Production of parathyroid hormone-related protein by normal and neoplastic keratinocytes

Abstract

Production of parathyroid hormone-related protein (PTHrP) by normal and neoplastic keratinocytes was compared in vitro to determine if the malignant phenotype alters the regulation of PTHrP expression and secretion. Three cell types consisted of normal human foreskin keratinocytes (NHFK), a well differentiated canine squamous carcinoma cell line (SCC 2/88), and a poorly differentiated human squamous carcinoma cell line (A253). The cells were grown in monolayers and on substrates (artificial dermis or collagen) to induce 3-dimensional layering of the cells and to permit medium collection from the basal and apical regions of the cells. Secretion of PTHrP was measured by N-terminal radioimmunoassay and mRNA production was measured by RNase protection assay. Secretion and mRNA production of PTHrP by NHFK and SCC 2/88 cells in monolayer culture was maximal at confluence (1–5 ng/ml) and decreased thereafter. In contrast, A253 cells continued to produce PTHrP after confluence in monolayer cultures. NHFK, SCC 2/88, and A253 cells in 3-dimensional, multilayered cultures produced high levels of PTHrP after confluence. Secretion of PTHrP was greatest from the basal cells compared to the apical cells of the cultures. PTHrP secretion was inhibited by monensin (1 µM). Transforming growth factor-β (1 to 5 ng/ml) increased PTHrP mRNA in NHFK at 6 hours and in SCC 2/88 cells at 1-24 hours. These data indicate that PTHrP is produced preferentially by proliferating keratinocytes and regulation of PTHrP production is different in normal keratinocytes compared to squamous carcinoma cells.