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A PHYSIOLOGICAL ROLE FOR CALCITONIN: PROTECTION OF THE MATERNAL SKELETON
769 A PHYSIOLOGICAL ROLE FOR CALCITONIN: PROTECTION OF THE MATERNAL SKELETON
CARMEL IAIN MACINTYRE
JOHN C. STEVENSON Endocrine Unit,
J.
HILLYARD
Royal Postgraduate Medical School, London W12 0HS
HILARY COOPER
MALCOLM I. WHITEHEAD
Department of Obstetrics, King’s College Hospital Medical School, London SE5 8RX
Blood-samples were taken around midday from the women in these two groups in order to coincide with the known peak secretion of calcitonin.3 The third group consisted of 10 healthy lactating women (age 22-28 years). Blood-samples were taken immediately before and 30 min after the midday breast feed, thus minimising the influence of the circadian variation of calcitonin. These samples were taken on two occasions, the first on the seventh day after childbirth and the second at 6 weeks after delivery. All the women were lactating throughout the study. No drugs other than iron and folic acid during pregnancy were taken by any of the subjects. Methods
Summary
Plasma-calcitonin levels, measured with an established and reliable extraction
radioimmunoassay technique, were significantly higher throughout normal pregnancy and lactation than in normal non-pregnant women, and were not immediately influenced by the acute stimulation of breast-feeding. Thus, more calcitonin circulates at times of physiologically increased calcium need. It is suggested that an important function of calcitonin is the protection of the healthy maternal skeleton from excessive resorption by opposing the resorptive action of 1,25-dihydroxycholecalciferol on bone.
Introduction THE actions of calcitonin have been
clearly established,l but it is still not known whether the hormone has an important physiological function in man. One reason for our ignorance is that measurement is difficult : circulating calcitonin levels are extremely low, especially in women.2 Another problem is that the striking circadian variation in plasma levels3 is often ignored, although such an omission may invalidate any conclusions. However, by the correct timing of blood-sampling and by use of an extraction radioimmunoassay technique,3 we can reliably measure physiological levels of calcitonin at multiple plasma dilutions reading from the most sensitive part of the standard curve. We have applied our techniques to the study of calcitonin in normal women during pregnancy and lactation. Patients Three groups of healthy volunteers were studied. The first group consisted of 40 normal premenopausal women (age 17-43 years). The second group comprised 51 healthy pregnant women (age 17-34), 19 in the first trimester of pregnancy (6-12 weeks), 12 in the second trimester (13-28 weeks), and 20 in the third trimester (29-40 weeks).
12 Needleman P, Bronson SD, Wyche A, Sivakoff M, Nicolaou KC. Caridiac and renal prostaglandin I2. Biosynthesis and biological effects in isolated perfused rabbit tissues.J Clin Invest 1978; 61: 839-49.
13. Pace-Asciak, CR, Rangaray, G. Distribution of prostaglandin biosynthetic pathways in several rat tissues. Formation of 6-keto-prostaglandin F1&agr; Biochem Biophys Acta 1977; 486: 579-82. 14. Gryglewski RJ, Korbut R, Ocetkiewicz A. Generation of prostacyclin by lungs in vivo and its release into the arterial circulation. Nature 1978; 273: 765-67. 15. Moncada S, Korbut R, Bunting S, Vane JR. Prostacyclin is a circulating hormone. Nature 1978; 273: 767-68. 16. Hamberg M, Svensson J, Samuelsson B. Thromboxanes: a new group of biologically active compounds derived from prostaglandin endoperoxides. Proc Natn Acad Sci U.S.A. 1975; 72: 2994-98. 17. O’Regan S, Rivard GE, Robitaille PO. Aspirin improves platelet dysfunction in Bartter’s syndrome. Proceedings of the International Congress of Nephrology. Montreal: 1978; 1.
