Accepted Manuscript

Hypocalcemic disorders

Erin Bove-Fenderson, PhD, Michael Mannstadt, MD

PII: S1521-690X(18)30074-5
DOI: 10.1016/j.beem.2018.05.006
Reference: YBEEM 1216

To appear in: Best Practice & Research Clinical Endocrinology & Metabolism

Please cite this article as: Bove-Fenderson E, Mannstadt M, Hypocalcemic disorders, Best Practice &
Research Clinical Endocrinology & Metabolism (2018), doi: 10.1016/j.beem.2018.05.006.

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 ACCEPTED MANUSCRIPT

Hypocalcemic disorders

Erin Bove-Fenderson, PhD

Michael Mannstadt, MD

Endocrine Unit, Massachusetts General Hospital and Harvard Medical School,

Thier 1101, 50 Blossom St, Boston MA 02114, USA
Corresponding author: Michael Mannstadt; mannstadt@mgh.harvard.edu

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Abstract

Calcium is vital for life, and extracellular calcium concentrations must constantly be maintained within a

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precise concentration range. Low serum calcium (hypocalcemia) occurs in conjunction with multiple disorders
and can be life-threatening if severe. Symptoms of acute hypocalcemia include neuromuscular irritability,
tetany, and seizures, which are rapidly resolved with intravenous administration of calcium gluconate.

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However, disorders that lead to chronic hypocalcemia often have more subtle manifestations.
Hypoparathyroidism, characterized by impaired secretion of parathyroid hormone (PTH), a key regulatory
hormone for maintaining calcium homeostasis, is a classic cause of chronic hypocalcemia. Disorders that

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disrupt the metabolism of vitamin D can also lead to chronic hypocalcemia, as vitamin D is responsible for
increasing the gut absorption of dietary calcium. Treatment and management options for chronic
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hypocalcemia vary depending on the underlying disorder. For example, in patients with hypoparathyroidism,
calcium and vitamin D supplementation must be carefully titrated to avoid symptoms of hypocalcemia while
keeping serum calcium in the low-normal range to minimize hypercalciuria, which can lead to renal
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dysfunction. Management of chronic hypocalcemia requires knowledge of the factors that influence the
complex regulatory axes of calcium homeostasis in a given disorder. This chapter discusses common and rare
disorders of hypocalcemia, symptoms and workup, and management options including replacement of PTH in
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hypoparathyroidism.

Keywords
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Parathyroid hormone
Hypocalcemia
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Hypoparathyroidism
Vitamin D
Pseudohypoparathyroidism
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Introduction
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Calcium is the most abundant mineral in the human body and is vital for life. Muscle function, nerve activity,
and bone mineralization depend on a precise balance of extracellular and intracellular calcium. Under normal
circumstances, serum calcium is maintained at a concentration of 8.5-10.5 mg/dL [1]. Concentrations outside
of this small window result in significant health consequences. Therefore, a sensitive mechanism of hormonal
regulation is required. This is achieved by the parathyroid gland, an endocrine organ with the specific function
of sensing small fluctuations in extracellular calcium and responding with changes in production of parathyroid
hormone (PTH), which acts in conjunction with active vitamin D to regulate calcium.

PTH is an 84-amino acid peptide hormone that increases serum calcium by three distinct mechanisms: it
increases reabsorption of calcium from the primary filtrate in the kidneys, increases bone turnover to release
calcium (and phosphate) from the bones, and enhances the conversion of 25-hydroxyvitamin D (25(OH)D) to
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the active 1,25-dihydroxyvitamin D (1,25(OH)2D), which potently increases calcium absorption in the gut. All of
these actions of PTH are mediated in target organs by the PTH/PTHrP receptor (PTH1R), a family B G-protein
coupled receptor. The PTH1R signals through several different heterotrimeric G-proteins, most strongly
through Gs and Gq [2] which triggers the activation of multiple signaling pathways including the
Gsα/cAMP/protein kinase A (PKA) and the Gqα/PLC-β/PKC pathways, respectively. Key downstream effects of
this activation include changes in expression and localization of ion transporters, activation of bone
resorption, and increased expression of CYP27B1 (1-α hydroxylase), the enzyme that converts 25(OH)D to
1,25(OH)2D.

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Hormonal regulation of blood calcium concentrations is achieved through a classic negative feedback loop [3].
Extracellular calcium binds to and activates the calcium sensing-receptor (CASR), a family C G-protein coupled
receptor with high expression on the surface of parathyroid and kidney cells. When calcium concentrations

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are high, the activated CASR engages G-protein Gi and Gq/11 signaling, which leads to the inhibition of PTH
secretion [4, 5]. When serum calcium concentrations are low, the activity of the calcium-sensing receptor is
reduced, leading to the release of PTH into circulation. This system is extremely sensitive to small fluctuations

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in serum calcium within the physiological range, resulting in large changes in PTH. When the PTH response is
plotted against changes in serum calcium, a steep inverse sigmoidal curve is demonstrated [3]. Low or falling
levels of serum calcium trigger the release of PTH within minutes; conversely, high levels lead to rapid
suppression of PTH secretion. In addition to regulating secretion of PTH, longer-term responses to

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hypocalcemia include decreased intraparathyroidal PTH degradation, increased PTH mRNA stability and
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expression, and proliferation of parathyroid cells [6]. When hypocalcemia becomes a chronic condition, for
example in a case of dietary deficiency of calcium or vitamin D, these changes can lead to secondary
hyperparathyroidism.
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A second major calciotropic hormone is 1,25(OH)2D, the active form of vitamin D. In contrast to PTH, the
regulation of serum calcium by vitamin D has a slow response time. PTH stimulates conversion of 25(OH)D to
the active 1,25(OH)2D in the proximal renal tubule of the kidney; in target organs, 1,25(OH)2D binds to its
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receptor (VDR) in the cell cytoplasm, which then translocates to the nucleus. There, the ligand-receptor
complex increases the transcription of target genes leading to the production of new proteins. In the
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intestines, these are proteins necessary for calcium transport. [7, 8] Like calcium, 1,25(OH)2D suppresses PTH
forming a negative feedback loop [6].

