Characterization of the mucin differentiation in human lung adenocarcinoma cell lines

Four human lung adenocarcinoma cell lines were established in serum-free F12 medium supplemented with insulin, transferrin, hydrocortisone, cholera toxin, selenium, epidermal growth factor, bovine hypothalamic extract, and retinoic acid. Histochemical analyses with periodic acid-Schiff with and without diastase treatment (PAS-D technique) and immunocytochemistry with a mucin-specific monoclonal antibody demonstrated that three of the cell lines (CL2, CL3, and NCL2) were capable of mucin production. Biochemical characterizations of mucin produced by adenocarcinoma cells were focused on one of the cell lines, CL2 cells, which showed the most prominent reactivity with mucin-specific monoclonal antibody. Biochemical analysis using the mucin precursors [3H]glucosamine and [14C]serine indicated that CL2 cells can synthesize high-molecular-weight (M(r) greater than 200 kD) glycoprotein molecules that can be immunoprecipitated by this mucin-specific monoclonal antibody. The high-molecular-weight glycoproteins isolated from CL2 cells specifically reacted with mucin-specific monoclonal antibody by Western blot analysis, and composition analyses showed high levels of serine and threonine and a low level of aromatic amino acids, which are similar to human airway mucin. These observations suggest that lung adenocarcinoma CL2 cells cultured in this serum-free medium can retain function of airway mucin synthesis. Cell kinetic studies of these four cell lines showed that the cell line (CL1) without the mucin differentiation had a higher proliferative index and a shorter population doubling time as compared with the other three cell lines (CL2, CL3, and NCL2) with mucin differentiation. Examination of the retinoblastoma protein expressions in these adenocarcinoma cell lines revealed a phosphorylated pattern that correlated inversely with the mucin synthesis status of these cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)