Human Hepatocellular Carcinoma Cell Bel-7402 Research Articles

Emodin Induced SREBP1-Dependent and SREBP1-Independent Apoptosis in Hepatocellular Carcinoma Cells.

Reynoutria multiflora (Thunb.) Moldenke (He Shou Wu) has been used for about 20 centuries as a Chinese medicinal herb for its activities of anticancer, anti-hyperlipidemia, and anti-aging. Previously, we found that He Shou Wu ethanol extract could induce apoptosis in hepatocellular carcinoma cells, and we also screened its active components. In this study, we investigated whether lowering lipid metabolism of emodin, a main active component in He Shou Wu, was associated with inhibitory effects in hepatocellular carcinoma cells. The correlation of apoptosis induction and lipid metabolism was investigated. The intrinsic apoptotic cell death, lipid production, and their signaling pathways were investigated in emodin-treated human hepatocellular carcinoma cells Bel-7402. The data showed that emodin triggered apoptosis in Bel-7402 cells. The mitochondrial membrane potential (ΔΨm) was reduced in emodin-treated Bel-7402 cells. We also found that emodin activated the expression of intrinsic apoptosis signaling pathway-related proteins, cleaved-caspase 9 and 3, Apaf 1, cytochrome c (CYTC), apoptosis-inducing factor, endonuclease G, Bax, and Bcl-2. Furthermore, the level of triglycerides and desaturation of fatty acids was reduced in Bel-7402 cells when exposed to emodin. Furthermore, the expression level of messenger RNA (mRNA) and protein of sterol regulatory element binding protein 1 (SREBP1) as well as its downstream signaling pathway and the synthesis and the desaturation of fatty acid metabolism-associated proteins (adenosine triphosphate citrate lyase, acetyl-CoA carboxylase alpha, fatty acid synthase (FASN), and stearoyl-CoA desaturase D) were also decreased. Notably, knock-out of SREBP1 in Bel-7402 cells was also found to induce less intrinsic apoptosis than did emodin. In conclusion, these results indicated that emodin could induce apoptosis in an SREBP1-dependent and SREBP1-independent manner in hepatocellular carcinoma cells.

Design of a Nile red-based NIR fluorescent probe for the detection of hydrogen peroxide in living cells.

In this article, a novel fluorescent probe (NRBE) for detecting H2O2 was developed using benzyl boronic ester as the H2O2-recognized group and Nile red as the matrix. The probe has several advantages, such as good selectivity, high sensitivity (LOD = 75 nM), good water solubility and emission in the near-infrared region (ex/em:585/670 nm). With the NRBE probe, the endogenous H2O2 in human hepatocellular carcinoma cells BEL-7402, was detected, and the H2O2 generated during the ischemia-reperfusion of the cells was imaged. These results show that NRBE can be applied for real-time detection of H2O2 in biological systems.

Differentiation of bel-7402 human hepatocarcinoma cells induced by aqueous extracts of fresh gecko (AG) and its anti-tumor activity in vivo

