• Journal of Yeungnam Medical Science
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• v.15 no.2
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• pp.286-296
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• 1998

Different techniques for culturing respiratory epithelial cells have been developed to overcome the limitations of studies on in vivo and on bioptic material. Traditionally, culture systems are divided into organ cultures, explant cultures and dissociated cell cultures. The first two contain both epithelial and non-epithelial cells. However, in monolayer cultures of dissociated cells only epithelial cells are present, the effects observed are caused by a pure epithelial responses. The purpose of this study is to establish primary culture method of human nasal epithelium (HNEC) by monolayer culture of dissociated cells to evaluate the role of the epithelial cells in the allergic and non-allergic nasal inflammatory reactions. HNEC was prepared by primary culture method of monolayer culture of dissociated cells from human inferior nasal turbinate mucosa of septal deviation patients. Primary cultured cells were characterized by indirect immunofluorescence assay and transmission electron microscopy. The immunoreactivities of cytokeratin-pan and cytokeratin No. 8 were observed in cultured HNEC. However, the immnoreactivities of vimentin and von Willebrand factor were not observed in cultured HNEC. The tonofilaments and desmosome were observed in cultured HNEC. The cultured epithelial cells were identified to be pure nasal epithelial cells. The monolayer culture of dissociated cells could successfully be employed for further study to investigate the role of the epithelial cells in allergic or non-allergic nasal inflammatory diseases.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.6
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• pp.777-784
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• 2001

Evaluation of the granulosa cell (GC) monolayers developed from small (<5 mm) and large (>5 mm) follicles on the meiotic competence of caprine oocytes during in vitro maturation was done in this study in comparison to the granulosa cell coculture. Ovaries were collected from the local abattoir and follicular contents were aspirated for the monolayer culture. For IVM the oocytes were collected by puncturing the nonatretic follicles (>4 mm). Results revealed that at the same seeding rate, small follicular granulosa cell monolayer achieved confluence 24-48 h earlier than large follicular granulosa cell monolayer. GC monolayers significantly p (<0.05) improved the rate of meiotic resumption and nuclear maturation (84.76% vs 74.74%) after 27 h of culture in comparison to GC coculture. Statistically there was no significant difference in the maturation rate between the caprine oocytes matured over small or large follicular GC monolayers. It is concluded from the present study that GC monolayers support better nuclear and cytoplasmic maturation of growing caprine oocytes which is evident by better maturation rate over GC monolayer as compared to the oocytes matured with GC coculture. Granulosa cells from small and large follicles can be used for IVM with more or less in the same efficiency after conditioning them with maturation media in 18-24 h before the onset of culture.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.39 no.1
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• pp.23-26
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• 2006

Effects of sera from several fish species and insulin on the development of cultured Silurus asotus hepatocytes were investigated. Hepatocytes with high viability (95%) were obtained from the livers of male catfish by two step collagenase perfusion. Isolated hepatocytes, initially showed a typical round-shape, firmly attached to the culture dish within 24 h. In the presence of catfish serum, hepatocytes attached each other, spread well on the dish and developed into monolayer after 3-4 days of incubation. Cells within the established monolayer became polygonal in shape and their nuclei and boundaries being clearly visible under the microscope. In contrast, when incubated in FBS-supplemented or serum-free medium, cells managed to form small clusters, each made of 2-10 cells. Cells in FBS-supplemented medium further developed into larger clusters. However, these clusters failed to develope into monolayer. In addition, when insulin was deprived from culture medium, formation of monolayer also failed. From these data, it can be concluded that the presence of both catfish serum and insulin is necessary for the formation of monolayer of catfish hepatocytes and the functional role of fish serum may differ from that of insulin and can not be displaced by FBS-supplementation.

• Development and Reproduction
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• v.13 no.4
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• pp.217-226
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• 2009

Effects of sera from several fish species on monolayer formation, viability and functions of catfish hepatocytes were investigated to establish a primary hepatocyte culture system for screening endocrine disruptors. Hepatocytes of Korean catfish (Silurus asotus) were attached and formed monolayer using the media supplemented with their own serum or sera from eel and tilapia, but not with fetal bovine serum (FBS). The amount of fish sera (0.5~3%) for monolayer culture of the catfish hepatocytes was less than 1/10 of FBS (5~20%) that is commonly used for primary culture of hepatocytes of other species. The results indicate that FBS can be replaced with sera from some fish species and the fish sera are more effective than FBS in maintaining the shape and functions of the hepatocytes. The primary culture of catfish hepatocytes was maintained monolayer with fish sera for at least 10 days, which makes possible to be used for screening the activities of endocrine disruptors. In conclusion, the primary culture system of hepatocytes with fish sera in the present study could be a useful tool for screening and studying endocrine disruptors.

• Reproductive and Developmental Biology
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• v.29 no.4
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• pp.241-245
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• 2005

Porcine oviduct epithelial cells (POEC) are widely used in co-culture experiments to improve early embryonic development, in vitro fertilization in embryo transfer programs for domestic animals and in vitro maturation of immature germ cells. POEC were mechanically isolated and cultured in tissue culture medium 199. Cells grew continuously, and confluent monolayers were formed after 7 days. After forming confluent monolayer of epithelial cells, supernatant was collected as the condition medium for maturing round spermatids in vitro. Round spermatids were also separated mechanically and cultured in the POEC condition medium. In this study we observed that $20\%$ of round spermatid cultured were matured into elongating spermatid after 24 h, and about $10\%$ of round spermatid cultured showed complete elongation (elongated spermatid) within $24\~48$ h of in vitro culture. No further development was observed within $50\~72$ h and transformed cells lost their viability after 72 h. These preliminary findings suggest that the condition medium from POEC may be possible to overcome the round spermatid block by improving the milieu of culture system.

