• Toxicological Research
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• v.11 no.2
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• pp.181-185
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• 1995

To know the mechanism of biphenyl dimethyl dicarboxylate (DDB) in the protection of chemically induced hepatotoxicity, the activity of glutamic pyruvic tran.saminase (GPT) and the level of lipid peroxidation metabolite (malondialdehyde, MDA) and ATP content in hepatocytes were determined in serum and primarily cultured hepatocytes. For in vibo study, rats were pretreated with DDB (300 mg/ kg, p.o.)for 7 days. DDB pretreatment efficiently reduced the elevation of serum GPT activity induced by carbon tetrachloride (1.6 ml/kg, s.c.) and acetaminophen administration (1500 mg/kg, i.p.). In ex vivo study, hepatocytes were isolated from the rats pretreated with DDB (300 mg/kg, p.o.)for 7 days and cultured for 12 hrs before inducing cytotoxicity with chemicals. The MDA formation and the GPT release induced by adriamycin $(1\times10^{-4} mg/ml)$ and cisplatin $(2\times10^{-4} mg/ml)$ were markedly decreased in the hepatocytes from the rats pretreated with DDB as compared to vehicle only. However, DDB pretreatment did not prevent the decrease of ATP contents of hepatocytes induced by cisplatin and adriamycin. In in vitro experiment, DDB was pretreated in primary cultured hepatocytes for 3 days. DDB enhanced the decreases of ATP contents induced by cisplatin and adriamycln. These results suggest that DDB may protect the hepatocytes from injury induced by hepatotoxlcants through inhibiting the lipid peroxidation.

• Korean Journal of Veterinary Research
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• v.50 no.2
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• pp.79-84
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• 2010

The molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) protein is a novel member of the $Ikappa{\beta}$ family. In the present study, we examined the effect of norepinephrine (NE) on MAIL mRNA expression in primary cultured mouse hepatocytes and non-parenchymal liver cells. MAIL mRNA expression in hepatocytes and non-parenchymal liver cells was not directly influenced by NE. However, MAIL mRNA expression in hepatocytes was significantly induced by incubation with a culture medium of non-parenchymal liver cells, treated with NE. Pretreatment with an interleukin (IL)-1 receptor antagonist significantly attenuated the stimulatory effect of the medium. Moreover, exogenous $IL-1{\beta}$ induced MAIL mRNA expression in hepatocytes, while IL-6 and tumor necrosis factor $\alpha$ did not. The concentration of $IL-1{\beta}$ in the medium of non-parenchymal liver cells was significantly increased after NE-treatment. These results suggest that NE can induce MAIL mRNA expression in hepatocytes through $IL-1{\beta}$, released from non-parenchymal liver cells.

• YAKHAK HOEJI
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• v.42 no.1
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• pp.108-113
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• 1998

G009, a polysaccharide isolated from the mycelia of Ganoderma lucidum IYO09, showed a hepatoprotective activity against $CCl_4$ induced cytotoxicity in primary cu ltured rat hepatocytes. Incubation of $CCl_4$-intoxicated hepatocytes with G009 significantly reduced the levels of glutamic pyruvic transaminase and sorbitol dehydrogenase released from hepatocytes in the medium. G009 showed antioxidative effect by elevating the activities of glutathione reductase and superoxide dismutase, and the content of glutathione in $CCl_4$-intoxidcated primary cultured rat hepatocytes. Furthermore, G009 significantly elevated glutathione-S-transferase activity in $CCl_4$-intoxicated primary cultured rat hepatocytes. G009 also reduced the production of malondialdehyde, a byproduct of lipid peroxidation. From these results, it could be concluded that G009 exerted hepatoprotective activity against $CCl_4$-induced cytotoxicity through antioxidation.