Calcitonin Measurements For each test, 25 ml of
venous
blood
was
taken from the
subject and placed in cooled heparinised tubes. The plasma was immediately separated and stored at -20°C until extraction and assay.3 Intra-assay and inter-assay variations were <10% and <14%, respectively. The sensitivity of the assay was 4-8 ng/1 with a 24 h incubation. For comparison, the sensitivity of the antiserum in a direct assay with a 7-day incubation period is approximately 2 pg/tube under optimum conditions.
StatisticalAnalyses Analysis of variance, two-tailed paired and unpaired Student’s t tests, and the Wilcoxon two-samples test were used as appropriate. Results The much
plasma calcitonin levels in pregnant women were, higher than those found in normal women (p <0.001) and were maintained throughout pregnancy;
the
mean
levels (ng/1±SEM) of 3745+_7.67, and for the first, second, and 41.79::!:6.41 55.19±11-02, third trimester, respectively, did not differ significantly from each other. Plasma-calcitonin levels were also raised during lactation (p<0.001); the increase was less striking but the levelsdid not differ significantly from those in pregnancy (figure). No significant difference was found between the mean levels before (23-02±2-51) and after (21.63±2.63) the midday breast feed, or at 1 week (2335+_2.49) and 6 weeks (21.30+_2.64) after delivery. There is a wide range of calcitonin levels in normal individuals, as has also been noted by Parthemore and Deftos;4 the reason is unknown. However, it does mean that there is some overlap between the levels found in the different groups. Discussion Our results show conclusively that plasma-calcitonin levels are raised throughout pregnancy. The physiological mechanisms responsible for this increase are not clear. Although there is evidence to suggest that oestrogen may be partly responsible,2,5 it can be seen from our results that the levels of calcitonin do not parallel those of oestrogen throughout pregnancy, so this cannot be the sole explanation. Previous reports of raised calcitonin levels in lactating animals6,7 accord well with our results in women. The mechanism is again unknown, but the acute stimulation of a breast feed appears to have no immediate effect on plasma calcitonin. It is unlikely that the calcitonin levels in lactating women represent a slow decline from pregnancy levels
770 Bone loss at that time could be both excessive and poorly tolerated because of an inadequate secretion of calcitonin and perhaps a slightly weakened skeleton. We thank Mr P. Needham for providing some of the samples obtained from pregnant patients. This work was supported in part by the Arthritis and Rheumatism Council and by the Medical Research Council. We thank Ciba-Geigy for synthetic human calcitonin. M.R.C. standard preparation 70/50 was donated by the National Institute for Biological Standards and Control. REFERENCES
TJ, Hunt NH, Ellison M, Underwood, JCE, Michelangeli VP. Physiology and biochemical pharmacology of calcitonin action. In: MacIntyre I, Szelke M, eds. Molecular endocrinology: proceedings of Endocrinology ’77. Amsterdam: Elsevier/North-Holland Biomedical Press, 1977.
1. Martin
179-92. Stevenson JC, MacIntyre I. Relative deficiency of plasma-calciin normal women. Lancet 1978; i: 961-62. 3. Hillyard, CJ, Cooke TJC, Coombes RC, Evans IMA, Maclntyre I. Normal Plasma calcitonin. Circadian variation and response to stimuli. Clin Endocr 1977; 6: 291-98. 4. Parthemore JG, Deftos LJ. Calcitonin secretion in normal human subjects. J Clin Endocr Metab 1978; 47: 184-88. 5. Delorme ML, Digioia Y, Fandard J, Merceron R, Raymond JY, Klotz HP. Oestrogenes et calcitonine. Ann Endocr 1976; 37: 503-04 6. Garel JM, Care AD, Barlet JP. A radioimmunoassay for ovine calcitonin an evaluation of calcitonin secretion during gestation, lactation and foetal
2.
