Tải bản đầy đủ - 0 (trang)
3 The Calcium-Sensing Receptor and Its Implication in the Control of PTH Secretion

3 The Calcium-Sensing Receptor and Its Implication in the Control of PTH Secretion

Tải bản đầy đủ - 0trang


M.E. Rodríguez-Ortiz et al.

Although Ca is the main agonist of the CaR, other ligands are also capable of

activate it. They are classified as a) type I agonists, including other divalent (Mg2+,

Sr2+, Ba2+) and trivalent (La3+, Gd3+) cations, and b) type II agonists, as polyamines

(putrescine, spermine, spermidine) and amino acids [10]. The ability of these molecules to activate the CaR has prompted the development of calcimimetics, drugs

capable of inhibiting the secretion of PTH upon the activation of the receptor.

The human CaR gene is located in the long arm of chromosome 3, at position 13

(3q13) whereas in the rat and mouse it resides on chromosomes 11 and 16, respectively. The protein of CaR has a molecular weight of 120 KDa. It possesses a large

N-terminal extracellular domain, a cysteine-rich domain linking the extracellular

domain to the first transmembrane helix, a seven transmembrane domain, and an

intracellular C-terminal domain. Although it also has been reported a binding site

for Ca in the transmembrane region of the receptor, the site located in the extracellular domain is required for the fully activation. The interaction between Ca and the

receptor induces a conformational change in its structure, the activation of transduction signals and the mobilization of intracellular Ca.

Mutations in CaR gene are responsible for several disorders. Inactivating mutations cause familial hypocalciuric hypocalcemia (FHH, OMIM 14598) as well as

neonatal severe hyperparathyroidism (NSHPT, OMIM 239200), whereas autosomal

dominant hypoparathyroidism (ADH, OMIM 601298) is associated to gain-offunction mutations in the CaR gene [11].

The CaR is not only expressed in organs directly involved in mineral homeostasis, such as parathyroid cells, C cells of the thyroid gland, bone or kidney, but also

in other tissues as gastrointestinal tract (esophagus, stomach, small intestine, and

colon), skin, cardiac and smooth muscle, and nervous system. The presence of the

CaR in the surface of parathyroid cells allows them to respond exquisitely to small

changes in the concentration of extracellular Ca. The activation of the receptor activates several signaling pathways (phospholipase A2 and C, protein kinase C, and

several mitogen-activated protein kinases) that eventually lead to the inhibition of

PTH [12].

The activation of the CaR has also been shown to regulate the synthesis of

PTH. Indeed, in a state of hypocalcemia the stability of PTH mRNA is increased by

the binding of proteins to the specific region in the 3ʹ-untranslated region (UTR) that

protects the transcript from degradation. Naveh-Many et al. identified the A+U-rich

element binding factor (AUF1) as a protein that may confer stability to the PTH

mRNA transcript [13].

The expression of the CaR is modulated by several factors. Vitamin D augments

the expression of the receptor at parathyroid level. In an in vivo study, Brown and

collaborators found that administration of 1,25(OH)2D3 in rats fed with diets deficient in vitamin D stimulated the expression of the parathyroid CaR dosedependently [14]. Of note, the expression of CaR in these vitamin D-deficient

animals was already diminished. The effect of vitamin D, as a modulator of the

parathyroid CaR, was confirmed in a subsequent work by Canaff and Handy. They

identified the transcriptional start sites of promoters P1 and P2 in the CaR and the

existence of vitamin D responsive elements (VDREs) in both sites [15].

7 Vitamin D and Parathyroid Hormone Regulation in Chronic Kidney Disease


In addition to vitamin D, other modulators of parathyroid function have also

been shown to regulate the CaR expression. For instance, moderately elevated

magnesium (Mg) upregulated parathyroid CaR expression in vitro [16]. Similarly,

the addition of FGF23 to incubated parathyroid glands produced an increase in the

CaR [17].

So far, the results obtained concerning the role of Ca itself as a regulator of the

CaR are contradictory. While some authors have not found an independent effect of

Ca [18], we observed that high Ca significantly increased the expression of CaR in

rat parathyroid glands culture in vitro [17]. On the other hand, several studies have

shown that calcimimetics up-regulate the CaR in parathyroid glands from uremic

rats, where the expression of the receptor is already decreased [19]. Moreover, this

effect appears to be independent of a possible effect on proliferation, as observed in

rats shortly after acute administration of calcimimetic [20].

Importantly, a novel mechanism that modulates CaR trafficking from cytosol to

the cell membrane has been recently described. The agonist-driven insertional signaling implies that the maturation, trafficking, and insertion of the CaR in the

plasma membrane are mediated by the ligand binding [21].


The Vitamin D Receptor

The biological effects of 1,25(OH)2D3, the most active vitamin D metabolite, are

mediated by the VDR. The VDR is a ligand-activated transcription factor belonging

to the superfamily of steroid/thyroid hormone receptors nuclear receptor that, when

bound to vitamin D, acts as a transcription factor.

