CHAPTER 39

Disorders of Bone and

Mineral Metabolism
 Calicum
• Calcium is the most common element in the body and the most
prevalent cation
• Bone contains (1) nearly all of the calcium (99%), (2) most of the
phosphate (85%), and (3) much of the magnesium (55%) of the body.
• Calcium exists in three physicochemical states in plasma with
approximately (1) 50% free (ionized), (2) 40% bound to plasma
proteins, primarily albumin, and (3) 10% complexed with small anions
• The free calcium fraction is the biologically active form. Its
concentration in plasma is tightly regulated by the calcium regulating
hormones PTH and 1,25(OH)2D.
• Growth hormone (GH), glucocorticoids e.g. cortisol), oestrogens,
testosterone and thyroid hormones (thyroxine (T4) and tri‐
iodothyronine (T3)) also influence calcium metabolism.
• Intracellular calcium has key roles in many important physiological
functions, including (1) muscle contraction, (2) hormone secretion, (3)
glycogen metabolism, and (4) cell division.
• Extracellular calcium is needed or (1) bone mineralization, (2) blood
coagulation, and (3) other functions .A decrease in the serum free
calcium concentration causes increased neuromuscular excitability
and tetany. An increased concentration reduces neuromuscular
excitability.
• In infancy and childhood, there is usually a positive balance, especially at
times of active skeletal growth. In older age, calcium output may exceed
input, and a state of negative balance then exists; this negative external
balance is particularly marked in women after the menopause, and is
important in the development of post‐menopausal osteoporosis. In
women, calcium is lost during pregnancy to the foetus, and by lactation
during breastfeeding
• There is a constant state of turnover in the skeleton associated with
deposition of calcium in sites of bone formation and release at sites
• of bone resorption (~5% per year of the adult skeleton is remodelled).
Calcium in the bone also acts as a reservoir that helps to stabilise ECF Ca2+
 Hypocalcemia
• If potentially misleading hypocalcaemia due to either contamination of
the sample with EDTA (from a full blood count tube) or decreased
serum albumin is first excluded, then the hypocalcaemia must be
pathological and must result from a decrease in plasma Ca2+ .
• Hypoalbuminemia is the most common cause o decreased total calcium
with normal free calcium (sometimes called pseudohypocalcemia);
Common clinical conditions associated with low serum albumin include
(1) chronic liver disease, (2) nephrotic syndrome, (3) congestive heart
failure, and (4) malnutrition. In such states it is preferable to measure
free calcium with a specific assay or, i such is not available, to use an
adjustment calculation or the total calcium concentration
• Tetany is the symptom that classically suggests the presence of a low
plasma Ca2+. and may also be caused by a rapid fall in plasma H+ (e.g.
acute respiratory alkalosis produced by hyperventilation or IV infusion
of NaHCO3). Occasionally it is due to a low plasma Mg2+ , and rarely
it is due to a sudden increase in plasma phosphate. Neuropsychiatric
symptoms and cataract are other possible consequences of
hypocalcaemia
 Causes of hypocalcemia
• Vitamin D deficiency
• The most common pathological cause of hypocalcaemia is defective calcium absorption due
to inadequate plasma levels of 1,25‐DHCC. Deficiency of 1,25‐DHCC may result from lack of
vitamin D or failure at any stage in its conversion to 1,25‐DHCC; rarely, the action of 1,25‐
DHCC is defective at the receptor level.
• Vitamin D defeciency is characterized by low serum 25(OH)D, high PTH (secondary
hyperparathyroidism), and high serum alkaline phosphatase (ALP).