Most (99%) of total body calcium is stored as hydroxyapatite in bone; the remaining 1% is present in the
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extracellular fluid and soft tissues [9]. Of the small fraction of total calcium circulating in the blood,
approximately 50% is bound to proteins (primarily albumin), and only about 50% is in a biologically available
ionized form [10]. Measuring ionized calcium is, in theory, the more precise method for identifying
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hypocalcemia. However, the measurement of ionized calcium requires special handling of the samples
immediately after blood is drawn, which is difficult to accomplish in many circumstances and can be a source
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of inaccurate measurements. A practical and reasonable alternative in most clinical settings is to determine
albumin concentrations at the same time as total calcium and to use this measurement to correct for albumin.
The commonly used formula (adjusted [Ca](mg/dL) = total [Ca] (mg/dL) + 0.8(4 – [albumin] (g/dL)))[11] is
reasonably accurate but has limited utility in special situations, for example acute and chronic renal failure,
critical illness, or other conditions associated with abnormal blood pH [12, 13].

Signs and symptoms of hypocalcemia

Hypocalcemia can be asymptomatic, especially if it is mild or if the patient is used to chronically low calcium
levels [14]. Large interindividual differences in the threshold below which symptoms might occur have been
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observed; both the absolute level of ionized (or albumin-corrected) calcium and the rate of decline can
influence that threshold. Symptoms can range from mild complaints (which may not raise the suspicion of
hypocalcemia) to severe and potentially life-threatening presentations such as laryngeal spasm and seizures
[15]. The most common clinical presentation of hypocalcemia is increased neuromuscular irritability, classic
symptoms of which include perioral numbness, tingling in the hands and feet, and muscle spasms.
Hypocalcemia can also be associated with prolongation of the QTc intervals on ECG, and, when profound, can
lead to reversible cardiomyopathy on rare occasions [16-18].

Chvostek’s and Trousseau’s signs, classic signs of neuromuscular irritability, can be elicited during examination

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when hypocalcemia is suspected [19, 20]. A positive Chvostek’s sign results in characteristic twitching of the
muscles of the face in response to tapping of the ipsilateral facial nerve. Trousseau’s sign is evoked by inflating
a blood pressure cuff on the upper arm above systolic blood pressure for 3 minutes, and manifests as carpal

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spasm, with wrist and thumb flexion. The sensitivity and specificity of Chvostek’s sign for identification of
hypocalcemia are not high, as studies report that 25% of healthy individuals have a positive Chvostek sign, and
29% of patients with hypocalcemia were negative [19]. Trousseau’s sign is reported to be more specific, with

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only 1% of healthy individual showing a positive Trousseau.

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1. PTH-related disorders
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This group encompasses disorders that are characterized by hypocalcemia due to impairment in PTH
production (hypoparathyroidism), calcium sensing (autosomal dominant hypocalcemia), or target organ
resistance to PTH (pseudohypoparathyroidism). These disorders are summarized in table 1.
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Hypoparathyroidism
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Etiologies
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1.1 Post-surgical hypoparathyroidism

One of the most common causes of hypocalcemia is hypoparathyroidism, which can be transient or persistent.
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Transient hypoparathyroidism can be seen in some patients immediately after thyroid or parathyroid surgery
and is characterized by reversible hypocalcemia, often with hyperphosphatemia, and PTH levels that are
inappropriately low. It can affect up to 30% of patients following total thyroidectomy [21]. Recovery of
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parathyroid function often occurs within the first days after surgery, but it may take months [22]. If the
condition persists for more than 6 months, it is usually defined as chronic hypoparathyroidism. In the United
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States, the prevalence of chronic hypoparathyroidism based on diagnosis of insured patients was estimated to
be about 60,000; adjustments to include the uninsured population yield an estimate of 77,000 persons [23].
The most common cause of chronic hypoparathyroidism is surgery, which accounts for about 75% of cases
[24]. Postsurgical hypoparathyroidism is caused by the accidental removal of the parathyroid glands or by
intraoperative damage to the parathyroids and/or their blood supply [14]. Chronic hypoparathyroidism is a
rare outcome of neck surgery; it occurs in up to 3% of patients who have undergone total thyroidectomy [25].
Risk factors for the development of hypoparathyroidism after surgery include low surgical volume of the
surgeon, re-operation, extent of the surgery, failure to identify the parathyroid glands during surgery, and
Graves’ disease [26, 27]. Several strategies have been developed over the last few years to decrease the risk of
postsurgical hypoparathyroidism. These include improved preoperative imaging modalities to locate
parathyroid adenomas for surgical parathyroidectomy, intraoperative PTH monitoring, and intraoperative
autotransplantation of damaged parathyroid glands [28-30].
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Non-surgical causes of hypoparathyroidism, listed in table 1 and described further below, include autoimmune
disorders and genetic disorders that affect the development of the parathyroid glands or impair PTH
production.

