This presentation deals with the physiological aspect of Calcium and phosphate metabolism, it's relationship with the various types of rickets and possible remedies

1. Calcium & Phosphorus
Metabolism; Role of
Phosphatonins
Dr. Shinjan Patra

2. Contents
• History & Basic physiological functions
• Sources & RDA
• Calcium Distribution & Variations
• Calcium Metabolism & Determinants
• Phosphate Distribution & Variations
• Phosphate Metabolism & Determinants
• Phosphatonins: Examples & basics
• Diseases related to them

3. Minerals
Macro Nutrients Micro Nutrients
Calcium Iron
Magnesium Iodine
Phosphorus Copper
Sodium Manganese
Potassium Zinc
Chloride Molybdenum
Sulphur Fluoride

4. Calcium: History
• Most abundant mineral in the human body
• Ancient Romans prepared lime- calcium oxide
in 975 AD
• Plaster of Paris (calcium sulfate) used for
setting broken bones.
• Sir Humphry Davy Isolated calcium in 1808
• Latin ‘Calx’ or ‘Calcis’ means Lime

5. Phosphate: History
• Second most abundant essential mineral in
the human body
• The first form of elemental phosphorus to be
produced was white phosphorus by Greeks
• Hennig Brand discovered in 1669 in Germany
• Means: Bringer of Light

6. Role of Calcium
• Affects nerve and muscle physiology
• Intracellular signal transduction pathways
• Co-factor in blood clotting cascade
• Constituent of bone and teeth
• Major structural element in the skeleton in the
form of calcium phosphate (Ca10(PO4)6(OH)2
known as hydroxyapatite
• Essential component in production of enzymes
and hormones that regulate metabolism

7. Role of phosphorus
• Key constituent of bone and teeth
• Component of intra cellular buffering : Forms energy
rich bonds in ATP
• Forms co-enzymes
• Regulates blood and urinary pH
• Forms organic molecules like DNA & RNA
• Cellular energy metabolism
• Constituent of molecules like phospholipids and
phosphoproteins

8. Sources of Calcium
Best Source Fair Source Good Source
Hard Cheese Beans Baked Beans
Milk Eggs Dried Legumes
Dark Green Leafy
Vegetables
Breads Dried Figs
Broccoli

9. Sources of Phosphate
Rich Source Moderate Sources
Milk Cereals
Meat Pulses
Fish Nuts
Poultry Legumes
Eggs

11. Homeostasis disturbances
• Disturbances in extracellular calcium concentration cause a
variety of symptoms; the most common of which reflect
abnormal neuromuscular activity
• Hypercalcemia may lead to muscle weakness, areflexia,
anorexia, constipation, vomiting, drowsiness, depression,
confusion, other cognitive dysfunction and coma
• Hypocalcemia, conversely, may cause anxiety, seizures,
muscle twitching, epilepsy, tetany, Chvostek’s and
Trousseau’s signs, carpal or pedal spasm, stridor,
bronchospasm, and intestinal cramps, as well as cataracts,
skeletal malformations and abnormal dentition

12. Hypercalciuria
• Hypercalcemia leads to hypercalciuria, which
can result in Nephrocalcinosis, Nephrolithiasis
and impaired kidney function

13. Calcium Distribution & its
variations

14. Basic balance of Calcium
• Between the amounts that are absorbed from
the gut, deposited into bone and into cells,
and excreted from the kidney
• Chief Control:
Gut
Bone
Kidney

15. Calcium Distribution
• Ninety-nine percent of total body calcium resides in bone, of which
99% is located within the crystal structure of the mineral phase
• The remaining 1% of bone calcium is rapidly exchangeable with
extracellular calcium
• Calcium is equally distributed between the intracellular and
extracellular fluids
• In blood, approximately 50% of total calcium is bound to proteins,
mainly albumin and globulins
• Ionized calcium concentration in serum approximately 5 mg/dL and
it is this ionized fraction that is biologically active and that is tightly
controlled by hormonal mechanisms

18. Ionized Calcium
• The usual 2:1 ratio of total to ionized calcium
may be disturbed by disorders such as
metabolic acidosis (calcium binding to
proteins is reduced at acid pH)
• By changes in serum protein concentrations,
as in starvation, cirrhosis, dehydration, or
multiple myeloma

19. Intracellular Calcium
• More than 99% of intracellular calcium exists in the
form of complexes within the mitochondrial
compartment, bound to the inner plasma membrane
• Release of calcium from membrane-bound
compartments transduces cellular signals and tightly
regulated
• Better understood with the identification of specific
receptors for calciotropic signaling molecules such as
the Inositol Triphosphate (IP3) receptor and Ryanodine
receptors