• Serum phosphate is often low, partly through impaired absorption, but also as a result of the
secondary hyperparathyroidism (renal disease is an exception).
• Serum ALP activity is often increased, reflecting increased osteoblastic activity. However, its
measurement is of limited diagnostic value in childhood because of the marked physiological
variations in activity that normally occur in this age group
• In chronic renal ailure, (1) hypoproteinemia, (2) hyperphosphatemia,
and (3) low serum 1,25(OH)2D (caused by reduced renal synthesis)
contribute to hypocalcemia
• Rapid remineralization o bone a er (1) surgery or primary
hyperparathyroidism (hungry bone syndrome), (2) treatment or
hyperthyroidism, or (3) treatment or hematological malignancy may
result in hypocalcemia. Acute pancreatitis is requently complicated by
hypocalcemia
• Hypoparathyroidism
• Primary hypoparathyroidism is rare. The combination of a reduced serum calcium
and an increased serum phosphate in a patient who does not have renal disease
suggests the diagnosis of hypoparathyroidism; serum ALP activity is usually
normal. Measurement of intact PTH confirms the diagnosis; levels are reduced
and are sometimes undetectable even by the most sensitive assays. Failure to
secrete PTH may be a secondary complication of thyroid or parathyroid surgery,
or it may be familial or sporadic in origin. Also, the parathyroid glands may be
destroyed by an autoimmune process, or as a result of infiltration by carcinoma of
the thyroid or other neoplasms.
• • Secondary hypoparathyroidism may occasionally be observed in patients with
magnesium deficiency
• • Pseudohypoparathyroidism is a rare but interesting condition in which the end‐
organ receptors in the bone and kidneys fail to respond normally to PTH. Patients
with pseudohypoparathyroidism have increased serum PTH.
 hypercalcemia
•  Increased plasma Ca2+ is a potentially serious problem that can lead to renal
damage, cardiac arrhythmias and general ill‐health.

• Hypercalcemia is caused by (1) increased intestinal absorption, (2) increased renal