Table 1.
Etiologies of Nonsurgical Hypoparathyroidism
Autoimmune
Autoimmune Polyendocrine Syndome Type 1 (APS1)
Genetic

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Familial Isolated Hypoparathyroidism
GCM2
PTH

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X-linked
Autosomal Dominant Hypocalcemia (ADH)
ADH1

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ADH2
Complex disorders
DiGeorge Syndrome Type 1

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Hypoparathyroidism, Sensory Neural Deafness, Renal Dysplasia Syndrome (HDR)
Kenny-Caffey Syndrome Type 1
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Kenny-Caffey Syndrome Type 2
Kearns-Sayre Syndrome (Mitochondrial)
Mitochondrial Encephalomyopathy with Lactic Acidosis and Stroke-like Episodes (MELAS) Syndrome
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Mitochondrial Trifunctional Protein (MTP) Deficiency Syndrome

Hypo-/Hypermagnesemia
Other
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Wilson's Disease
Post-radioactive
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Hemochromatosis
Metastases
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1.2 Non-surgical hypoparathyroidism

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Autoimmune disorders
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Hypoparathyroidism is a classic part of autoimmune polyendocrine syndrome type 1 (APS1), also called
autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome (APECED). This autosomal-
recessive disorder is caused by loss-of-function mutations in the autoimmune regulator (AIRE) gene [31, 32],
which codes for a protein that is expressed in the thymus and involved in the negative selection of auto-
reactive immune cells [33]. The classic triad of the disease includes mucocutaneous candidiasis, Addison’s
disease, and hypoparathyroidism. The presence of any two of these components clinically defines the disease.
In patients with APS1, hypoparathyroidism is treated with calcium and vitamin D supplementation. When
chronic malabsorption is part of APS1, the management of hypoparathyroidism with oral medication can be
challenging.
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Idiopathic hypoparathyroidism in patients with autoimmune disorders without proven APS1 is thought to be
the consequence of autoimmune destruction of the parathyroid glands. However, no diagnostic criteria exist
to define this disorder.

Genetic disorders

Familial Isolated Hypoparathyroidism

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Familial isolated hypoparathyroidism (FIH) is a term used to describe cases of inherited forms of
hypoparathyroidism without features of complex diseases. The molecular genetics of FIH are diverse and

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include mutations in genes for PTH, the transcription factor glial cell missing 2 (GCM2), and the transcription
factor SOX3 [34]. Mutations of the PTH gene are relatively rare. They can impair translocation and/or cleavage
of the preproPTH hormone, which in turn impairs trafficking and secretion and leads to a cellular stress

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response [35-40]. GCM2 is a transcription factor that is essential for parathyroid gland development. Gcm2
knockout mice that lack parathyroid glands, and humans with homozygous inactivating or heterozygous
dominant-negative GCM2 mutations develop hypoparathyroidism at an early age [41-45]. X-linked
hypoparathyroidism is a rare form of the disease that is associated with a deletion-insertion near the SOX3

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gene on the X chromosome, which may have a key role in parathyroid gland development [46].
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Autosomal-dominant hypocalcemia (ADH) type 1

ADH is one of the most common causes of genetic hypoparathyroidism and includes type 1 (ADH1) and type 2
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(ADH2), each with distinct molecular etiologies. ADH1 is the more common of the two and is caused by
activating mutations of the calcium sensing receptor [47]. These mutations result in a heightened sensitivity of
the CASR to extracellular calcium, with consequent suppression of PTH secretion at lower blood calcium levels
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than normally required. This change in sensitivity produces a calcium response curve that is shifted to the left
in vitro and in mouse models bearing ADH-associated mutations [48]. ADH1 can occur sporadically or in the
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setting of autosomal dominant disease. More than 70 mutations have been found, a compilation of which can
be found at on the online Calcium Sensing Receptor Database (http://www.casrdb.mcgill.ca). Some of these
mutations are predicted to increase responsiveness of the receptor to the binding of calcium in the
extracellular domain, others increase cell surface expression of the CASR, or change the receptors’ affinity for
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intracellular binding partners such as calmodulin (CaM) [49]. Crystal structures of the extracellular domain of
the CASR predict that some activating mutations could affect cooperative binding of L-amino acids, inducing a
conformational switch that lowers the threshold for extracellular calcium activation [49, 50].
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Patients with ADH1 present with hypocalcemia, hyperphosphatemia, hypercalciuria, and PTH levels that are
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often detectable, but inappropriately low. Many patients may be asymptomatic, however severe
manifestations of hypocalcemia have been reported [51-53]. Treatment of ADH1 has some important caveats.
While the lack of PTH in the kidneys leads to a higher fractional excretion of calcium and hypercalciuria in
most hypoparathyroid patients who receive calcium supplementation, this problem is accentuated in patients
with ADH1 because of the additional hypercalciuric effect of the activated CASR expressed in the kidneys [47].
Monitoring, rather than treatment, is therefore recommended for patients with ADH who are asymptomatic,
as treatment with calcium and activated vitamin D further exacerbates the high risk of renal disease [53, 54].
In patients with symptoms who do require treatment, the target serum calcium should be as low as tolerated.
Thiazides can be used to lower urinary calcium [55], but should be used with caution as diuretics may increase
the risk of hypomagnesemia in some ADH1 patients [56].
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PTH replacement therapy is also an option for ADH1 patients and has been successfully used in trials of PTH(1-
34) [57, 58]. Patients with activating CASR mutations were excluded in the phase 3 RCT using PTH(1-84), so
the effects of treatment with the full-length peptide in this patient population are unknown. Calcilytics, a class
of negative allosteric modulators of the CASR, reduce receptor activity [59] and can increase PTH in animals
and humans. Calcilytics were initially developed for use in osteoporosis as an “endogenous PTH therapy”but
showed no significant effect on bone mineral density in clinical trials. These small molecules do have potential
use for patients with ADH1, as they directly target the molecular defect of the disorder [60, 61]. In mouse
models bearing disease-relevant mutations in the CASR gene, calcilytics were shown to increase serum
calcium without incurring hypercalciuria [62, 63], and a small trial in humans with ADH1 demonstrated a dose-

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dependent increase in PTH secretion [64]. However, further studies are needed to demonstrate a concomitant
increase in serum calcium in humans.

