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At least 19 recordsLinked to original sources
PubMed1993

Comparative effects of eel calcitonin, salmon calcitonin and [Asu1,7]eel calcitonin on hypophyseal and osteoblastic function.

Three different calcitonins: salmon calcitonin, eel calcitonin and the semi-synthetic analog [Asu1,7]eel calcitonin have been evaluated for their ability to affect phosphoinositide hydrolysis in primary cultures of anterior pituitary cells and in the osteoblast-like UMR-106 cells. In both cellular systems a repeated treatment with any form of calcitonin induced an inhibition of inositol phospholipid turnover. Eel calcitonin and its analog were always more potent than salmon calcitonin, but the efficacy of the three polypeptides was comparable. In cultured anterior pituitary cells, the inhibitory effect on phosphoinositide hydrolysis observed after chronic treatment with calcitonin was accompanied by a reduction of prolactin release. In contrast, a single treatment of cultured anterior pituitary cells with eel calcitonin or its analog [Asu1,7]eel calcitonin induced an increase of inositol phosphate accumulation, while salmon calcitonin was inactive. Accordingly, eel and [Asu1,7]eel calcitonin, but not salmon calcitonin, induced a slight but significant stimulation of prolactin secretion. In UMR-106 cells, the three calcitonins exhibited similar potency and efficacy in reducing parathyroid hormone-stimulated 4 beta[3H]-phorbol-12,13-dibutyrate ([3H]PdBu) binding, an indirect index of protein kinase C activation. Taken together, these results suggest that, either at the pituitary or in osteoblast-like cells, some of the effects exerted by calcitonin may be ascribed to an interference with the intracellular events initiated by modulation of phosphoinositide turnover.

Analysis of Variance↗
PubMed1989

Effects of calcitonin, calcitonin analogues, and calcitonin gene-related peptide on basal in vitro renin secretion.

We investigated the relation of calcitonin as a calcium-active hormone to its more recently described effects on peripheral vascular tone. Basal renal renin secretion in vitro in rat kidney slices was studied in the presence of salmon calcitonin (SCT, 4400 U/mg), amino acid substituted analogues of SCT, 16-alanine SCT (6200 U/mg) and 12,16,19 tri-alanine SCT (350 U/mg), and of rat calcitonin gene-related peptide (rCGRP). All calcitonin species at the same hypocalcemic activity (1 U/mL) modestly but significantly suppressed renin secretion from control levels (9.79 +/- 0.44 to 7.51 +/- 0.53, 7.70 +/- 0.72, and 7.78 +/- 0.90 Goldblatt units/g/h for SCT, 16-ala SCT, and tri-ala SCT, P less than .05 for all calcitonins v control), whereas rCGRP had no effect. Thus, on a molar basis, the renin suppressing effects of the various calcitonin species paralleled their bioassay-defined calcium sequestering activity, 16-ala SCT greater than SCT much greater than tri-ala SCT. Lower concentrations of SCT (10(-2) U/mL and 10(-4) U/mL, approximately 6 X 10(-10) and 6 X 10(-12) mol/L, respectively) had virtually identical effects. Moreover, verapamil (5 X 10(-6) mol/L) blocked the SCT-induced suppression of renin secretion (9.79 +/- 0.44 v 9.36 +/- 1.05 GU/g/h, P = NS). We conclude that the juxtaglomerular apparatus is a calcitonin-responsive system, in which calcitonin and its analogues act to suppress basal renin secretion in vitro. This effect seems to depend on and may be mediated by modulating cellular calcium uptake, and suggests a wider, calcium-related role for calcitonin than had previously been suspected.

Animals↗
PubMedDate not supplied

Deletions of genes encoding calcitonin/alpha-CGRP, amylin and calcitonin receptor have given new and unexpected insights into the function of calcitonin receptors and calcitonin receptor-like receptors in bone.

