Establishment of a Skin Cell Line from Red Tilapia (Oreochromis spp.) and Its Potential Applications in Hyperosmotic Stress and Immune Response Research.

In this study, a skin-derived cell line, designated RTS, was established from red tilapia (Oreochromis spp.) using the tissue explant culture method. Cell outgrowth from skin explants was first observed on day 6 of primary culture, and the cells could be continuously subcultured with stable adherent growth. To date, the cell line has been maintained for more than 88 passages in vitro. Growth curve analysis demonstrated that RTS cells proliferated in DMEM, MEM, and L-15 media supplemented with 15% fetal bovine serum, with DMEM exhibiting the most favorable support for cell proliferation. Chromosome analysis showed that the diploid chromosome number of RTS cells was 2n = 44. Molecular identification based on a partial mitochondrial COI fragment indicated that RTS belonged to the genus Oreochromis and was phylogenetically closest to O. mossambicus. In addition, RTS cells were amenable to plasmid transfection, with a transient transfection efficiency of 35.2 ± 3.8% using pEGFP-N3. In response to Streptococcus agalactiae challenge, immune-related genes in RTS cells including MyD88, NF-κB, NOD2, TLR2, and IL-2 showed dynamic transcriptional changes. Under hyperosmotic treatment (532 ± 13 mOsm/kg), ostf1, HSP70, NKCC1, and nka exhibited distinct temporal expression patterns, indicating osmotic stress-responsive transcriptional changes. These results demonstrate that RTS is a stable and manipulable red tilapia skin-derived cell line and may serve as a useful in vitro model for studies of immunity and osmotic stress in red tilapia.