• Asian-Australasian Journal of Animal Sciences
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• 제12권8호
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• pp.1258-1262
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• 1999

With an attempt to elucidating the lipid metabolism of Tsaiya ducks, thirty ducks at growing (8 weeks of age) and laying periods (10 weeks after the onset of laying) were examined, respectively. The ducks were randomly allocated into ad libitum feeding and 3-day fasting groups, to investigate their in vitro hepatocytes lipogenesis capacity and adipocytes lipolysis rate. Results indicate that (1) the capacity of hepatocytes incorporation of glucose and acetate into total lipid and metabolite of $^{14}CO_2$ production during the laying period was greater than during the growing period. Approximately 50% of the glucose or acetate converted into triacylglycerol (TG) by the hepatocytes were recovered as fatty acid during the growing period, while it was 65-70% during the laying period. (2) Acetate used for lipogenesis ability was superior to glucose in both periods. (3) The adipocytes lipolysis rate was increased significantly (p<0.05) by fasting. In contrast, the capacity of incorporated glucose or acetate into total lipid, triacylglycerol, fatty acid and glycerol by hepatocytes was reduced significantly (p<0.05) by fasting.

• Toxicological Research
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• 제3권2호
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• pp.73-80
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• 1987

Primary cultures of adult rat hepatocytes were used to study the cytotoxicity of D-galactosamine. Hepatocytes were isolated by a collagenase perfusion technique and maintained as monolayers in serum-free medium on collagen-coated culture dishes. Treatment of galactosamine to the culture markedly inhibited the uptake of ${\alpha}$-aminoisobutyric acid (AIB) inducible with glucagon and dexamethasone. At0.1 mM of galactosamine, AIB uptake was inhibited significantly when treated for 12 hr. At higher doses (0.25, 0.5 and 1.0mM), a significant inhibition was noticed after 1 hr exposure. Generally the magnitude of the inhibition was related to the dose and treatment time of galactosamine. Treatment of galactosamine also produced a dose- and treatment time-related suppression of the tyrosine aminotransferase (TAT) induction caused by dexamethasone. Meanwhile, uptake of ouabain was not affected by the treatment of galactosamine. The viability of the hepatocytes was decreased only slightly by the treatment of galactosamine; more than 87% of the cells excluded tryphane blue when treated 1 mM galactosamine for 12 hr. Galactosamine induced depressions of AIB uptake and TAT activity were prevented by the simultaneous addition of uridine to the culture. D-Galactosamine, cytotoxicity, hepatocytes culture, ${\alpha}$-aminoisobutyric acid uptake, tyrosine aminotransferase.

• Toxicological Research
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• 제11권2호
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• pp.199-203
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• 1995

The focus of this study was to investigate that cellular parameters and glucose uptake might be altered by extracellular calcium and starvation. Addition of 1 mM $Ca^{++}$ to hepatocytes (equalling to the free calcium concentration of blood) significantly increased intracellular $Na^+$ and decreased $Na^+$ & LDH leakage. This pertains to the hepatocytes of control rats as well as those of rats fasted for 24 and 48. hr. These effects might be come from the membrane-stabilizing effects of calcium. But calcium had no effects on cell volumes, superoxide-formation and glucose uptake. Actually hepatocytes of starved rats showed changes in several cellular parameters. Starvation increased LDH leakage, glucose uptake and the total concentration of $Na^+$ and $Na^+$ whereas it markedly decreased cell volumes. Since total tonicity remained unchanged, intracellular $Na^+$ and $Na^+$ could contribute to a higher share of total osmolarity in starvation. Starvation increased the cytoplasmic pH because $R-NH^{3+}$ions and their corresponding counterions disappeared. This increase may be related to suppress the protonization of amino groups in proteins. Starvation decreased hepatic glycogen, a major compound that affects cytosolic volume of hepatocytes. The data indicate that starvation increases the glucose transport activity. The possible molecular basis will be discussed.

