1. Unit II – Endocrine Section Calcium Metabolism Daylily S Ooi MBBS, FRCPC (Med Biochemistry) 3973: Describe the function of parathyroid hormone 3974: Explain the physiological actions of PTH on bone kidneys and intestines 3975: Describe Vitamin D action on target tissues 3976: Describe the regulation of 1, 25 di-OH vitamin D 3977: Explain the regulation of serum calcium 3978: Describe the physiological action of calcitonin

2. You may only access and use this presentation for educational purposes. You may not post this presentation online or distribute it without the permission of the author. Slides marked with ✪ are modified from Dr. D Liu’s lecture 2014. Disclosure Unit II – Calcium Metabolism – DS Ooi

3. Objectives Additional topics added to this lecture are in blue Unit II – Calcium Metabolism – DS Ooi Distribution of calcium, phosphate and magnesium in the body 3977: Explain the regulation of serum calcium Organs involved Hormonal and other regulators Parathyroid hormone Production and regulation 3973: Describe the function of parathyroid hormone 3974: Explain the physiological actions of PTH on bone, kidneys and intestines Vitamin D Forms Production 3976: Describe the regulation of 1, 25 di-OH vitamin D 3975: Describe Vitamin D action on target tissues Calcitonin Production 3978: Describe the physiological action of calcitonin Clinical uses Disturbances of calcium homeostasis – causes, symptoms, management Hypercalcemia Hypocalcemia Secondary hyperparathyroidism Disturbances of Magnesium and Phosphate

4. Distribution of Calcium Total body calcium ~ 1kg (25.5 moles) 99% in Bone (25 moles). With phosphorus, constitutes 65% of bone by weight. Soft tissue - intracellular (25 mmoles) – Very little as cytosolic free calcium (100 n moles) – 99% is within cellular compartments – bound to inner plasma membrane of mitochondria, or endoplasmic reticulum Extracellular fluid (23 mmoles) Unit II – Calcium Metabolism – DS Ooi

5. Distribution of calcium in blood 45% protein bound (80% albumin, 20% globulins ) 10% complexed (citrate, lactate, phosphate, bicarbonate) 45% free ionized form (physiologically active) Varies with pH alkaline pH  binding   free ionized form acid pH  binding   free ionized Ca Unit II – Calcium Metabolism – DS Ooi

6. Calcium Function: Extracellular Excitation-contraction in muscles Synaptic transmission Platelet aggregation and coagulation Intracellular Secretion of hormones and other regulators by exocytosis Secondary messenger in cell division, cell motility ✪ Unit II – Calcium Metabolism – DS Ooi

7. Magnesium Distribution: Total body magnesium 1 mole (Total body Ca = 25 moles) – Bone predominantly – Cells – Serum - 30% protein bound (Serum Ca 40% bound) Function: Neuromuscular conduction Parathyroid hormone secretion Unit II – Calcium Metabolism – DS Ooi

8. Phosphate Distribution: Total body phosphate 700g (24 moles) Bone 83% - hydroxyapatite (calcium phospate) Cells 16% - organic phosphates (nucleic acid, ATP, phospholipids) Extracellular 1% - inorganic phosphates Fluid (H 2 PO 4 - : HPO 4 = 1:4) Function: High energy phosphate bonds Buffer Unit II – Calcium Metabolism – DS Ooi

9. Hormones: Organs involved: Intestine Kidneys Bone Regulation of calcium metabolism: Unit II – Calcium Metabolism – DS Ooi

10. Calcium balance over 24 hours PTH 14 mmoles/d Vit D Soft Tissue 25 mmoles (1000 mg) Bone 25 moles (1000 mg) Extracellular Fluid 23 mmoles (920 mg) GI Tract Kidney 20 mmoles (800 mg) 8 mmoles (320 mg) 16 mmoles (640 mg) 4 mmoles (160 mg) 4 mmoles Vit D Glomerular filtrate 270 mmoles PTH Unit II – Calcium Metabolism – DS Ooi

11. Calcium Intestine Absorption process: Regulated saturable transcellular absorption Nonsaturable paracellular absorption (dependent on mineral concentration in lumen) Main sites: Duodenum and jejunum Unit II – Calcium Metabolism – DS Ooi

12. Factors affecting intestinal calcium absorption Unit II – Calcium Metabolism – DS Ooi GIT: Gastro-intestinal tract; diOH: dihydroxy

13. Kidney: Calcium handling Filtered Ca 270 mmol/24h Prox Convoluted Tubule (passive) Distal Convoluted Tubule (active) Collecting Duct Thick Asc Loop of Henle 70% 20% 8% (GF:180L Conc: 1.5 mmol/L) 98% reabsorbed ~ 5 mmol excreted in 24h <5% Unit II – Calcium Metabolism – DS Ooi

