ALTERED PARATHYROID HORMONE-RELATED PROTEIN SECRETION AND MESSENGER-RNA EXPRESSION IN SQUAMOUS CARCINOMA-CELLS IN-VITRO

Parathyroid hormone-related protein (PTHrP), a major factor in the pat hogenesis of humoral hypercalcemia of malignancy, is produced by many squamous carcinoma cells (SCCs). Two SCC fines were grown in multilaye red culture systems and compared to cells grown as monolayers to evalu ate the effects of cell proliferation, confluence and differentiation on PTHrP secretion and mRNA expression. Well-differentiated (SCC 2/88) and poorly differentiated (SCC-A253) SCCs were grown as monolayer and three-dimensional cultures on collagen-coated membranes to compare th e regulation of PTHrP expression and secretion by the cell lines in vi tro. Parathyroid hormone-related protein secretion was evaluated by ra dioimmunoassay and immunohistochemistry. Messenger RNA expression was analyzed by RNase protection assay and in situ hybridization. Secretio n of PTHrP was greatest in preconfluent SCC 2/88 cells in monolayer cu lture and decreased after confluence, which was the result of decrease d PTHrP mRNA expression In contrast, PTHrP secretion in cultures of SC C-A253 cells reached maximal levels after confluence. In multilayered cultures, total PTHrP secretion and mRNA expression remained high in b oth SCC 2/88 and SCC-A253 cells, and secretion by the multi-layered cu ltures was principally in the basal direction. Parathyroid hormone-rel ated protein was present in all cell layers in three-dimensional cultu res as determined by immunohistochemistry. These results indicated tha t multilayered cultures of SCCs produced PTHrP continuously, whereas d ecreased proliferation in monolayer cultures was associated with decre ased PTHrP production. Multilayered cultures represent a better system to investigate PTHrP secretion and mRNA expression in vitro and PTHrP secretion by SCCs in vivo may be greatest by proliferating cells.