Does salmon calcitonin cause cancer? A review and meta-analysis.

Recently an association between the use of calcitonin and cancer has been postulated. We reviewed the biological rationale and performed an additional analysis of historical data with respect to the possibility. An association cannot be excluded, but the relationship is weak and causality is unlikely. The purpose of the present study is to review the strength of association and likelihood of a causal relationship between use of calcitonin and cancer. We reviewed the evidence for this association, including the molecular signaling mechanisms of calcitonin, preclinical data, an "experiment of nature," and the results of a previous meta-analysis which showed a weak association. We performed an additional meta-analysis to incorporate the data from a novel investigational oral formulation of salmon calcitonin. Review of the literature did not identify a cellular signaling mechanism of action which might account for a causal relationship or toxicologic or postmarketing data to support the thesis. Additional clinical results incorporated into previous meta-analyses weakened but did not completely negate the possibility of association. A causal association between calcitonin use and malignancy is unlikely, as there is little biological plausibility. The preponderance of nonclinical and clinical evidence also does not favor a causal relationship.

Efficacy and safety of oral recombinant calcitonin tablets in postmenopausal women with low bone mass and increased fracture risk: a randomized, placebo-controlled trial.

UNLABELLED: The effect of an investigational oral calcitonin tablet upon bone mineral density (BMD) of the spine was investigated in postmenopausal women with low bone mass and at increased risk of fracture. Compared to placebo, calcitonin tablets increased lumbar spine BMD. This agent may provide an additional choice for patients. INTRODUCTION: An investigational oral salmon calcitonin preparation was previously shown to increase lumbar spine BMD in postmenopausal women with osteoporosis. Our objective was to evaluate the use of this agent in postmenopausal women with low bone mass and at increased fracture risk but not meeting BMD criteria for osteoporosis. METHODS: Treatment-naïve women were randomized to receive oral recombinant salmon calcitonin tablets or placebo once daily for 1 year. Dual-energy X-ray absorptiometry was performed at baseline and at study weeks 28 and 54. CTx-1, a bone resorption marker, was obtained at the same time points. Subjects returned periodically for tolerability assessment and adverse event (AE) recording. RESULTS: One hundred twenty-nine women in the USA were randomized, 86 to calcitonin and 43 to placebo. Calcitonin recipients experienced a significant increase from baseline in lumbar spine BMD; the difference compared with placebo was significant. Dosing at bedtime or with dinner was equally effective. CTx-1 was suppressed in calcitonin recipients but not in placebo subjects. Gastrointestinal AEs were common, but the overall safety profile was comparable between groups. CONCLUSIONS: Oral calcitonin may provide a useful therapeutic alternative for some women with low bone mass.

Short-term aluminum administration in the rat. Effects on bone formation and relationship to renal osteomalacia.

Aluminum may be pathogenic in the osteomalacia observed in some patients receiving hemodialysis. To study the early effects of Al on bone growth, bone formation, mineralization, and resorption were measured during short-term Al exposure in the tibial cortex of pair-fed control (C, n = 10), aluminum-treated (AL, n = 9), subtotally nephrectomized control (NX-C, n = 7), and subtotally nephrectomized aluminum-treated (NX-AL, n = 8) rats using double tetracycline labeling of bone. Animals received 2 mg/d of elemental Al intraperitoneally for 5 d/wk over 4 wk. Total bone and matrix (osteoid) formation, periosteal bone and matrix formation, and periosteal bone and matrix apposition fell by 20% in AL from C, P less than 0.05 for all values, and by 40% in NX-AL from NX-C, P less than 0.01 for all values. Moreover, each measurement was significantly less in NX-AL than in AL, P less than 0.05 for all values. Osteoid width did not increase following aluminum administration in either AL or NX-AL. Resorption surface increased from control values in both AL and NX-AL; also, resorptive activity at the endosteum was greater in NX-AL than in NX-C, P less than 0.05. Thus, aluminum impairs new bone and matrix formation but does not cause classic osteomalacia in the cortical bone of rats whether renal function is normal or reduced. These findings may represent either a different response to aluminum administration in cortical bone as contrasted to trabecular bone or an early phase in the development of osteomalacia. Aluminum may increase bone resorption and contribute to osteopenia in clinical states associated with aluminum accumulation in bone.

Dose effect of gestational ethanol exposure on placentation and fetal growth.