Hillyard CJ, tonin
life. J Endocr 1974; 62: 497-509. 7. Toverud SU, Cooper CW, Munson PL. Calcium metabolism during lactation: elevated blood levels of calcitonin. Endocrinology 1978; 103:
472-79. 8. DeLuca HF. The
kidney as an endocrine organ for the production of 1,25-dihydroxyvitamin D3, a calcium-mobilizing hormone. N Eng J Med
1973; 289: 359-65. 9. Kumar R, Cohen WR, Silva P, Epstein FH. Elevated 1,25-dihydroxyvitamin-D plasma levels in normal human pregnancy and lactation. J Clin In-
Plasma-calcitonin levels in normal, pregnant and lactating women.
Horizontal bars are group means. Plasma-calcitonin levels were significantly higher in pregnant and lactating women than in normal women (p<0.001). because they appear to remain constant for up to 6 weeks during lactation. Our results are entirely consistent with the view that an important function of calcitonin is the protection of the skeleton against resorption.2In both pregnancy and lactation, there is an increased physiological demand on the mother for calcium. And this demand is normally satisfied by enhanced intestinal absorption of calcium produced by increased plasma levels of 1,25-dihydroxycholecalciferol (1,25[OH)2D3),8 the active hormonal form of vitamin D3’ Indeed, increased levels of this hormone are also found throughout all trimesters of pregnancy and during lactation,9 regardless of the variations in actual calcium need. It should be recalled that both 1,25(OH)2D3 and parathyroid hormone (PTH) are bone-resorbing hormones. However, 1,25(OH)2D3 causes bone resorption in physiological amounts, 10 whereas this may not be the case with PTH.11-13 Since it appears that increases in 1,25(OH)2D3 are accompanied by similar increases in calcitonin, we suggest that an important function of calcitonin in man is to oppose the action of 1,25(OH)2D3 on bone and thus protect the healthy maternal skeleton from excessive resorption during pregnancy and lactation. Some of the low levels in a number of the pregnant women may represent an inadequate response of calcitonin secretion to the increased 1,25(OH)2D3 levels in pregnancy. Thus, they may have increased skeletal resorption at this time compared with pregnant women with adequate calcitonin levels. While such bone loss would be insufficient to pose an immediate threat to the integrity of the skeleton, it may leave these women at some disadvantage when they reach the menopause.
10. 11.
vest 1979; 63: 342-44. Reynolds JJ, Holick MF, DeLuca HF. The role of vitamin D metabolites in bone resorption. Calcif Tiss Res 1973; 12: 295-301. Parsons JA. Parathyroid physiology and the skeleton. In: Bourne GH, ed. Biochemistry and physiology of bone. London: Academic Press, 1976; 4:
159-225. 12. Gaillard PJ, Wassenaar AM, Van Wijke-Wheeler ME. Effects of parathyroid hormone and a synthetic fragment (PTH 1-34) on bone in vitro. Proc Kon Ned Akad 1977; C80: 267-80. 13. Raisz LG, Canalis EM, Dietrich JW, Kream BE, Gworek SC. Hormonal regulation of bone formation. Recent Prog Horm Res 1978; 34: 335-56.
STRONG ASSOCIATION BETWEEN IDIOPATHIC MEMBRANOUS NEPHROPATHY AND HLA-DRW3.
J. MANOS P. A. DYER R. HARRIS N. P. MALLICK
P. T. KLOUDA E. J. ACHESON F. S. GOLDBY W. LAWLER
G. WILLIAMS
Department of Medical Genetics, St Mary’s Hospital, Manchester, Department of Renal Medicine, Royal Infirmary, and Department
of Pathology, University of Manchester
HLA A, B, and DR antigens were determined in a homogeneous group of patients with idiopathic membranous nephropathy. The frequency of HLA-DRw3 was significantly higher in the patients than in a control population. The frequencies of HLA B8 and B18, which are in linkage disequilibrium with DRw3, were also increased.