• 한국환경과학회:학술대회논문집
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• 한국환경과학회 2003년도 International Symposium on Clean Environment
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• pp.159-164
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• 2003

Effects of Al on vitellogenin (VTG) and VTG mRNA induction by estradiol-17 $\beta$($E_2$) were examined in primary hepatocyte culture of rainbow trout. Hepatocytes were precultured for 2 days and then E2 ($2{\times}10^{-6}$M) and Al ($10^{-6}-10^{-4}$M) were simultaneously added to the incubation medium. Hepatocytes were cultured for 5 more days. Media and hepatocytes were then analyzed by SDS-PAGE and Northern blotting for VTG and VTG mRNA, respectively. These metal had no appreciable effect on the viability of hepatocytes in culture. However, Al interfered with VTG production and VTG mRNA expression. Al reduced VTG production in a concentration-dependent way and a significant reduction accurred at Al concentrations greater than $5{\times}10^{-5}$M. VTG mRNA expression also decreased with a negative correlation with Al concentration (r＝-0.98). These results suggest that Al inhibit VTG production at the transcriptional level to reduce VTG mRNA expression.

• KSBB Journal
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• 제19권3호
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• pp.210-214
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• 2004

본 연구에서는 생분해성 고분자 담체인 PGA 담체에 부착된 간세포의 이식을 통해서 이식된 간세포가 괴사하지 않고 남아 있으며 간 조직 구조의 일종인 담세관 유사구조를 확인하였다. 조직공학적인 간세포 이식 방법의 개발은 간 질환에 새로운 치료방법 개발의 가능성을 열어줄 수 있다.

• 약학회지
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• 제40권1호
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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.

• Animal Bioscience
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• 제34권7호
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• pp.1210-1220
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• 2021

Objective: Unlike mammals, goose fatty liver shows a strong tolerance to fatty acids without obvious injury. Stearyl-coenzyme A desaturase 1 (SCD1) serves crucial role in desaturation of saturated fatty acids (SAFs), but its role in the SAFs tolerance of goose hepatocytes has not been reported. This study was conducted to explore the role of SCD1 in regulating palmitic acid (PA) tolerance of goose primary hepatocytes. Methods: 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide was examined to reflect the effect of PA on hepatocytes viability, and quantitative polymerase chain reaction was used to detect the mRNA levels of several genes related to endoplasmic reticulum (ER) stress, inflammation, and apoptosis, and the role of SCD1 in PA tolerance of goose hepatocytes was explored using RNA interfere. Results: Our results indicated that goose hepatocytes exhibited a higher tolerant capacity to PA than human hepatic cell line (LO2 cells). In goose primary hepatocytes, the mRNA levels of fatty acid desaturation-related genes (SCD1 and fatty acid desaturase 2) and fatty acid elongate enzyme-related gene (elongase of very long chain fatty acids 6) were significantly upregulated with 0.6 mM PA treatment. However, in LO2 cells, expression of ER stress-related genes (x box-binding protein, binding immunoglobulin protein, and activating transcription factor 6), inflammatory response-related genes (interleukin-6 [IL-6], interleukin-1β [IL-1β], and interferon-γ) and apoptosis-related genes (bcl-2-associated X protein, b-cell lymphoma 2, Caspase-3, and Caspase-9) was significantly enhanced with 0.6 mM PA treatment. Additionally, small interfering RNA (siRNA) mediated downregulation of SCD1 significantly reduced the PA tolerance of goose primary hepatocytes under the treatment of 0.6 mM PA; meanwhile, the mRNA levels of inflammatory-related genes (IL-6 and IL-1β) and several key genes involved in the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT), forkhead box O1 (FoxO1), mammalian target of rapamycin and AMPK pathways (AKT1, AKT2, FoxO1, and sirtuin 1), as well as the protein expression of cytochrome C and the apoptosis rate were upregulated. Conclusion: In conclusion, our data suggested that SCD1 was involved in enhancing the PA tolerance of goose primary hepatocytes by regulating inflammation- and apoptosis-related genes expression.

