ABSTRACT
The cytoplasmic spreading of osteoclasts has been used to assess responsiveness to agents such as calcitonin and associated signal transduction mechanisms. Although cyclic AMP and intracellular calcium are known mediators of calcitonin effects in osteoclasts, the role of protein kinase C (PKC) is less clear. We have used time-lapse videomicroscopy of isolated rat osteoclasts to characterize shape changes induced by calcitonin, forskolin, and phorbol 12-myristate-13-acetate (PMA) in the absence and presence of PKC blockers. Treatment with calcitonin reduced cytoplasmic plan area but increased perimeter length, resulting in a characteristic "stellate" appearance, whereas forskolin produced "nonstellate" contraction. The response of osteoclasts to PMA was dose dependent. High concentrations (10(-7)-10(-6) M) produced biphasic responses with transitory, calcitonin-like "stellate" contraction followed by sustained expansion, whereas low concentrations (10(-11)-10(-9) M) produced expansion only. The effects of low-concentration PMA could be prevented by pretreatment with a PKC blocker, whereas the effects of high concentrations were only partially inhibited. The effects of forskolin were unchanged by pretreatment with the PKC blocker. Treatment with calcitonin in the presence of various PKC blockers resulted in paradoxical transient expansion followed by contraction. These results indicate that calcitonin-induced shape change in osteoclasts is a complex process involving protein kinase C in addition to cyclic AMP-dependent mechanisms and possibly other factors.
Subject(s)
Analgesics/toxicity , Calcitonin/toxicity , Osteoclasts/drug effects , Protein Kinase C/metabolism , Animals , Carcinogens/toxicity , Cell Separation , Cell Size/drug effects , Colforsin/toxicity , Cyclic AMP/metabolism , Cytoplasm/drug effects , Cytoplasm/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Naphthalenes/pharmacology , Osteoclasts/enzymology , Protein Kinase C/antagonists & inhibitors , Rats , Rats, Sprague-Dawley , Tetradecanoylphorbol Acetate/toxicityABSTRACT
Zinc is an important element in biology yet little is understood of its role in bone cell metabolism and function. This study examined the effects of zinc on osteoclast (OC) function in cultures derived from neonatal rats and in cocultures of OC and UMR 106-01 osteoblast-like cells (UMR/OC cocultures). Treatment with zinc (10(-12)-10(-4) mol/L) had no effect on either bone resorption or the number of multinucleate cells positive for tartrate-resistant acid phosphatase (TRACP + ve MNC) in OC cultured for 24 h on bone slices. However, in UMR/OC cocultures, 10(-4) mol/L zinc (but not lower concentrations) decreased resorption pit formation by approximately 50% and increased TRACP + ve MNC number by approximately 40%. When osteoblast-like cells were pretreated with zinc prior to, but not during, coculture with OC, effects on TRACP + ve MNC and pit number persisted, although the effect was reduced. Zinc treatment also inhibited resorption and stimulated TRACP and calcitonin receptor (CTR) + ve MNC numbers in long-term (96-120 h) UMR/OC cocultures. Our results indicate that zinc increases TRACP + ve CTR + ve MNC numbers yet inhibits bone-resorbing activity, and that these effects are dependent on the presence of osteoblastic cells. Zinc is abundant in bone and may act as a local regulator of bone cells.