Summary
Introduction IDIOPATHIC membranous nephropathy (IMN) is associated clinically with moderate to heavy proteinuria. It is an important cause of the nephrotic syndrome and progressive renal failure. The histological diagnosis is
CARMEL IAIN MACINTYRE
JOHN C. STEVENSON Endocrine Unit,
J.
HILLYARD
Royal Postgraduate Medical School, London W12 0HS
HILARY COOPER
MALCOLM I. WHITEHEAD
Department of Obstetrics, King’s College Hospital Medical School, London SE5 8RX
Blood-samples were taken around midday from the women in these two groups in order to coincide with the known peak secretion of calcitonin.3 The third group consisted of 10 healthy lactating women (age 22-28 years). Blood-samples were taken immediately before and 30 min after the midday breast feed, thus minimising the influence of the circadian variation of calcitonin. These samples were taken on two occasions, the first on the seventh day after childbirth and the second at 6 weeks after delivery. All the women were lactating throughout the study. No drugs other than iron and folic acid during pregnancy were taken by any of the subjects. Methods
Summary
Plasma-calcitonin levels, measured with an established and reliable extraction
radioimmunoassay technique, were significantly higher throughout normal pregnancy and lactation than in normal non-pregnant women, and were not immediately influenced by the acute stimulation of breast-feeding. Thus, more calcitonin circulates at times of physiologically increased calcium need. It is suggested that an important function of calcitonin is the protection of the healthy maternal skeleton from excessive resorption by opposing the resorptive action of 1,25-dihydroxycholecalciferol on bone.
Introduction THE actions of calcitonin have been
clearly established,l but it is still not known whether the hormone has an important physiological function in man. One reason for our ignorance is that measurement is difficult : circulating calcitonin levels are extremely low, especially in women.2 Another problem is that the striking circadian variation in plasma levels3 is often ignored, although such an omission may invalidate any conclusions. However, by the correct timing of blood-sampling and by use of an extraction radioimmunoassay technique,3 we can reliably measure physiological levels of calcitonin at multiple plasma dilutions reading from the most sensitive part of the standard curve. We have applied our techniques to the study of calcitonin in normal women during pregnancy and lactation. Patients Three groups of healthy volunteers were studied. The first group consisted of 40 normal premenopausal women (age 17-43 years). The second group comprised 51 healthy pregnant women (age 17-34), 19 in the first trimester of pregnancy (6-12 weeks), 12 in the second trimester (13-28 weeks), and 20 in the third trimester (29-40 weeks).
12 Needleman P, Bronson SD, Wyche A, Sivakoff M, Nicolaou KC. Caridiac and renal prostaglandin I2. Biosynthesis and biological effects in isolated perfused rabbit tissues.J Clin Invest 1978; 61: 839-49.
13. Pace-Asciak, CR, Rangaray, G. Distribution of prostaglandin biosynthetic pathways in several rat tissues. Formation of 6-keto-prostaglandin F1&agr; Biochem Biophys Acta 1977; 486: 579-82. 14. Gryglewski RJ, Korbut R, Ocetkiewicz A. Generation of prostacyclin by lungs in vivo and its release into the arterial circulation. Nature 1978; 273: 765-67. 15. Moncada S, Korbut R, Bunting S, Vane JR. Prostacyclin is a circulating hormone. Nature 1978; 273: 767-68. 16. Hamberg M, Svensson J, Samuelsson B. Thromboxanes: a new group of biologically active compounds derived from prostaglandin endoperoxides. Proc Natn Acad Sci U.S.A. 1975; 72: 2994-98. 17. O’Regan S, Rivard GE, Robitaille PO. Aspirin improves platelet dysfunction in Bartter’s syndrome. Proceedings of the International Congress of Nephrology. Montreal: 1978; 1.