INTRODUCTION: Prenatal ethanol exposure compromises fetal growth by impairing placentation. Invasive trophoblastic cells, which mediate placentation, express the insulin-IGF regulated gene, aspartyl-asparaginyl ß-hydroxylase (ASPH), which has a critical role in cell motility and invasion. The aims of this study were to characterize effects of ethanol on trophoblastic cell motility, and assess ethanol dose-dependent impairments in placentation and fetal development. METHODS: Pregnant Long Evans dams were fed with isocaloric liquid diets containing 0%, 8%, 18% or 37% ethanol (caloric content) from gestation day (GD) 6 to GD18. Fetal development, placental morphology, density of invasive trophoblasts at the mesometrial triangle, as well as placental and mesometrial ASPH and Notch-1 protein expression were evaluated. Directional motility of control and ethanol-exposed HTR-8/SVneo cells was assessed by ATP Luminescence-Based assay. RESULTS: Severity of fetal growth impairment correlated with increasing doses of ethanol. Ethanol exposure produced dose-dependent alterations in branching morphogenesis at the labyrinthine zone, and inhibited physiological transformation of maternal arteries. ASPH and Notch-1 protein expression levels were reduced, corresponding with impairments in placentation. DISCUSSION: Prenatal ethanol exposure compromises fetal growth and placentation in a dose-responsive manner. Ethanol's adverse effects on placental development are mediated by: (1) altered branching morphogenesis in labyrinthine zone; (2) suppression of invasive trophoblastic precursor cells; and (3) inhibition of trophoblastic cell adhesion and motility, corresponding with reduced ASPH and Notch-1 protein expression.

Targeted expression of calcitonin gene-related peptide to osteoblasts increases bone density in mice.

The neuropeptide calcitonin gene-related peptide (CGRP) is concentrated in fine sensory nerve endings innervating all tissues, including bone. CGRP inhibits osteoclasts, stimulates insulin-like growth factor I and inhibits tumor necrosis factor alpha production by osteoblasts in vitro. To investigate the role of CGRP in bone in vivo, mice were engineered to express CGRP in osteoblasts by placing the human CGRP gene under the control of the rat osteocalcin promoter (Ost-CGRP tg+ mice). Calvaria cultures from transgene positive (tg+), but not tg- mice, produced bioactive CGRP. Trabecular bone density and bone volume, determined by peripheral quantitative computed tomography and bone histomorphometry, respectively, were higher in tg+ than tg- littermates. This increase in bone volume was associated with an increased bone formation rate. Trabecular bone density decreased in tg+ mice as a result of ovariectomy, but remained higher than in sham tg- mice. Targeting CGRP to osteoblasts appears to favor the establishment of a higher trabecular bone mass in mice.

Multiple forms of peptidyl alpha-amidating enzyme: purification from rat medullary thyroid carcinoma CA-77 cell-conditioned medium.

Conditioned medium from cultures of rat medullary thyroid carcinoma CA-77 cells was used as a source for purification of the secreted forms of peptidyl alpha-amidating enzyme. The alpha-amidating enzyme activity was partially purified using a combination of weak anion exchange and gel filtration chromatography. Subsequent strong anion exchange chromatography at pH 6.0 resolved this partially pure enzyme into four distinct peaks of activity, termed Ia, Ib, II, and III. Peaks Ia and Ib exhibited broad pH optima between pH 6.0-8.5, whereas peaks II and III both exhibited pH optima at approximately pH 5.0. The peak III activity was further purified to electrophoretic homogeneity using hydrophobic interactive chromatography followed by strong anion exchange chromatography at pH 8.0. The enzyme exhibited an apparent molecular mass of 75K, a pH optimum of approximately pH 5.0, and a maximal turnover number of 580 min-1 in the presence of L-ascorbate. Kinetic studies demonstrated that the enzyme probably functions through a ping-pong mechanism with respect to the binding of the glycine-extended peptide substrate and the L-ascorbate cofactor. The peak III enzyme exhibits several distinctive characteristics compared to amidating enzymes isolated and characterized by other laboratories.

Synthesis of collagen in cardiac and vascular walls.

Collagen synthesis and content have been shown to be elevated in the arterial wall and heart of hypertensive animals. These increases in collagen synthesis and content have been implicated in the maintenance of raised blood pressure. Inhibitors of collagen synthesis, beta APN and 3,4-dihydroproline, were shown to delay the onset of hypertension and reverse established hypertension. Therapeutic intervention with antihypertensive drugs resulted in a decrease in arterial collagen synthesis and a reversal of cardiac hypertrophy. In some instances (i.e., DOCA-salt), chronic administration of antihypertensive drugs caused a restructuring of the arterial wall and resulted in an apparent remission of the hypertensive disease.

Endothelin-I-mediated vasoconstriction: specific blockade by verapamil.

