Calcium induces differentiation of primary human salivary acinar cells

Cultivation of human parotid glands in serum-free medium (Ca2+ concentration, 0.2 mM) with growth supplements resulted in isolation of a homogeneous population of epithelial cells without any mesenchymal cells. The isolated cells showed an undifferentiated phenotype with scant cytoplasmic organelles and low levels of alpha-amylase expression. The cells remained viable and undifferentiated for up to 24 passages when subcultured at 80% confluence in 0.2 mM Ca2+ medium with a 1:3 split ratios. There was little cell-cell contact. A Ca2+ switch from 0.2 to 1 mM induced cell-cell contact with translocation of desmosomal proteins from the cytoplasm to the cell membrane, and sequential differentiation of serous acinar cells with a glandular arrangement, well-developed cytoplasmic organelles, and an increased level of a-amylase expression. These morphological changes and desmosome assembly were blocked by treatment with non-specific PKC inhibitor. Moreover, the addition of PKC activator, tetradecanoylphorbol 13-acetate (TPA), to 0.2 mM Ca2+ medium caused transient assembly of desmosome-like structure, but did not induce cell-cell contact or morphological differentiation. Cultivation of the cells in 1.5 mM Ca2+ medium resulted in increased stratification of the cells and reduced a-amylase expression. These findings provide the first demonstration that continuous cultivation in 1.0 mM Ca2+ medium is required for cellular differentiation of salivary gland acinar cells, and maintenance of the differentiated state. (C) 2002 Wiley-Liss, Inc.