Calcitonin

Calcitonin, a peptide hormone of 32 amino acids having a regulatory function in calcium and phosphoms metabolism, is used in various bone disorders such as osteoporosis. Salmon, human, pig and eel calcitonin are used therapeutically. Species differences may be significant - salmon calcitonin is ten times more potent than human calcitonin, for example. Human calcitonin (hCT) has a tendency to associate rapidly in solution and, like insulin, form fibrils, resulting in a viscous solution. The fibrils are 8 nm in diameter and often associate with one another. Heating fibrillated hCT solutions in 50% acetic acid-water converts the system back to soluble monomers. hCT has a pK of 8.7, while salmon calcitonin (SCT) has a pK of 10.4. This accounts for the high stability of SCT at pH 7.4, as electrostatic interactions between calcitonin monomers play a role. At pH 7.4, SCT monomers will be charged and will repel each other. At 20 mg moH in Tris buffer at 22°C and pH 7.4 the fibrillation time for hCT is 1 minute, whereas for SCT it is of the order of 21 days. 

Calcitonin is secreted continuously under conditions of normocalcemia, and the synthesis of calcitonin is increased when the calcium concentrations in plasma and intracellular fluids increase. Hypermagnesemia has a similar effect on calcitonin production. In hypocalcemia, the production of calcitonin falls. The gastrointestinal hormones—gastrin, glucagon, cholecystokinin, and secretin—and high dietary calcium also stimulate calcitonin production. Long-term hypercalcemia may cause hyperplasia of the C cells. 

Calcitonin acts on receptors in bone osteoclasts with a resulting reduction of bone resorption, and it also acts on the renal tubular reabsorption of phosphate. The phosphaturic effects are accompanied by diuresis and increased excretion of other electrolytes. Calcitonin and parathyroid hormone act as a dual negative feedback mechanism in controlling calcium in intra- and extracellular fluids. The range of calcitonin assays suitable for laboratory animals is limited, but the hormone can be measured by two-site radioimmunometric assays (Moukhtar et al. 2005). 

Comparative vertebrate endocrinology, 3rd ed. Cambridge Cambridge University Press. 

Braverman, L. E., and R. D. Utiger, eds. 2004. Werner and Ingbar s the thyroid A fundamental and clinical text, 9th ed. Philadelphia Lippincott-Williams Wilkins. 

Connors, J. M. 1997. Physiology of the thyroid gland and agents affecting its secretion. In Endocrine toxicology. Target Organ series, ed. J. A. Thomas, pp. 43-68. Boca Raton, FL CRC Press. 

3 Calcitonin Calcitonin is secreted by the C cells of the thyroid gland in response to hypercalcemia. Its primary action is to oppose the actions of penathyroid hormone by suppressing osteoclast actions. It also stimulates 

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E. Flanigan, Fligh Performance Eiquid Chromatography in the Production and Quality Control of Salmon Calcitonin, Purdue University, West Lafayette, Ind., 1991, p. 207. 

CGRP has a wide distribution in the nervous system (19) and was the first peptide to be localized to motoneurons (124). It is also found in primary sensory neurons where it is colocalized with substance P (125). CGRP is derived from a precursor stmcturaHy related to the calcitonin precursor. The latter precursor produces two products, calcitonin itself and katacalcin, while the CGRP precursor produces one copy of CGRP (123). Like other peptides, CGRP is cleaved from its precursor by tryptic breakdown between double basic amino acid residues. 

Factors controlling calcium homeostasis are calcitonin, parathyroid hormone(PTH), and a vitamin D metabolite. Calcitonin, a polypeptide of 32 amino acid residues, mol wt - SGOO, is synthesized by the thyroid gland. Release is stimulated by small increases in blood Ca " concentration. The sites of action of calcitonin are the bones and kidneys. Calcitonin increases bone calcification, thereby inhibiting resorption. In the kidney, it inhibits Ca " reabsorption and increases Ca " excretion in urine. Calcitonin operates via a cyclic adenosine monophosphate (cAMP) mechanism. 

One method of treatment is to inject calcitonin, which decreases blood Ca " concentration and increases bone calcification (33). Another is to increase the release of calcitonin into the blood by increasing the blood level of Ca " ( 4). This latter treatment is accompHshed by increasing Ca " absorption from the intestine requiring dietary calcium supplements and avoidance of high phosphate diets. The latter decrease Ca " absorption by precipitation of the insoluble calcium phosphate. 

Other Calcium Disorders. In addition to hypocalcemia, tremors, osteoporosis, and muscle spasms (tetary), calcium deficiency can lead to rickets, osteomalacia, and possibly heart disease. These, as well as Paget s disease, can also result from faulty utilization of calcium. Calcium excess can lead to excess secretion of calcitonin, possible calcification of soft tissues, and kidney stones when combined with magnesium deficiency. 

Calcitonin Gene-Related Peptide. Calcitonia gene-related peptide (CGRP) [83652-28-2] (74) is a 37-amino acid peptide (Fig. 4) (32). 