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ADH type 2

ADH type 2 is a rare disorder caused by mutations in GNA11, which encodes the alpha subunit of G11, an

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intracellular signaling molecule used by the CASR [65-69]. These mutations induce gain-of-function of G11 and
lead to consequences similar to activating mutations in the calcium-sensing receptor [65, 66, 68]. ADH2 is
characterized by hypocalcemia, hyperphosphatemia, and inappropriately low PTH levels, making it very similar
to ADH1 in presentation. Patients with ADH2 appear to have less pronounced elevations in calcium excretion,

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although there are few reported cases of the disease in which this has been observed [47]. Some case studies
report short stature compared to unaffected family members, implying that G11α may have functions in
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skeletal development [66, 69]. Patients are usually treated with calcium and calcitriol to alleviate
hypocalcemia. This disease is rare and no clinical studies on different treatment modalities have been
published. However, in mouse models of ADH2, calcilytics [70-72] and Gq/11-specific inhibitors[71] have been
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shown to be effective in raising serum calcium levels.

Hypoparathyroidism as part of complex disorders

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In addition to APS1 (discussed above), hypoparathyroidism is a dysfunction that is associated with several
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other syndromic disorders (Table 1). 22q11.2 deletion syndrome (DiGeorge syndrome type 1) is a relatively
common autosomal dominant disorder caused by a heterozygous microdeletion in chromosome 22 [73]. It can
lead to immunodeficiency, cardiac outflow tract abnormalities, and hypoparathyroidism. The loss of the
transcription factor TBX1, which is essential for the development of the parathyroid glands, is thought to be
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responsible for the hypoparathyroidism seen in this syndrome [74]. Hypoparathyroidism, sensorineural
deafness and renal abnormality (HDR) syndrome is an autosomal-dominant disorder caused by a loss-of-
function mutation in the gene for GATA3, a transcription factor with a prominent role in organogenesis of the
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parathyroid glands [75]. Kenny-Caffey syndrome is a rare disorder characterized by short stature,
osteosclerotic bone dysplasia, and hypoparathyroidism [76, 77]. The autosomal recessive (type 1) form is
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caused by mutations in tubulin-specific chaperone E (TBCE) [78] and the dominant form (type 2) by mutations
in FAM111A [79]. The products of both genes are likely involved in embryonic development of the parathyroid
glands.
Several rare mitochondrial disorders can also be associated with hypoparathyroidism (Table 1). The
mechanisms by which these genetic disorders lead to hypoparathyroidism are currently not well understood.

Hypo- and Hypermagnesemia

Both hypo- and hypermagnesemia can lead to low serum calcium, and measurement of serum magnesium is
therefore recommended in patients who present with hypocalcemia without known etiology [80, 81].
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Hypermagnesemia is rare and most often occurs in the setting of renal failure and/or with exogenous
administration of magnesium, for example as treatment for premature labor. Like calcium, magnesium acts as
an agonist for the CASR, although with 2-3-fold lower potency [82, 83]. In the parathyroid glands activation of
the CASR represses PTH secretion. Combined with functions of activated CASR in other tissues, including
decreased calcium reabsorption in the kidneys, hypermagnesemia can lead to functional hypoparathyroidism
and hypocalcemia. Predictably, in vitro studies have shown the elevation of extracellular magnesium results in
a decrease in PTH secretion in cultured parathyroid cells [84, 85]. Pregnant women who are supplemented
with magnesium sulfate to treat premature labor or toxemia of pregnancy can display decreased PTH and
frank hypocalcemia [86-89]. For patients with healthy kidneys cessation of magnesium supplementation along

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with administration of calcium gluconate is sufficient to relieve symptoms [89].

Hypomagnesemia is a more common biochemical finding in clinical settings. It can be a complication of

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gastrointestinal, renal and metabolic disorders, or a side effect of drug use (e.g. diuretics, proton pump
inhibitors) [90]. Rarely, it is associated with genetic disorders that directly disrupt maintenance of magnesium
homeostasis [91]. Paradoxically, hypomagnesemia, like hypermagnesemia, has been shown to impair

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secretion of PTH and lead to peripheral PTH resistance and the development of hypocalcemia [90, 92, 93].
Several hypotheses have been suggested to explain how low serum magnesium levels can contribute to the
inhibition of PTH secretion. Low magnesium concentrations may act intracellularly by directly stimulating G-
protein activation downstream of the CASR. Some data support this hypothesis; in cells expressing the CASR

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the presence of either pertussis toxin or a mutant form of Giα with a low affinity for magnesium is sufficient to
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block activation in the presence of low magnesium concentrations [94]. Treatment with calcium is
recommended for patients with hypomagnesemia who present with acute symptomatic hypocalcemia,
followed by intravenous administration of magnesium to restore levels to a normal range [90]. Conditions that
lead to magnesium wasting should be followed by biochemical assessment and possible continuing oral
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therapy in the form of magnesium supplements.

Several genetic disorders are known to cause hypomagnesemia with concurrent hypocalcemia [95]. Mutations
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in claudin-16 and -19 inhibit the reabsorption of calcium and magnesium in the thick ascending loop of the
kidney, causing familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC) [96-99].
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Hypocalcemia occurs despite elevated PTH levels, implicating renal wasting of calcium as the cause of low
serum calcium in this disease. FHHNC is a progressive disorder leads to nephrocalcinosis and end-stage renal
disease. Supplementation with magnesium and the use of thiazides to minimize calcium excretion has minimal
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effect; patients will eventually require renal replacement therapy or transplant [96, 100]. Hypomagnesemia
with secondary hypocalcemia (HSH) is a rare autosome recessive disorder caused by a mutation in the renal
magnesium transport protein TRPM6, which is responsible for both intestinal absorption and renal
reabsorption of magnesium [101-103]. Patients with HSH have low PTH levels secondary to significant
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hypomagnesemia. Therapy with oral supplementation of high amounts of magnesium citrate restores
magnesium levels and corrects functional hypoparathyroidism and hypocalcemia in these patients [93].
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Others

Rare causes of hypoparathyroidism include infiltrative diseases that can affect the parathyroid glands such as
Wilson’s disease, hemochromatosis, and tumor infiltration [15]. A few cases of radiation-induced
hypoparathyroidism have been reported, but the parathyroid glands are relatively radiation-resistant [25].
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Long-term complications of hypoparathyroidism

