Calcium PTH Vitamin D basics

1. Calcium PTH Vitamin D basics Amani Alhozali Endocrine fellow R5

2. Objective • Calcium homeostasis • PTH structure and function • CaR structure and actions • Vitamin D metabolism and action • Calcitonin.

3. Calcium metabolism • Calcium ions are of critical importance for variety of vital bodily function. • Intracellular calcium is a key intracellular second messenger play role in controlling various cellular processes such as secretion ,differentiation , proliferation , motility ,and cell death. • Extracellular calcium is crucial for proper functioning of many tissue »excitation-contraction coupling in the heart &other muscles. »synaptic transmission and other CNS function »platelet aggregation and coagulation. »hormones secretion

4. Calcium metabolism • Extracellular and intracellular calcium level are tightly controlled within normal range. • About 50% of total Ca in the serum is present in ionized form, the remainder 40% bound to albumin and 10% complexed with PO4 or citrate. • It is the ionized ca that is regulated in extracellular fluid. • The concentration of ionized ca 1.25±0.07mmol/L • Serum ionized calcium maintained within a verynarrow range by the close relationship of serum ionized calcium and PTH. • This relationship is described by an inverse sigmoidal curve.

7. PTH response to hypocalcaemia • Second to minuets : exocytosis of PTH from secretory vesicle into the extracellular fluid. • Minutes to one hour: reduction in the intracellular degradation of PTH. • Hours to days : increased in PTH gene expression. (also stimulated by low serum calcitriol) • Days to weeks : proliferation of parathyroid cells. ( also stimulated by low serum calcitriol)

8. PTH Glands • PTH secreted from four parathyroid glands located adjacent to thyroid gland. • The glands weigh 40 mg of each. • Location is variable, the tow superior glands found near posterior aspect of thyroid gland. • The inferior glands located near the inferior thyroid margin • 12-15%of normal persons have 5th gland • Parathyroid gland arise from 3rd&4th branchial pouches.

10. PTH • PTH is 84- amino –acid peptide with a molecular weight of 9300. • PTH half-life 2-4 minutes after secretion. • PTH cleaved to produce an amino terminal fragment and a carboxyl terminal fragment. • Activities of PTH are encoded in the amino terminal. • PTH is cleared in the liver and kidney.

14. PTH assay

15. PTH Receptors. • There are two mammalian receptor for PTH. • PTH-1 receptor binds PTH and PTHrP with equal affinity. • PTH1R binds intact PTH and N-terminal residues. • Activation of PTH1R activates multiple cellular pathways and release intracellular calcium stores. • PTH1R heavily expressed in bone and kidney,and also present in other tissue such as breast ,skin ,heart ,blood vessels and pancreas . • PTH2R selectively binds PTH only. • PTH2R expressed heavily in the CNS,CVS ,GIT, lung and testes. • New PTH receptors (C-PTHRs) with specificity to carboxyl-terminal region of PTH,PTH 7-84 and shown to possess hypocalcemic activity ,that is reserved by PTH1-34and PTH 1-84.the C-PTHRs are present in different tissue but expressed heavily in bone .

18. PTH • PTH regulate ionized ca level by effect on 3 target organ: bone ,intestine, and kidney. • PTH has direct effect on tubular reabsorption of calcium ,phosphate and bicarbonate. • In the kidney PTH increase the reabsorption of calcium from distal convoluted tubule. • PTH inhibit reabsorption of phosphate in renal proximal tubule. • mild hyperchloremic metabolic acidosis in hyperparathyrodism due to impaired bicarbonate reabsorption.

19. Actions of PTH • Classical effect of PTH mediated through PTH1R, a G-coupled receptor expressed in kidney and bone. elevation of serum calcium phosphaturia calcitriol synthesis

20. Skeletal actions of PTH • PTH acts on bone to release calcium in two phases: 1- immediate effect to mobilize calcium from skeletal store. 2-later PTH stimulates release of calcium by activation of bone resorption. • Osteoblasts express PTH receptors. • PTH stimulate osteoblasts ,which stimulate the transformation of preosteoclast to mature osteoclast. Osteoclast dissolve the mineralized collagen matrex in bone. • Chronic hyperparathyroidism result in bone resorption. • Intermittent administration of PTH stimulate bone formation more than resorption and decrease risk of both vertebral and non vertebral fracture in patient with osteoprosis. • Positive effect of intermittent PTH on bone mediated through PTH1R.

21. Renal actions of PTH • Reabsorption of calcium 1- calcium reabsorbed passively in the proximal tubule and loop of Henle. 2- calcium transport actively according to chang in calcium balance in distal tubule under control of PTH. • PTH inhibit phosphate reabsorption mostly in proximal tubule . • PTH stimulates the synthesis 1- hydroxylase in proximal tubules and thus conversion of calcidiol to calcitriol.

