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B S Moonga

Publications and source records attributed to B S Moonga.

63 records · Page 4Linked to original sources
PubMed1991

Correlates of osteoclast function in the presence of perchlorate ions in the rat.

The effects of perchlorate anion (ClO4-) on osteoclast properties were investigated through a number of independent in vivo and in vitro procedures. Intravenous infusion of ClO4- significantly reduced plasma [Ca] in young (50 g) Wistar rats, in the absence of changes in plasma [Mg] or [albumin]. This effect was maximal at 20 min after administration, and at a dose of 600 mumol/rat. Scanning electron-microscope images suggested that the presence of 10 mM-perchlorate reduced both the total area of cortical bone resorbed by freshly disaggregated rat osteoclasts, and the number of osteoclastic excavations in vitro. Similar effects were observed in the presence of 5 mM [Ca2+]. The effects of Ca2+ were potentiated by otherwise ineffective (1 mM) doses of perchlorate. Indo-1 dual-emission microspectrofluorimetry indicated a transient sixfold elevation of cytosolic free [Ca2+], in isolated cultured osteoclasts, with addition of 10 mM-perchlorate. Records of time-lapse video images indicated that this was followed by a marked and sustained cell retraction, by up to 70% of control cell area. Such effects were not observed at thiocyanate concentrations (10 mM) that would have produced comparable lyotropic effects as perchlorate. However, perchlorate did not alter morphometric measures for pseudopodial motility and cell migration. Nor did it influence supernatant concentrations of tartrate-resistant (osteoclastic) acid phosphatase in cultures of resorbing osteoclasts. These findings suggest that perchlorate is a potent inhibitor of osteoclast function, and acts through an influence on intracellular [Ca2+], and in turn upon the degree of cell retraction.

Acid Phosphatase↗
PubMed1990

Structure-activity relationship of human calcitonin-gene-related peptide.

The calcitonin-calcitonin-gene-related peptide (CGRP) gene complex encodes a small family of peptides: calcitonin, CGRP and katacalcin. Calcitonin is a circulating hormone that prevents skeletal breakdown by inhibiting the resorption of bone by osteoclasts. CGRP, a potent vasodilator, is involved in normal regulation of blood flow. The calcitonins structurally resemble the CGRP peptides, and both are known to cross-react at each others' receptors. The present study was undertaken to examine the structural prerequisites for biological activity of the intact CGRP molecule. We therefore prepared eight chymotryptic and tryptic fragments of CGRP and synthesized its acetylated and S-carboxyamidomethylcysteinyl analogues. The analogues were purified by h.p.l.c. and their structures were confirmed by fast-atom bombardment mass spectrometry. We have examined the effects of structurally modified analogues and fragments of human CGRP in a calcitonin-receptor-mediated assay, the osteoclast bone resorption assay, and in one or two CGRP-receptor-mediated assays, the rabbit skin blood flow assay and the oedema formation assay. The results showed that (1) in the osteoclast bone resorption assay, both CGRP peptides, alpha and beta, were equipotent, and were both at least 1000-fold were both approx. 1000-fold more potent than salmon calcitonin; human calcitonin had no effect; (3) the bis- and N-acetylated CGRP analogues retained reduced levels of biological activity in all assays, whereas S-carboxyamidomethylcysteinyl-human CGRP was without activity; and (4) all tryptic and chymotryptic fragments of CGRP were without biological activity, with the exception of hCGRP-(Ala1-Lys35): this fragment had much reduced activity compared with the intact peptide in inhibiting osteoclastic bone resorption and increasing blood flow in the rabbit skin. The results suggest that: (1) calcitonin and CGRP act at distinct receptors to mediate different physiological effects; (2) minor amino acid substitutions, as between the alpha and beta forms of CGRP (these two forms have 94% structural similarity) do not result in differences in biological activity; (3) the intact peptide is required for full biological activity of the CGRP molecule, and even the loss of two amino acids at the C-terminus of the molecule results in a marked decrease in activity; (4) the disulphide bridge appears to play an important role in the interaction of the intact CGRP molecule with its receptor; and (5) the C-terminal region is probably necessary for the peptide to assume the right conformation in the interaction with the receptor.

