감성돔(Acanthopagrus schlegelii) 근육 세포주 확립에 근육 조직의 저온 보관이 미치는 영향 평가

Abstract

The establishment of fish muscle cell lines is critical for the development of cellular aquaculture and biotechnology. However, immediate processing of fresh tissue after sampling remains a limitation in practical scenarios. In this study, we evaluated the effect of cold storage duration on the isolation and in vitro cultivation of skeletal muscle cells from black sea bream (Acanthopagrus schlegelii). Muscle tissues were stored at 4℃ for 0, 2, 5, 10, 20, and 30 days before initiating primary culture through explant or dispersed culture methods, with the dispersed method not applied to the 30-day group. The potential for cell line establishment depending on the duration of cold storage was evaluated, and subsequently the established cell lines were characterized for their growth and differentiation at early (passage 5) and late (passage 20) stages of culture. To further investigate the relationship between cold storage and successful cell line establishment, the effects of cold storage on cell yield and viability after tissue dissociation, as well as changes in histology and ultrastructure, were assessed at the tissue level. Culture results demonstrated that cell lines could be established from tissues cold-stored up to 10 days via explant culture, and up to 5 days via dispersed culture. All cell lines established through dispersed culture method exhibited the ability to differentiate into myotubes. Tissue level analysis revealed that cold storage did not significantly affect cell viability for up to 5 days, maintaining levels above 80%. However, the number of isolated cells decreased with prolonged storage. Histological analysis revealed gradual expansion of interstitial spaces and increased tissue disorganization, particularly after 5–10 days. Transmission electron microscopy also showed progressive ultrastructural damage, with an increasing number of vacuolated structures observed in a time-dependent manner. These findings suggest that short-term cold storage (≤5 days) may preserve the potential for muscle cell line establishment, though tissue quality progressively deteriorates with longer storage durations.