• Asian-Australasian Journal of Animal Sciences
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• v.19 no.9
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• pp.1328-1334
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• 2006

This paper aimed to study the toxicity of arsanilic acid on rat primary hepatocytes in vitro by a modification of the perfusion method. The conditions included concentrations of 0, 1.085, 10.85, 108.5, 1,085 and 10,850 mg/kg arsanilic acid in RPMI 1,640 medium at rat hepatocytes plates respectively, each group had five repeats at $37^{\circ}C$ for 48 h. The rat primary hepatocytes survival ratio, DNA Ladder, activities of glutathione peroxidase (GSH-px), superoxide dismutase (SOD) and catalase (CAT) in hepatocytes, activity of SOD in the medium and the expression of gene bax in hepatocytes were measured at 12 h, 24 h and 48 h respectively. The results showed that arsanilic acid decreased the activities of GSH-px and SOD, and increased the activity of CAT in all dosages, and affected as positive DNA ladder. Although the SOD activities of both hepatocytes and medium in 1.085 mg/L arsanilic acid were significantly lower than the base line at 12 h, CAT activity in 10.85 mg/L arsanilic acid was significantly higher than the base line at 48 h, and all of the DNA ladders were positive, which means 1.085 mg/L arsanilic acid induced apoptosis at 24 h. The gene expression of bax was significantly upregulated in 1.085 mg/L arsanilic acid or higher for 24 h.The parameters in 1,085 mg/L and 10,850 mg/L arsanilic acid had more severe changes than the others at any time indicating that these levels of arsanilic acid were toxic hazards for hepatocyte survival. It was concluded that arsanilic acid induced a dosage- and time-dependent gene expression of bax, 1.085 mg/L arsanilic acid could be involved in rat liver cell apoptosis at 24 h. Arsanilic acid as additives in livestock feed could present potential toxic implications for farm animals.

• Journal of Physiology & Pathology in Korean Medicine
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• v.27 no.5
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• pp.617-624
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• 2013

The purpose of this study is to investigate the protective effects of 25 herbal formulas on acetaminophen (APAP) or D-galactosamine (D-GalN)-induced hepatotoxicity in primary rat hepatocytes. Cell viability was measured using by Cell Counting Kit-8. 15 kinds of herbal formulas significantly reversed the cell viabilities of D-GalN-treated rat hepatocytes compared with D-GalN alone (p<0.05). In particular, 9 herbal formulas (Bangpungtongseong-san, Bojungikgi-tang, Galgeun-tang, Gumiganghwal-tang, Guibi-tang, Sagunja-tang, Samsoeum, Pyeongwi-san and Yijin-tang) showed the potent protective effects. However, 8 herbal formula exerted weak protective effects and 2 herbal formula did not exert effects on hepatotoxicity by D-GalN. On APAP-induced hepatotoxicity, 7 kinds of herbal formulas increased the viabilities of hepatocytes compare with APAP alone (p<0.05). These results could be provide a valuable information for the future in vivo or clinical studies to predict the hepatoprotective effects of herbal formulas.

• Korean Journal of Clinical Laboratory Science
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• v.39 no.3
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• pp.190-195
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• 2007

In the present study, the author investigated to the cytotoxocity in cultured rat hepatocytes of Viscum album lectin. The cytotoxcity effect in Viscum album lectin on the activity of LDH was also investigated. Viscum album lectin significantly increased LDH leakage into medium of hepatocytes treated or untreated with $CCl_4$ (p<0.001). However, Viscum album lectin significantly increased LDH leakage from $CCl_4$-induced hepatocyte (p<0.001). There was a significant increase in LDH levels relative to the control group. Histological observation basically supported the result obtained from LDH assay. The livers of rats challenged with $CCl_4$ produced a marked increased cytoplasmic vacuoles and inflammatory cells in number, while the number of necrotic cells and swollen hepatocytes did not change significnatly. Rats administered DMSO alone did not alter the normal hepatic architecture. Histological observation of liver section in rat treated 72 hrs with either Viscum album lectin $CCl_4$-induced liver damage showed number of cytoplasmic vaculoe and necrotic cell. The number of inflammatory cell increased markedly. This results suggest to the conclusion that Viscum album lectin has a effect of hepatotoxicity activator.

• 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.