Of central concern in this patient population is the increased risk of renal disease, believed to be due to
hypercalciuria, which is a combined result of diminished renal reabsorption of calcium and standard treatment
with calcium and active vitamin D, which can further increase the filtered load [104, 105]. A large retrospective
cohort study with 120 patients with a mean follow-up time of 7.4 years showed that 38% of patients with a
24-hour urine result had hypercalciuria [105]. Roughly half of the patients had renal imaging done, and 31% of
those had kidney stones or nephrocalcinosis on imaging. Rates of chronic kidney disease were two to 17-fold
higher compared to age-appropriate historical controls from the NHANES study. Two large case-control

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studies from a national registry in Denmark confirmed the high rates of renal disease. In patients with post-
surgical hypoparathyroidism, the hazard ratio for developing kidney stones and renal insufficiency (after
adjustment for prior diabetes and renal disease) was 4.02 and 3.10, respectively [104]. In patients with non-

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surgical hypoparathyroidism, the hazard ratio for developing renal insufficiency was 6 [106].

Other findings, such as basal ganglia calcifications [107] and development of cataracts [108, 109], appear to be

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complications of long-term hypoparathyroidism and are usually permanent. These may be related to elevated
calcium-phosphate product as a result of chronic hyperphosphatemia, or to transient hypercalcemia - perhaps
occurring as little as several hours every week in patients on treatment [110].

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Bone health in hypoparathyroidism
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Although hypocalcemia itself does not generally lead to changes in skeletal health in adult patients,
hypoparathyroidism is associated with a distinct bone phenotype due to low or absent PTH, which is a key
hormone for stimulation of bone turnover. Bone markers tend to be at the lower end of normal in patients
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with hypoparathyroidism, indicating a decrease in resorption and formation [111-113], and tetracycline
labeling demonstrates a strong reduction in bone formation [114]. Hypoparathyroid patients often have
increased areal bone mineral density (BMD) compared to control groups, as analyzed by dual-energy X-ray
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absorptiometry (DXA) [105, 115]. This difference in BMD has been shown in both post-surgical and non-
surgical hypoparathyroidism [116-118]. Hypoparathyroidism is associated with structural abnormalities,
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detected by peripheral quantitative computed tomography (pQCT), which include an increase in cancellous
bone volume, trabecular and cortical width, and a decrease in cortical porosity and bone formation and
resorption rates [114, 119]. Other changes in microarchitecture include an increase in plate-like structure in
trabecular bone [120]. Women with hypoparathyroidism seem to be protected against the increased high
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bone turnover seen after menopause [121, 122]. However, it is unclear how these potential changes in bone
metabolism and microstructure impact fracture risk in either pre- or post-menopausal populations [25]. A
large, case-controlled study on a population in Denmark yielded conflicting results; no overall increased risk of
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any fracture was found in patients with postsurgical hypoparathyroidism [123], while there was lower risk for
upper extremity fracture. For the smaller group of non-surgical hypoparathyroid patients there was also no
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difference in the overall risk of fractures, but an increased risk for upper extremity fracture [118]. Predictions
of bone strength using finite element analysis (FEA) based on HRpQCT predicts no difference in bone strength
between hypoparathyroid patients and the general population [118, 119].

Conventional treatment and monitoring

Conventional treatment for hypoparathyroidism includes oral administration of calcium and active vitamin D
to increase serum calcium levels and decrease symptoms [25, 81, 124]. In addition to minimizing symptoms of
hypocalcemia, treatment is also focused on avoidance of long-term complications. Serum calcium targets are
low to low-normal because patients are at risk for renal complications from hypercalciuria, as discussed above
[104, 105]. The major challenge of this treatment goal is maintaining a calcium level that is high enough to
avoid significant symptoms of hypocalcemia, but low enough to avoid hypercalciuria and renal complications
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over time. Monitoring 24-hour urinary calcium excretion and targeting it below 250 in women and 300 mg/day
in men to minimize adverse renal effects are essential components of disease management. Measurement of
24-hour urinary calcium should be taken annually, or more frequently if changes in medications or other
circumstances require it. Thiazide diuretics are often prescribed to lower urinary calcium excretion. These are
most effective when paired with a low sodium diet [125]. Calcium supplements should be given spaced
throughout the day to avoid transient hypercalcemia [81]. Calcium carbonate is often the supplement of
choice for patients without impaired gastrointestinal absorption, as it contains more elemental calcium per
calcium salt and can minimize the number of tablets. Other goals include avoidance of an elevated calcium-
phosphate product, which can increase the risk of extraskeletal calcifications and might be responsible for

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basal ganglia calcifications.

Treatment with PTH

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Because of the risk for long-term complications, and the inability of oral calcium and calcitriol to replace the
functions of PTH, investigators studied the effects of PTH treatment in hypoparathyroid patients. Initially,

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these studies used human PTH(1-34), a truncated form for the full-length peptide. PTH(1-34) retains all known
functions of PTH, and is an approved anabolic agent for the treatment of severe forms of osteoporosis [126].
The rationale for this treatment approach is the hypothesis that PTH is superior to calcium and activated
vitamin D, by virtue of its ability to decrease urinary calcium associated with nephrolithiasis and

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nephrocalcinosis, and by minimizing the high serum calcium-phosphate product that may be responsible for
other extraskeletal complications. In initial studies, once-a-day injection of PTH(1-34) was shown to maintain
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serum calcium at target levels while decreasing urinary calcium [127]. Twice daily injection led to less
variability in serum calcium levels than once daily, and a lower daily dose of PTH was required to maintain
target levels [58]. Long-term studies comparing twice-daily PTH(1-34) injection to treatment with calcium and
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calcitriol demonstrated 24-hour urine calcium excretion within the normal range in the treatment group [128].
Both once-daily and twice-daily injection of PTH(1-34) resulted in a fluctuation of serum calcium levels and a
corresponding fluctuation in urinary calcium excretion over a 24-hour period, potentially due to the relatively
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short half-life of PTH(1-34) in circulation. In an attempt to create a more physiological calcium response, a
pump system for delivery of PTH(1-34) was tested against twice-daily injection [129]. Compared to injection,
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the pump yielded steadier serum calcium levels, with a 50% reduction in urine calcium excretion. The dose of
PTH required using the pump system was 65% lower compared to the dose needed for twice daily injections.