It has been suggested that skeletal nerves fibers may play important roles in neuro-osteogenic interactions. This view is partly based upon information obtained from immunohistochemical studies, chemical and surgical denervation experiments and clinical observations in patients with stroke and spinal cord injury, indicating the presence of a network of nerve fibers in the skeleton and that defective signalling in skeletal nerve fibers affects remodelling of bone. This view is also supported by data showing that functional receptors for signalling molecules in skeletal nerve fibers are expressed in bone cells and that activation of these receptors leads to profound effects on bone forming osteoblasts and bone resorbing osteoclasts. Convincing evidence for a role of neuronal signalling in bone metabolism has been provided by gene deletion approaches in which it has been shown that leptin-sensitive and neuropeptide Y-sensitive receptors in hypothalamus are important for bone remodelling in mice. Recently, gene deletion experiments have shown that calcitonin gene-related peptide (CGRP), one of the neuropeptides present in skeletal nerve fibers, is an important physiological regulator of bone formation at the level of osteoblast activity. CGRP belongs to the calcitonin (CT) family of peptides also including CT, amylin and adrenomedullin, as well as the recently described intermedin and calcitonin receptor-stimulating peptide. These peptides utilize two seven transmembrane G protein-coupled receptors - the calcitonin receptor (CTR) and the calcitonin receptor- like receptor (CRLR) - which can dimerize with three different single transmembrane proteins, making up the RAMP family. Associations between RAMPs and either CTR or CRLR give rise to seven distinct, molecularly characterized, receptors for CT, CGRP, amylin and adrenomedullin. Deletions of the genes for ligands in the CT family of peptides and for one of the receptors have revealed unexpected findings that have changed our view on the role of these peptides in bone remodelling. It was anticipated that deletions of the CT/alpha-CGRP and CTR genes would lead to bone loss, since CT has been shown to inhibit bone resorption in vitro and in vivo and has been used to treat patients with excessive bone resorption. Surprisingly, it was found that CT/alpha-CGRP-/- and CTR+/- mice have increased bone mass due to increased bone formation. Mice with deletion of the amylin gene, however, exhibited bone loss due to enhanced bone resorption. Selective deletion of the alpha-CGRP gene also leads to bone loss, but due to decreased bone formation. Thus, our understanding of the role of the CT family of peptides has been changed dramatically and much more data have to be gained before we fully understand the roles these peptides have in bone biology.

Amyloid↗
PubMed1991

The neurotransmitter role of calcitonin gene-related peptide in the rat and guinea-pig ureter: effect of a calcitonin gene-related peptide antagonist and species-related differences in the action of omega conotoxin on calcitonin gene-related peptide release from primary afferents.

In the rat and guinea-pig isolated ureter electrical field stimulation of intrinsic nerves (10 Hz for 10 s) produces transient inhibition of evoked (20 mM KCl or 0.1-1 microM neurokinin A) rhythmic contractions by releasing transmitter(s) from peripheral endings of capsaicin-sensitive primary afferents. The C-terminal fragment of human calcitonin gene-related peptide (8-37) blocked the inhibitory effect of electrical field stimulation as well as that produced by exogenous calcitonin gene-related peptide, while leaving unaffected the inhibitory response to isoprenaline. Human calcitonin gene-related peptide (8-37) was devoid of any inhibitory activity of its own but enhanced the amplitude and frequency of KCl-evoked rhythmic contractions in the rat ureter, probably by antagonizing the inhibitory effect of endogenous calcitonin gene-related peptide released by KCl. Omega conotoxin fraction GVIA, a peptide which possesses a potent blocking activity of N-type voltage-sensitive calcium channels, prevented the inhibitory response to electrical stimulation in the guinea-pig ureter, while leaving the response unaffected in the rat ureter. Conotoxin had no effect toward the inhibition produced by exogenous calcitonin gene-related peptide indicating its prejunctional site of action, demonstrated previously in the guinea-pig ureter [Maggi et al. (1990) Neurosci, Lett. 114, 203-206]. Dermorphin, an amphibian peptide with potent agonist activity on mu-type opioid receptors, inhibited the response to electrical stimulation in the guinea-pig ureter but had no effect in the rat ureter.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2003

Molecular cloning and expression of equine calcitonin, calcitonin gene-related peptide-I, and calcitonin gene-related peptide-II.