14. Bones: PTH 14 mmoles/d Vit D Soft Tissue 25 mmoles (1000 mg) Bone 25 moles (1000 mg) Extracellular Fluid 23 mmoles (920 mg) GI Tract Kidney 20 mmoles (800 mg) 8 mmoles (320 mg) 16 mmoles (640 mg) 4 mmoles (160 mg) 4 mmoles Vit D Glomerular filtrate 270 mmoles PTH Unit II – Calcium Metabolism – DS Ooi

15. Regulation of calcium: Hormones involved  Parathyroid hormone (PTH, parathyrin)  Bone  Kidneys  Intestines  Vitamin D  Intestines  Bone  Kidneys  Calcitonin  Bone  Kidneys Unit II – Calcium Metabolism – DS Ooi

16. Parathyroid hormone (PTH)  Synthesized as preprohoromone that undergoes cleavage before secretion  Continuously synthesized, minimal storage in parathyroid glands  Metabolized by liver & kidneys, plasma t 1/2 2 min  Laboratory assays measure:  C terminal (long t½), N-terminal (short t½ )  Mid-terminal (long half-life)  Intact PTH – most intact assays, also measure a 7-84aa fragment which accumulates in renal failure Unit II – Calcium Metabolism – DS Ooi ✪

17. Regulators of Plasma PTH  Plasma calcium  Hypocalcemia  PTH biosynthesis  Ca-sensing receptor (CaSR) on parathyroid cells Calcium binding results in   PTH synthesis & secretion   degradation of stored PTH  Calcitriol 1,25(OH) 2 D 3   PTH gene transcription  Hypocalcaemia overrides calcitriol effect on PTH production  Less important regulators:  Catecholamines  Magnesium - low Mg can cause hypocalcaemia  Prostaglandins Unit II – Calcium Metabolism – DS Ooi ✪

18. PTH secretion vs. Plasma calcium From Williams Textbook of Endocrinology Unit II – Calcium Metabolism – DS Ooi Secretion rate of PTH varies inversely with plasma calcium concentation, in a sigmoid fashion ✪

19. PTH Actions: Maintains ionized calcium concentration within narrow range Bones: Initiates osteoclastic bone resorption – release of calcium from bones At high concentrations (e.g. 1 o or 2 o hyperparathyroidism) –  bone resorption >>  bone formation –  cortical bone mass At lower concentrations, especially if episodic release –  bone formation >>  bone resorption –  trabecular bone mass Unit II – Calcium Metabolism – DS Ooi ✪

20. PTH Actions on kidneys: Kidneys: –  calcium reabsorption in distal convoluted tubules Note: 90% of filtered Ca reabsorbed in proximal tubule and loop of Henle independent of PTH, mostly via passive paracellular route –  phosphate reabsorption in proximal and distal convoluted tubules –Stimulates 1  -hydroxylase (  calcidiol to calcitriol) Intestines: effect through vitamin D Unit II – Calcium Metabolism – DS Ooi

21. Vitamin D Unit II – Calcium Metabolism – DS Ooi Action: Binds to nuclear Vitamin D Receptor (VDR) – resulting in regulation of DNA transcription Calcitriol has highest affinity

22. Vitamin D 7-dehydrocholesterol Cholecalciferol 25-OH vitamin D Calcitriol PTH 24,25 - D Intestines  Ca & PO 4 absorption Parathyroids  PTH Secretion  Cell proliferation  Gene transcription Bone Multiple effects Muscle (deficiency associated with myopathy) Unit II – Calcium Metabolism – DS Ooi ✪

23. Hormonal Regulation of Blood Calcium  Calcium Absorption Gastrointestinal Tract  Ca excretion  P excretion  Blood Ca  mineralization  Bone loss  Bone resorption PTH Parathyroid Glands  Blood calcium Unit II – Calcium Metabolism – DS Ooi 1,25 diOH Vit D Cholecalciferol (Vit D3) 7-Dehydrocholesterol Effect of UV on Skin 25-OH cholecalciferol 25 hydroxylase 1 hydroxylase Legend: Stimulate Inhibit Delayed effects (Stimulation)

24. Regulation of Blood Calcium When blood calcium falls:  Bone resorption PTH  Blood Ca ++ Parathyroid Glands  Ca excretion   Serum Ca  P excretion   Serum P T½ 10 m Metabolised in liver to n-terminal (active, t½ 1-2m) c-terminal fragments (t½1-2h) which are cleared by kidney  Ca  P  Serum Ca  Serum P Unit II – Calcium Metabolism – DS Ooi