Ethnopharmacological relevanceGecko, a kind of reptile, has been widely used as a traditional Chinese medicine to treat various diseases including cancer in China for thousands of years. The aim of this study was to investigate the anti-tumor effect of AG (aqueous extracts of fresh gecko) on human hepatocellular carcinoma cell Bel-7402 in vitro and mouse H22 hepatocellular in vivo. Further to underlie the molecular mechanism of AG inducing the differentiation of Bel-7402 cells. Materials and methodsAG was obtained by water extracting method and qualitatively analyzed through High Performance Liquid Chromatography. The total protein concentration of AG was measured by BCA (bicinchoninic acid disodium) assay. The anti-tumor activities in vivo were analyzed through H22 (mouse hepatocellular carcinoma cell line H22) tumor xenografts mice. The cytotoxic activity of AG on Bel-7402 cells was evaluated by MTT assays. AFP (alpha fetoprotein) was detected by radioimmunoassay. ALB (albumin), ALP (alkaline phosphatase) and γ-GT (γ-glutamyl transpeptidase) were detected by biochemical methods with commercial kits. While morphological changes were observed through an inverted microscope. Moreover, the expression level of the proteins involved in MAPK (mitogen-activated protein kinase) signal pathway which was closely related to cellular differentiation was assessed by Western blot. ResultsAG showed obviously anti-tumor activity in vivo and anti-proliferative activity on Bel-7402 cells in vitro both dose-dependently. The number of clones of Bel-7402 cells treated with AG reduced and the cells were displaying differentiation state such as relatively bigger size and dispersed growth. The biochemical function markers of the cells were significantly changed after being treated with AG. The data showed that AFP secretion of the cells decreased 42.5%, ALB secretion increased 58.9%, the activity of ALP and γ-GT markedly decreased 67.0% and 48.5% separately when the concentration of AG was 10μl/ml, and those effects were all in a dose-dependent manner. The major original and phosphorylated signal proteins (ERK1/2 (extracellular sigal-regualted kinase 1/2), P38 (p38 MAPK) and JNK1/2 (c-Jun N-terminal kinase 1/2)) involved in MAPK signal pathway were measured and the results showed that AG activated the ERK1/2 of Bel-7402 cells. ConclusionsAG has anti-tumor activity in vivo and inhibits Bel-7402 cell proliferation in vitro through inducing cell differentiation, and the mechanism involves the activation of ERK1/2.

Celastrol exerts synergistic effects with PHA-665752 and inhibits tumor growth of c-Met-deficient hepatocellular carcinoma in vivo

PHA665752 (PHA), a selective small molecule c-Met Inhibitor, potently inhibited HGF-stimulated and constitutive c-Met phosphorylation, as well as HGF and c-Met-driven phenotypes of a variety of tumor cells including hepatocellular carcinoma cells. However, these effects were impaired in c-Met-deficient cancer cells. In the present study, we investigated the potential anti-human c-Met-deficient hepatocellular carcinoma effects of Celastrol, a novel triterpene, and its combination with PHA. Human hepatocellular carcinoma cells BEL-7402 (c-Met-positive) and Huh7 (c-Met-deficient) were treated with different dose of PHA with or without equal dose of Celastrol, and cell growth, cell cycle and apoptosis were evaluated, respectively, by MTT assay, flow cytometry and Caspase3/7 activity. Nude mice bearing Huh7 xenografts were used to assess the in vivo anti-tumor activity. Our results showed that Celastrol at high concentration (>1.0 μM) induced G2/M arrest and apoptosis with the activation of Caspase3/7 in Huh7 cells whereas at low concentration (<1.0 μM) had no obvious effects. Low concentration Celastrol presented significant combined effects with PHA on Huh7 cells and Huh7 xenografts in terms of growth inhibition, migration inhibition and apoptosis induction. These results suggest that Celastrol and its combination with PHA present the therapeutic potential on c-Met-deficient hepatocellular carcinoma, and deserve further preclinical and clinical studies.

Effects of transmembrane protease serine 4 on the proliferation and invasion of hepatocellular carcinoma

Objective This study investigates the effects of transmembrane protease serine 4 (TMPRSS4) on the proliferation and invasion of hepatocellular carcinoma (HCC).Methods Stable TMPRSS4 over-expression cell lines were established using human HCC cells BEL-7402 which were transfected by lentiviral vectors.Expression of TMPRSS4 was measured by Western blot and RT PCR.Cell proliferation was measured by MTT analysis and HCC cell invasion was evaluated by tran swell invasion assay.Results RT-PCR and Western blot analysis showed that compared to control HCC cells (control group) and cells transfected with blank plasmid (blank plasmid group),the expression of TMPRSS4 in cells transfected with TMPRSS4 plasmid (TMPRSS4 plasmid group) was significantly increased.There were no significant differences among these three groups regarding the proliferation of HCC cells (P＞0.05).The number of cells in the TMPRSS4 plasmid group that passed through the transwell inserts was 49.3 ± 7.4,significantly higher than that of control group and blank plasmid group,which was 23.3 ± 5.0 and 19.3 ± 3.2 respectively (P＜0.05).Conclusion Therefore,the expression of TMPRSS4 can be detected in HCC,and over-expression of TMPRSS4 promotes the invasion of HCC but has no effect on cell proliferation. Key words: Hepatocellular carcinoma; Ttransmembrane protease serine 4; Invasion; Proliferation