The capacity of three vasodilators that act by distinct mechanisms to reverse endothelin-I-mediated vasoconstriction was studied in 11 healthy nonsmoking male subjects (mean age +/- SEM, 26 +/- 2 years; mean weight +/- SEM, 74 +/- 2 kg) by use of brachial artery infusion and forearm strain-gauge plethysmography. Isoproterenol (cyclic adenosine monophosphate-mediated vasodilation), sodium nitroprusside (cyclic guanosine monophosphate-mediated vasodilation), and verapamil (L-type calcium channel blocker) were compared for capacity to reverse endothelin-I-mediated increase in forearm vascular resistance (FVR). Endothelin-I infusion increased FVR 1.9-fold in the control state. Isoproterenol infusion decreased FVR with or without concurrent endothelin-I infusion; however, at comparable isoproterenol infusion rates, endothelin-I increased FVR similar to the control state (for 5 ng/min isoproterenol, endothelin-I increased FVR 1.85-fold; for 12.5 ng/min isoproterenol, endothelin-I increased FVR 2.03-fold). Similarly, sodium nitroprusside infusion decreased FVR with or without concurrent endothelin-I infusion; however, at comparable sodium nitroprusside infusion rates the endothelin-I increase in FVR was similar to control (for 0.48 micrograms/min sodium nitroprusside, endothelin-I increased FVR 1.89-fold; for 0.96 micrograms/min sodium nitroprusside, endothelin-I increased FVR 2.36-fold). In contrast, verapamil infusion decreased FVR with or without endothelin-I infusion. At a verapamil infusion rate of 19.1 microns/min, endothelin-I increase in FVR was comparable to control (for 19.1 microns/min verapamil, endothelin-I increased FVR 1.36-fold, less than the 1.0-fold increase in the control state; p < 0.05). Isoproterenol and sodium nitroprusside decreased FVR during concurrent endothelin-I infusion but did not reverse the endothelin-I effect. In contrast, verapamil reversed endothelin-I--induced vasoconstriction to control FVR, suggesting a specific antagonism of endothelin-I--mediated increase in FVR.

Hypercalcemia associated with parenteral amino acid and dextrose infusion.

A patient receiving parenteral nutrition with an amino acid dextrose solution developed hypercalcemia which seemed related to the rate of nutrient infusion. In a retrospective study of patients receiving parenteral nutrition over a 3-month period six of 72 (8%) developed hypercalcemia. After changes in infusion rate there were corresponding changes in the serum calcium concentration. There was a significant correlation between the serum calcium concentration and the average infusion rate over the preceding four days (p = 0.012). This was even more significant (p less than 0.005) when ionized calcium was calculated to diminish the effects of calcium binding by proteins.

Calcitonin for osteoporosis and bone pain.

Calcitonin has been approved for the treatment of osteoporosis and other diseases involving accelerated bone turnover for approximately 25 years. The most commonly studied and prescribed form is salmon calcitonin, which has a greater efficacy in clinical use. A wealth of well-controlled clinical studies have demonstrated that calcitonin preserves or increases bone mineral density (BMD) and reduces the risk of vertebral fractures in osteoporosis. Recent studies have indicated that while a low BMD is correlated with an increase in fracture risk, increases in BMD alone do not explain the antifracture efficacy of antiresorptive therapies such as calcitonin. Therapies that moderately increase BMD may reduce fracture risk by reducing the rate of bone turnover and maintaining the integrity of the trabecular architecture, resulting in the preservation of bone strength and quality in osteoporotic patients. An advantage of calcitonin that is not shared by other antiresorptive therapies is its direct analgesic effect on bone pain. Calcitonin has been demonstrated to be clinically useful in improving pain, not only from the acute vertebral fractures of osteoporosis, but also in Paget's disease, bone malignancies, and other sources of musculoskeletal pain. Drugs containing calcitonin may be approved for additional indications in the near future, and as more convenient routes of administration such as the oral route become available, the demand for the calcitonin peptide is expected to increase.

Effects of calcitonin gene-related peptide on bone turnover in ovariectomized rats.