In this study, we describe the cloning and tissue expression of equine calcitonin (CT), calcitonin-gene related peptide (CGRP)-I, and CGRP-II cDNA. We also describe a novel divergent form of CGRP (CGRP-I). Equine CT has greatest homology (>85%) to human, rat and mouse subgroups of calcitonins. Equine CGRP-I has low homology (<59%) to CGRPs of other species. The signal and N-terminal peptides for equine CT and CGRP-I were identical, indicating that these peptides are encoded by a gene equivalent to the human CALC-I gene. Equine CGRP-II has >80% homology to chicken, human, rat, ovine, swine, and bovine CGRPs. The homology between equine CGRP-I and CGRP-II is low (56%). The high homology of equine CGRP-II and the low homology of equine CGRP-I to CGRP in other species were unexpected findings. Northern blot analysis revealed that CT mRNA expression was restricted to the thyroid gland; however, RT-PCR revealed that CT mRNA expression was also present in the pituitary gland and in the liver. CGRP-I and CGRP-II mRNA expression was present in several regions of the nervous system and other tissues of neuroectodermal origin. An unexpected finding was CGRP-I expression in the kidney by both Northern analysis and by RT-PCR. Based on these results, CT gene expression in the horse was not restricted to the thyroid gland, and CT may be important in regulating pituitary cell function. CGRPs are widely expressed in tissues of the central and peripheral nervous system. Information from this study will be valuable to study the role of CT, CGRP-I, and CGRP-II in equine health and disease.

Animals↗
PubMed1990

International Standards for salmon calcitonin, eel calcitonin, and the Asu1-7 analogue of eel calcitonin: calibration by international collaborative study.

Regulatory specifications in most countries require that the potency of salmon calcitonin (sCT) clinical products be expressed in International Units (IU) defined by the World Health Organization (WHO) International Standard. The first ampouled standard was prepared in 1972 and has been distributed world-wide since then. A batch of ampoules to serve as the replacement standard is now required. Other piscine calcitonins, eel calcitonin (eCT) and an amino-suberic acid analogue of eCT (Asu1-7 eCT) are now clinical products in some countries and international standards are required for these peptides which are similar to, but not identical with, sCT. This paper describes the preparation of three new ampouled standards and their biological calibration by international collaborative study comprising 17 participants from 10 countries. Following the recommendations in the final report of the collaborative study, the 2nd International Standard (IS) for sCT, the 1st IS for eCT and the 1st IS for Asu1-7 eCT were recently established by WHO, each with an assigned potency in IU, and are now available for issue.

Animals↗
PubMed1987

Calcitonin, calcitonin gene-related peptide and renal calcitonin receptors in the Zucker rat.

In this study we investigated (a) whether genetically obese Zucker rats exhibit higher levels of calcitonin gene-related peptide (CGRP) in the thyroid and plasma than lean controls, (b) if CGRP, like calcitonin (CT), increases in plasma in response to iv Ca or iv pentagastrin and (c) whether or not renal CT receptors exhibit 'downregulation' in fat rats compared to leans. Fat and lean rats 8-12 months old were used. Compared to leans, fat rats exhibited higher circulating levels of CGRP as well as CT. Also, the total thyroid content (micrograms per gland) and concentration (ng/mg wet weight) of CGRP were higher in fat rats. One minute after iv pentagastrin, both lean and fat rats showed a marked increase in plasma CT but not plasma CGRP. In another experiment involving only lean rats, iv Ca given 4 min earlier produced a massive increase in plasma CT but no detectable increase in plasma CGRP. Using renal membranes and 125I-sCT as the ligand, we found that renal CT receptors in fat rats appeared more abundant than in lean rats and exhibited the same KD (3-6 nM) for both. The findings show that, compared to leans, fat rats have higher levels of CGRP as well as CT in the thyroid gland and blood. However, stimulation of C-cell release of CT was not accompanied by a corresponding rise in plasma CGRP, suggesting that either (a) the plasma CGRP is derived largely from non-thyroidal sources and the adult rat thyroid releases only trivial amounts of CGRP or (b) Ca and pentagastrin are secretagogues for thyroidal CT but not CGRP. Examination of renal CT receptors in fat and lean rats provided no evidence for downregulation of receptors in fat rats exposed chronically to higher circulating levels of CT than leans.

