Human pulp cells respond to calcitonin gene-related peptide in vitro

Cell monolayers derived from human pulpal explants were passaged 3 to 4 times before characterization of the response of the cells to calcitonin gene-related peptide (CGRP). Northern blot analysis of mRNA revealed the presence of transcripts for bone morphogenetic protein-2 (BMP-2). Stimulation with CGRP produced a 1.8-fold increase in BMP-2 mRNA expression by the cells. Analysis of binding of CGRP to whole cells indicated that unlabeled human CGRP competed with labeled CGRP in a dose dependent fashion with a K_D, estimated from the EC₅₀, in the range of 5 × 10_-8M. Binding of labeled CGRP was greatly reduced by the presence of 10^-6M CGRP but was unaffected by the presence of 10^-6M Human PTH(1-34) or 10^-6M calcitonin. CGRP produced a 2.8-fold increase in cyclic AMP over basal levels, which was similar to the increase produced by PTH(1-34) (2.6 fold) but slightly more than the increase produced by calcitonin (1.9 fold). The pulp-derived cells displayed a high basal level of alkaline phosphatase enzyme activity, which was not altered by treatment with CGRP or either PTH{1-34) or 1,25(OH)₂D₃. Stimulation with 2.5 × 10_-8M 1,25(OH)²D₃ did produce a 7.6-fold increase in osteocalcin. These results indicate that pulp cells possess the cellular machinery to respond to CGRP and that stimulation of the production of BMP-2, a factor known to be associated with induction of dentin formation, is a component of the response.