The full-length PTH molecule, PTH(1-84), has been shown to have a longer half-life when injected
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subcutaneously in the thigh as opposed to the abdomen and may have a longer half-life than PTH(1-34) [130].
PTH(1-84) was tested in 33 patients with hypoparathyroidism in a six-year open-label study [131]. The initial
dosing of a subcutaneous injection of 100 mcg every other day was switched to daily injections when lower
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doses of the drug became available, allowing the drug to be titrated to meet the treatment goals for each
patient. Compared with baseline, calcitriol and calcium doses were significantly lower (by 67% and 53%,
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respectively), serum calcium was within the target range, and serum phosphate and 24-hour urinary calcium
excretion decreased at several time-points during the study. A randomized, placebo-controlled 6-month study
giving a fixed dose of 100 mcg of PTH(1-84) or placebo to 62 patients showed a 73% and 75% reduction in
calcitriol and calcium, respectively, but urinary calcium did not decrease [132]. The pivotal phase 3 trial, a
randomized multicenter trial comparing PTH(1-84) with placebo (in a 2:1 ratio), demonstrated that 53% of
patients achieved the primary endpoint of reducing oral calcium and calcitriol by at least 50% while
maintaining serum calcium within the target range [133]. Urinary calcium did not change, serum phosphate
and the calcium-phosphate product were lower in the PTH group, and quality-of-life measures showed a
tendency of improvement in the PTH group, but not in the placebo group, compared to baseline [134]. PTH(1-
84) was the first parathyroid hormone replacement therapy to be approved by the FDA for the treatment of
hypoparathyroidism.
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When should PTH(1-84) be used in hypoparathyroidism? The FDA approved the drug as an adjunct to calcium
and vitamin D for the treatment of adults who cannot be well-controlled on conventional therapy. The
definition of the term “not well-controlled” is subject to discussion. The First International Conference on the
Management of Hypoparathyroidism defined six groups of patients for whom replacement therapy with
recombinant human PTH as an alternative treatment option should be considered [81]. These include: cases
where the amount of calcium and vitamin D supplements have to be extremely high in order to reach
treatment targets; frequent erratic swings in calcium concentration not controlled by modification of
treatment program; renal complications; persistent hyperphosphatemia; gastrointestinal malabsorption
disorders; and severely reduced quality of life [25].

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While standard calcium and calcitriol treatment for hypoparathyroidism does not improve bone turnover in
hypoparathyroid patients, PTH replacement therapy, which restores the major hormonal driver of bone

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resorption, could be expected to increase bone turnover, restoring normal bone density and
microarchitecture. Studies of PTH replacement therapy have reported increases in bone turnover markers
with PTH(1-34) [127, 128], and PTH(1-84) injection [131, 135]. During the 6-year PTH(1-84) treatment study,

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bone turnover markers increased from baseline, peaking within the first year of treatment before receding to
a steady state at the end of the study period [131]. Areal bone mineral density (BMD) changes were observed,
including an increase in lumbar spine and total hip BMD, and a decrease in one-third distal radius BMD. A
histomorphometric assessment of iliac crest bone biopsies from patients with hypoparathyroidism showed

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changes in trabecular bone architecture over a 2-year period of PTH(1-84) treatment [136]. These changes
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included an increase in trabecular width and a decrease in trabecular number. Cortical porosity was shown to
increase during the treatment period, in agreement with a trend toward increased porosity observed in an
earlier, short-term study [132]. The use of computational methods (FEA) to estimate bone strength on µCT
data have indicated that there may be a transient increase in bone strength in patients treated with PTH(1-84)
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at 3 months and 1 year after treatment, however PTH did not appear to change the plate-rod characteristics of
trabecular bone compared to baseline [137]. No studies comparing fracture risk between treated or untreated
hypoparathyroid patients and the euparathyroid population have been published.
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Pseudohypoparathyroidism (PHP)

Pseudohypoparathyroidism (PHP), a disease of PTH resistance, was first described by Fuller Albright in 1942
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[138]. The classic features of PHP are hypocalcemia with hyperphosphatemia and elevated serum PTH, but
resistance to other hormones can also occur (see below); the presence of additional features is dependent on
the genetic or epigenetic cause of the disorder [25, 139]. Renal resistance at the level of the proximal tubule
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to PTH is a hallmark of PHP and can be tested by administering exogenous PTH(1-34) to the patient and
monitoring changes in cAMP and phosphate excretion (the Ellsworth-Howard test) [140-142].
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PHP falls into several categories based on molecular features of the disorder. Here, we discuss PHP types 1a
and 1b, classic types of the disease caused by genetic or epigenetic mutations in GNAS, which encodes for the
G-protein subunit Gsα. GNAS is biallelically expressed in most tissues, but in the proximal renal tubule, gonads,
thyroid, and pituitary, only the maternal allele is expressed [143]. Inactivating mutations on the maternal
allele (PHP1a) [144-148], or methylation changes that inactivate it (PHP1b) [149-151], lead to a functional loss
of Gsα and result in PTH resistance in these tissues. Patients with PHP1a and 1b therefore often have multiple
hormone resistances, for example to TSH and GHRH, consistent with imprinting of the gene in select tissues,
and with the pervasive role of Gsα as a signaling protein for several GPCRs [143]. PHP1a is associated with an
additional, distinctive developmental phenotype called Albright’s Hereditary Osteodystrophy (AHO), which is
thought to be caused by haploinsufficiency of Gsα in the skeleton. Clinical features of AHO include round
facies, short stature, brachydactyly, subcutaneous ossifications, and in some cases cognitive impairment [152].
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Patients with inactivating mutations on the paternal allele (pseudopseudohypoparathyroidism (PPHP)) have
AHO but lack the hallmark resistance to PTH and other hormones that characterizes PHP [153]. Because only
the maternal allele is expressed in the proximal renal tubule, individuals with PPHP maintain normal PTH-
regulated mineral homeostasis. This pattern of genomic imprinting makes it possible for members of the same
family to have PHP or PPHP, depending on which parent the trait was inherited from [146].