25. Regulation of Blood Calcium concentrations Intermediate/Long term PTH 25-OH D 1,25 diOH Vit D Cholecalciferol (Vit D3)  Calcium Absorption Parathyroid Glands 7-Dehydrocholesterol Effect of UV on Skin  mineralization Unit II – Calcium Metabolism – DS Ooi

26. Calcitonin Unit II – Calcium Metabolism – DS Ooi

27. Calcitonin Production:  32-amino acid peptide hormone  Produced by thyroid parafollicular C-cells Unit II – Calcium Metabolism – DS Ooi ✪

28. Calcitonin Action:  Inhibits osteoclast-mediated bone resorption (counteracts action of PTH)  Renal (at higher concentrations):  Inhibits P reabsorption   P excretion  Some natriuretic effect  mildly  Ca excretion Non-essential & less important than PTH Total thyroidectomy does not result in hypercalcaemia High calcitonin in medullary thyroid cancer does not result in hypocalcaemia Unit II – Calcium Metabolism – DS Ooi ✪

29. Clinical uses of calcitonin  Tumour marker for medullary thyroid carcinoma  Therapeutic applications:  Hypercalcemia - administration quickly lowers plasma Ca through reduced osteoclast activity  Osteoporosis - reduces fracture risk & pain associated with fractures (no longer used due to increased risk of malignancy)  Paget disease of bone (bisphosphonates preferred) Unit II – Calcium Metabolism – DS Ooi ✪

30. Other hormones affect bone Growth hormone & IGF-1  bone remodeling Glucocorticoids  Ca absorption Long term administration  bone formation Hyperthyroidism  skeletal growth in children  bone resorption in adults InsulinRequired for normal growth Gonadal hormones Critical for skeletal development & maintenance Unit II – Calcium Metabolism – DS Ooi IGF – Insulin-like Growth Factor

31. Local bone regulators Cytokines e.g., interleukins, TNF- , TNF-   bone resorption,  formation TGF-  & EGF Produced by neoplasms  bone resorption Prostaglandins Synthesized by many skeletal cells Affects bone resorption & formation Unit II – Calcium Metabolism – DS Ooi TNF: Tissue Necrosis Factor, TGF: Transforming Growth Factor, EGF: Epidermal Growth Factor

32. Hormonal Regulation of Blood Calcium Cholecalciferol (Vit D3) 7-Dehydrocholesterol Effect of UV on Skin 25-OH Cholecalciferol (Calcidiol) 25  hydroxylase 1,25 diOH Vit D (calcitriol) 1  hydroxylase PTH Parathyroid Glands Parafollicular C cellsCalcitonin  Osteoclast inhibition  bone remodelling Diet Ergocalciferol (D2) 25-OH Ergocalciferol 1, 25 diOH D2 Ca release from bone (permissive) Activates remodelling  Ca and phosphate reabsorption  Intestinal Ca and P transport Inhibition of PTH synthesis   Ca release from bones Bone remodelling (RANKL)  DCT Ca reabsorption  PCT and DCT P reabsorption Renal 1  hydroxylase activation Unit II – Calcium Metabolism – DS Ooi

33. Disorders of Calcium homeostasis Unit II – Calcium Metabolism – DS Ooi

34. Mechanisms for hypercalcemia  Bone resorption  Gastrointestinal absorption of calcium  Renal excretion of calcium Unit II – Calcium Metabolism – DS Ooi

35. PTH Mediated Primary hyperparathyroidism –Sporadic –Inherited variants Multiple endocrine neoplasia (MEN1, 2a) Familial isolated hyperparathyroidism –Hyperparathyroid-jaw tumour syndrome Familial hypocalciuric hypercalcemia CaSR mutation (AD inheritance) Tertiary hyperparathyroidism Following prolonged stimulation, part of the parathyroid gland escapes feedback control Unit II – Calcium Metabolism – DS Ooi

36. Primary Hyperparathyroidism F > M, up to 0.4% of F>60y may be affected Pathology: –Solitary adenoma 80% –Hyperplasia 15% –Parathyroid carcinoma 1-2% Unit II – Calcium Metabolism – DS Ooi ✪

37. Non-PTH Mediated Malignancy –PTH-related peptide (PTHrP) –Osteolytic bone metastasis and local cytokines –Activation of extrarenal 1  hydroxylase Vitamin D –Chronic granulomatous disorders (sarcoidosis, TB) 1  hydroxylase –Exogenous vitamin D intake Drugs: –Milk-alkali syndrome (  Ca absorption, alkalosis  renal Ca excretion) –Lithium (  renal Ca excretion, ? block Ca feedback on parathyroids) –Thiazides (  Renal Ca excretion) –Vitamin A toxicity –Theophylline toxicity Miscellaneous: –Hyperthyroidism –Acromegaly –Adrenal insufficiency –Immobilization Unit II – Calcium Metabolism – DS Ooi