20. Calcium variations
• Higher during infancy/childhood and adolescence than in the adult
• Does not change at puberty or, in women, during the menstrual
cycle
• During pregnancy, total serum calcium and albumin decline
progressively, but ionized calcium is minimally affected
• Despite daily losses of 200 to 300 mg/day of calcium in breast milk,
lactating women maintain normal levels of ionized calcium in blood
by increasing intestinal calcium absorption
Krabbe S, Transbol I, Christiansen C. Bone mineral homeostasis, bone growth, and mineralization during years of pubertal growth: A unifying
concept. Arch Dis Child. 1982;57:359–363.
Pitkin RM, Reynolds WA, et al. Calcium metabolism in normal pregnancy: A longitudinal study. Am J Obstet. 1979;133:781– 790

21. Fetal variations
• Fetal calcium content rises dramatically during
the third trimester (to about 30 g at term)
• Fetal serum total and ionized calcium
concentrations are higher than maternal levels,
consistent with active placental transport of
calcium
Hillman L, Sateesha S, Haussler M, et al. Control of mineral homeostasis during lactation: interrelationships of
25-hydroxyvitamin D, 24,25-dihydroxyvitamin D, 1,25-dihydroxyvitamin, parathyroid hormone, calcitonin,
prolactin, and estradiol. Am J Obstet Gynecol. 1981;139:471–476.
Greer FR, Tasang RC, Searcy JE, et al. Mineral homeostasis during lactation-relationship to serum 1,25-
dihydroxyvitamin D, 25-hydroxyvitamin D, parathyroid hormone, and calcitonin. Am J Clin Nutr.
1982;36:431–437

22. CaSR concept
• Expressed in parathyroid, renal, epithelial, and
numerous other cells
• Calcium can act as an extracellular ligand to
directly control cellular function
• Suppression of PTH secretion
• In the thick ascending loop of the renal tubule,
inhibition of calcium, magnesium, and NaCl
reabsorption

24. Calcium Metabolism &
Determinants

25. Calcium GI Absorption
• Intestinal calcium absorption ranges broadly between 20%
and 70%
• Factors:
 Overall calcium balance
 Vitamin D
 Declines with age
 Previous calcium intake
 Other nutrients
 Pregnancy, lactation
• Calcium balance cannot be maintained if dietary calcium
consistently falls below 200 to 400 mg/day

26. Sites
• Absorbed throughout the intestine
• Absorption most efficient in the duodenum and proximal
jejunum
• Highest levels of vitamin D–dependent calcium-binding
proteins and in lower luminal pH=5
• Absorbed by both a passive paracellular route (at higher
levels of calcium) and by an active, transcellular mechanism
(upregulated by Vit-D in low calcium)
• TRPV5 & TRPV6 predominantly regulated
Gallagher JC, Riggs BL, Eisman J, et al. Intestinal calcium absorption and serum vitamin D metabolites in normal subjects and
osteoporotic patients. J Clin Invest. 1979;64:729–736

27. Vit-D physiology related to Ca
BALANCE
• Unlike the 25-hydroxylase, the 1α-hydroxylase is
tightly regulated
• Inducers: PTH & Hypophosphatemia
• Inhibitors: Calcium, 1,25(OH)2D, and FGF23
• Other factors: Estrogen, Calcitonin, GH and
prolactin have been shown to increase
• Ketoconazole has been shown to decrease

28. Vit-D in gut absorption of Ca
• Saturable transcellular pathway is dependent
on 1,25(OH)2D3
• Substantial evidence exists that the hormone
enhances para-cellular pathway as well
Even L, Weisman Y, Goldray D, Hochberg Z. Selective modulation by vitamin D of renal response to parathyroid hormone: a study
in calcitriol-resistant rickets. J Clin Endocrinol Metab. 1996;81:2836–2840

30. Renal Calcium Excretion
• Urinary calcium excretion (and net intestinal calcium absorption)
approximately 200 mg/day
• Approximately 60% to 70% of tubular calcium reabsorption occurs
in the proximal tubule mainly via passive diffusion along
paracellular pathways
• Another 20% to 25% of the filtered calcium load reabsorbed by
paracellular diffusion in the (cTAL)
• 8% to 10% of filtered calcium is reabsorbed in more distal
segments of the tubule (PTH regulated)
• Additionally more distal nephron segments, such as cortical and
medullary collecting ducts
Quamme GA. Effect of hypercalcemia on renal tubular handling of calcium and magnesium. Canadian J Physiol Pharmacol.
1982;60:1275–1280.