PHP1a and 1b are diagnosed when hypocalcemia and hyperphosphatemia are accompanied by elevated PTH
in the setting of normal 25(OH)D and magnesium levels. The Ellsworth-Howard test can be used to confirm the
diagnosis [139, 140]. The molecular diagnosis of PHP1a is made by sequencing exons 1-13 of GNAS; PHP1b is

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diagnosed on a molecular level by detecting characteristic methylation patterns at differentially methylated
regions (DMR) in the GNAS locus, which is currently only performed in research laboratories.

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The goal of treatment of patients with PHP1a and 1b with calcium and calcitriol differs from treatment goals
for hypoparathyroidism. In PHP, therapy aims at normalizing serum calcium and PTH levels, rather than
maintaining calcium in the low/low-normal range. Because Gsα is biallelically expressed at the distal renal

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tubule, calcium reabsorption under the influence of PTH is normal in PHP patients, which prevents the
hypercalciuria seen in hypoparathyroid patients [112, 141, 154].

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2. Vitamin D-related disorders

This group of disorders includes the relatively common nutritional vitamin D deficiency, impaired metabolism
of vitamin D, and very rare defects in the vitamin D receptor.
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Vitamin D deficiency
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The two main sources of vitamin D are diet (consumed as a supplement or in vitamin D-rich foods like fatty
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fish) and endogenous production in epithelial cells under exposure to UVB light [155]. Vitamin D is
hydroxylated at the C-25 position in the liver, producing 25-hydroxyvitamin D (25(OH)D), a fat-soluble vitamin
that circulates in the blood and is stored in adipose tissue [156]. 25(OH)D is hydroxylated in position 1 by 1-α
hydroxylase (CYP27B1) to produce the active 1,25-hydroxyvitamin D (1,25(OH)2D). This conversion takes place
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primarily in the proximal tubules of the kidneys, with control of CYP27B1 gene expression moderated by PTH
(activating) and the hormone FGF23 (repressing) [157]. 1,25(OH)2D is released from the kidneys into
circulation, where it plays a host of roles through interaction with the vitamin D receptor (VDR) in target
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tissues. Prominent among these is the active facilitation of calcium absorption in the intestine. 1,25(OH)2D
increases the efficiency of absorption of ingested calcium, making it a secondary key hormone in calcium
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homeostasis [155].

Vitamin D deficiency is common and can be associated with bone disease and other negative health effects.
The National Health and Nutrition Examination Survey (2005-2006) determined that its prevalence was 41.6%
in the United States (deficiency in this case was defined as < 20 ng/ml serum 25(OH)D) [158]. The primary
cause for vitamin D deficiency is thought to be a lack in dietary sources and a lack of exposure to sunshine;
other potential causes are drugs (such as those taken for epilepsy [159-161]), malabsorption [20], obesity in
children [162], and chronic kidney disease [163]. Vitamin D deficiency often leads to an increase in PTH
(secondary hyperparathyroidism) in response to decreased serum calcium. Elevated PTH can lead to urinary
phosphate losses resulting in hypophosphatemia [164]. The classical biochemical constellation of vitamin D
deficiency, therefore, is low normal calcium with increased PTH and hypophosphatemia.
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Severe vitamin D deficiency (<10 ng/ml) can lead to rickets in growing children or osteomalacia in adults.
Rickets causes deformation of weight-bearing limbs, growth retardation, delayed dentition, pain associated
with exercise, and muscle weakness [165]. Abnormalities of the growth plate associated with rickets, which
include expansion of hypertrophic chondrocytes and an increase in demineralized osteoid, appear to be linked
to hypophosphatemia. In three different animal models of the disease, low phosphate was shown to induce
apoptosis of hypertrophic chondrocytes in the growth plate, leading to a rachitic phenotype [166]. Severe
vitamin D deficiency after the growth plates have fused can lead to the expansion of unmineralized osteoid
(osteomalacia) [167, 168].

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Treatment for rickets and osteomalacia due to vitamin-D deficiency is relatively simple and has not changed
since the preventative component of cod liver oil was discovered nearly a century ago [169]. Vitamin D
supplementation is provided, with dosage varying depending on the patient’s age, extent of deficiency,

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symptoms, and the physician’s preference [165, 170]. For adults with osteomalacia or severe vitamin D
deficiency, high weekly oral doses of vitamin D (50,000 units) are usually given, followed by a maintenance
dose of vitamin D, typically 1,000 or 2,000 units per day.