38. Malignancy-associated hypercalcemia The most common cause of hypercalcemia in hospitalized patients Incidence: 15 cases/100,000/yr Common cancers –squamous cell cancer of lung, breast –renal cell carcinoma –myeloma, lymphoma Rare in colon, gastric and thyroid cancers Unit II – Calcium Metabolism – DS Ooi ✪

39. Hypercalcemia – Symptoms/Signs Unit II – Calcium Metabolism – DS Ooi Bones Stones Groans Moans

40. Management of Hypercalcemia: IV fluids Loop diuretics (furosemide) Calcitonin Steroids Bisphosphonates Dialysis Calcium sensor receptor agonist (Cinacalcet) – for primary hyperparathyroidism Unit II – Calcium Metabolism – DS Ooi ✪

41. Hypocalcemia Causes: 1. Insufficient PTH activity –Hypoparathyroidism (post thyroid surgery) –Hypomagnesemia (Mg required for PTH release) –Pseudoparathyroidism (PTH resistance) 2. Insufficient Vitamin D action –Insufficient Dietary/Exposure to UV rays –  1  -hydroxylase Chronic renal failure Vitamin D dependent rickets –Vitamin D resistant rickets 3. Sequestration of calcium –Acute pancreatitis 4. Drugs – calcitonin, furosemide Unit II – Calcium Metabolism – DS Ooi

42. Hypocalcemia – Symptoms/Signs Unit II – Calcium Metabolism – DS Ooi

43. Management of hypocalcemia Acute:  Replace calcium  Calcium gluconate IV  Oral calcium  Treat hypomagnesemia, if present  May require vitamin D  Correct underlying cause Long-term  High dose vitamin D (D2 50,000 IU daily, calcitriol – up to 2 ug daily)  Adequate calcium intake ✪ Unit II – Calcium Metabolism – DS Ooi

44. Secondary hyperparathyroidism  PTH caused by other conditions Vitamin D disorders – Deficiency or malabsorption – Rickets Phosphate disorders – Chronic kidney disease – Phosphate depletion – Malabsorption – Aluminium toxicity Calcium deficiency ✪ Unit II – Calcium Metabolism – DS Ooi

45. Chronic kidney disease Parathyroid glands  Ca absorption (passive P absorption) PTH  P N or  Ca 2+ 1,25 D + PTH + +    Ca, P P ✪ Unit II – Calcium Metabolism – DS Ooi

46. Hypermagnesemia Causes: Chronic renal failure Intravenous MgSO 4 - as antihypertensive, sedative during parturition Effects: Usually does not rise to critical concentrations, and not clinically important. Sedation  Neuromuscular activity Unit II – Calcium Metabolism – DS Ooi

47. Hypomagnesemia Causes: 1. Reduce intake – malabsorption –  intake (alcoholics) 2.  renal loss – diuretics – alcohol – Renal tubular defects – drugs - gentamicin, amphotericin B Effects:  PTH release (hypocalcemia) Unit II – Calcium Metabolism – DS Ooi

48. Hyperphosphatemia Cause: Most often seen in chronic renal failure Effect:  serum calcium  Secondary hyperparathyroidism Management: Oral Phosphate binders Dialysis for CRF patients Unit II – Calcium Metabolism – DS Ooi

49. Hypophosphatemia Cause: 1.  intake - starvation, malabsorption, Al(OH) 3 2.  loss renal tubular leaks hyperparathyroidism Vit D resistant rickets (impaired tubular Phosphate transport) Effect: 1.Loss of RBC membrane integrity (hemolysis) 2.Muscle weakness Unit II – Calcium Metabolism – DS Ooi

50. Key points  Blood calcium is tightly regulated, primarily by PTH & vitamin D. Calcitonin plays a far less important role.  PTH acts on kidneys and bones; Vit D on bones, intestines, kidneys  Common clinical conditions: Hypercalcemia  Hypercalcemia in malignancy  Primary hyperparathyroidism Hypocalcemia  Post thyroid, parathyroid surgery Secondary hyperparathyroidism  Vitamin D deficiency  Renal failure  Remember! Always adjust serum total calcium for albumin concentration (0.2 mmol Ca for every 10 g of albumin) Unit II – Calcium Metabolism – DS Ooi

51. Appendix: Parathyroid hormone From Endocrinology: An Integrated Approach, 2001 Unit II – Calcium Metabolism – DS Ooi 84 amino acid peptide hormone Synthesis: 7-84aa Blocks PTH activity Accumulates in CKD Measured by most intact PTH assays ✪

52. Vitamin D: Chemical structures D2D2 D3D3 25-OH D 3 1,25(OH) 2 D 3 From www.chm.bris.ac.uk ✪ Unit II – Calcium Metabolism – DS Ooi