• KSBB Journal
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• v.11 no.4
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• pp.453-461
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• 1996

We compared the effects of two different systems of collagen matrix protein application on the survival and the biological functions of cultured primary hepatocytes. The rat liver primary hepatocytes were grown for approximately 40 days in vitro either on single collagen gel or between collagen sandwich gels. The morphological changes were observed for this culture period. While the hepatocytes grown on single gel began to die around at 7 days of culture, the cells grown between collagen gels still maintained their viability and began to die after 15 days. As markers for liver hepatic functions, we determined the biochemical activities of hepatocytes such as the secretions of albumin, fibronectin, fibrinogen, urea, and the reduction of secreted ammonia. We found that the rat hepatocytes cultured between collagen gels maintained fairly good biochemical functions than the hepatocytes cultured on single gel did. Therefore, the application of an extracellular matrix protein, collagen, in sandwich form was confirmed as a better choice for maintaining the functional hepatocytes culture for long term in vitro.

• Archives of Pharmacal Research
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• v.26 no.4
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• pp.338-343
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• 2003

The purpose of the present study was to investigate the bidirectional transport of 1-anilino-8-naphthalene sulfonate (ANS) using isolated rat hepatocytes. The initial uptake rate of ANS by isolated hepatocytes was determined. The uptake process of ANS was saturable, with a $K_m of 29.1\pm3.2 \mu M and V_{max} of 2.9\pm0.1$ mmol/min/mg protein. Subsequently, the initial efflux rate of ANS from isolated hepatocytes was determined by resuspending preloaded cells to 3.0% (w/v) BSA buffer. The efflux process for total ANS revealed a little saturability. The mean value of the efflux clearance was $2.2\pm0.1 \mu$ L/min/mg protein. The efflux rate of ANS from hepatocytes was markedly decreased at $4^{\circ}C$, indicating that the apparent efflux of ANS might not be attributed to the release of ANS bound to the cell surface, but to the efflux of ANS from intracellular space. The efflux clearance was furthermore corrected for the unbound intracellular ANS concentration on the basis of its binding parameters to cytosol. The relation between efflux rate and unbound ANS concentration was fitted well to the Michaelis-Menten equation with a saturable and a nonsaturable components. The $V_{max} and K_m$ values were 0.54 mmol/min/mg protein, and 10.0 $\mu$ M, respectively. Based on the comparison of the ratios of $V_{max} to K_m (V_{max}/K_m)$ corresponding to the transport clearance, the influx clearance was two times higher than the efflux clearance. Together with our preliminary studies that ATP suppression in hepatocytes substantially inhibited ANS influx rate, we concluded that the hepatic uptake of ANS is actively taken up into hepatocytes via the carrier mediated transport system.

• Toxicological Research
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• v.4 no.1
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• pp.37-45
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• 1988

Primary cultures of adult rat hepatocytes were used to study in vitro cytotoxic effects of T-2 toxin on liver cells. When T-2 toxin was added to the culture, a significant depression of the hormonal induction of ${\alpha}$-aminoisobutyric acid (AIB) uptake and tyrosine aminotransferase (TAT) activity was observed. However, T-2 toxin did not affect the uptake of ouabain into hepatocytes. Protein synthesis was inhibited by T-2 toxin, but RNA synthesis was not severely affected. The inhibitory effects of T-2 toxin on protein synthesis was diminished rapidly with culture time and the hepatocytes culture maintained control level of protein synthesis within 24 hrs.

• Journal of Life Science
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• v.10 no.5
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• pp.475-483
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• 2000

Normally, aerobic cells are protected from the damage of free radicals by antioxidative enzymes such as catalase, superoxide dismutase (SOD), glutathione (GSH) peroxidase and GSH-S-transferase. In this study, we have investigate the effect of egg yolk protein hydrolysates on antioxidative activity and the activity of antioxidative enzyme in cultured hepatocytes (Chang). Without the pretreatment with hydrolysate, about 50% of the hepatocytes were killed within 2h by 225$\mu$M tert-butyl hydroperoxide (t-BHP). By contrast, fewer than 20% of the 5 K hydrolysate (permeate from 5 kDa membrane and not passed through 1 kDa membrane)-pretreated hepatocytes were killed by the same concentrations of t-BHP. In addition, the activities of catalase, GSH peroxidase and GSH-transferase were significantly increasing with the treatment of 5 K hydrolysate. These results suggest that 5 K hydrolysate exerts antioxidative effect by increasing activity of antioxidative enzymes.