Calcitonin Measurements For each test, 25 ml of
venous
blood
was
taken from the
subject and placed in cooled heparinised tubes. The plasma was immediately separated and stored at -20°C until extraction and assay.3 Intra-assay and inter-assay variations were <10% and <14%, respectively. The sensitivity of the assay was 4-8 ng/1 with a 24 h incubation. For comparison, the sensitivity of the antiserum in a direct assay with a 7-day incubation period is approximately 2 pg/tube under optimum conditions.
StatisticalAnalyses Analysis of variance, two-tailed paired and unpaired Student’s t tests, and the Wilcoxon two-samples test were used as appropriate. Results The much
plasma calcitonin levels in pregnant women were, higher than those found in normal women (p <0.001) and were maintained throughout pregnancy;
the
mean
levels (ng/1±SEM) of 3745+_7.67, and for the first, second, and 41.79::!:6.41 55.19±11-02, third trimester, respectively, did not differ significantly from each other. Plasma-calcitonin levels were also raised during lactation (p<0.001); the increase was less striking but the levelsdid not differ significantly from those in pregnancy (figure). No significant difference was found between the mean levels before (23-02±2-51) and after (21.63±2.63) the midday breast feed, or at 1 week (2335+_2.49) and 6 weeks (21.30+_2.64) after delivery. There is a wide range of calcitonin levels in normal individuals, as has also been noted by Parthemore and Deftos;4 the reason is unknown. However, it does mean that there is some overlap between the levels found in the different groups. Discussion Our results show conclusively that plasma-calcitonin levels are raised throughout pregnancy. The physiological mechanisms responsible for this increase are not clear. Although there is evidence to suggest that oestrogen may be partly responsible,2,5 it can be seen from our results that the levels of calcitonin do not parallel those of oestrogen throughout pregnancy, so this cannot be the sole explanation. Previous reports of raised calcitonin levels in lactating animals6,7 accord well with our results in women. The mechanism is again unknown, but the acute stimulation of a breast feed appears to have no immediate effect on plasma calcitonin. It is unlikely that the calcitonin levels in lactating women represent a slow decline from pregnancy levels
770 Bone loss at that time could be both excessive and poorly tolerated because of an inadequate secretion of calcitonin and perhaps a slightly weakened skeleton. We thank Mr P. Needham for providing some of the samples obtained from pregnant patients. This work was supported in part by the Arthritis and Rheumatism Council and by the Medical Research Council. We thank Ciba-Geigy for synthetic human calcitonin. M.R.C. standard preparation 70/50 was donated by the National Institute for Biological Standards and Control. REFERENCES
TJ, Hunt NH, Ellison M, Underwood, JCE, Michelangeli VP. Physiology and biochemical pharmacology of calcitonin action. In: MacIntyre I, Szelke M, eds. Molecular endocrinology: proceedings of Endocrinology ’77. Amsterdam: Elsevier/North-Holland Biomedical Press, 1977.
1. Martin
179-92. Stevenson JC, MacIntyre I. Relative deficiency of plasma-calciin normal women. Lancet 1978; i: 961-62. 3. Hillyard, CJ, Cooke TJC, Coombes RC, Evans IMA, Maclntyre I. Normal Plasma calcitonin. Circadian variation and response to stimuli. Clin Endocr 1977; 6: 291-98. 4. Parthemore JG, Deftos LJ. Calcitonin secretion in normal human subjects. J Clin Endocr Metab 1978; 47: 184-88. 5. Delorme ML, Digioia Y, Fandard J, Merceron R, Raymond JY, Klotz HP. Oestrogenes et calcitonine. Ann Endocr 1976; 37: 503-04 6. Garel JM, Care AD, Barlet JP. A radioimmunoassay for ovine calcitonin an evaluation of calcitonin secretion during gestation, lactation and foetal
2.