Amino Acid Sequence↗
PubMed1990

A new approach for calcitonin determination based on target cell responsiveness.

We report the development and validation of three microbioassays for calcitonin based on calcitonin-induced inhibition of the activity of isolated osteoclasts. Having precisely quantified osteoclast motility, spreading and bone resorptive activity, we have applied stringent analytical procedures to define assay characteristics. We have found that the appropriately transformed responses significantly regress on log dose of the peptides. Furthermore, potency estimates obtained using calcitonins from three species (human, salmon and a synthetic analogue of eel calcitonin) have been found to be consistent with those obtained using conventional calcitonin bioassays. In addition, the assays are remarkably sensitive (detection limit 10(-15) M), highly specific and precise. We have determined plasma levels of bioactive calcitonin on samples from patients with medullary thyroid carcinoma; these are several-fold lower than those obtained using our routine calcitonin radioimmunoassay. Our study thus, forms the basis of an entirely new approach for the determination of 'biologically active' calcitonin, and we envisage that such target cell-specific assays could become useful microanalytical methods.

Animals↗
PubMed1990

Intracellular regulation of enzyme secretion from rat osteoclasts and evidence for a functional role in bone resorption.

1. Osteoclasts are known to secrete acid phosphatase, an iron-containing phosphohydrolase. We have investigated (a) the possibility that acid phosphatase has a functional role in bone resorption and (b) the factors controlling enzyme secretion from isolated rat osteoclasts. 2. Osteoclasts were freshly disaggregated from neonatal rat long bones and dispersed at low densities on devitalized cortical bone slices or on plastic substrate. The levels of acid phosphatase in culture medium were measured spectrophotometrically using 4-nitrophenyl phosphate as hydrolysable substrate. The total plan area of bone resorbed was quantified by scanning electron microscopy in combination with image processing and analysis. 3. Ninety-three per cent of the total enzyme activity detected in the supernatant exposed to bone-osteoclast preparations was resistant to inhibition by D-tartaric acid and was bound to an antibody known to be highly specific for the osteoclast-derived isoenzyme, showing that it originated from osteoclasts. 4. A diminution in the level of supernatant enzyme activity achieved by incubating bone-osteoclast preparations with an antiserum specifically binding the osteoclast isoenzyme, or with a non-competitive inhibitor, molybdate or with competitive inhibitors, disphosphonates, led to a marked reduction of osteoclastic bone resorption. 5. The rate of the enzyme released into the culture supernatant, whether from resorbing (cultured on bone) or non-resorbing (cultured on plastic) osteoclasts declined gradually over 22 h, but that from the former was significantly depressed within the first 30 min of incubation. The supernatant enzyme concentration increased linearly up to 3 h; the levels released from resorbing osteoclasts remained consistently lower than those from non-resorbing cells. 6. Exposure of osteoclasts for 18 h to elevated [Ca2+]o levels produced a concentration-dependent inhibition of supernatant acid phosphatase levels. In the presence of 20 mM [Ca2+]o enzyme secretion from resorbing osteoclasts was significantly lower than that from non-resorbing cells. 7. Exposure of bone-osteoclast preparations to pertussis toxin produced no significant change of acid phosphatase release, while cholera toxin, dibutyryl cyclic AMP and forskolin produced a marked elevation of enzyme secretion. Ionomycin was found to inhibit enzyme release and this was less marked when osteoclasts were incubated on plastic substrate.(ABSTRACT TRUNCATED AT 400 WORDS)

Acid Phosphatase↗
PubMed1990

Evidence that the action of calcitonin on rat osteoclasts is mediated by two G proteins acting via separate post-receptor pathways.

Calcitonin inhibits osteoclastic bone resorption and its action involves two separate acute effects on the osteoclast, both essential to the action of the hormone: abolition of cell motility (Q) and marked cellular retraction (R). The former was mimicked by dibutyryl cyclic AMP and cholera toxin and the latter by pertussis toxin, ionomycin and increases in ambient calcium. Aluminium fluoride ions produced both Q and R effects, while lithium prevented both. In addition, calcitonin elicited a biphasic elevation of cytosolic-free calcium in single isolated osteoclasts. We propose that the action of calcitonin is mediated by at least two G proteins, one responsible for the Q effect and the other for the R effect. In addition, two second messengers, cyclic AMP and calcium, are involved. These findings may help to explain the potency of calcitonin in inhibiting bone resorption, and may allow the rational design of new therapeutic agents designed to alter osteoclast behaviour.

