Bidirectional regulation of osteoclast function by nitric oxide synthase isoforms.

Abstract

Nitric oxide (NO) produces rapid osteoclast detachment and contraction in vitro, and this effect is accompanied by a profound inhibition of bone resorption. Work by others has confirmed these findings in vivo: inhibition of NO synthase [NOS; L-arginine, NADPH: oxygen oxidoreductase (NO-forming), EC 1.14.13.39] in normal rats is followed by increased bone resorption reflected by a marked loss in bone mineral density. In our present study, immunocytochemistry and Northern blotting show the presence of the constitutive calcium-sensitive NOS isoform (cNOS) in normal rat osteoclasts and in the human preosteoclast cell line (FLG 29.1 by normal rat osteoclasts treated with LPS or IFN-'y. In contrast, the nonselective NOS inhibitor NG-monomethyl-Larginine inhibits resorption by untreated neonatal rat osteoclasts. Thus, osteoclast function may require intermittent calcium-stimulated increases in NO production by cNOS against a basal inhibitory background activity of the iNOS isoform. However, bone resorption depends on precursor replication and on the activity of the mature cells, and we found that the NO donor 3-morpholinosydnonimine (SIN-1) (50 ,uM) profoundly depressed replication in the human preosteoclast line. Taken together, these results strongly suggest that NO maintains a central control of bone resorption in both avian and mammalian species by exerting a powerful tonic restraint of osteoclast numbers and activity. The presence of NOS in human cells implies a similar function in man and that conventional views of calcium homoeostasis and skeletal metabolism will need substantial revision. Since NO also influences behavior of the osteoblast, the boneforming cell, in vitro, a similar effect in vivo might imply a general influence on bone remodeling.Nitric oxide, or NO, secreted by the endothelium exerts an overriding control of blood pressure and blood flow by producing a dominant tonic dilatation of arterial muscle. To this crucial physiological role is added a number of other important functions. In the central nervous system, NO potentiates long-term memory; in the periphery it is the transmitter in the nonadrenergic noncholinergic (NANC) inhibitory nerves of the gastrointestinal and urogenital tracts. In contrast, the macrophage uses NO to kill invading microorganisms. NO is produced by several NO-synthase [NOS; L-arginine, NADPH: oxygen oxidoreductase (NO-forming), EC. 1.14.13.39] isoforms that require the same cofactors but vary in their response to calcium, mode of regulation, and tissue specificity (1). Constitutive isoforms of NOS have been identified in brain (2) and endothelium (3), while in macrophages (4) and hepatocytes (5) NOS transcripts of inducible NOS (iNOS) isoforms are detected only after treatment with cytokines or lipopolysaccharide (LPS). However, this gas radical now appears to be involved in still another function-that of control of osteoclastic activity. The osteoclast is the only cell in the body that can resorb bone and, together with the kidney, this cell plays the major role...

Endothelial Nitric Oxide Synthase Gene-Deficient Mice Demonstrate Marked Retardation in Postnatal Bone Formation, Reduced Bone Volume, and Defects in Osteoblast Maturation and Activity

“…In the absence of significant differences in osteoclast numbers between eNOSϩ/ϩ and eNOSϪ/Ϫ mice, this might indicate that eNOS is involved in the control of osteoclast activity and is supported by studies showing that osteoclasts contain eNOS and constitutive (eNOS) activity might be required to stimulate osteoclast activity. 9,16 However, it would seem that by comparison to the effects on osteoblast function and bone formation the overall impact of eNOS gene deficiency on the osteoclast and bone resorption is less important. Indeed, by 9 and 18 weeks of age there were no significant differences in eroded surface or osteoclast number.…”

“…9 -16 However, con-stitutive eNOS activity may also be of consequence, stimulating osteoclast function. 9,16 Basal, constitutive NO synthesis within osteoblasts supports proliferation and activity of these cells. 17 Moreover, osteoblast NO synthesis/activity is augmented by osteogenic hormones such as estrogen 18 and involves up-regulation of eNOS.…”

Aseptic Loosening Of Total Hip Replacement: Macrophage Expression Of Inducible Nitric Oxide Synthase And Cyclo-oxygenase-2, Together With Peroxynitrite Formation, As A Possible Mechanism For Early Prosthesis Failure

“…This rapid NO production implies the presence of a constitutive form of NOS within the bone cells and our recent studies have indeed shown that eNOS isoform is present in osteoblasts, osteocytes and osteoclasts. 25,34 It thus appears that NO may play a significant role during the early stages of mechanical strain-induced formation of new bone. 35,36 It is therefore possible to speculate that low levels of NO, as produced by the constitutive calcium-dependent isoforms of NOS, may be important in the initial stages of bone-implant adaptation.…”

“…Other studies on osteoclast function and models of bone resorption, however, have suggested that it acts on osteoclasts in a biphasic manner regulating osteoclast motility and resorption activity and may have a preferential role as an inhibitor of bone resorption. 24,25 NO is by definition a free radical, but it is not particularly reactive or toxic at relatively low levels, since it reacts with haemoglobin to form an inactive metabolite, methaemoglobin. At high levels, however, its cytotoxicity is much increased by its reaction with superoxide anion to form peroxynitrite.…”

Organic nitrates for osteoporosis: an update

“…In vitro studies demonstrate that NO has a biphasic effect on osteoclast activity and bone resorption; [14][15][16][17][18] low concentrations (as produced with activation of eNOS) potentiate bone resorption whereas high concentrations (as produced with activation of iNOS) inhibit activity. [19][20][21] The mechanism by which NO influences osteoclast activity may, in part occur via the receptor activator of NF-kappaB ligand (RANKL)/osteoprotegerin (OPG) pathway: high levels of NO stimulates OPG, OPG binds to RANKL that prevents the binding of RANKL to the receptor activator of NF-kappaB (RANK) and decreases osteoclast activity. 22 The effects of NO on osteoblasts are less well-characterized.…”