• YAKHAK HOEJI
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• v.40 no.1
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• pp.52-58
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• 1996

Solvent fractions were prepared from traditional herbal drugs which of methanol extracts inhibited $CCl_4$-induced cytotoxicity in primary cultured rat hepatocytes and c ontinuously assayed their effects. Ethylacetate and n-buthanol fractions from Cibotii Rhizoma and chloroform fraction from Gelatina Nigra inhibited the release of LDH and GPT from $CCl_4$-treated hepatocytes, respectively. Water fraction (WAR) among solvent fractions from Astragali Radix showed the most potent inhibitory effect on the release of GOT or GPT by treatment with $CCl_4$. All of solvent fractions prepared from Eucommiae Cortex had no effect on $CCl_4$-induced cytotoxicity. Chloroform and ethylacetate fractions from Rehmanniae Radix Preparata increased the release of GPT from $CCl_4$-treated hepatocytes. n-Hexan, chloroform or ethylacetate fraction from 5 herbal drugs increased the release of LDH, GOT or GPT from normal hepatocytes at the dose of 1.Omg/ml. Administration of WAR suppressed the elevation of GOT, ALP activities and MDA contents in the serum as well as in the liver tissue of $CCl_4$-intoxicated rats. Based on these results, isolation of antihepatotoxic substances from WAR is under the process.

• Toxicological Research
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• v.7 no.2
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• pp.173-181
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• 1991

Membrane transport of brazilin was investigated i primary cultured rat hepatocytes. Brazilin was transported into hepatocytes very slowly and reached plateau at about 60 minutes. Saturation of transport process was not observed and the transport was not affected by ouabain, metabolic inhibitors, and its structural analog. The amount of brazilin transported into hepatocytes was decreased when the environmental temperature was decreased. These results suggest that brazilin might be transported into hepatocytes by simple diffusional process.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.2
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• pp.121-127
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• 2000

Intracellular accumulation of bile acids in the hepatocytes during cholestasis is thought to be pathogenic in cholestatic liver diseases. The objective of this study was to determine the role of lipid peroxidation and glutathione on the bile acid-induced hepatic cell death mechanism in primary cultured rat hepatocytes. To induce hepatic cell death, we incubated primary cultured rat hepatocytes with glycochenodeoxycholic acid $(GCDC;\;0{\sim}400\;{\mu}M)$ for 3 hours. In electron microscopic examination and agarose gel electrophoresis, low concentration of GCDC treatment mainly induced apoptotic feature. Whereas $400\;{\mu}M$ GCDC treated cells demonstrated both apoptosis and necrosis. Lipid peroxidation was increased dose-dependently in GCDC treated hepatocyte. And this was also accompanied by decreased glutathione. Therefore, oxygen free radical damage may play a partial role in GCDC-induced hepatic cell death.

• Journal of Pharmaceutical Investigation
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• v.25 no.4
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• pp.365-369
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• 1995

The acute cytotoxicities of chloroquine sulfate, propranolol, ascorbic acid, acetylsalicylic acid and acrylamide on cultured adult rat hepatocytes were evaluated by the use of LDH leakage and NR uptake test. On the basis of $IC_{50}$ values, the rank order of cytotoxicities of these drugs in both tests was chloroquine sulfate > propranolol > acetylsalicylic acid > ascorbic acid. The $IC_{50}$ of LDH test was very similar to that of NR uptake test. Thus, we concluded that both tests are reliable and sensitive methods in detecting toxicity in adult cultured rat hepatocytes.

• Journal of Embryo Transfer
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• v.31 no.3
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• pp.169-177
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• 2016

There is a growing interest in the application of primary hepatocytes for treatment of liver diseases in humans and for drug development. Several studies have focused on long-term survival and di-differentiation blocking of primary hepatocytes in an in vitro culture system. Therefore, the present study also aimed to optimize an in vitro culture system using primary rat hepatocytes. Primary rat hepatocytes from 6-week-old male Crl:CD rats were isolated using a modified two-step collagenase perfusion. Healthy $3.5{\times}10^6$ primary rat hepatocytes were seeded into a 2 dimensional (2D) culture in a 25T culture flask coated with collagen type I or into a 3D culture in a 125-ml spinner flask for 7 days. Production of plasma protein (ALB and TF), apoptosis (BAX and BCL2), and CYP (CYP3A1) related genes were compared between the 2D and 3D culture systems. The 3D culture system had an advantage over the 2D system because of the relatively high expression of ALB and low expression of BAX in the 3D system. However, the level of CYP3A1 did not improve in the 3D culture with and without the presence of a dexamethasone inducer. Therefore, 3D culture has an advantage for albumin production and primary rat hepatocyte survivability, but a low expression of CYP3A1 indicated that primary rat hepatocytes require a high-density culture for stress reduction by continuous flow.