts were observed in HepG2 cells, and the 50% inhibitory concentration was determined. Cytoprotective effects of sublethal doses of HSCF extracts were evaluated using a tert-butyl hydroperoxide (tBHP)-induced cellular damage model. We also assessed whether NFE2-related factor-2 (Nrf2) was transactivated by HSCF extracts. The antioxidant capacity of HSCF extracts was evaluated with superoxide dismutase (SOD) and catalase (CAT) activities and the expression of the antioxidant genes glutamate cysteine ligase catalytic subunit (GCLC), hemeoxygenase-1 (HO1), and NAD (P)H dehydrogenase quinone 1 (NQO1). HSCF extracts up to 1,000 μg/mL showed no cytotoxic effect in HepG2 cells. Indeed, 300 and 1,000 μg/mL of HSCF extracts significantly protected HepG2 cells from oxidative stress-mediated cell death by tBHP. As a molecular mechanism, HSCF extracts at 1,000 μg/mL significantly increased Nrf2 transactivation and induced expression of its target genes (GCLC, HO1, NQO1). Furthermore, 1,000 μg/mL of HSCF extracts enhanced SOD activity. Although treatment with 300 and 1,000 μg/mL of HSCF extracts tended to slightly increase CAT activity, the increases were not statistically significant. These findings provide direct evident that HSCF extracts have favorable hepatoprotective effects against oxidative stress through Nrf2-mediated antioxidant gene induction.

A novel anti-alpha-fetoprotein single-chain variable fragment displays anti-tumor effects in HepG2 cells as a single agent or in combination with paclitaxel.

Human hepatocellular carcinoma (HCC) has a high rate of tumor recurrence and metastasis, resulting in shortened survival time. The function of alpha-fetoprotein (AFP) as a regulatory factor in the growth of HCC cells has been well defined. The aim of this study was to investigate the use of a novel AFP-specific single-chain variable fragment that blocked AFP and inhibited HCC cell growth. The results indicated that the anti-AFP single-chain variable fragment (scFv) induced growth inhibition of AFP-expressing HCC cell lines in vitro through induction of G1 cell cycle arrest and apoptosis. The mechanism of apoptosis probably involved with blocking AFP internalization and regulation of the PTEN/PI3K/Akt signaling network. Moreover, the anti-AFP-scFv also effectively sensitized the HepG2 cells to paclitaxel (PTX) at a lower concentration. The combination effect of PTX and anti-AFP-scFv displayed a synergistic effect on HepG2 cells both in vitro and in vivo. Our results demonstrated that targeting AFP by specific antibodies has potential immunotherapeutic efficacy in human HCC.

In vitro Genotoxic Effect of Arsenical Compounds in HepG2 Liver Cells

Aims: The aim of this study was to determine the genotoxic effect in HepG2 cells of the four arsenic compounds by measuring DNA damage, depletion of cellular glutathione (GSH) as a measure of oxidative stress as well as antioxidant agent and apoptotic and necrotic events. Study Design: Arsenic is an environmental chemical of toxicological concern today since it has been shown to be a human genotoxin and carcinogen. This project investigated four forms of inorganic arsenic: arsenate (As+5), sodium arsenite (As+3), arsenic trioxide (As2O3) and dimethyl arsenic acid (DMA+5); a major metabolite of arsenate. Methodology: HepG2 cells (1.5x105/ml) treated with the four arsenics (10 µM) for 24hr. Harvested cells were analysed for GSH concentration by reverse phase HPLC with fluorescence detection. Cells were investigated for DNA damage using the Comet assay. Cells were analysed by flow cytometry to detect apoptosis and necrosis, all running in parallel. Results: The DNA damage for cells dosed with DMA or As2O3 was not significantly different from control. However, significant DNA damage was seen for cells treated with arsenite and arsenate (p&lt;0.001, p&lt;0.05 respectively). Reduced glutathione was increased by arsenic compounds compared to control. However, this was only statistically significant for arsenite (p&lt;0.001) and arsenate (p&lt;0.05). Whereas, results not indicated apoptosis and necrosis in viable adherent cells although this was not statistically different. Conclusions: All four arsenicals appeared to increase GSH content and DNA damage compared to control especially both arsenite and arsenate significantly are different. Further experiments are required to assess the mechanism(s) of genotoxicity of the arsenicals in GSH depletion.

In vitro study of the cytotoxic and genotoxic effects of indomethacin-loaded Eudragit(®) L 100 nanocapsules.

Indomethacin is a non-steroidal anti-inflammatory agent included in one of the most commonly used drug classes worldwide. The use of this drug results in certain side effects, including gastrointestinal complications. Therefore, there exists a need to develop better methods for the delivery of such drugs into the body, such as those employing nanoparticles. The aim of the present study was to evaluate the cytotoxic and genotoxic effects of indomethacin-loaded Eudragit(®) L 100 nanocapsules (NI; based on methacrylic acid and methyl methacrylate) on cells unable (lymphocytes) and able to metabolize drugs (HepG2 cells), using comet and cytokinesis-block micronucleus (CBMN) assays in vitro. Cells were exposed to NI at concentrations of 5, 10, 50, 125, 250, and 500 μg/mL. The comet assay showed that NI induced no significant DNA damage in either cell type at any of the concentrations tested. The CBMN test confirmed these results; however, the highest concentration of 500 μg/mL resulted in a small but statistically significant clastogenic/aneugenic effect in HepG2 cells. These findings should encourage the development of new investigations of this nanomaterial as a delivery vehicle for anti-inflammatory drugs, such as indomethacin.