Animals↗
PubMed1986

Effect of intracerebroventricular administration of rat calcitonin gene-related peptide (CGRP), human calcitonin and [Asu1,7]-eel calcitonin on gastric acid secretion in rats.

The effect of intracerebroventricular injection of rat calcitonin gene-related peptide (CGRP), human calcitonin (CT) and [Asu1,7]-eel CT on the volume and acidity of gastric juice was examined in the pylorus-ligated male rats. These 3 peptides were effective in suppressing both the volume and acidity of secreted gastric juice. Their potency on a molar basis, however, was markedly different; [Asu1,7]-eel CT was most potent, followed by human CT and finally by rat CGRP. These finding suggest that CGRP could not substitute for [Asu1,7]-eel or human CT in exerting the suppressive effect of gastric acid secretion.

Animals↗
PubMed1992

N-terminal truncation of salmon calcitonin leads to calcitonin antagonists. Structure activity relationship of N-terminally truncated salmon calcitonin fragments in vitro and in vivo.

Structural requirements for binding to the bone calcitonin (CT) receptor and for CT bioactivity both in vitro and in vivo were assessed for a series of N-terminally truncated, N alpha-acetylated, fragments of salmon calcitonin (sCT). Sequential deletion of amino acid residues from the amino-terminus of [Ala7]sCT-(2-32) peptide amide first led to partial agonists and, upon deletion of residues 1 to 7, to a high affinity antagonist, N alpha-acetyl-sCT-(8-32)-NH2. The presence of two separate domains within the sCT sequence is proposed: (I) a binding domain comprising residues 9-32 and (II) an activation domain requiring residues 3 to 6. N alpha-acetyl-sCT-(8-32)-NH2, in several bioassays including plasminogen activator release from LLC-PK1 cells (pA2 = 7.31), cAMP production in UMR-106-06 cells (pA2 = 7.81) and in the fetal rat long bone resorption assay showed potent antagonistic properties.

Acetylation↗
PubMed1985

F.9 embryonal carcinoma cell calcitonin autocrine system: correlation between immunoreactive calcitonin secretion and calcitonin receptor number.

Mouse teratocarcinoma cells in culture offer an in vitro system to study the initial steps of embryogenesis. It has been suggested that, at such early stages, cell functions are regulated by an autocrine process in which embryonic cells produce factors that in turn act on themselves. F.9 cells possess specific membrane receptors for calcitonin (CT) (120 fmol/mg of protein, Ka, = 3.5 X 10(8) M-1). These cells produce CT detected by heterologous radioimmunoassay in serum-free culture-conditioned medium (75 pg/10(7) cells/12 h). When F.9 cells are incubated in serum-free medium, CT binding and secretion concomitantly drop by 50% within the first 2 h, then increase progressively to an upper plateau after the sixth hour. Preincubation with 10(-6) M CT leads to disappearance of CT receptors and CT secretion in the culture medium up to 6 h. Avoiding accumulation of CT in the medium by a continuous flow rate for 6 h leads to a progressive decrease of CT receptors. In addition, retinoic acid treatment of cells induces a parallel progressive decrease of CT receptor number and of total CT synthesis. These results suggest a reciprocal regulation of CT receptors and CT secretion, or a close relationship between their regulations.

Animals↗
PubMed2000

Preoperative calcitonin levels are predictive of tumor size and postoperative calcitonin normalization in medullary thyroid carcinoma. Groupe d'Etudes des Tumeurs a Calcitonine (GETC).