• Fisheries and Aquatic Sciences
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• v.19 no.2
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• pp.9.1-9.7
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• 2016

This study was conducted to identify the general conditions for the isolation and in vitro culture of ovary-derived cells in rockfish (Sebastes schlegeli). The effects of three different enzymes on cell retrieval from ovarian tissues were evaluated first, and then the ovary-dissociated cells were cultured under various culture conditions, with varying basal media and culture temperatures, addition of growth factors, and/or culture types. We found that collagenase type I treatment was effective for cell isolation from ovarian tissues. From a total of 42 trials to evaluate the effects of basal media and culture temperatures on cell culture of ovary-dissociated cells, we observed that Leibovitz's L15 medium was more supportive than Dulbecco's modified Eagle's medium for culture, and the cells could grow at all three temperatures tested, 15, 20, and $25^{\circ}C$, at least up to passage 2. However, growth factor addition did not improve cell growth. Introduction of suspension culture after monolayer culture expanded the culture period significantly more than did monolayer culture alone. Our results may provide a basis for developing an in vitro system for S. schlegeli germline cell culture, which will ultimately lead to improvement of the species.

• Journal of Microbiology and Biotechnology
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• v.15 no.6
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• pp.1299-1303
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• 2005

Human articular chondrocytes (HAC) were cultivated as a monolayer in a serum-free medium for primary culture (SFM-P). An optimized SFM-P provides $95\%$ proliferation rate of that obtainable from primary and secondary chondrocyte cultures grown in a control medium with serum. The gradual decrease in the amounts of synthesized glycosaminoglycan and type II collagen was improved by coating the culture dishes with type IV collagen and fibronectin. A significant improvement in the expression of type II collagen and aggrecan mRNA could be achieved. In addition, the monolayer cultures showed better synthesis of the extracellular matrices than alginate-bead cultures in SFM-P.

• Journal of Microbiology and Biotechnology
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• v.17 no.5
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• pp.739-744
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• 2007

Insect cell lines and the control of infection for obtaining the maximum amount of polyhedrin-Cry1Ac-polyhedrin fusion protein from Bactrus in monolayer and suspension culture systems were tested. Growth rates of the Trichoplusia ni(High-Five) cell line in both culture systems were better than the other insect cell lines, Spodoptera frugiferda(Sf-9, Sf-21), Trichoplusia ni(Tn5), and Spodoptera exigua(Se301). The expression of the fusion protein in a monolayer culture showed that Se301 cells were 2.3-4.8 times more productive on a per cell basis than the other cell lines. However, in suspension culture, only High-Five cells were productive. High-Five cells infected with Bactrus at a multiplicity of infection(MOI) of 5 and a cell density of $3.0{\times}10^5$ cells per ml were more productive than the other infection condition in a suspension culture suitable for a large-scale production of baculovirus. In conclusion, for the large-scale production of Bactrus in vitro, High-Five cells showing good growth and high productivity are suitable.

• Journal of the Korean Society of Food Science and Nutrition
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• v.26 no.6
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• pp.1173-1180
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• 1997

Hepatotoxicity of caffeine and acetaminophen was investigated in this study. Special attention was paid to the effect of vitamins on the reduction of hepatotoxicity caused by the chemicals. Rat hepaocytes isolated by two-step perfusion method were cultured in two differents methods-suspension, monolayer cultures-, and exposed to caffeine and/or acetaminophen for 24hrs. Caffeine or acetaminophen exhibited no significant hepatotoxicity in terms of intracellular glutathione(GSH) level and lipid peroxidation(MDA), but GSH level was significantly decreased after administrated acetaminophen, and the toxicity caused by the chemicals showed a dose-dependent manner. The synergistic effect of caffeine and acetaminophen was observed when both caffeine and acetaminophen were supplemented to culture medium. At the concentration 1mM, caffeine enhanced the intracellular GSH depletion and MDA formation by 63% and 64%, respectively, compared to single supplementation of 10mM acetaminophen in culture medium. This hepatotoxicity induced membrane integrity loss was observed by lightmicroscope on the simultaneous administration of caffeine and acetaminophen in monolayer cultured hepatocytes. Co-supplementation of vitamins with caffeine/acetaminophen to culture medium results in the protection of hepatocytes from hepatotoxic attach by caffeine/acetaminophen. Especially, vitamin E was superior to vitamin C and $\beta$-carotene from the standpoints of GSH depletion and MDA formation. From this results, it has been speculated that vitamin E may play a role of antioxidant scavenging radicals produced from acetaminophen. Taken all together, in vitro culture system like monolayer culture of hepatocytes may be a useful tool for the evaluation of hepatotoxicity or protection ability of food ingredients.

• KSBB Journal
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• v.7 no.4
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• pp.278-283
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• 1992

Rat hepatocytes were isolated and cultured on the petri dishes treated with various coating materials. Untreated or collagen coated petri dish gave monolayer culture of hepatocyte and proteoglycan, dermatan sulfate, and BSA treated petri dish gave hemispheroid. The untreated Primaria petri dish gave spheroid type of hepatocyte, and heparin and hyaluronic acid treatment gave multilayers. To sustain high cell viability, monolayer cultured hepatocytes was more useful, while it was found that the hemispheroid or spheroid type hepatocytes was more active in the hepatic functions such as ammonia metabolism and albumin synthesis.