23. Calcium sensing receptors • CaR is a120-KDa G protein-coupled receptor. It is member of family C of the superfamily of seven transmembrane (7TM). • It is expressed abundantly in parathyroid, thyroid C cells and kidney. • Activation of the CaR by increased extracellular Ca2+ inhibits parathyroid hormone (PTH) secretion, stimulates calcitonin secretion, and promotes urinary Ca2+ excretion, and thereby maintains the extracellular Ca2+ at the normal level . • CaR has seven membrane- spanning domain, the intracellular loops are directly involved in coupling the receptor to G protein.

25. Calcium sensing Receptors • In Parathyroid gland: • The CaSR is normally expressed at high levels on the surface of the parathyroid chief cells . High extracellular ionized calcium activate CaSR which in turn promote calcium released from endoplasmic reticulum (ER) and elevation of intracellular calcium which inhibit PTH secretion ,synthesis and parathyroid cellular proliferation.

27. CaSR action in the kidney • CaSR is an important regulator of urinary calcium excretion. • CaSR expressed on the basolateral membrane on the cells of the thick ascending limb of the loop henle. • Calcium binding to the receptor lead to the generation of arachidonic acid metabolite that then inhibits K channel in the luminal membrane and the Na-K ATPase pump in the basolateral membrane • Inhibition of K recycling reduces Na-Cl reabsorption via the Na-K-2Cl transporter, diminishing the generation of the lumen positive electrical gradient and therefor passive reabsorption of ca and mg. • Inhibition of the Na pump reduces the driving force for Na and Cl entry from tubular fluid by Na-K2Cl cotransporter.

29. CaSR action in the Kidney. • Inorganic phosphate (Pi) is absorbed by proximal tubules through a cellular pathway that is inhibited by parathyroid hormone (PTH). • The calcium-sensing receptor (CaSR) is expressed on apical membranes of proximal tubule. • CaSR activation blocks PTH-inhibitable phosphate absorption.

31. case • A 41 old woman was noted to have an elevated serum ionized calcium level of 1.39mmol/l(1.1-1.3) during an evaluation for infertility. her only complaints were fatigue and occasional headaches. She did not have any history of constipation, nausea, vomiting ,kidney stones or fractures. she was not aware of any family history of hypercalcemia. Her physical exam was unremarkable Laboratory blood test; Total calcium =2.6 mmol/l Ionized calcium = 1.36 mmol/l Po4 = 1.26 mmol/l Albumin ,creatinine normal PTH intact = 5.5 pmol/L 24 hour calcium = 3.23 mmol/l(129 mg)

33. Familial Hypocalciuric Hypercalcaemia • FHH result from an inactivation mutation in the calcium sensing receptor gene. • The mutation in FHH makes the receptor less sensitive to calcium. • In the parathyroid glands a higher than normal serum calcium required to reduce PTH . • In the kidney increase tubular calcium and magnesium reabsorption. • The net effect is hypercalcemia , hypocalciuria , and frequently hypermagnesemia.

34. FHH • FHH is a bening cause of hypocalcaemia characterized by AD inheritance. • Affected heterozygous patient typically present with hypercalcaemia ,hypocalciuria and mild to moderate hypermagnesemia. • Homozygous state lead to more sever neonatal hyperparathyroidism and sever hypocalcaemia. • FHH have normal or very slightly high serum PTH. • Patient usually asymptomatic or mild symptom and sign of hypercacemia.

35. Distinction from primary hyperparathyroidism • Absence of osteopenia ,osteitis fibrosa ,nephrolithiasis ,polyuria ,or mental changes; however pancreatitis and gallstones is associated in some cases of FHH. • The presence of hypercalcemia in family members. • Reduce urinary excretion of cacium. • Normal excretion of urinary cyclic AMP. • No evidence of abnormal parathyroid tissue on ultrasound or scan. • Benign disease and no surgical parathyroidectomy.

36. Calcium excretion — Measurements of 24-hour urinary collection for calcium (Ca) and creatinine (Cr) can confirm the diagnosis of FHH and distinguish it from primary hyperparathyroidism. Approximately 40 percent of patients with hyperparathyroid have hypercalciuria (24-hour calcium excretion above 250 mg [6.2 mmol] in women and 300 mg [7.5 mmol] in men) . Calcium excretion is typically below 200 mg/day (5 mmol/day) in patients with FHH Calculation of the Ca/Cr clearance ratio most useful test to differentiat 2 disorders. The ratio of calcium clearance to creatinine clerance is less than 0.01 (1%) in patient with FHH and generally between 0.02 to 0.05 (2 -5%) in patient with primary hyperparathyroidism. This ratio is calculated from the following formula:   Ca/Cr clearance ratio  =   [Urine Ca  x  serum Cr]  ÷  [Serum Ca  x  Urine Cr]

37. Autosomal Dominant Hpocalcemic Hypercalciuria. • Autosomal dominant hypocalcemia is the mirror image of FHH: familial hypocalcemia with urinary calcium excretion which is inappropriately high-normal or elevated in the basal state. • The serum calcium concentration is usually in the range of 6 to 8 mg/dL (1.5 to 2.0 mmol/L). • This disorder is associated with an activating mutation in the calcium-sensing receptor; as a result, a low serum calcium concentration is perceived as normal . • Serum PTH concentrations are normal and, in contrast to other causes of hypocalcemia, urinary calcium excretion is normal or high, presumably due to increased activation of the calcium-sensing receptor in the loop of Henle.

