• Biomedical Science Letters
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• v.11 no.1
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• pp.57-61
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• 2005

The number of sites at which a protein can be readily cleaved by a proteolytic enzyme is greatly influenced by its three-dimensional structure. For native, properly-folded proteins both the rate of cleavage and number of sites at which cleavage takes place are usually much less than for the denatured protein. In order to compare the tertiary structure of recombinant HepG2 type glucose transporter with that of its native counterpart in the erythrocyte, the pattern of tryptic cleavage of the protein expressed in insect cell membranes was therefore examined. After 30 minutes digestion, a fragment of approximate Mr 19,000-21,000 was generated. In addition to this, there were two less intensely stained fragments of apparent Mr 28,000 and 17,000. The pattern of labelling was similar up to 2 hours of digestion. However, the fragments of Mr 19,000-21,000 and Mr 17,000 were no longer detectable after 4 hours digestion. The observation of a very similar pattern of fragments yielded by tryptic digestion of the HepG2 type transporter expressed in insect cells suggests that the recombinant protein exhibits a tertiary structure similar if not identical to that of its human counterpart. Also, the endogenous sugar transporter(s) present in Sf21 cells did not bind cytochalasin B, the potent transporter inhibitor. Therefore, the baculovirus/Spodoptera frugiperda (Sf) cell expression system could be very useful for production of large amounts of human glucose transporters, heterologously.

• Korean Journal of Pharmacognosy
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• v.38 no.2 s.149
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• pp.176-180
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• 2007

Five liglans, sauchinone (1), di-0-methyltetrahydrofuriguaiacin B (2), manassantin A (3), manassantin B (4) and saucerneol B (5), have been isolated from the MeOH extract of Saururus chinesis roots. The evaluation for protective effect of compounds 1-5 against tacrine-induced cytotoxicity in human liver-derived Hep G2 cells was conducted. Compounds 1,2, and 5 showed significant protective effects with the EC$_{50}$ values of74.2${\pm}$0.9, 111.3${\pm}$0.8,64.3${\pm}$0.8 ${\mu}$M, respectively. Silybin, one of the well-known hepatoprotective agents, used as a positive control, and also showed protective effect with an EC$_{50}$ value of 86.2${\pm}$0.5 ${\mu}$M.

• BMB Reports
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• v.31 no.4
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• pp.333-338
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• 1998

The sphingomyelin cycle and ceramide generation have been recognized as potential growth suppression signals in mammalian cells. Ceramide has been shown to induce differentiation, cell growth arrest, senescence, and apoptosis. Although the intracelluar target for the action of ceramide remains unknown, recent studies have demonstrated the role of cytosolic ceramideactivated protein phosphatase(CAPP). In this study, the cytotoxic effect of C2-ceramide, a synthetic cellpermeable ceramide analog, on HEp-2 cells and the mechanism by which ceramide induces cell death were investigated. The addition of exogenous C2-ceramide resulted in a concentration dependent cell death. Okadaic acid, a potent inhibitor of CAPP, enhanced ceramide-mediated cell death, which suggests that CAPP is not involved in this process. To understand the mechanism of action of ceramide, we studied the relationship between ceramide and c-Myc and pRb which are defined components of cell growth regulation. Western blot analyses revealed that C2-ceramide (10${\mu}M$) induced c-Myc down-regulation, but there were no significant changes in pRb. However, treatment of okadaic acid (10 nM) enhanced c-Myc and pRb down-regulation. Reduction of the amount of c-Myc and pRb occurred during HEp-2 cell death. These results suggest that the cytotoxic effect of ceramide in HEp-2 cells may not be mediated through the action of CAPP and that the downstream target for ceramide is c-Myc and pRb.

• Korean Journal of Pharmacognosy
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• v.38 no.4
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• pp.400-402
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• 2007

Tacrine is an acetylcholinesterase inhibitor that is approved for the treatment of Alzheimer's disease. However, tacrine treatment for Alzheimer's disease results in reversible hepatotoxicity in 30-50% of patients, which seriously limits its clinical use. Accordingly, the identification of constituents in natural products that have protective effects on tacrine-induced hepatotoxicity would be valuable. In the present study, an immortalized human hepatoma cell line, HepG2 was employed to screen for agents that protect against tacrine-induced hepatotoxicity. The bioassay-guided fractionation of water extract of Cassiae Semen furnished two anthraquinones, aurantio-obtusin (1) and obtusifolin (2). Compounds 1 and 2 showed hepatoprotective effects with the protection ratio values of 55.3 +/- 0.5% and 41.2 +/- 0.8% at $160{\mu}M$, respectively.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.3
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• pp.1193-1196
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• 2014

Senescence marker protein 30 (SMP30), a hepatocellular carcinoma (HCe) associated antigen had been identified by our research group. To study its mechanisms of regulation and associations with the occurrence and development of HCe, we inhibited expression by RNAi technique, and observed effects on the biological characteristics of Hep G2 cells. In cell viability assays, cell growth in the experimental group (with siRNA transfection) was elevated. In Transwell invasion assays, compared with blank and control groups, numbers of invading cells in the experimental group were significantly increased, whereas in apoptosis assays, the percentage apoptosis demonstrated no differences, but after UV irradiation, that in the experimental group was higher than the other two groups. In a word, SMP30 can inhibit the proliferation and invasion of human hepatoma cells and thus can be regarded as a cancer suppressive factor.

