The mouse submandibular gland (SMG) is an excellent model for the study of many important biological phenomena such as hormonal regulation of differentiation, neurotransmitter control of secretion, epithelial transport, exocytosis and endocytosis as well as the regulation of mouse SMG specific gene expression, in particular, NGF, EGF and renin. The postnatal development and sexual dimorphism of the mouse gland permits the isolation of male SMGs of different ages, corresponding to different stages of differentiation, particularly with respect to the cytodifferentiation of ductal cell types. We have immortalized SMG epithelial cell lines using mice transgenic for the large T antigen of SV40 or polyoma viruses. Epithelial clusters from the dissected glands were placed in culture and cell lines were established from the immortalized population. Two cell lines, SIMS and SIMP, which retain structural and functional characteristics, are described here. The cell lines are immortalised but not transformed, as judged by the absence of anchorage independent growth potential and the lack of tumour formation in athymic nude mice. Confocal and electron microscopy examination demonstrate that SIMP and SIMS cells express E-cadherin and ZO-1 and have features of polarised epithelial cells. In addition, they form spherical cysts with a wide lumen when grown in type I collagen gels. When grown on a filter support SIMS cells form a tight monolayer, exhibit vectorial transport function and show exclusive Na+, K(+)-ATPase localisation to the basolateral domain. We determined the cell type restricted expression of cytokeratin markers in the mouse SMG in vivo and we demonstrate that SIMS and SIMP cell lines express duct-specific cytokeratins. Finally, the expression of a set of differentiation markers, including EGF, NGF and renin, was detected by RT-PCR and by indirect immunofluorescence staining in these lines. Thus, these polarised ductal cell lines, as well as having important intrinsic properties, represent well characterised mouse epithelial models which, until now, have not been readily available for cellular studies.
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