Endothelin inhibits osteoclastic bone resorption by a direct effect on cell motility: implications for the vascular control of bone resorption.
The abundance of endothelin (ET)-producing endothelial cells in bone marrow and the proximity of these cells to bone-resorbing osteoclasts prompted us to evaluate the action of ET-1 on osteoclast function. Osteoclasts disaggregated from neonatal rat long bones were settled onto devitalized cortical bone substrate, and resorption was quantified by morphometry. The supernatant tartrate-resistant acid phosphatase activity was determined by a spectrophotometric method using paranitrophenol phosphate as substrate. Cell motility was quantified by time lapse video- and computer-assisted image processing using an empirical procedure for morphometric analysis. Cytosolic free calcium levels ([Ca2+]i) were measured in single cells by an indo 1-based microspectrofluorimetric method. Using the area of bone resorbed per slice as response, we found that ET-1 caused a significant (P = 0.011) concentration-dependent inhibition of osteoclastic bone resorption (EC50 = 2.5 nM) without inhibiting acid phosphatase secretion. Exposure of isolated osteoclasts to ET-1 also led to a marked concentration-dependent inhibition of osteoclast motility (EC50 = 7.9 nM; P = 0.013; t1/2 = 18 min) without significant effects on cell spread area. These effects of ET-1 were reversible after removing the peptide, and the cells remained viable during the experiments. In addition, ET-1 did not elevate [Ca2+]i at the concentrations tested. The results suggest that ET-1 specifically interacts with an osteoclast receptor to inhibit osteoclastic bone resorption and cell motility. As the concentration of ET-1 required for osteoclast inhibition was similar to that reported for smooth muscle contraction, it is possible that ET-1, produced locally from the bone marrow endothelial cell, might play a primary role in osteoclast regulation.
Alam
A S
AS
Department of Cellular and Molecular Sciences, St. George's Hospital Medical School, London, United Kingdom.
Gallagher
A
A
Shankar
V
V
Ghatei
M A
MA
Datta
H K
HK
Huang
C L
CL
Moonga
B S
BS
Chambers
T J
TJ
Bloom
S R
SR
Zaidi
M
M
D000135:Acid Phosphatase / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D001862:Bone Resorption; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D016232:Endothelins / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000502:physiology; D010010:Osteoclasts / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D051381:Rats; D011919:Rats, Inbred Strains
D016232:Endothelins; D000135:Acid Phosphatase; D002118:Calcium