• Journal of Nutrition and Health
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• v.27 no.9
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• pp.910-923
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• 1994

The effects of individual fatty acids, differing in their degree of unsaturation(18:0, 18:1, 18:2 and 18:3) on the biosynthesis and secretion and lipids were investigated in Hep-G2 cells. Synthesis of apolipoprotein was measured by the incorporation of 3H-leucine into apolipoprotein(d<1.21g/ml) and synthesis of lipids was measured by the incorporation of 3H-glycerol and 14C-acetate into various lipid classes. Inclusion of 1.0mM of each fatty acids into the culture medium significantly increased the synthesis of total apolipoprotein and Apo B(p<0.05). However, addition of fatty acid did not affect the synthesis of cellular and medium protein. Among different fatty acids tested, oleic acid had the greatest effect on Apo B synthesis. While stearic, linoleic and linolenic acid, all had similar effects. The secretion of triglyceride into the medium markedly increased in all fatty acid groups being 5-6 times over the albumin control. The triglyceride secretion was the highest int he oleic acid group. The secretion of phospholipid and cholesterol also increased with triglyceride output. A positive relationship existed between the output of lipoprotein-triglyceride and Apo B. Since the synthesis of Apo B was significantly increased when various fatty acids were included into the culture medium, part of the apparently stimulated synthesis of the apolipoprotein may be in response to the increased formation and secretion of lipoprotein lipids.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.3
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• pp.1829-1832
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• 2013

In the present study, investigations were carried out to screen the anticancer activities of fenugreek seed oil against cancer cell lines (HEp-2, MCF-7, WISH cells), and a normal cell line (Vero cells). Cytotoxicity was assessed with MTT and NRU assays, and cellular morphological alterations were studied using phase contrast light microscopy. All cells were exposed toi 10-1000 ${\mu}g/ml$ of fenugreek seed oil for 24 h. The results show that fenugreek seed oil significantly reduced the cell viability, and altered the cellular morphology in a dose dependent manner. Among the cell lines, HEp-2 cells showed the highest decrease in cell viability, followed by MCF-7, WISH, and Vero cells by MTT and NRU assays. Cell viability at 1000 ${\mu}g/ml$ was recorded as 55% in HEp-2 cells, 67% in MCF-7 cells, 75% in WISH cells, and 86% in Vero cells. The present study provides preliminary screening data for fenugreek seed oil pointing to potent cytotoxicity against cancer cells.

• Journal of the Korean Society of Food Science and Nutrition
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• v.30 no.1
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• pp.80-85
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• 2001

Various lines of evidence suggest that dietary components protect the initiation step of carcinogenesis. In this study, the ethylacetate (PGMEA), ethylether (PGMEE), butanol (PGMB) and aqueous (PGMA) soluble fractions of Punica granatum L. (PG) were screened for their growth inhibition using the MTT assay on HepG2, HeLa, C6, MCF-7 and HT-29 cells and for their activity to induce quinone reductase (QR) in HepG2 cells. Among various fractions of Punica granatum L., the PGMEE showed the strongest growth inhibition at 500 $\mu\textrm{g}$/mL which resulted 92.5% on Hela cell lines and 97.8% on C6 cell lines. The PGMEA and PGMB also showed significant growth inhibition. The assay of QR induction on HepG2 cells, grown in the presence of PGMEE at the concentration of 50$\mu\textrm{g}$/mL, was 1.4 times more effective compared with the control value of1.0. These results suggested that useful cancer chemoprevention materials could be isolated from PGMEE fraction of Punica granatum L.

• Preventive Nutrition and Food Science
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• v.7 no.4
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• pp.353-357
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• 2002

We have previously demonstrated that kudzu root extracts have a hypocholesterolemic effect on rats fed diets high in fat and cholesterol. To further elucidate the mechanism involved, in this study we investigated the effect of water extracts of kudzu root, Pueraria radix, on the production of apolipoprotein B$_{100}$ (APo B$_{100}$) in HepG$_2$ liver cells and secretion of apolipoprotein B$_{48}$ (Apo B$_{48}$) in Caco$_2$ cells. Human cell lines, HepG$_2$ liver cells and Caco$_2$ intestinal epithelial cells, were grown with various concentrations (0%, 0.5%, 1.0%, 1.5%, 2.0%) of water extracts of kudzu root in the media. The kudzu root extract decreased Apo B$_{100}$ production and secretion. Treatment of HeP G$_2$ cells with the kudzu root extract also significantly decreased the intracellular total and free cholesterol concentration, and also decreased esterified cholesterol but was only significant at the highest dose of 2%. Apo B$_{48}$ production, but not secretion, from enterocytes was lowered by the kudzu root extracts. This research provided evidence that the hypocholesterolemic properties of kudzu root may be a consequence of decreased production and secretion of Apo B$_{100}$ in the liver and Apo B$_{48}$ in the intestine.

• Journal of Environmental Health Sciences
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• v.29 no.3
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• pp.9-15
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• 2003

The modification of CYP2El activity is a matter of considerable interest because of its role in the metabolic activation of a variety of environmental toxicants. In the present study, the time-course of changes in human CYP2El activities was determined following treatment with solvents (acetone, dimethylsulphoxide or pyridine) using intact HepG2 cells transfected by human CYP2El. Hydroxylation of chlorzoxazone was used for the measurement of CYP2El activity. CYP2E1 protein level was increased upon cultivation of cells in the presence of the solvents for 24 hr. Determination of CYP2El activities after 24 ht cultivation with the solvents demonstrated that acetone or dimethylsulphoxide increased, whereas pyridine inhibited the activities. This differential effect of the solvents on CYP2El activities persisted to subsequent 24 ht. Competitive inhibition study suggested that pyridine has stronger binding affinity to CYP2E1 than acetone or dimethylsulphoxide. These results demonstrate that different binding affinity of the solvents to CYP2El plays important role in determining real CYP2El activity in intact cells after exposure to the solvents. Present study would be helpful in precise understanding of human CYP2El-mediated toxicity.