AJP - Cell Physiology, Vol 263, Issue 1 C130-C139, Copyright © 1992 by American Physiological Society
Protein kinase C affects microfilaments, bone resorption, and [Ca2+]o sensing in cultured osteoclasts
A. Teti, S. Colucci, M. Grano, L. Argentino and A. Zambonin Zallone
Institute of Human Anatomy, School of Pharmacy, University of Bari, Italy.
The effects of protein kinase C (PKC) in the control of osteoclast activity
are still unknown. We investigated the role of the enzyme in the control of
microfilament organization, podosome assembly, bone resorption, and
extracellular Ca2+ sensing in chicken and rabbit osteoclasts treated with
agents known to affect PKC activity. Cells were treated for 20 min with a
PKC activator [phorbol 12-myristate 13-acetate (PMA)], a PKC inhibitor
(staurosporine), a protein kinase A (PKA) inhibitor (H-9), a guanosine
3',5'-cyclic monophosphate-dependent protein kinase-PKA-PKC inhibitor
(H-7), or with the inactive phorbol, 4 alpha-phorbol, to examine
microfilaments by decoration with rhodamine-phalloidin. In PMA-treated
osteoclasts, the number of microfilament-containing adhesion structures
(podosomes) per cell decreased. However, enlarged microfilamentous cores in
podosomes and stress fiber-like filaments, otherwise absent in controls,
appeared. Whereas H-7 induced increase of the number of podosomes,
staurosporine, H-9, and 4 alpha-phorbol failed to change microfilament
organization. Chicken osteoclasts received also long-term treatment with
the agents in the presence of [3H]proline-prelabeled chicken or rat bone
particles to measure bone resorption. PMA, as well as staurosporine and
H-7, stimulated the resorbing activity, whereas cells were insensitive to
H-9 and 4 alpha-phorbol. Measurement of cytosolic free calcium
concentration in PMA-treated fura-2-loaded single osteoclasts demonstrated
a synergistic effect of PKC activation on the inhibitory extracellular
calcium concentration-sensing mechanism, which was, by contrast, blocked by
H-7, staurosporine, and H-9 and was insensitive to 4 alpha-phorbol. These
results indicate that PKC regulates osteoclast activity inducing both
morphological and functional modifications.