retention, (3) increased skeletal resorption, or (4) a combination o mechanisms.
• The most common causes are primary hyperparathyroidism and malignant disease
• In asymptomatic ambulatory patients with hypercalcaemia, primary
hyperparathyroidism may account for up to 80% of cases whereas in sick
hospitalised hypercalcaemic patients, malignancy‐associated hypercalcaemia is
more likely
• these two disorders account or 90% to 95% o all cases o hypercalcemia
 Causes of hypercalcemia
• Primary hyperparathyroidism
• Primary hyperparathyroidism is most often caused by an adenoma, but may be
caused by hyperplasia involving multiple parathyroid glands, or, rarely, by parathyroid
carcinoma.
• Greater than 80% o hyperparathyroid patients are relatively free of symptoms on
presentation because of the early detection o this disorder by the widespread use of
chemistry testing panels that include calcium
• Chronic hypercalcemia with hypercalciuria has been known to lead to formation of
calcium-containing kidney stones, which, in some cases, leads to slowly developing
renal failure.
• The majority o patients with primary hyperparathyroidism (>60%) are
postmenopausal women
• Both serum calcium and albumin should be measured, and may need
to be repeated, since the hypercalcaemia can be intermittent. Serum
phosphate is sometimes low as a result of the phosphaturic effect of
PTH, though this is not a reliable finding. A mild metabolic acidosis
may be present, since PTH increases urinary HCO3 losses.
• . Measurement o intact PTH (with concomitant measurement o
calcium) is the most sensitive and specific test or parathyroid unction
and is central to the differential diagnosis o hypercalcemia
• 90% of patients with primary hyperparathyroidism have an elevated
level. Approximately 10% of patients with primary
hyperparathyroidism may have serum intact PTH in the upper part of
the reference range. However, in the setting of persistent
hypercalcaemia, such levels are considered to be inappropriately high
since serum intact PTH should be suppressed if hypercalcaemia is
unrelated to increased parathyroid activity. If serum intact PTH is
within reference limits or is only marginally elevated in an
asymptomatic hypercalcaemic patient, the diagnosis of familial benign
hypocalciuric hypercalcaemia (FBHH) should also be considered
• Serum 1,25(OH)2D concentration is usually above the middle of the
reference interval in primary hyperparathyroidism, as PTH stimulates its
production. By contrast, 1,25(OH)2D (like P H) is low-normal or
suppressed in nonparathyroid hypercalcemia, except in sarcoidosis,
other granulomatous diseases, and certain lymphomas in which the
pathological tissues contain the 25-hydroxyvitamin D-1α-hydroxylase
required to produce 1,25(OH)2D.
• PTH increases the renal clearance of bicarbonate and phosphate. In
hyperparathyroidism, a mild hyperchloremic metabolic acidosis is often
present, whereas in nonparathyroid hypercalcemia a mild
hypochloremic metabolic alkalosis is likely.
• Hypercalcaemia of malignancy
• Hypercalcemia occurs in 5% to 30% o individuals with cancer. tumors
most commonly cause hypercalcemia by (1) producing parathyroid
hormone–related protein (PTHrP), which is secreted into the
circulation and stimulates bone resorption and/or (2) invasion o the
bone by metastatic tumor, which produces local factors that stimulate
bone resorption.
• Serum intact PTH is usually suppressed in patients with malignancy‐
associated hypercalcaemia, although it sometimes falls within the
lower part of the reference range.
• Solid tumours that have metastasised to bone may cause hypercalcaemia by paracrine
activation of osteoclasts. The tumour cells may also increase bone resorption directly.
• • Some solid tumours (e.g. carcinoma of the lung, head and neck), in the absence of
bony metastases, may give rise to hypercalcaemia. An important factor in this humoral
hypercalcaemia of malignancy is PTHrP, a peptide with marked sequence homology
with PTH that also acts through the PTH receptor. PTHrP may also be secreted by some
tumours that metastasise to bone, so that humoral and local osteolytic mechanisms
may combine to produce hypercalcaemia. True ectopic production of PTH appears to
be rare.
• • In multiple myeloma, hypercalcaemia appears to result from the release of local
cytokines that promote local bone resorption. Lymphomas may also cause
hypercalcaemia.
• Vitamin D excess
• Increased serum 1,25‐DHCC and hypercalcaemia may result from excessive vitamin
D intake or if overdosage with 25‐HCC, 1α‐HCC or 1,25‐DHCC occurs. Measurement
of serum 25‐HCC or 1,25‐DHCC confirms the diagnosis
• Drugs
• thiazide diuretics; these interfere with renal calcium excretion. Long‐term lithium
therapy may also be a cause, possibly by stimulating PTH secretion. Serum calcium
should be monitored on a regular basis in patients receiving lithium therapy
• Sarcoidosis
• About 10–20% of patients with sarcoidosis may have hypercalcaemia, often only
intermittently. More often, they have hypercalciuria. The unregulated conversion of
25‐HCC to 1,25‐DHCC (and consequently increased intestinal absorption of calcium)
by sarcoid tissue macrophages is responsible. Hypercalcaemia may show seasonal
variation in parallel with the production of vitamin D3 sunlight.
• 
• Tertiary hyperparathyroidism
• This description refers to the development of parathyroid hyperplasia
as a complication of previously existing secondary
hyperparathyroidism. This diagnosis needs to be considered in patients
with renal failure or intestinal malabsorption if they develop
hypercalcaemia that is not attributable to treatment with vitamin D or
one of its hydroxylated derivatives (usually 1α‐HCC or 1,25‐DHCC).
Serum calcium is almost always increased and serum PTH
inappropriately high. Unlike primary hyperparathyroidism, however,
fasting serum phosphate may be increased, especially if tertiary
hyperparathyroidism develops in a patient with renal failure
• Familial benign hypocalciuric hypercalcaemia (FBHH)
• inactivating mutation in the calcium‐sensing receptor gene in the parathyroid gland,
kidney and other organs, and results in a high plasma Ca2+ that is sensed as ‘normal’,
• with normal or marginally elevated plasma PTH. It is important that FBHH is distinguished
from primary hyperparathyroidism since parathyroidectomy does not reduce plasma Ca2+
• , and no active treatment is indicated. While there is no single biochemical test that can
always distinguish, unequivocally, FBHH from primary hyperparathyroidism, in patients
with FBHH urinary calcium is usually low and serum magnesium tends to be high normal.
Therefore, a combination of family studies and the measurement of calcium excretion
together with serum magnesium is helpful in identifying the condition. Calcium excretion
is measured on a second void spot urine and blood sample obtained after an overnight
fast, and is calculated by multiplying the urine calcium : creatinine ratio (both in mmol/L)
by the serum creatinine (in µmol/L). Calcium excretion ≤14 µmol/L of glomerular filtrate
suggests FBHH whereas calcium excretion ≥0.27 µmol/L suggests PHPT. If uncertainty
remains after these initial investigations, testing for mutations in the calcium‐sensing
receptor gene may be performed.
• Endocrine disorders
• Hypercalcaemia has been reported occasionally in association with
hypoadrenalism, phaeochromocytoma and thyrotoxicosis.