Medullary thyroid carcinoma (MTC) is a calcitonin (CT)-secreting endocrine tumor. Although plasma CT level is a specific and sensitive marker of MTC, its preoperative usefulness in predicting tumor size and postoperative CT normalization has not been documented. From a nationwide database set up by the French CT Tumor Study Group, 226 MTC patients were selected according to the following criteria: preoperative CT level determination by an immunoradiometric assay (normal value, < 10 pg/mL) within the 6 months prior to surgery, total thyroidectomy and diagnosis of MTC ascertained by histological report including tumor size. Patients were 129 females and 97 males (female/male ratio, 1.3). One hundred and twelve patients (49.6%) had the sporadic variety of the disease, 74 (32.7%) had multiple endocrine neoplasia 2A, three (1.3%) had multiple endocrine neoplasia 2B, and 37 (16.4%) had familial MTC. Median age at diagnosis was 44.8 yr (range, 4.9-80.1 yr). Complete neck dissection was performed in 159 patients (70.4%). Postoperative CT normalization was ascertained by negative response of CT to pentagastrin stimulation (< 10 pg/mL) in 94 patients. Seventy-one patients were considered as not cured because of residual tumor tissue and/or elevated CT levels. Median tumor size was 11.0 mm (range, 0.2-80.0 mm), significantly larger in females (15.0 vs. 8.0 mm, P < 0.05), and in sporadic forms (15.0 vs. 7.0 mm, P < 0.05). Tumor size was significantly correlated (r2 = 0.52, P < 0.01) with preoperative CT levels, the relationship being more straight in familial (r2 = 0.71) than in sporadic (r2 = 0.36) forms. Furthermore, preoperative CT levels under 50 pg/mL appeared to be predictive of postoperative CT normalization (44 of 45 patients). However, higher CT levels did not mean absence of postoperative CT normalization (50 of 120 patients). We conclude that low preoperative CT levels are predictive of tumor size and postoperative CT normalization.

Adolescent↗
PubMed1998

Uterine relaxation responses to calcitonin gene-related peptide and calcitonin gene-related peptide receptors decreased during labor in rats.

OBJECTIVES: Our purpose was to investigate (1) whether uterine relaxation responses to calcitonin gene-related peptide are differentially regulated during pregnancy and labor, (2) the involvement of nitric oxide in smooth muscle relaxant action of calcitonin gene-related peptide in the rat uterus, (3) whether receptors for calcitonin gene-related peptide are expressed in rat uterus, and if so (4) whether the concentrations of these receptors are differently regulated during pregnancy and labor. STUDY DESIGN: Rats were killed on day 18 of gestation, at the time of spontaneous labor, or postpartum day 2. The uteri were removed for in vitro contractility measurements, nitric oxide production, and calcitonin gene-related peptide receptor binding assay. RESULTS: (1) Calcitonin gene-related peptide induced a dose-dependent relaxation in spontaneously contracting uterine strips from pregnant rats on day 18 of gestation; (2) the relaxation effects of calcitonin gene-related peptide on the uterus were decreased during spontaneous delivery at term and post partum compared with that during pregnancy; (3) calcitonin gene-related peptide-induced relaxation was inhibited by pretreatment of the uterine tissue with a calcitonin gene-related peptide receptor antagonist, calcitonin gene-related peptide(8-37); (4) nitric oxide synthesis inhibitor (N(G)-nitro-L-arginine methyl ester) and soluble guanylate cyclase inhibitor (LY83583) significantly decreased calcitonin gene-related peptide-induced relaxation of the rat uterus during pregnancy; (5) calcitonin gene-related peptide increased the uterine nitric oxide production in pregnant rats, and this increase was obliterated in the presence of calcitonin gene-related peptide(8-37); and (6) calcitonin gene-related peptide receptors are present in rat uterus, and the concentration of these receptors dramatically increases during pregnancy and decreases during labor at term. CONCLUSIONS: Calcitonin gene-related peptide inhibits uterine spontaneous contractions in rats during pregnancy but not during labor and post partum. The inhibitory effects of calcitonin gene-related peptide on uterine contractility appear to be modulated, at least in part, by the activation of nitric oxide generation in the rat uterus. Changes in calcitonin gene-related peptide receptors could contribute to the changes in calcitonin gene-related peptide-mediated uterine relaxation during pregnancy and labor.