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Impaired 1-α hydroxylase (CYP27B1) activity

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Individuals with vitamin D dependent rickets type 1 (VDDR-1, also known as pseudovitamin D deficiency
rickets or PDDR) have a clinical presentation that resembles vitamin D deficiency [164, 171]. This can include
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rickets, growth retardation, hypocalcemia, secondary hyperparathyroidism, and hypophosphatemia. These
individuals carry homozygous (or compound heterozygous) mutations that impair the expression or activity of
1-α hydroxylase, the enzyme that converts 25(OH)D to 1,25(OH)2D [172]. The hallmark of these disorders,
therefore, is the presence of normal 25(OH)D levels with low to undetectable 1,25(OH)2D, which distinguishes
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VDDR-1 from other forms of vitamin D deficiency [173]. Treatment consists of long-term daily replacement
doses of calcitriol, which results in rapid and complete correction of rachitic phenotype [164, 174, 175].
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Loss-of-function of the VDR

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Homozygous loss of function mutations in the VDR have been described but are exceedingly rare [176-178].
These mutations produce vitamin D dependent rickets type 2 (VDDR 2, aka hereditary vitamin D resistant
rickets or HVDRR), a disorder that is characterized by target organ resistance to 1,25(OH)2D leading to
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hypocalcemia, early onset rickets, secondary hyperparathyroidism and in some cases alopecia. 1,25(OH)2D is
elevated in these individuals, while 25(OH)D is often normal [175].
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Treatment for VDDR 2 is not trivial due to its nature as a disease of end-organ resistance. The primary goal of
treatment is correction of severe hypocalcemia and secondary hyperparathyroidism initiated by poor
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absorption of dietary calcium. For individuals with mutations that change the affinity of the VDR ligand binding
domain for 1,25(OH)2D, aggressive vitamin D and calcitriol supplementation can overcome partial resistance of
the receptor [175]. Generally, however, calcium balance is restored using high amounts of calcium, either by
oral supplementation or by IV (or a combination) [179]. After adolescence and with resolution of rachitic
phenotype, IV treatment is often no longer necessary, and oral calcium seems to support normal bone health
and serum calcium levels [180].
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3. Other causes of hypocalcemia

Impaired bone resorption

Drugs that effectively block bone resorption, such as denosumab and IV bisphosphonates, can lead to
hypocalcemia, particularly when vitamin D levels are insufficient, oral calcium intake or absorption low, or in
cases where the patient suffers from another disorder that interferes with PTH-mineral balance, such as CKD
[181-183]. This undesirable side effect can be controlled by proper monitoring of patients’ serum calcium and
supplementation with calcium and vitamin D during the course of treatment as a preventative measure [182].

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Hungry bone syndrome

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“Hungry bone” syndrome is characterized by hypocalcemia that often is profound, and can occur after
parathyroidectomy in patients with hyperparathyroidism or thyroidectomy in hyperthyroid patients [14]. The
etiology is attributed to massive calcium fluxes into the bone to mineralize osteoid.

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Critical illness

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Hypocalcemia is common during critical illness or postoperatively and is often mild [184, 185]. The cause is
thought to be multifactorial, including the effects of circulating cytokines which are associated with impaired
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PTH secretion and production of 1,25(OH)2D, and PTH end-organ resistance [186].

Others
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Medications can lead to hypocalcemia by chelating or precipitating calcium (eg citrate in massive blood
transfusions, phosphate administration, foscarnet, EDTA), accelerated enzymatic processing of vitamin D (eg
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anti-convulsants), decreasing PTH sensitivity (eg magnesium sulfate, calcitonin), increasing the sensitivity of
the CASR (cinacalcet) or increased calcium excretion (loop diuretics) [187].
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Practice points

Hypocalcemic disorders are very common in clinical practice. Most cases are due to disturbances in one of the
two major hormonal systems that regulate calcium homeostasis; PTH or vitamin D. Symptoms include
neuromuscular irritability but life-threatening sequelae such as seizures can occur. The diagnosis of the
underlying cause is straightforward in most cases, and treatment is geared towards the underlying cause.
Several important practical considerations include
• Measurement of total serum calcium should be accompanied by simultaneous measurement of
albumin for correction.

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• In patients with hypoparathyroidism, monitoring urinary calcium and instituting measures to
reduce hypercalciuria are important to prevent long-term kidney damage. These measures include
lowering serum calcium, adding thiazide diuretic together with low-salt diet, and distributing

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calcium supplements evenly throughout the day to avoid peaks in serum calcium.
• PTH replacement therapy in hypoparathyroidism is an emerging concept; defining the target
population is a work in progress.

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Research agenda
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• PTH replacement therapy for hypoparathyroidism is still in the early stages of development but has
shown demonstrable success in treating hypoparathyroid patients in recent clinical trials. Optimization
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of this therapeutic approach is a key goal for future research, for example studies will test different
dosing regimens and longer-acting PTH analogs.
• More clinical outcome data are needed to define the role of serum phosphate in hypoparathyroidism.
•
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Home monitoring devices for serum calcium could lead to better management and calcium control in
patients with chronic hypocalcemia.
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Figure legend
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Figure 1. Summary of the key organs and known actors that influence the development of disorders of
hypocalcemia. The parathyroid glands produce parathyroid hormone (PTH), which stimulates calcium
reabsorption and production of active vitamin D (1,25(OH)2D) in kidneys and resorption of calcium from bone.
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1,25(OH)2D enhances absorption of dietary calcium in the intestines. Calcium in turn regulates PTH production
in a negative feedback loop essential for maintaining physiological concentrations of extracellular calcium. Any
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interference within this regulatory system can lead to imbalance in serum calcium levels, which can give rise to
hypocalcemia.
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 ACCEPTED MANUSCRIPT

PT
Kidney
1α-hydroxylase mutations (VDDR1)

RI
Chronic kidney disease
PTH Cldn-16,-19 mutations (FHHNC)

SC
TRPM6 mutations (HSH)
PTH resistance (PHP1a,1b)

U
Ca2+

AN
Parathyroid glands
Iatrogenic (removed or damaged)
Bone Autoimmune (APS1)

M
Resorption-blocking drugs Developmental (FIH) Ca2+ 1,25(OH)2D
“Hungry bone’ syndrome CaSR or G11-related (AHD1, ADH2)
Complex disorders

D
Magnesium

TE
Infiltrative diseases

PTH
EP
Intestines
Ca2+ Malabsorption
C

VDR mutations (VDDR2)

TRPM6 mutations (HSH)
AC