• Milk–alkali syndrome
• Milk consumption may be excessive in patients with symptoms of
peptic ulceration; calcium intake is correspondingly increased. If this is
accompanied by excessive intake of alkali (e.g. NaHCO3), as an antacid
hypercalcaemia may develop. The alkali is thought to reduce urinary
calcium excretion and to be important in the pathogenesis of the
condition.
 Measurement of Calcium
• The term ionized calcium is a misnomer, because all plasma or serum
calcium is ionized, whether or not it is associated with protein or small
anions
• Measurement of Total Calcium
• include (1) spectrophotometric methods, (2) ionspecific electrode (ISE)
methods , and (3) atomic absorption methods.
• Spectrophotometric Methods: Although less accurate than atomic
absorption spectrometry, they have been easier to automate.
• 1- o-Cresolphthalein Complex one Method.
• 2- Arsenazo III Method: Unlike CPC, arsenazo III is stable as a single reagent.
 Specimen Requirements
• Serum and heparinized plasma are the preferred specimens or the
measurement of total calcium. Citrate, oxalate, and
ethylenediaminetetraacetic acid (EDTA) anticoagulants should not be
used because they interfere by forming complexes with calcium
• Urine specimens should be preserved by adding 20 to 30 mL of 6
mol/L HCl per 24-hour specimen (1 to 2 mL or a random specimen) to
prevent calcium salt precipitation. Addition of acid after the collection
may not completely dissolve precipitated calcium salts.
 Interferences
• Hemolysis, icterus, lipemia, paraproteins, magnesium, and gadolinium
chelates in contrast agents have been reported to interfere with
photometric methods.
• hemolysis causes a negative error because red blood cells contain lower
concentrations of calcium than does serum, more significant errors may be
caused by the spectral interference of hemoglobin. Depending on the
method, hemoglobin has been reported to produce either negative or
positive interference.
• Gadolinium magnetic resonance contrast agents cause a significant error
(usually underestimation) in spectrophotometric methods
• 
 Adjus ted or Correc ted Total Ca lcium

• Adjusted Total Calcium (mg/dL) = Total Calcium (mg/dL) + 0.8 [4 −

Albumin (g/dL)]
• Adjusted Total Calcium (mmol/L) = Total Calcium (mmol/L) + 0.02 [40
− Albumin (g/L)]
• Some factors limiting the ability o total and adjusted calcium to
predict free calcium are listed in Box 39-3. When possible,
mathematical adjustments or corrections should be replaced by direct
determination of free calcium.
 Measurement o Free Calcium
• ISEs are widely used or the rapid measurement of free calcium,
electrolytes, and blood gases . Calcium ISEs contain a calcium-
selective membrane and an internal reference electrode. Modern
calcium ISEs use liquid membranes containing the ion-selective
calcium sensor dissolved in an organic liquid trapped in a polymeric
matrix. Neutral carriers (e.g., E H 1001) are the most commonly used
calcium sensors, ollowed by ion exchangers, such as
organophosphate. temperature affects electrode response and the
extent of calcium binding by protein and small anions. Most free
calcium analyzers adjust and maintain samples at 37 °C,
 Interferences
• Wide variations, however, in the concentration of Na+ and high
concentrations of Mg2+ and Li+ may influence the apparent
concentration of free calcium
• Physiological anions, including (1) protein, (2) phosphate, (3) citrate,
(4) lactate, (5) sulfate, and (6) oxalate, and chemicals, such as (7) EDTA
and (8) EGTA, Form complexes with calcium and reduce Free calcium.
 Effect of pH

• Increasing the pH of a specimen in vitro increases the ionization and negative

charge on albumin and other proteins, leading to an increase in protein bound
calcium and a decrease in free calcium. Decreasing pH in vitro decreases
ionization and negative charge, decreasing protein-bound calcium and increasing
free calcium. Free calcium changes by about 5% or each 0.1 unit change in pH

• Effects of Anticoagulants
• Heparin is the only acceptable anticoagulant for free calcium determinations
• Citrate, oxalate, and EDTA bind calcium and unacceptably decrease free calcium
concentrations.
 Specimen Requirements
• Specimens or free calcium measurement must be collected and handled anaerobically
and promptly to minimize alterations in pH and free calcium due to the loss of CO2 and
the metabolism of blood cells. Syringes and evacuated tubes should be filled completely
and sealed to prevent the loss of CO2 (increase in pH).
• Free calcium is measured in (1) heparinized whole blood, (2) heparinized plasma, or (3)
serum
• specimens are analyzed within 30 minutes, heparinized whole blood may be preferable
because it reduces processing time and specimen volume requirements and avoids the
alteration in pH associated with centrifugation at temperatures other than 37 °C. Free
calcium is reported to be stable in whole blood specimens for 1 hour at room
temperature and for 4 hours at 4 °C
• Aerobic handling of specimens and correction of the free calcium to pH 7.4 may be
misleading in patients with alkalosis or acidosis and should be avoided
 Patient Preparation and Sources of Preanalytical Error for Total
and Free Calcium Measurements