• Animal Bioscience
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• 제35권11호
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• pp.1787-1799
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• 2022

Objective: Choline deficiency, one main trigger for nonalcoholic fatty liver disease (NAFLD), is closely related to lipid metabolism disorder. Previous study in a choline-deficient model has largely focused on gene expression rather than gene structure, especially sparse are studies regarding to alternative splicing (AS). In modern life science research, primary hepatocytes culture technology facilitates such studies, which can accurately imitate liver activity in vitro and show unique superiority. Whereas limitations to traditional hepatocytes culture technology exist in terms of efficiency and operability. This study pursued an optimization culture method for duck primary hepatocytes to explore AS in choline-deficient model. Methods: We performed an optimization culture method for duck primary hepatocytes with multi-step digestion procedure from Pekin duck embryos. Subsequently a NAFLD model was constructed with choline-free medium. RNA-seq and further analysis by rMATS were performed to identify AS events alterations in choline-deficency duck primary hepatocytes. Results: The results showed E13 (embryonic day 13) to E15 is suitable to obtain hepatocytes, and the viability reached over 95% by trypan blue exclusion assay. Primary hepatocyte retained their biological function as well identified by Periodic Acid-Schiff staining method and Glucose-6-phosphate dehydrogenase activity assay, respectively. Meanwhile, genes of alb and afp and specific protein of albumin were detected to verify cultured hepatocytes. Immunofluorescence was used to evaluate purity of hepatocytes, presenting up to 90%. On this base, choline-deficient model was constructed and displayed significantly increase of intracellular triglyceride and cholesterol as reported previously. Intriguingly, our data suggested that AS events in choline-deficient model were implicated in pivotal biological processes as an aberrant transcriptional regulator, of which 16 genes were involved in lipid metabolism and highly enriched in glycerophospholipid metabolism. Conclusion: An effective and rapid protocol for obtaining duck primary hepatocytes was established, by which our findings manifested choline deficiency could induce the accumulation of lipid and result in aberrant AS events in hepatocytes, providing a novel insight into various AS in the metabolism role of choline.

• Asian-Australasian Journal of Animal Sciences
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• 제19권9호
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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.

• Animal Bioscience
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• 제36권11호
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• pp.1693-1699
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• 2023

Objective: Cows that are nursing get around 80% of their glucose from liver gluconeogenesis. Propionate, a significant precursor of liver gluconeogenesis, can regulate the key genes involved in hepatic gluconeogenesis expression, but its precise effects on the activity of enzymes have not yet been fully elucidated. Therefore, the aim of this study was to investigate the effects of propionate on the activity, gene expression, and protein abundance of the key enzymes involved in the gluconeogenesis of dairy cow hepatocytes. Methods: The hepatocytes were cultured and treated with various concentrations of sodium propionate (0, 1.25, 2.50, 3.75, and 5.00 mM) for 12 h. Glucose content in the culture media was determined by an enzymatic coloring method. The activities of gluconeogenesis related enzymes were determined by enzyme linked immunosorbent assay kits, and the levels of gene expression and protein abundance of the enzymes were detected by real-time quantitative polymerase chain reaction and Western blot, respectively. Results: Propionate supplementation considerably increased the amount of glucose in the culture medium compared to the control (p<0.05); while there was no discernible difference among the various treatment concentrations (p>0.05). The activities of cytoplasmic phosphoenolpyruvate carboxylase (PEPCK1), mitochondrial phosphoenolpyruvate carboxylase (PEPCK2), pyruvate carboxylase (PC), and glucose-6-phosphatase (G6PC) were increased with the addition of 2.50 and 3.75 mM propionate; the gene expressions and protein abundances of PEPCK1, PEPCK2, PC, and G6PC were increased by 3.75 mM propionate addition. Conclusion: Propionate encouraged glucose synthesis in bovine hepatocytes, and 3.75 mM propionate directly increased the activities, gene expressions and protein abundances of PC, PEPCK1, PEPCK2, and G6PC in bovine hepatocytes, providing a theoretical basis of propionate-regulating gluconeogenesis in bovine hepatocytes.