Aluminum↗
PubMed1990

The calcitonin gene peptides: biology and clinical relevance.

The calcitonin/CGRP multigene complex encodes a family of peptides: calcitonin, its C-terminal flanking peptide, katacalcin, and a third novel peptide, calcitonin gene-related peptide (CGRP). The 32-amino acid peptide calcitonin inhibits the osteoclast, thereby conserving skeletal mass during periods of potential calcium lack, such as pregnancy, growth, and lactation. This hormonal role is emphasized by observations that lower circulating calcitonin levels are associated with bone loss and that calcitonin replacement prevents further bone loss. Structurally, CGRP resembles calcitonin and has been implicated in neuromodulation and in the physiological regulation of blood flow. Here we review the molecular genetics, structure, and function of the calcitonin-gene peptides as analyzed in the laboratory and focus on more recent clinical studies relating to disorders and therapeutics.

Amino Acid Sequence↗
PubMedDate not supplied

Role of the endothelial cell in osteoclast control: new perspectives.

The osteoclast is of central importance in the process of bone remodeling. Its function is regulated by hormones and locally produced factors. Endothelial cells occur in close proximity to the osteoclast. Some endothelial cell-derived products, including endothelins, nitric oxide, and reactive oxygen species, have been recently implicated as modulators of osteoclast function. Endothelins inhibit bone resorption and osteoclast margin ruffling (quiescence or Q effect) at concentrations similar to those effective for their primary vasoconstrictive action. Contrary to expectations, however, it has been shown that endothelin action on the osteoclast is not mediated through an elevation of cytosolic Ca2+. Nitric oxide (NO) produces marked cell retraction (retraction or R effect), but its detailed mode of action is unknown. However, it is clear that the effects of this autocoid are not due to enhanced cyclic guanosine monophosphate (cGMP) production, a transduction system commonly used by NO. Finally, the reactive oxygen species H2O2 has been shown recently to enhance osteoclastic activity. Thus, the reported effects of the endothelial cell-derived products on the osteoclast are generally consistent with a regulatory role for endothelial cells in osteoclast control and suggest the existence of unique activation pathways, well worth exploring further. Unravelling the responsible mechanisms may also help understand the pathophysiology of a range of bone and joint diseases. For example, in rheumatoid arthritis, there is increased H2O2 production from activated neutrophils, and bone resorption is a major pathophysiological feature.

Bone Remodeling↗
PubMedDate not supplied

Novel mechanisms of calcium handling by the osteoclast: A review-hypothesis.

The osteoclast is a cell that is unique in its ability to resorb bone and, in doing so, becomes exposed to unusually high millimolar Ca2+ concentrations. It is generally accepted that, during resorption, osteoclasts can "sense" changes in their ambient Ca2+ concentration. This triggers a sharp cytosolic Ca2+ increase through both Ca2+ release and Ca2+ influx. The change in cytosolic Ca2+ is transduced finally into inhibition of bone resorption. It has been shown that a type 2 ryanodine receptor isoform, expressed uniquely in the plasma membrane, functions as a Ca2+ influx channel and possibly as a Ca2+ sensor. Ryanodine receptors are ordinarily Ca2+ release channels that have a microsomal membrane location in a wide variety of eukaryotic cells, including the osteoclasts. However, only recently has it become obvious that ryanodine receptors are also expressed in osteoclast nuclear membranes, at which site they probably gate nucleoplasmic Ca2+ influx. Nucleoplasmic Ca2+ in turn regulates key nuclear processes, including gene expression and apoptosis. Here, we review the potential mechanisms underlying the recognition, movement, and effects of Ca2+ in the osteoclast. We will also speculate on the general biological significance of the unique processes used by the osteoclast to handle high Ca2+ loads during bone resorption.

Animals↗