31. PTH dependant channels
• Apical epithelial calcium channel (TRPV5)
• Cytosolic calcium-binding protein calbindin-
D28K
• Basolateral plasma membrane calcium
ATPase(s) (PMCAs)
• Basolateral Na+/Ca++ exchanger-1 (NCX1)
Shimizu T, Nakamura M, Yoshitomi K, Imai M. Interaction of trichlormethiazide or amiloride with PTH in stimulating Ca2+ absorption in rabbit.
CNT Am J Physiol. 1991;261:F36–F43

33. CaSR control
• Hypercalcemia suppresses renal tubular calcium
reabsorption, even in the absence of PTH, an
effect attributable to direct activation of
basolateral CaSRs in the cTAL
• CaSR activation in the cTAL also antagonizes PTH-
stimulated increases in calcium reabsorption,
possibly by impairing cyclic AMP generation
Scoble JE, Mills S, Hruska KA. Calcium transport in canine renal basolateral membrane vesicles. Effect of
parathyroid hormones. J Clin Invest. 1985;75:1096–1105
Bouhtiauy I, LaJeunesse D, Brunette MG. The mechanism of parathyroid hormone action on calcium
reabsorption by the distal tubule. Endocrinology. 1991;128:251–258

34. Vit-D dependent mechanisms
• 1,25(OH)2D is required for the calcium-
reabsorptive response to PTH
• Accelerates the increase in DCT cell calcium
entry initiated by PTH
• Directly increases DCT calcium reabsorption,
apparently by driving higher expression of
TRPV5, Calbindin D28K, and PMCA

35. Vit-D over PTH
• 1,25(OH)2D3 has been shown to regulate
gene transcription and cell proliferation in the
parathyroids
• Shown to decrease the transcription of the
PTH gene both in vivo and in vitro

37. PTH structure & function
• PTH gene transcription (as well as PTH peptide
secretion) dependent upon the extracellular
calcium and phosphate
• Secretion of mature PTH regulated through
the CaSR
• PTH/PTHrP receptor is highly expressed in
kidney and bone

39. PTH actions on Bone
• PTH acts on a number of cell types both
directly and indirectly
• PTH administration by any route increases
both bone resorption by increasing osteoclast
number and bone formation by increasing
osteoblast number

41. Calcitonin
• When given in large doses, acutely reduces proximal tubular
calcium reabsorption by a mechanism independent of PTH
• Impairs osteoclast-mediated bone resorption by a direct action on
osteoclasts
• Physiologic regulation of calcium reabsorption is thought to be
unlikely
• Long-term hypercalcitoninemia secondary to (MCT) and in patients
with subtotal thyroidectomy resulting in lack of calcitonin secretory
reserve: no influence on the bone density at the lumbar spine and
distal radius
Wimalawansa SJ. Long- and short-term side effects and safety of calcitonin in man: a prospective study. Calcif Tissue Int.
1993;52(2):90–93

42. Other hormones
• Hypercalciuria observed in states of excess growth
hormone or cortisol seems likely to be secondary to an
increased filtered load of calcium rather than to direct
tubular actions of these hormones
• Estrogen treatment of normal postmenopausal women
lowers urinary calcium excretion by increasing tubular
calcium reabsorption
Chipman JJ, Zerwekh J, Nicar M, et al. Effect of growth hormone administration: Reciprocal changes in serum
1a,25-dihydroxyvitamin D and intestinal calcium absorption. J Clin Endocrinol Metab. 1980;51:321–324
Hahn TJ, Halstead LR, Baran DT. Effects of short-term glucocorticoid administration on intestinal calcium
absorption and circulating vitamin D metabolite concentrations in man. J Clin Endocrinol Metab.
1981;52:111–114
Gallagher JC, Nordin BEC. Treatment with oestrogens of primary hyperparathyroidism in post-menopausal
women. Lancet. 1972;1:503–507

43. Calcium Loading state
• Parathyroid suppression
• Inhibition of renal 1,25(OH)2D3 synthesis
• Decreased intestinal active transport of calcium
• Increased renal excretion of calcium
• Decreased renal excretion of phosphate (secondary to
functional hypoparathyroidism)
• Decrease in bone resorption sufficient to allow positive
skeletal calcium balance
 Decline in intestinal calcium absorption the major
safeguard against calcium overload

