Characterization of cultivated adult lacrimal gland epithelial cells

1. Tetsuya Kawakita
2. Shinya Kobayashi
3. Motoko Kawashima
4. Naoko Okada
5. Kazuo Tsubota

Department of Ophthalmology, Keio University School of Medicine, Tokyo, Japan

Objective: Previous report showed that mouse lacrimal gland epithelial cells were cultured successfully only in case of newborn mouse lacrimal gland. This study was performed to cultivate and characterize adult mouse lacrimal gland epithelial cells, and investigate regeneration potential of these cells by sphere culture method.

Materials and Methods: Lacrimal glands were removed from newborn mice (C57B/6) and isolated lacrimal gland epithelial cells were seeded on a tissue culture-treated or non-coating dish in Cnt-07 culture medium with or without cholera toxin. Cultivated cells were characterized by pan-cytokeratin, α-smooth muscle actin, and lactoferrin immunostaining. Lacrimal gland cells from 7-weeks old GFP and non-GFP (C57B/6) mice were seeded on non-coating dish or on 3T3 feeder layer for colony forming potential. Sphere which was generated from newborn lacrimal gland epithelial cells were embedded in collagen and injected into lacrimal gland recession mouse model to evaluate their regeneration potential.

Results: The lacrimal gland epithelial cells of newborn mice could be selected by cholera toxin, which was confirmed by pan-cytokeratin (+) and α-smooth muscle actin (-). With this medium and low adherent culture dish, adult mice generated sphere formation from single cells. Such cells could also form colony formation on 3T3 feeder cells. Spheres generated from new born lacrimal gland could be transplanted into injured lacrimal gland, and tended to recover corneal injured score, but not secretory function.

Conclusion: Adult mouse lacrimal gland epithelial cells were successfully cultivated and these cells could generate the sphere formation. Such spheres could be transplantable to injured lacrimal gland.