Establishment of immortalized mesenchymal stem cells derived from the submandibular glands of tdTomato transgenic mice

Transgenic mice that overexpress the red fluorescent protein tdTomato (tdTomato mice) are well suited for use in regenerative medicine studies. Cultured cells from this murine model exhibit strong red fluorescence, enabling real-time in vivo imaging through the body surface of grafted animals. Mesenchymal stem cells (MSCs) have marked potential for use in cell therapy and regenerative medicine; however, the mechanisms that regulate their dynamics in vivo are poorly understood. In the present study, an MSC line was derived from the submandibular gland fibroblasts of tdTomato mice. The fluorescent signal from this cell line was observed in organs throughout the body, as well as in salivary glands. Primary culture cells derived from the submandibular gland were immortalized with SV40 large T antigen (GManSV cells); these cells exhibited increased migratory ability, as compared with those isolated from the sublingual gland. GManSV cells were tdTomato-positive and exhibited spindle-shaped fibroblastic morphology; they also robustly expressed mouse MSC markers: Stem cell antigen-1 (Sca-1), CD44, and CD90. This cell line retained multipotent stem cell characteristics, as evidenced by its ability to differentiate into both osteogenic and adipogenic lineages. These results indicate that Sca-1+/CD44+/CD90+-GManSV cells may be useful for kinetic studies of submandibular gland-derived MSCs in the context of in vitro co-culture with other types of salivary gland-derived cells. These cells may also be used for in vivo imaging studies, in order to identify novel cell therapy and regenerative medicine for the treatment of salivary gland diseases.