Human osteoblast-like cell proliferation induced by calcitonin-related peptides involves PKC activity.
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 [(3)H]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 (EC(50) = 3.4 x 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.
Villa
I
I
Bone Metabolic Unit, Scientific Institute H San Raffaele, 20132 Milan, Italy.
Dal Fiume
C
C
Maestroni
A
A
Rubinacci
A
A
Ravasi
F
F
Guidobono
F
F
D000682:Amyloid / Q000494:pharmacology; D002116:Calcitonin / Q000737:chemistry / Q000494:pharmacology*; D058286:Calcitonin Receptor-Like Protein; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D058228:Islet Amyloid Polypeptide; D010006:Osteoblasts / Q000166:cytology*; D010446:Peptide Fragments / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D018003:Receptors, Calcitonin / Q000378:metabolism; D019311:Staurosporine / Q000494:pharmacology
D000682:Amyloid; C545578:CALCRL protein, human; D058286:Calcitonin Receptor-Like Protein; D004791:Enzyme Inhibitors; D058228:Islet Amyloid Polypeptide; D010446:Peptide Fragments; D018003:Receptors, Calcitonin; D015240:Phorbol 12,13-Dibutyrate; D002116:Calcitonin; D000242:Cyclic AMP; D011493:Protein Kinase C; D019311:Staurosporine