• Biomolecules & Therapeutics
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• v.4 no.3
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• pp.271-274
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• 1996

The antihepatotoxic activity of Bezoar Bovis and Moschus was investigated by in vitro assay method using galactosamine and carbon tetrachloride-induced cytotoxicity in primary-cultured rat hepatocytes. The antihepatotoxic activity was evaluated by measuring the level of glutamate pyruvate transaminase and sorbitol dehydrogenase which were released from the necrotic hepatocytes to the culture medium. In galactosamine-intoxicated hepatocytes, the chloroform fraction of Bezoar Bovis reduced the level of glutamate pyruvate transaminase and sorbitol dehydrogenase resulting in 65% and 59% protection, respectively. The n-Hexane fraction of Moschus resulted in 45% and 40% protection, respectively in this system. In the case of carbon tetrachloride-intoxicated rat hepatocytes, Bezoar Bovis did not have significant effect and only the aqueous fraction of Moschus showed 42% and 40% protection, respectively.

• Korean Journal of Pharmacognosy
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• v.23 no.4
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• pp.268-275
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• 1992

We studied to screen medicinal plants having hepatoprotective activity with the primary cultured rat hepatocytes intoxicated with carbon tetrachloride cytotoxicity. The lowest concentration and treatment time of carbon tetrachloride giving the greatest intoxication to the primary cultured hepatocytes were observed in 10mM and 60 minutes, respectively. GTP and GOT activity of culture broth of the primary cultured rat hepatocytes intoxicated by $CCl_4$ cytotoxicity at this condition were increased 135.9% and 178.3% compared with that of the primaries cultured hepatocytes not treated with $CCl_4$, respectively. This increased GPT activity was inhibited by glycyrrizin, which was known to have hepatoprotective activity, and the inhibition activity was dependent on the concentration of glycyrrhizin. Forty species among the extracts obtained from 117 species of medicinal plants were shown to have the hepatoprotective activity. Among these 40 species, Prunus persica, Scutellaria baicalensis, Astragalus membranaceus, Tribulus terrestris, Caragana chamlagu, Acanthopanax sessiliflorum and Achyranthes japonica were indicated a lower GPT activity than that of Glycyrrhiza uralensis containing glycyrrhizin and GPT activity of these were indicated 75.5%, 70.0%, 59.0%, 77.5%, 60.0%, 75.0% and 79.0%, respectively.

• Biomolecules & Therapeutics
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• v.11 no.3
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• pp.174-177
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• 2003

The present experiment was performed to investigate the protective effects of paeonol isolated from Moutan Cortex Radicis on primary cultured rat hepatocytes exposed to Br-A23187 ($Ca^{2+}$ ionophore). Br-A23187 is frequently used as a model of cell killing as inducing both necrotic and apoptotic cell death. Hepatocytes were isolated by collagenase perfusion from livers of fasted male Sprague Dawley rats and cultured overnight. Cell viability was determined by propidium iodide using fluorocytometry in Krebs-Ringer-HEPES buffer at pH 7.4. In addition, intracellular calcium was measured by excitation at 340 and 380 nm and emission at 505 nm using a luminescence spectrophotometer. Paeonol (20-100 ${\mu}M$) inhibited cell killing induced by 10 ${\mu}M$ Br-A23187, in a dose-dependent manner. Paeonol also reduced increased intracellular calcium level when hepatocytes were exposed to Br-A23187. Therefore, the present results suggest that paeonol protects the hepatocytotoxicity induced by Br-A23187, via inhibiting the influx of calcium into into rat hepatocytes.