Calcitonin gene-related peptide (CGRP) is a neuropeptide abundantly concentrated in sensory nerve endings innervating bone metaphysis and periosteum, which indicates that it plays a local role in bone metabolism. CGRP-alpha and -beta share structural and functional homology with calcitonin (CT) and have been shown to inhibit bone resorption in vitro and to induce hypocalcemia in vivo. We recently reported that CGRP stimulates the production of the growth factor insulin-like growth factor-I and inhibits that of the cytokine tumor necrosis factor-alpha by osteoblasts, suggesting that CGRP may control bone cell activity. To investigate this possibility, we used ovariectomized (ovx) rats as a high bone turnover model and compared the effects of CGRP to those of CT. ovx young female rats were injected daily starting the day after surgery with either phosphate-buffered saline, CGRP-alpha (1.15 mg/kg per day), or CT (3 micrograms/kg per day) for 28 days. Ovariectomy induced an increase in bone turnover associated with a 60% loss in trabecular bone volume of the proximal tibia. CGRP inhibited bone resorption but not bone formation, and was nevertheless less efficient than CT in preventing bone loss, since CGRP-treated rats had a loss of 46% of cancellous bone, whereas CT-treated rats had a loss of 21%. This suggests that CGRP is either less potent than CT at inhibiting bone resorption or else very rapidly degraded. These data indicate that CGRP can control bone cells through a mechanism that is in part different from that of CT, and further suggest that CGRP may play a local role in bone metabolism.

Effect of removal of atherogenic diet on protein synthesis and cholesterol retention in rabbit aorta and lung.

The purpose of these studies was to determine the effect of cholesterol feeding in rabbits on the synthesis of collagen and non-collagen proteins in both aortic and lung tissues. Rabbits were fed a 2% cholesterol diet for 30 or 60 days, followed by 30 days of a low cholesterol diet (i.e. 30-30 or 60-30). After 30 days of cholesterol feeding non-collagen protein synthetic rates were significantly elevated in aortic tissues, but not in the lung. After 60 days of cholesterol feeding, both collagen and non-collagen synthetic rates were elevated in the aorta but not in the lung. Both tissues demonstrated significant increases in cholesterol content. When cholesterol was removed from the diet, cholesterol continued to accumulate in the aorta but decreased in the lung. The 60-30 group demonstrated both the largest increase in aortic cholesterol, and the largest increase in the per cent of collagen being synthesized in the aorta. These data therefore demonstrate that cholesterol feeding stimulates both collagen and non-collagen protein synthesis and suggests that there may be some differences in the lag phase before significant changes are apparent in both parameters. Following removal of cholesterol from the diet the per cent collagen synthesized in the aorta increased further, due to an apparent reduction in non-collagen protein synthesis rather than a further acceleration of collagen synthesis. These changes may be important in explaining how intermittent-cholesterol feeding produces a more fibrous aortic lesion.

Utility of pharmacodynamic measures for assessing the oral bioavailability of peptides. 1. Administration of recombinant salmon calcitonin in rats.

Salmon calcitonin (sCT) is a therapeutic peptide used in the treatment of Paget's Disease, postmenopausal osteoporosis, and hypercalcemia due to malignancy. In this study, recombinant sCT (rsCT) was administered intravenously (iv), subcutaneously (sc), and intraduodenally (id.) in rats to evaluate pharmacodynamic (PD) response as a measure of rsCT bioavailability (F) and to test the feasibility of delivering rsCT orally. rsCT pharmacokinetics were linear throughout the range of iv and sc doses studied. Following sc administration, F ranged from 11.2% to 23.1% and was linear. The absorption of rsCT after id. administration was low (0.022%); however, a significant lowering of serum calcium concentrations was observed. Serum calcium lowering was nonlinear and saturable after sc administration with the minimum dose required for maximum calcium lowering (Dmin/max) equal to 10.2 ng and a maximal response of 426.8 mg min/dL. Using Dmin/max as the reference dose, absolute Fs were recalculated using PD response after id. administration of 1 and 2 mg of rsCT and were 0.040% and 0.029%, respectively. Substantial overestimates of F were obtained when the reference dose was not properly selected. While the absorption of rsCT was low, the significant lowering of serum calcium levels suggests that oral delivery of sCT is feasible. The results of these studies also suggest that PD response is useful in assessing the oral bioavailability of peptides; however, when PD response is saturable, as is the case for rsCT, the reference dose should be carefully selected in order to avoid overestimates of F.

Modulation of carrageenan-induced hind paw edema by substance P.

Substance P has been implicated as a mediator of inflammation. The involvement of this neuropeptide in carrageenan-induced hind paw edema in the rat was assessed. Subcutaneous injection of carrageenan into the rat paw caused a significant increase in substance P levels, which preceded the onset of inflammation. While injection of substance P alone caused mild edema, coadministration of submaximal doses of carrageenan and substance P resulted in a synergistic exacerbation in the degree of inflammation. This synergistic response was not detected when the nonamidated precursor of substance P was coinjected with carrageenan. The effects of substance P depletion on inflammation were also evaluated. In animals pretreated with capsaicin followed by injection with carrageenan, no significant increase in either the levels of substance P or the extent of edema was observed when compared to capsaicin-treated controls. These results indicate that substance P may play an important role in the early stages of carrageenan-induced paw edema and that a reduction in the biosynthesis of substance P may lessen the severity of this inflammatory response.