Aminoquinolines↗
PubMedDate not supplied

Effects of cerebral lesions on binding sites for calcitonin and calcitonin gene-related peptide in the rat nucleus accumbens and ventral tegmental area.

Binding site densities of [125I]-labelled salmon calcitonin and human calcitonin gene-related peptide were investigated in the rat nucleus accumbens and ventral tegmental area by means of quantitative autoradiography following selective brain lesions. [125I]salmon calcitonin and [125I]human calcitonin gene-related peptide binding sites were highly concentrated in the accumbens, whereas the ventral tegmental area only contained [125I]salmon calcitonin binding sites. Unilateral injection of 6-hydroxydopamine into the ventral tegmental area did not alter [125I]salmon calcitonin and [125I]human calcitonin gene-related peptide binding site densities in the ipsilateral accumbens, while it produced a significant decrease in [125I]salmon calcitonin binding sites in the lesioned ventral tegmental area (-50%). In contrast, following unilateral injection of quinolinic acid into the accumbens, the densities of [125I]salmon calcitonin and [125I]human calcitonin gene-related peptide binding sites were significantly decreased in the lesioned accumbens (-57% and -56%, respectively), while [125I]salmon calcitonin binding site densities were not modified in the ipsilateral ventral tegmental area. The present study clearly suggests that [125I]salmon calcitonin and [125I]human calcitonin gene-related peptide binding sites are located on intrinsic neurons but not on the dopaminergic nerve terminals in the accumbens. Moreover, a certain proportion of [125I]salmon calcitonin binding sites could be present on dopaminergic cell bodies in the ventral tegmental area.

Animals↗
PubMed1985

Calcitonin and calcitonin gene-related peptide interact with the same receptor in cultured LLC-PK1 kidney cells.

Calcitonin and calcitonin gene-related peptide stimulate adenylate cyclase activity and plasminogen activator production in cultured renal tubular LLC-PK1 cells. Salmon [125I]calcitonin and human [125I]calcitonin gene-related peptide bound specifically to the cells. Salmon [125I]calcitonin binding was reduced at lower concentrations of non-radioactive salmon calcitonin than of human calcitonin gene-related peptide. For the stimulation of adenylate cyclase activity and plasminogen activator production, the potency of salmon calcitonin was higher than that of human calcitonin and calcitonin gene-related peptide. In a subclone of LLC-PK cells lacking salmon calcitonin binding sites, no specific binding of [125I]CGRP occurred, and adenylate cyclase activity and plasminogen activator production was not increased by the peptides. Thus, in LLC-PK1 cells the stimulation of adenylate cyclase activity and plasminogen activator production by calcitonin gene-related peptide is probably mediated by the calcitonin receptor.

Adenylyl Cyclases↗
PubMed1989

Calcitonin (but not calcitonin gene-related peptide) increases mouse bone cell proliferation in a dose-dependent manner, and increases mouse bone formation, alone and in combination with fluoride.

Previous in vitro studies have shown that salmon calcitonin had direct effects to increase parameters associated with embryonic chicken bone formation and to increase mouse and chicken osteoblast-line cell proliferation. The current studies demonstrate increased cell proliferation (i.e., [3H]-thymidine incorporation into DNA and tetrazolium salt reduction/deposition) in the osteoblastic murine cell line MC-3T3-E1 in response to salmon calcitonin (P less than 0.005) and to human calcitonin (P less than 0.005), but not to human calcitonin gene-related peptide. The current studies also show that salmon calcitonin increased several indices of murine bone formation. We found that 72 hours of exposure to salmon calcitonin [at 5 mU/ml-about 0.37 nM; mU/ml = milliunits of calcitonin activity/ml incubation medium (at 4,000 U/mg protein)] increased net 45Ca deposition (121% of control, P less than 0.05), net [3H]-proline incorporation 149% of control, P less than 0.001), and alkaline phosphatase activity (146% of control, P less than 0.01), in neonatal mouse half-calvaria. The calcitonin-dependent increase in alkaline phosphatase activity was not affected by co-incubation with 1 nM parathyroid hormone. Co-incubation with fluoride (which also increased net [3H]-proline incorporation and alkaline phosphatase activity in neonatal mouse half-calvaria, P less than 0.05, for each) enhanced the osteogenic response to low-dose calcitonin, (i.e., co-incubation with fluoride shifted the biphasic calcitonin dose-response curve to a range of lower calcitonin concentrations). The calcitonin-fluoride combinations had proportional effects on net [3H]-proline incorporation and alkaline phosphatase in the treated mouse calvaria (r = 0.78, P less than 0.005).