44. Ca deprived people
• High serum concentrations of PTH and
1,25(OH)2D3
• Increased intestinal calcium absorption
• Increased bone resorption and progressive
osteopenia
• Increased renal tubular calcium reabsorption
• Low urinary calcium excretion
• Decreased renal tubular phosphate reabsorption

46. Phosphate Distribution & Variation

47. Distribution
• 85% of body phosphate is in the mineral phase of bone, and the
remainder is located in inorganic or organic form
• Exists almost entirely in ionized form as either H2PO4– or HPO42–
• Only 12% of serum phosphate protein bound
• Additional small fraction loosely complexed with calcium,
magnesium and other cations
• Carbohydrate ingestion may markedly reduce serum phosphate by
moving serum phosphate from the extracellular to the intracellular
space
• Serum phosphate undergoes diurnal variation of as much as 1.5
mg/dL (0.5 mmol/L) with a nadir between 8 am and 11 am.

48. Pregnancy/Lactation/New born
• Fasting serum phosphate remains stable throughout
the menstrual cycle and during pregnancy
• Lactation: remains same
• Placenta actively transports phosphate into the fetus,
as reflected in the higher phosphate concentrations of
newborn
• Typically increases in women after menopause but
decreases in older adults

49. Intra Cellular Phosphate
• Free phosphate concentrations are generally
comparable to those in the extracellular
• Inside-negative electrical potential of the cell
creates a significant energy requirement for
translocation of phosphate into cells
• Accomplished through sodium phosphate co-
transport

50. Phosphate Metabolism &
Determinants

51. Intestinal absorption
• Avidly absorbed throughout the small
intestine, but especially in the jejunum
• Saturable, sodium-dependent process
responsive to vitamin D
• A non-saturable, sodium-independent
mechanism thought to represent para-cellular
diffusional transport : Mediated by NPT2b
transporters

52. Renal Regulation
• Controlled primarily by the rate of proximal renal
tubular phosphate reabsorption
• Sensitive to dietary phosphate intake also
• Integrated activity of the major sodium-dependent co-
transporters (NaPi-IIa and NaPi-IIc) : strongly down-
regulated by PTH and FGF23
• Hypophosphatemia due to increased urinary losses due
to decreased net renal tubular phosphate reabsorption

54. Role of PTH
• PTH rapidly (15 to 60 minutes) reduces the
number of NPT-IIa co-transporters on the apical
surface of the cells in the renal proximal tubule
• After parathyroidectomy twofold to threefold
increase in both protein and messenger RNA
(mRNA) levels of NPT-IIa, which correlates with a
striking increase in phosphate reabsorption

58. Phosphatonins

59. Phosphatonins basics
• Circulating factor that induces phosphaturia through PTH
independent mechanisms leading to hypophosphatemia
• Examples:
 FGF23
 MEPE
 SFRP4
 FGF7
• Hyperphosphaturia, hypophosphatemia and
rickets/osteomalacia
Phosphatonins. Peter J. Tebben, Theresa J. Berndt, Rajiv Kumar. Osteoporosis. http://dx.doi.org/10.1016/B978-0-12-415853-
5.00016-9

60. FGF-23
• Osteocytes & Osteoclasts expresses FGF23
• Low phosphate diet have significantly lower
serum FGF23 concentrations
• Effects of FGF23 are mediated through
fibroblast growth factor receptors (FGFR)
found on the cell surface
• Klotho proteins as co-factors

62. FGF23 actions
• Indirectly decreases phosphate transport in
the intestine by reducing serum
1α,25(OH)2D3 concentrations
• Mineralization defect with widened osteoid

63. Various Diseases caused by
abnormal FGF-23 metabolism

64. Autosomal Dominant
Hypophosphatemic Rickets (ADHR)
• Mutant FGF23 resistant to proteolytic cleavage
between residues 176 and 180
• Features:
 Hypophosphatemia and impaired bone mineralization
 Increased renal phosphate excretion, elevated ALP and
inappropriately low serum 1α,25(OH)2D3 relative to
the degree of hypophosphatemia
 Reduced Intestinal phosphate absorption as well

65. X-Linked Hypophosphatemic rickets
(XLHR)
• Mutations in the gene encoding PHEX, an
endo-peptidase on the X chromosome
• FGF23 production is increased in the absence
of functional PHEX
• However, not all patients with XLH have
elevated concentrations of FGF23 implying
that other factors may also be important
• Burosumab- Established therapy