Risperidone in chronic schizophrenia: a detailed audit, open switch study and two-year follow-up of patients on depot medication.

Little information exists on the medium- to long-term outcome of switching patients with schizophrenia from traditional depot to atypical oral antipsychotic agents. By detailed clinical audit, we identified a representative group of 102 patients of an Irish psychiatric service with DSM-IV chronic schizophrenia and on depot neuroleptics for a mean of 15 years. Of 69 eligible to participate, 33 entered a 6-month switch study of risperidone, with limited follow-up of consenters and non-consenters at 1 and 2 years. At 6 months, 23 of 33 were still on risperidone and had small significant improvements in clinical and extrapyramidal side effects, QOL and adjunct medication measures over baseline. At 12 months, 19 of 33 were still on risperidone, reducing to 13 of 33 at 2 years. At 2 years, of 32 surviving consenters to switch, 19 had suffered clinically detrimental events and were no longer on risperidone, compared to none of the 33 surviving non-consenters, who were all still on depot. These findings suggest that switching from depot to risperidone may encounter high rates of refusal and attrition subsequent to switch. While a majority of switched patients may improve to least 6 months, audit plus switch may have clinically unfavourable effects on others over a 2-year follow-up period [corrected].

Camel racing: a new cause of extradural haemorrhage in Australia.

Camel racing is a relatively new sport in Australia. A 52 year old woman fell from her camel during a country race. Although she was wearing an approved equestrian helmet, she suffered a skull fracture and a life-threatening extradural haematoma. Her treatment highlights the key issues of management of head injuries in remote places. A paramount requirement is close collaboration between country medical practitioner, neurosurgeon and retrieval specialist.

Square pegs in round holes: has psychometric testing a place in choosing a surgical career? A preliminary report of work in progress.

Methods of selection of candidates for training in surgery has long been regarded as lacking explicit criteria and objectivity. Our purpose was to discover the aptitudes and personality types of applicants for surgical posts at the outset, in order to discover which were most likely to result in a satisfactory progression through training and which were associated with career difficulties. This longitudinal predictive validation study has been undertaken in a London Teaching Hospital since 1994. After short-listing, but immediately before interview, all candidates for senior house officer posts in basic surgical training and in geriatric medicine were asked to undertake psychometric tests of numerical (GMA) and spatial (SIT7) reasoning, personality type (MBTI), and self-rating of competency. There were no differences in ability scores between surgeons or geriatricians. Personality differences were revealed between the surgeons and the geriatricians, and between male and female surgeons. This study suggests that while there are no differences in ability between surgeons and geriatricians at the start of training, there are differences in personality. Long-term follow-up of the career development of this cohort of surgical SHOs is required to determine whether the psychometric measures described correlate with achievements of milestones in their surgical careers.

Production of recombinant salmon calcitonin by in vitro amidation of an Escherichia coli produced precursor peptide.

Salmon calcitonin (sCT) is a 32 amino acid peptide hormone that requires C-terminal amidation for full biological activity. We have produced salmon calcitonin by in vitro amidation of an E. coli produced precursor peptide. Glycine-extended sCT, the substrate for amidation, was produced in recombinant E. coli as part of a fusion with glutathione-S-transferase. The microbially produced soluble fusion protein was purified to near homogeneity by affinity chromatography. Following S-sulfonation of the fusion protein, the glycine-extended peptide was cleaved from the fusion by cyanogen bromide. The S-sulfonated peptide was recovered and enzymatically converted to the amidated peptide in a reaction with recombinant peptidylglycine alpha-amidating enzyme (alpha-AE) secreted from Chinese hamster ovary (CHO) cells. After reformation of the intramolecular disulfide bond, the sCT was purified with a step yield of 60%. The ease and speed of this recombinant process, as well as its potential for scale-up, make it adaptable to production demands for calcitonin, a proven useful agent for the treatment of post-menopausal osteoporosis. Moreover, the relaxed specificity of the recombinant alpha-AE for the penultimate amino acid which is amidated allows the basic process to be applied to the production of other amidated peptides.

Precision-cut slice culture method for rat placenta.

Primary trophoblasts, placental explants, and cell line cultures are commonly used to investigate placental development, physiology, and pathology, particularly in relation to pregnancy outcomes. Organotypic slice cultures are increasingly used in other systems because they maintain the normal three-dimensional tissue architecture and have all cell types represented. Herein, we demonstrate the utility of the precision-cut placental slice culture model for studying trophoblastic diseases.