Alkaline Phosphatase↗
PubMed2003

Regulation of osteoclasts by calcitonin and amphiphilic calcitonin conjugates: role of cytosolic calcium.

The peptide hormone calcitonin is a potent inhibitor of osteoclastic resorption, but it is unstable and poorly absorbed following oral administration. Conjugates of salmon calcitonin covalently linked to low-molecular-weight amphiphilic polymers show improved stability and absorption. The purpose of this study was to investigate the biological activity of these conjugates in vitro using rat osteoclasts and HEK-293 cells transfected with the C1a isoform of the calcitonin receptor. Salmon calcitonin or its conjugates (10 pM-10 nM) caused rapid arrest of osteoclast membrane ruffling and subsequent retraction. The same amphiphilic polymer attached to an unrelated protein had no effect on osteoclast morphology or motility. Since calcitonin-induced retraction of osteoclasts is thought to be mediated by Ca2+ signaling, we investigated the effects of calcitonin and its conjugates on cytosolic free Ca2+ concentration ([Ca24]i). In HEK-293 cells transfected with the calcitonin receptor, these agents induced transient elevations of [Ca2+]i. However, the rise of [Ca2+]i in HEK-293 cells occurred at concentrations 100-1000-fold higher than those required to elicit osteoclast retraction. To investigate the role of Ca2+ in osteoclast retraction, we preloaded cells with BAPTA to buffer changes in [Ca2+]i. BAPTA decreased the initial rate of calcitonin-induced osteoclast retraction, but it did not affect the degree of retraction 2-3 hours following calcitonin, indicating that retraction is mediated primarily by Ca(2+)-independent processes. We conclude that calcitonin conjugates cause osteoclast retraction and [Ca2+]i signaling in a manner similar to that elicited by calcitonin. Thus, orally bioavailable calcitonin conjugates show potential for use as antiresorptive agents.

Animals↗
PubMed1995

Downregulation of calcitonin receptor mRNA expression by calcitonin during human osteoclast-like cell differentiation.

Calcitonin inhibits both osteoclast formation and bone resorption, and is a primary treatment for patients with hypercalcemia and increased bone turnover. However, the clinical utility of calcitonin is limited because patients become refractory to calcitonin after several days (the calcitonin "escape phenomenon"). The molecular basis for calcitonin "escape" is unclear. To determine the regulatory mechanisms controlling calcitonin receptor (CTR) expression in osteoclasts and their precursors, we treated immature mononuclear precursors for human osteoclast-like multinucleated cells (MNC) formed in vitro with 1,25-(OH)2D3, to induce their differentiation to committed mononuclear precursors, and mature multinucleated osteoclasts, and used reverse transcriptase (RT)-PCR to assess expression of CTR mRNA in both committed mononuclear precursors and MNC. The PCR fragment produced was cloned and sequenced to confirm that it was derived from CTR mRNA. CTR mRNA expression was detected in mononuclear MNC precursors after 7 d of 1,25-(OH)2D3 treatment. It was also present in osteoclast-like MNC and highly purified giant cells from osteoclastomas, but not in monocytes or macrophage polykaryons formed in vitro. Calcitonin markedly decreased CTR but not actin mRNA expression in giant cells and MNC after 12 h, and removal of calcitonin restored CTR mRNA expression. Similarly, calcitonin decreased calcitonin-induced adenylate cyclase activity. These data suggest: (a) downregulation of CTR gene expression by calcitonin may in part explain the calcitonin "escape phenomenon"; and (b) expression of CTR mRNA occurs in mononuclear osteoclast precursors within 7 d after exposure to 1,25-(OH)2D3.

Base Sequence↗