Human osteoblast-like cell proliferation induced by calcitonin-related peptides involves PKC activity

Abstract

The calcitonin peptides [calcitonin (CT), calcitonin gene-related peptide (CGRP), amylin] share many biological actions, including activity on bone cells. In the present study, CT (10−11 to 10−9 M) stimulated [3H]thymidine incorporation in primary cultures of human osteoblasts (hOB), as already demonstrated for CGRP and amylin. RT-PCR analysis showed that the calcitonin receptor and the calcitonin receptor-like receptor are both expressed in hOB. In these cells, CT (10−10 M) and amylin (10−9 M), in contrast to CGRP (10−8 M), did not increase cAMP production. All three peptides stimulated protein kinase C (PKC) activity. To evaluate PKC involvement in hOB proliferation, cells were incubated with phorbol 12,13-dibutyrate, a stimulator of PKC activity; cell proliferation was increased in a dose-dependent manner (EC50 = 3.4 × 10−8 M). Staurosporine (10−9 M), a PKC inhibitor, blocked phorbol 12,13-dibutyrate-induced PKC activity and cell proliferation. Inhibition of PKC by staurosporine also counteracted the stimulatory effect of CT, CGRP, and amylin on hOB proliferation. From these data, it is deduced that the activation of PKC is important for hOB proliferation and that it is involved in the anabolic effect of CT peptides on bone.

Endogenous Calcitonin Gene-related Peptide Protects Human Alveolar Epithelial Cells through Protein Kinase Cϵ and Heat Shock Protein

“…Although previous study has shown the activation of PKC by CGRP in osteoblast cells (38), our study is the first to clarify that the PKC⑀ subtype is selectively activated by CGRP in alveolar epithelial cells. Further studies are warranted to elucidate whether the CGRP/PKC⑀/HSP70 pathway also plays a role in the effect of CGRP in other cell types, especially the endothelial cells, cardiac myocytes, and neurons.…”

Calcitonin gene-related peptide elevates calcium and polarizes membrane potential in MG-63 cells by both cAMP-independent and -dependent mechanisms

“…3,19, and 28; Burns and Kawase, unpublished observations). In addition, we (18,20,21) and others (11,12,38,39) have recently reported that CGRP can stimulate cAMP-independent intracellular signaling pathways in several osteoblastic cell types. We thought it significant that mature osteoblast-like cells respond to CGRP by increasing intracellular Ca 2ϩ ([Ca 2ϩ ] int ) and/or altering membrane potential (E m ) while yielding small increases (if any) in cAMP production (18,20,21).…”

Calcitonin gene-related peptide stimulates BMP-2 expression and the differentiation of human osteoblast-like cells in vitro

“…BMP-2 promotes the proliferation of pre-osteoblast cells, induces the osteogenic or chondrogenic differentiation of mesenchymal cells [17,18] , and improves the osteogenic phenotype and capacity of stem cells through increased ALP activity and osteocalcin mRNA expression. Signal transduction studies have revealed that the Smad and P38MAPK pathways are immediately downstream of BMP receptors and that they play central roles in BMP signal transduction [4,[19][20][21] .…”