38. The diagnosis of autosomal dominant hypocalcemia should be suspected in hypocalcemic patients with the following features • Normal (or only slightly low) serum PTH concentrations • Frequently, few if any symptoms of hypocalcemia, despite reductions in the serum calcium concentration that would be expected to cause symptoms • High or high normal urinary calcium excretion rather than the expected low excretion • A family history of hypocalcemia • Recurrent nephrolithiasis • No previous normal serum calcium values • Low serum magnesium concentration

39. Treatment —  • As the serum calcium concentration increases, the activating mutation in the calcium-sensing receptor in the loop of Henle will lead to a marked increase in urinary calcium excretion, which can cause renal stones, nephrocalcinosis and renal insufficiency • Thus, the goal of therapy in symptomatic patients with autosomal dominant (or sporadic) hypocalcemia with hypercalciuria is to maintain a serum calcium concentration just sufficient to alleviate the symptoms. • A possible adjunct in patients who remain symptomatic despite hypercalciuria is to give a thiazide diuretic to reduce urinary calcium excretion and raise the serum calcium concentration.

41. Vitamin D • The term vitamin D (calciferol) refers to two secosteroids: vitamin D2 (ergocalciferol) and vitamin D3 ( cholecalciferol ). • Both are produced by photolysis from naturally occurring sterol precursor. • Vitamin D3 is formed in the skin from 7-dehydrocholesterol,wich distributed in epidermis and dermis . • The cleavage of the B ring of 7-dehydrocholesterol to form previtamin D3 requires ultraviolet light. • Previtamin D3 undergoes thermal isomerization to vitamin D3. • Vitamin D2 is manufactured through the ultraviolet irradiation of ergosterol from yeast ,and vitamin D3 lanolin. Both are used in over-the-counter vitamin D supplements. • Vitamin transported in the blood principally bind to DBP (85%) and albumin (15%). • Production of 1,25(OH)2D in the kidney stimulated by PTH and IGF-1 and inhibited by FGF23 and high levels of calcium and phosphate.

42. VITAMIN D synthesis and metabolism.

43. Mechanism of action of Vitamin D • Genomic action : 1,25 (OH)2D enter target cell and binds to its receptor VDR. The VDR then heterodimerized with the retinoid X receptor, RXR. The VDR – RXR complex then binds to specific regions within the regulatory portion of the genes called VDREs.the binding of the VDR-RXR complex to theVDREs attracts number of other proteins called coactivators to signal the beginning of transcription. • Non-Genomic action rapid action of vitaminD mediated through cell surface receptor.

45. Action of Vitamin D Intestinal action of Vitamin D • 1,25 (OH)2D enhance the efficacy of small intestine to absorbed calcium and phosphorus. • Both vitamin D and VDR are required for optimal absorption of calcium. • Vitamin D induce active cellular calcium uptake and transport mechanisms. • Calcium uptake required epithelial calcium channel TRPV6 and TRPV5. • Calcium uptake is the rate limiting step in intestinal calcium absorption, which is highly dependent on vitamin D. • Vitamin D increase active phosphorus transport.

46. Action of Vitamin D in Bone • Vitamin D is essential for the development &maintenance of mineralized skeleton. • Osteoblastic bone formation and osteoclastic bone resorption demand both vitamin D and VDR. • 1,25(OH)2D VDR system is critical in PTH induced osteoclastogenesis. • 1,25(OH)2D VDR increased the expression of RANKL on the surface of osteoblast ,RANK interaction with its receptor RANKL promotes maturation of osteoclast progenitor cell & mature osteoclast. • Vitamin D ,PTH and prostaglandin stimulate RANKL expression.

47. Action of Vitamin D in Kidney • The kidney expresses VDR, and 1,25 (OH)2D stimulate Ca²-ATPase in distal tubule as well as 24,25(OH)2D production in the proximal tubule. • 1,25 dihydroxyvitamin D decrease its own synthesis through negative feedback and decrease secretion and synthesis of PTH. • 1,25 dihydroxyvitamin D increase expression of 25-hydroxyvitamin D-24-hydroxylase to catabolize 1,25(OH)2D to the water-soluble ,biological inactive calcitroic acid .

49. Vitamin D Dependent Rickets (VDDR) type I is a rare AR disease due to mutation in 25(OH)D 1- hydroxylase gene result in rickets accompanied by low level of 1,25(OH)2D. • Vitamin D Dependant Rickets (VDDR) type II is a rare AR disease due to inactivating mutation in the VDR gene result in childhood rickets and high level of 1,25(OH)2D. many of these patient have alopecia

50. CAlCITONIN • Calcitonin is a 32-amino-acid peptide • Calcitonin secreted by parafollicular C cells of the thyroid. • Secretion of calcitonin is under the control of ionized ca. • CaSR expressed on C cell of thyroid ,high extracellular calcium increase secretion of calcitonin. • Hypocalcaemia inhibit calcitonin secretion.