• A common and important source of preanalytical error is the increase in

total, but not free, calcium associated with tourniquet use and venous
occlusion during sampling. Errors of 0.5 to 1.0 mg/dL or 0.12 to 0.25 mmol/L
in total calcium may result because of the increase in protein-bound calcium
caused by the efflux of water from the vascular compartment during stasis.
If a tourniquet is required, it should be applied just be ore sampling and
released within 1 minute. Fist clenching or other forearm exercise should be
avoided before phlebotomy because forearm exercise causes a decrease in
pH (lactic acid production) and an increase in free calcium. Changes in
posture cause fluid shifts and alter the concentration o cells and large
molecules, including albumin and total calcium (as part o it is protein-
bound) in the vascular compartment.
• Postural changes of about 10% in the concentrations of albumin and
other proteins are common. The hemodilution caused by recumbency
(along with hypoalbuminemia) contributes to the increased
prevalence of hypocalcemia (total, but not free calcium) often
observed in hospital patients.
• prolonged immobilization and bed rest lead to bone resorption and
increase total and free calcium in blood. Hyperventilation and
exercise decrease and increase the concentration of free calcium,
respectively, because of changes in serum pH.
 Reference Intervals
• The reference interval or total calcium in adults is approximately 8.6 to
10.3 mg/dL or 2.15 to 2.57 mmol/L.
• free calcium in adults is about 4.6 to 5.3 mg/dL or 1.15 to 1.33 mmol/L.
• total calcium declines in parallel with serum albumin during pregnancy,
whereas free calcium remains unchanged
• Because of the dependence of free calcium on pH, it is recommended
that pH be measured and reported with all free calcium determinations.
• The desirable analytical coefficients of variation, based on within-person
biological variation, are 0.9% and 1.0% or less, respectively, for free and
total calcium
 Phosphate metabolism
• Intracellular phosphate has vital functions in macromolecular
structure (e.g. in DNA), energy metabolism (e.g. energy‐rich
phosphates such as ATP), cell signalling and enzyme activation by
phosphorylation
 Hypophos phatemia
• usually <2.5 mg/dL or <0.81 mmol/L, is relatively common in hospitalized patients
(approximately 2%).
• Hypophosphatemia or phosphate depletion may be caused by (1) a shift of
phosphate from extracellular to intracellular spaces, (2) renal phosphate wasting,
(3) decreased intestinal absorption, and (4) loss from intracellular phosphate
• The antacids bind phosphate, thus hindering its absorption. The hypophosphatemia
and phosphate depletion in patients with malabsorption may be more closely
related to their secondary hyperparathyroidism (and resulting loss o phosphorus in
urine) than to malabsorption o phosphate.
• Diabetic ketoacidosis is associated initially with high- normal to increased serum
phosphate. treatment o the ketosis and acidosis with insulin and intravenous fluids,
however, results in a rapid decrease in the serum phosphate concentration
• Plasma concentrations <1.5 mg/dL or <0.48 mmol/L may produce clinical
manifestations
• Other conditions that may occur in phosphate depletion include (1) muscle
• weakness, (2) acute respiratory failure, and (3) decreased cardiac output. At very
low serum phosphate (<1 mg/dL or <0.32 mmol/L), rhabdomyolysis may occur.
Phosphate depletion in erythrocytes decreases erythrocyte 2,3-
diphosphoglycerate, which causes tissue hypoxia because of increased affnity of
hemoglobin for oxygen. Severe hypophosphatemia (serum phosphate
concentration <0.5 mg/dL or <0.16 mmol/L) may result in hemolysis of red blood
cellshypoxia.
• If hypophosphatemia is chronic, rickets (in children) and osteomalacia (in adults)
may develop.
 Hyperphos phatemia
• Hyperphosphatemia is usually secondary to the inability o the kidneys to
excrete phosphate, as in renal failure. Moderate increases of serum
phosphate occur in individuals with (1) low PTH (hypoparathyroidism), (2)
PTH resistance (pseudohypoparathyroidism), or (3) acromegaly (increased
growth hormone), caused by an elevated renal phosphate threshold. A
rapid increase in serum phosphate may be associated with hypocalcemia.
• There ore symptoms may include (1) tetany, (2) seizures, and (3)
hypotension. Long-term hyperphosphatemia may be associated with (1)
secondary hyperparathyroidism, (2) osteitis fibrosa, and (3) so tissue calcifi
cation of the kidneys, blood vessels, cornea, skin, and periarticular tissue.
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