Ceacam6 (carcinoembryonic antigen cell adhesion molecule) is a GPI-anchored membrane glycoprotein recently identified in human fetal lung undergoing in vitro hormone-induced differentiation of type II cells. The protein is a tumor biomarker in adult lung and selected other tissues, where it has antimicrobial activity; however, expression and function in lung have not been characterized. Undifferentiated epithelial cells were isolated by collagenase/trypsin digestion from human fetal lung (16-21 wk gestation) and cultured 1-5 d in the absence (control) or presence of DCI (dexamethasone, 10 nM; 8-Br-cAMP, 0.1 mM; isobutylmethylxanthine, 0.1 mM). Some cells in control media were treated with adenovirus Ad12A2 (0.5-6 pfu/cell) to overexpress thyroid transcription factor-1 (TTF-1) or were transfected by electroporation (AMAXA nucleofection technology) with siRNAs (200 nM) for TTF-1 or Ceacam6 to knockdown expression in the presence of DCI. CEACAM6 mRNA and protein were up-regulated ≈10-fold by exposure of fetal lung epithelial cells to either DCI or to a dose of Ad12A2 that produced a level of recombinant TTF-1 comparable with endogenous TTF-1 in DCI-treated cells. By confocal immunofluorescence, CEACAM6 localized to both intracellular vesicles, costaining with SP-B (ie, lamellar bodies), and to the plasma membrane of type II cells. Knockdown of TTF-1 (46 ± 4% of control) in DCI-treated cells reduced CEACAM6 mRNA and protein by 80% (n = 4). Knockdown of CEACAM6 expression by siRNA (88 ± 3%, n = 3) did not alter expression or staining patterns of TTF-1, SP-B or DC-LAMP (a lamellar body membrane protein). Secretion rates for CEACAM6 were similar for both DCI- and Ad12A2-treated cells, with no response to treatment with secretagogues that stimulate surfactant secretion. Treatment of cells with mannosamine, an inhibitor of GPI anchor synthesis, blocked ≈50% of CEACAM6 secretion, consistent with GPI-anchor involvement in the secretory mechanism. We conclude that CEACAM6 expression in primary fetal lung epithelial cells is regulated by glucocorticoid and cAMP and is dependent on TTF-1. CEACAM6 is associated with intracellular surfactant but is secreted primarily by a constitutive pathway independent of lamellar bodies. CEACAM6 may contribute to the innate defense mechanisms of the mature type II cell.
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