Isolation and in vitro culture of primary cell populations derived from ovarian tissues of the rockfish, Sebastes schlegeli

Abstract

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°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.

Isolation, in vitro culture, and characterization of black crappie, Pomoxis nigromaculatus and white crappie, P. annularis ovarian tissue primary cells

“…Cell growth was observed at all temperatures tested. Similar results were observed by Ryu et al (2016) when ovarian cells were dissociated and cultured from rockfish, Sebastes schlegeli . Ryu et al (2016) grew cells at 15°C, 20°C, and 25°C.…”

“…Similar results were observed by Ryu et al (2016) when ovarian cells were dissociated and cultured from rockfish, Sebastes schlegeli . Ryu et al (2016) grew cells at 15°C, 20°C, and 25°C. We found that primary cells consisted of a heterogenous population of epithelial and fibroblast‐like cells.…”

“…Ideally, it would be advantageous to reduce the supplementation of basal media with serum, as it is an animal-derived factor and an expensive cell culture product. Ryu et al (2016) have demonstrated that Leibovitz's L-15 medium is more supportive than Dulbecco's modified Eagles' medium for the culture of ovary dissociated cells. Therefore, two concentrations (10% and 20%) of FBS in L-15 media were evaluated to determine the optimum concentration for cell growth.…”

“…Three replicates were maintained for each treatment. Three days after seeding, each cell-cultured flask was washed twice with Ca 2+ /Mg 2+ Àfree Dulbecco's PBS (DPBS) (Gibco) to refresh the culture media (Ryu et al, 2016). Primary cell attachment and cell morphology were observed under a microscope, and images were taken before changing the culture medium.…”

Effects of Temperatures and Basal Media on Primary Culture of the Blastomeres Derived from the Embryos at Blastula Stage in Marine Medaka Oryzias dancena

“…But, in some cases like testis tissues, they are cultured in less than 37°C based on the anatomic difference (Gohbara et al, 2010). In contrast to mammalian species, however, fish can acclimate to a wide range of temperature as a poikilothermic vertebrate and thus the cultured cells from fish can also tolerate a wide range of temperature (Lannan et al, 1984;Qin et al, 2006;Ryu et al, 2016). However, it has been reported that the alteration of temperature in culture of fish cells is able to influence the physiology of the cells (Lannan et al, 1984).…”