Hillyard CJ, tonin
life. J Endocr 1974; 62: 497-509. 7. Toverud SU, Cooper CW, Munson PL. Calcium metabolism during lactation: elevated blood levels of calcitonin. Endocrinology 1978; 103:
472-79. 8. DeLuca HF. The
kidney as an endocrine organ for the production of 1,25-dihydroxyvitamin D3, a calcium-mobilizing hormone. N Eng J Med
1973; 289: 359-65. 9. Kumar R, Cohen WR, Silva P, Epstein FH. Elevated 1,25-dihydroxyvitamin-D plasma levels in normal human pregnancy and lactation. J Clin In-
Plasma-calcitonin levels in normal, pregnant and lactating women.
Horizontal bars are group means. Plasma-calcitonin levels were significantly higher in pregnant and lactating women than in normal women (p<0.001). because they appear to remain constant for up to 6 weeks during lactation. Our results are entirely consistent with the view that an important function of calcitonin is the protection of the skeleton against resorption.2In both pregnancy and lactation, there is an increased physiological demand on the mother for calcium. And this demand is normally satisfied by enhanced intestinal absorption of calcium produced by increased plasma levels of 1,25-dihydroxycholecalciferol (1,25[OH)2D3),8 the active hormonal form of vitamin D3’ Indeed, increased levels of this hormone are also found throughout all trimesters of pregnancy and during lactation,9 regardless of the variations in actual calcium need. It should be recalled that both 1,25(OH)2D3 and parathyroid hormone (PTH) are bone-resorbing hormones. However, 1,25(OH)2D3 causes bone resorption in physiological amounts, 10 whereas this may not be the case with PTH.11-13 Since it appears that increases in 1,25(OH)2D3 are accompanied by similar increases in calcitonin, we suggest that an important function of calcitonin in man is to oppose the action of 1,25(OH)2D3 on bone and thus protect the healthy maternal skeleton from excessive resorption during pregnancy and lactation. Some of the low levels in a number of the pregnant women may represent an inadequate response of calcitonin secretion to the increased 1,25(OH)2D3 levels in pregnancy. Thus, they may have increased skeletal resorption at this time compared with pregnant women with adequate calcitonin levels. While such bone loss would be insufficient to pose an immediate threat to the integrity of the skeleton, it may leave these women at some disadvantage when they reach the menopause.
10. 11.
vest 1979; 63: 342-44. Reynolds JJ, Holick MF, DeLuca HF. The role of vitamin D metabolites in bone resorption. Calcif Tiss Res 1973; 12: 295-301. Parsons JA. Parathyroid physiology and the skeleton. In: Bourne GH, ed. Biochemistry and physiology of bone. London: Academic Press, 1976; 4:
159-225. 12. Gaillard PJ, Wassenaar AM, Van Wijke-Wheeler ME. Effects of parathyroid hormone and a synthetic fragment (PTH 1-34) on bone in vitro. Proc Kon Ned Akad 1977; C80: 267-80. 13. Raisz LG, Canalis EM, Dietrich JW, Kream BE, Gworek SC. Hormonal regulation of bone formation. Recent Prog Horm Res 1978; 34: 335-56.
STRONG ASSOCIATION BETWEEN IDIOPATHIC MEMBRANOUS NEPHROPATHY AND HLA-DRW3.
J. MANOS P. A. DYER R. HARRIS N. P. MALLICK
P. T. KLOUDA E. J. ACHESON F. S. GOLDBY W. LAWLER
G. WILLIAMS
Department of Medical Genetics, St Mary’s Hospital, Manchester, Department of Renal Medicine, Royal Infirmary, and Department
of Pathology, University of Manchester
HLA A, B, and DR antigens were determined in a homogeneous group of patients with idiopathic membranous nephropathy. The frequency of HLA-DRw3 was significantly higher in the patients than in a control population. The frequencies of HLA B8 and B18, which are in linkage disequilibrium with DRw3, were also increased.
Summary
Introduction IDIOPATHIC membranous nephropathy (IMN) is associated clinically with moderate to heavy proteinuria. It is an important cause of the nephrotic syndrome and progressive renal failure. The histological diagnosis is