Peptide in Transfected Rodent Cell Lines Growth Factor-like Properties of Parathyroid Hormone-related

Parathyroid hormone-related peptide (PTHrP) was originally isolated from tumors associated with the development of hypercalcémie in vivo. Analyses of PTHrP gene expression have demonstrated that PTHrP is also produced in a wide variety of normal fetal and adult nonneoplastic tissues. The results of recent experiments have demonstrated that PTHrP is a growth factor-regulated gene, and different molecular forms of synthetic PTHrP display variable activities in assays for growth factor-like proper ties in vitro. We have studied the growth factor-like activity of PTHrP in cells transfected with a human PTHrP(hPTHrP) expression vector. Trans fected cell lines contained increased amounts of PTHrP mRNA transcripts as assessed by Northern blot analysis. The PTHrP mRNA transcripts were translated into immunoreactive hPTHrP as measured by radioimmunoassay, and conditioned medium from transfected cell lines stimulated cyclic AMP(cAMP) formation in ROS 17/2.8 osteosarcoma cells. The transform ing growth factor-ß-like properties of hPTHrP-producing NRK 49F clones were examined using the large-colony transformation assay in soft agar. PTHrP-producing NRK 49F clones did not form large colonies in the presence of epidermal growth factor. In contrast, PTHrP-producing and wild-type NRK 49F cells formed large colonies in the presence of epider mal growth factor and transforming growth factor-ß. No effect on cell growth was observed in PTHrP-producing NRK 49F rat kidney fibroblasts or mouse Mil 313 fibroblasts. In contrast, RGB 2.2 osteoblast cells expressing the hPTHrP cDNA were growth inhibited. Incubation of wildtype I«'II 2.2 cells with synthetic hPTHrP(l-34] (at concentrations of 1.0-10.0 nM) also produced growth inhibition. PTHrP increased cAMP formation in RCB 2.2 cells but not in NIH 3T3 or NRK 49F cells. Incu bation of RCB 2.2 cells with dibutyryl cAMP was also associated with inhibition of cell growth. The results of these studies demonstrate that PTHrP may function as an autocrine growth inhibitor of specific cell types, possibly through a cAMP-dependent pathway.