Capsaicin, an active ingredient in red pepper, is well known for its ability to release and deplete substance P in sensory C fibers. However, this action is not specific for substance P, as neurokinin A, calcitonin gene-related peptide (CGRP), and somatostatin also are released. 

The main role of the human thyroid gland is production of thyroid hormones (iodinated amino acids), essential for adequate growth, development, and energy metaboHsm (1 6). Thyroid underfunction is an occurrence that can be treated successfully with thyroid preparations. In addition, the thyroid secretes calcitonin (also known as thyrocalcitonin), a polypeptide that lowers excessively high calcium blood levels. Thyroid hyperfunction, another important clinical entity, can be corrected by treatment with a variety of substances known as antithyroid dmgs. 

Several years ago, it was discovered that the thyroid gland was also the source of a hypocalcemic hormone having effects in general opposition to those of the parathyroid hormone. This hormone is produced in mammals by the parafollicular C-ceUs and in other vertebrates by the ultimobrachial bodies (45). Originally called thyrocalcitonin, it is now referred to as calcitonin (CT). 

Calcitonins from several species have been characteri2ed and synthesi2ed. They are all single-chain 32-residue polypeptides (ca 3600 mol wt), although a disulfide link between the first and seventh cysteine residues results in a cycHc stmcture that is indispensable for activity (Eig. 6). 

Eig. 6. Amino acid sequence of human (H) and porcine (P) calcitonins. 

Calcitonin is secreted when abnormally high calcium levels occur in plasma. Although plasma concentrations are normally minute (<100 pg/mL), they increase two- to threefold after calcium infusion. Calcitonin has a short plasma half-life (ca 10 min). Certain thyroid tumors are the result of CT concentrations 50—500 times normal. The mechanism of action is a direct inhibition of bone resorption. Calcitonin is used clinically in various diseases in which hypercalcemia is present, eg, Paget s disease (46). 

Calcitonin. Calcitonin is available commercially from pork and salmon extracts (Calcimar, Armour) as well as by synthesis. Preparations are bioassayed on the basis of their calcium-lowering activity in comparison to the potency of pure pork calcitonin of which ca 4 p.g is equivalent to 1 MRC unit (Medical Research Council, U.K.). For clinical use, vials containing 400 units in 4 mL are available. The recommended daily dosage is 100 units to be adrninistered subcutaneously or intramuscularly because its plasma half-life is short (4—12 min). 

DHD = 24(R),25-dihydroxyvitamiQD2 250H = 25-hydroxyvitamia D PTH = parathyroid hormone CT = calcitonin F. = inorganic phosphoms. 

Vitamin D withdrawal is an obvious treatment for D toxicity (219). However, because of the 5—7 d half-life of plasma vitamin D and 20—30 d half-life of 25-hydroxy vitamin D, it may not be immediately successful. A prompt reduction in dietary calcium is also indicated to reduce hypercalcemia. Sodium phytate can aid in reducing intestinal calcium transport. Calcitonin glucagon and glucocorticoid therapy have also been reported to reduce semm calcium resulting from D intoxication (210). 

Three hormones regulate turnover of calcium in the body (22). 1,25-Dihydroxycholecalciferol is a steroid derivative made by the combined action of the skin, Hver, and kidneys, or furnished by dietary factors with vitamin D activity. The apparent action of this compound is to promote the transcription of genes for proteins that faciUtate transport of calcium and phosphate ions through the plasma membrane. Parathormone (PTH) is a polypeptide hormone secreted by the parathyroid gland, in response to a fall in extracellular Ca(Il). It acts on bones and kidneys in concert with 1,25-dihydroxycholecalciferol to stimulate resorption of bone and reabsorption of calcium from the glomerular filtrate. Calcitonin, the third hormone, is a polypeptide secreted by the thyroid gland in response to a rise in blood Ca(Il) concentration. Its production leads to an increase in bone deposition, increased loss of calcium and phosphate in the urine, and inhibition of the synthesis of 1,25-dihydroxycholecalciferol. 

Pha.rma.ceutica.ls. Neopentanoic acid derivatives are widely used in the preparation of pharmaceuticals, eg, as a means of introducing the tert-huty group into a molecule. More frequendy, however, derivatives have been prepared that exploit the enhanced hydrolytic stabiUty of the neopentanoate group. Eor example, when salmon calcitonin is treated with N-hydroxysuccinimide pivalate [42014-50-6], the resulting derivative retains the biological activity of the precursor, but gives an extended duration of activity (51). 

Figure 5.5 Trifluoroacetate determination in calcitonin acetate (a) without and (b) with heait-cut column switching. Reprinted from Journal of Chromatography, 602, S. R. Villasenor, Matrix elimination in ion cliromatography by heart-cut column switching techniques ,pp 155-161, copyright 1992, with permission from Elsevier Science.

The process for the manufacture of human calcitonin in pure form from C-cell rich medulla... 

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