2ND MESSENGER SIGNALING OF PTH-STIMULATED AND PTHRP-STIMULATED OSTEOCLAST-LIKE CELL-FORMATION FROM HEMATOPOIETIC BLAST CELLS

The second messenger signaling mechanisms of parathyroid hormone (PTH) - and PTH-related peptide (PTHRP)-stimulated osteoclast-like cell form ation were investigated in mouse hemopoietic blast cells that possesse d PTH binding sites. Human (h) PTH-(1-34) or hPTHRP-(1-34) resulted in a dose-dependent stimulation of tartrate-resistant acid phosphatase-p ositive multinucleated cells (MNC) formation. Pretreatment with [Nle8, 18Tyr34]hPTH-(3-34) significantly blocked hPTH-(1-34)- and hPTHRP-(1-3 4)-stimulated MNC formation. Dibutyryladenosine 3',5'-cyclic monophosp hate (10(-4) M) and forskolin (10(-5) M) as well as the stimulatory di astereoisomer of adenosine 3',5'-cyclic phosphorothioate (Sp-cAMPS), a direct activator of adenosine 3',5'-cyclic monophosphate (cAMP)-depen dent protein kinase (PKA) (10(-4) M), stimulated MNC formation, and Rp -cAMPS, an inhibitor of PKA activation (10(-4) M), almost completely i nhibited MNC formation stimulated by the aforementioned agents but not by 1,25-dihydroxyvitamin D3. Moreover, Rp-cAMPS significantly blocked PTH- and PTHRP-stimulated MNC formation. Treatment with calcium ionop hores (10(-8) and 10(-7) M) and phorbol 12-myristate 13-acetate, a pro tein kinase C (PKC) activator (10(-8) to 10(-6) M), but not 4alpha-pho rbol 12,13-didecanoate, a phorbol incapable of activating PKC, stimula ted MNC formation. Two PKC inhibitors [1-(5-isoquinolinylsulfonyl)-2-m ethylpiperazine dihydrochloride and staurosporine] equally blocked PTH - and PTHRP-stimulated MNC formation. The combined pretreatment with R p-cAMPS and PKC inhibitors completely blocked PTH- and PTHRP-stimulate d MNC formation. Present findings indicate that the activation of PKA and PKC is directly linked to PTH- and PTHRP-stimulated osteoclast-lik e cell formation from hemopoietic blast cells.