66. Tumour Induced Osteomalacia (TIO)
• Studies using Serial Analysis of Gene Expression (SAGE)
demonstrated that in addition to FGF23, other phosphaturic factors
including MEPE, SFRP4, and FGF7 also highly expressed
• This may explain why not all patients with TIO have elevated serum
concentrations of FGF23
• Serum FGF23 concentrations decline into the normal range shortly
after removal of the offending tumor
• Exhibit signs and symptoms of rickets/osteomalacia including bone
pain, fractures, and weakness
• Hemangiopericytoma is the most common histological type of
tumor associated
Jonsson KB, Zahradnik R, Larsson T, White KE, Sugimoto T, Imanishi Y, et al. Fibroblast growth factor 23 in oncogenic osteomalacia and X-
linked hypophosphatemia. N Engl J Med 2003;348(17):1656–63.

67. McCune Albright Syndrome
• Post-zygotic activating mutations in the
GNAS1 gene
• Some patients display renal phosphate
wasting
• Not related to elevated cAMP
• Intensity of FGF23 staining in bone tissue
negatively correlated with serum phosphorus
concentrations

68. CKD
• As renal function declines, serum phosphorus concentrations
increase and 1α,25(OH)2D3 concentrations decrease
• Serum FGF23 concentrations correlate positively with serum
phosphorus and with the fractional excretion of phosphorus in
some patients with CKD
• Decreased 1α,25(OH)2D3 may lead to increased PTH production
and contribute to secondary hyperparathyroidism
• FGF23 concentrations highly predictive of Adverse outcomes
Kazama JJ, Gejyo F, Shigematsu T, Fukagawa M. Role of circulating fibroblast growth factor 23 in the development of secondary
hyperparathyroidism. Ther Apher Dial 2005;9(4):328–30

69. Tumoral Calcinosis
• Mutations in the GALNT3 gene or the FGF23 gene
• Hyperphosphatemia, increased renal
reabsorption of phosphorus, and normal or
elevated 1α,25(OH)2D3
• Altered processing of the mutant form of FGF23
(S71G)
• Reduced detection of intact and N-terminal
FGF23 by western blotting
• Dramatic extra-skeletal mineral deposits

71. SFRP4
• Antagonize the Wnt pathway as demonstrated by
reduced β-catenin and increased phosphorylated β-
catenin expression
• Highly expressed in some TIO tumors
• SFRP4 inhibits sodium-dependent phosphate transport
in OK cells
• PTH is not necessary for sFRP4 to induce phosphaturia
• Expectedly low levels of 1,25 (OH)D3
Berndt T, Craig TA, Bowe AE, Vassiliadis J, Reczek D, Finnegan R, et al. Secreted frizzled-related protein 4 is a potent
tumor-derived phosphaturic agent. J Clin Invest 2003;112(5):785–94

72. MATRIX EXTRACELLULAR
PHOSPHOGLYCOPROTEIN (MEPE)
• Expressed in osteoblasts and osteocytes
• Correlated positively with serum phosphate concentrations
during skeletogenesis and during fracture repair within
fibroblast-like cells, chondrocytes and osteocytes
• Has been shown to play a role in XLHR
• Highly expressed protein in many tumors causing TIO
• 1,25 (OH)D3 don’t decline
• Releases a peptide containing an ASARM sequence that is
capable of inhibiting mineralization
Argiro L, Desbarats M, Glorieux FH, Ecarot B. MEPE, the gene encoding a tumor-secreted protein in oncogenic
hypophosphatemic osteomalacia, is expressed in bone. Genomics 2001;74(3):342–51

73. FGF7
• TIO tumors that abundantly expressed FGF7
• Exposure of OK cells to FGF7 reduced
phosphate transport in a dose dependent
manner
• FGF7 may be involved in normal phosphate
homeostasis and regulated by vitamin D
Carpenter TO, Ellis BK, Insogna KL, Philbrick WM, Sterpka J, Shimkets R. Fibroblast growth factor 7, an inhibitor of phosphate transport
derived from oncogenic osteomalacia-causing tumors. J Clin Endocrinol Metab 2005;90(2):1012–20

75. Take Home Messages
• Calcium & Phosphate metabolism is inter-linked &
determined by many factors
• PTH, Vit-D along with the Phosphatonins are the chief
metabolic determinants
• The various diseases linked to PTH functioning
abnormalities needed to be understood to diagnose &
treat properly
• Phosphatonins related diseases are not that
uncommon & pathophysiology needs to be understood
for treatment

76. Thank you………………