Schedule-dependent effects of sorafenib in combination with paclitaxel on human hepatocellular carcinoma cell line BEL-7402

Sorafenib is a multi-targeted antitumor drug. The monotherapy efficacy of sorafenib is relatively low. This study was to evaluate the schedule-dependent effect of sorafenib in combination with paclitaxel (TAX) on human hepatocellular carcinoma cell BEL-7402, and explore the underlying mechanism. BEL-7402 cells were treated with sorafenib or paclitaxel alone or in three different schedules: sorafenib was give prior to, after, or simultaneously with paclitaxel. The half maximal inhibitory concentration (IC50) of sorafenib and paclitaxel was estimated by MTT. Alteration of cell cycle and apoptosis were analyzed by flow cytometry. The protein level of Bcl-2 in BEL-7402 cells was measured by western blot. At 48 h, the IC50 of sorafenib and paclitaxel for BEL-7402 cells was (2.43+/-0.32) microg/mL and (1.89+/-0.72) microg/mL, respectively. Sorafenib caused cell cycle arrest at S phase, while paclitaxel blocked cells at G2/M phase. S and G2/M phases were extended and a higher apoptotic rate (36.43+/-2.29)% was induced when sorafenib was given after paclitaxel in comparision with other groups (P<0.01). The protein level of Bcl-2 was the lowest in BEL-7402 cells treated with sorafenib after paclitaxel. Administration of sorafenib after paclitaxel induces a higher apoptotic rate in BEL-7402 cells than administration before or simutanously with paclitaxel. This is probably due to that the two drugs act on different cell cycle phases and the expression of Bcl-2 might be involved.

Biological activities of procyanidins from the bark of Pinus caribaea Morelet

The extraction and biological activities of procyanidins from the bark of Pinus caribaea Morelet (P. caribaea) have been studied for the first time in our research. The main factors influencing the extraction, including solvent, temperature and liquid-to-solid (L/S) ratio, were discussed. The results illustrate that the suitable conditions for the extraction of procyanidins from the bark of P. caribaea are as follows: extraction temperature at 50°C, L/S ratio at 11 : 1 and ethanol volume fraction at 70%. The focus of this study is to investigate the biological activities of the procyanidins from the bark of P. caribaea. Free-radical scavenging activity of the procyanidins was measured by means of 2,2-diphenyl-1-picrylhydrazyl (DPPH) method, and it was clear that the procyanidins product had a strong radical scavenging ability. Furthermore, the inhibition effect of the procyanidins from the bark of P. caribaea on different human cancer cells cultured in vitro was investigated. The results indicated that the procyanidins extracted with water had a stronger inhibition on promycelocytic cells HL-60, an effective inhibition on human stomach adenocarcinoma cells BGC-823 and human hepatocellular carcinoma cells BEL-7402, but no effect on human lung carcinoma cells A549, whereas the procyanidins extracted with ethanol had an effective inhibition on HL-60 and BGC-823, but no effect on A549 or BEL-7402.

Heat shock protein 70 chaperoned alpha-fetoprotein in human hepatocellular carcinoma cell line BEL-7402

To investigate the interaction between heat shock protein 70 (HSP70) and ?-fetoprotein (AFP) in human hepatocellular carcinoma (HCC) cell line BEL-7402. The expression and localization of HSP70 and AFP in human HCC cell line BEL-7402 were determined by immunocytochemistry and indirect immunofluorescence cytochemical staining. The interaction between HSP70 and AFP in HCC cells was analyzed by immunoprecipitation and Western blot. Immunocytochemical staining detection showed that HCC cell BEL-7402 expressed a high level of HSP70 and AFP synchronously. Both were stained in cell plasma. AFP existed in the immunoprecipitate of anti-HSP70 mAb, while there was HSP70 in the immunoprecipitate of anti-AFP mAb. HSP70 chaperones AFP in human HCC cell BEL-7402. The interaction between HSP70 and AFP in human HCC cell can be a new route to study the pathogenesis and immunotherapy of HCC.