Growth and differentiation of primary tracheal epithelial cells in culture: Regulation by extracellular calcium

Abstract

Growth and differentiation of primary monkey tracheal epithelial (MTE) cells maintained on collagen gel substrata were studied in a defined serum-free culture medium containing 0.03 to 3.0 mM extracellular calcium. Cell attachment efficiency (40-60%) was not altered by different calcium levels. Growth of primary MTE cells on collagen gel substrata, which was vitamin A dependent, was enhanced 50% in the medium supplemented with high calcium (greater than 0.3 mM). High calcium medium also increased cell-cell interactions, formation of desmosomes, and multi-cell layering. The relative content of mucous cells, which were identified by a mucin-specific monoclonal antibody and the presence of mucus-secreting granules at the ultrastructural level, was greater in the high-calcium medium. Furthermore, the secretion of mucin into the medium, determined either by an ELISA or by the incorporation of 3H-glucosamine into mucous glycoprotein fractions, was also increased more than 5-fold in media containing high calcium content (greater than 0.6 mM). In contrast, MTE cells cultured in low calcium medium (less than 0.15 mM) were squamous-like with prominent tonofilaments, and their secretory product was mainly hyaluronate. These results demonstrate that media containing a high calcium content promote conducting airway epithelium to express mucous cell differentiation, while media with low calcium content promote squamous cell differentiation.

Growth and differentiation of conducting airway epithelial cells in culture

“…We observed that the expression of mucous cell function is dependent on the concentration of calcium in the medium. These studies demonstrate that >0.3 mM calcium in culture medium is needed to achieve mucous cell differentiation [24].…”

“…Furthermore, the quantitative kinetics and the time-course studies suggest that the mucous cell population in the hamster culture is derived from the undifferentiated cell type. Evidence of in vitro mucous cell differentiation has now been extended to cultures derived from monkey [23,24], human [6], rabbit [25], and guinea pig [26] airway tissues. Recently, the development of mucinspecific monoclonal antibodies allow us to use both the enzyme-linked immunosorbent assay (ELISA) [16] and immunohistochemical methods [6] to quantify mucin production and mucous cell population, respectively, in culture.…”

Modelling the Human Respiratory System: Approaches forin VitroSafety Testing and Drug Discovery

“…72 This allowed the development of ALI cultures originally with guinea pig tracheal epithelial cells. 118 ALI cultures were rapidly adopted for use with primate, 119 human, 120,121 rat, 122 bovine 123 and canine 124 tracheal epithelial cells.…”

Asthma: a comparison of animal models using stereological methods

“…The role of vitamin A in regulation of epithelial differentiation has been established for: mucin genes [47]; cytokeratins [48], markers of squamous cell differentiation [39]; small proline-rich protein [49][50][51]; an antioxidant, thioredoxin [52]; the cytokine, interleukin (IL)-8 [53]; epithelial growth factor [54]; and heterogeneous nuclear protein AI [55]. Extracellular calcium also plays a role in epithelial differentiation in vitro [56].…”