The VDR was discovered in 1969 in the intestine of vitamin D–deficient chicks

[22]. The VDR protein is composed of different structural regions with a certain

degree of variation between species. The primary amino acid sequence of the VDR

consists of six functional domains: the variable regions (A and B domains) which

include an activator region called AF-1, DNA binding (the C domain) is the most

conserved region and responsible for the recognition of the VDRE in the target

genes; this region contains two zinc fingers which are absolutely essential to stabilize the binding VDR-DNA. The hinge region (D domain) which gives to VDR an

ability of rotation enabling adequate link to VDREs, the ligand-binding region

(LBD, E domain) responsible for binding of vitamin D, and transcriptional activation (domain F) is the C-terminal region which contains a site for dimerization and

a AF-2 domain with regulatory function of transcription. To date, N terminal isoforms of the receptor have been identified that differ only in the length of their A/B

domains [23].

VDR is a DNA-binding transcription factor, which generates an active signaling

transduction complex. VDR control of gene transcription is mediated by several

stages including binding of vitamin D in the C-terminal portion of VDR, heterodimerization with retinoid X receptor (RXR) and nuclear translocation, binding of

VDR–RXR to specific DNA sequences (VDRE) in target gene promoter, and


M.E. Rodríguez-Ortiz et al.

recruitment of VDR-interacting nuclear co-regulators or co-factors resulting in activation or inhibition of gene transcription.

At the parathyroid level, the VDR directly down-regulates the transcription of

the gene encoding PTH through binding to VDRE. Negative VDREs have been

mapped both in the human and rat PTH promoters, resulting in a reduction of the

PTH synthesis [24, 25]. Results from VDR activation in other target tissues will not

be commented here.


Regulation of Vitamin D Receptor in Parathyroid Glands

An homologous regulation of VDR expression by 1,25(OH)2D3 amplifies the effect

of circulating 1,25(OH)2D3 on the PTH gene. This effect was described by NavehMany et al. [26]: the administration of 1,25(OH)2D3 increased the levels of the VDR

mRNA in rat parathyroid glands. Other studies have confirmed this finding. Denda

et al. [27] reported that low serum 1,25(OH)2D3 levels were, at least in part, responsible for the decrease in VDR content in parathyroid glands of uremic rats and that

treatment with 1,25(OH)2D3 prevented this decrease, ameliorating the development

of SHPT. In vitro studies also demonstrated a direct effect of 1,25(OH)2D3 to

increase VDR expression [18].

Of special importance in the regulation of parathyroid function is the cooperation

of Ca and vitamin D on reducing parathyroid function. Several experimental and

clinical data suggests that the 1,25(OH)2D3 system is relatively ineffective in controlling PTH when serum Ca concentration is low. A study by Naveh-Many et al. [5]

showed that the ability of exogenous 1,25(OH)2D3 to inhibit PTH mRNA in vitamin

D–deficient rats was largely prevented when serum Ca was low, suggesting that the

inhibition of PTH by 1,25(OH)2D3 was not possible if hypocalcemia was present. In

another study [28], the administration of calcitriol (CTR) in vitamin D–deficient

chickens for 3–6 days increased parathyroid VDR mRNA, and this effect was

enhanced when serum Ca levels were normal. In this same study it was observed

that 6 days of dietary Ca restriction decreased VDR mRNA and that 1,25(OH)2D3

administration only upregulated VDR expression when chickens were fed a normal

Ca diet. In rats fed for 6 weeks with diets containing different vitamin D, Brown

et al. [29] did not observe an independent effect of 1,25(OH)2D3, but upregulation

of VDR by 1,25(OH)2D3 appeared as being mediated primarily by an increased

serum Ca concentration.

A definitive evidence for an independent effect of Ca on parathyroid VDR

expression came from in vivo experiments by Garfia et al. [30]. By using a 6-h

hypercalcemic or hypocalcemic clamp in rats, it was found that 1,25(OH)2D3 produced an increase in VDR levels only when Ca concentration was elevated; however, 1,25(OH)2D3 did not increase VDR expression in rats maintained with

hypocalcemia during a 6-h period. In addition, the downregulation of VDR by

hypocalcemia resulted in impairment of the inhibitory action of 1,25(OH)2D3 on

parathyroid cell function. 1,25(OH)2D3 administered to rats after the induction of

7 Vitamin D and Parathyroid Hormone Regulation in Chronic Kidney Disease


hypocalcemia (when VDR was already downregulated) was not able to reduce PTH

mRNA, whereas it was downregulated in rats receiving 1,25(OH)2D3 before the

initiation of hypocalcemia. These results were also confirmed in vitro by Cañadillas

et al. [31] and by Carrillo et al. [18], who observed in rat parathyroid glands that the

higher the Ca, the higher VDR expression. Thus, beside the direct posttranscriptional effect on PTH mRNA, the low Ca may also stimulate PTH levels indirectly

through lowering the parathyroid VDR expression which makes 1,25(OH)2D3 less

effective. Of note, downregulation of parathyroid VDR caused by low Ca prevents

PTH inhibition by 1,25(OH)2D3. This is an important concept in the pathogenesis of

SHPT because hypocalcemia may not allow a normal inhibition of parathyroid cells

by 1,25(OH)2D3 even when 1,25(OH)2D3 levels are normal.

Finally, Cañadillas et al. [31] characterized in vitro the signaling system responsible for the stimulation of parathyroid VDR by extracellular Ca, which takes place

through the elevation of the cytosolic Ca level, and the subsequent stimulation of the

PLA2-AA-dependent ERK1/2- pathway. High extracellular Ca also increased the

VDR promoter activity through the activation of the Sp1 transcription factor in

VDR-transfected HEKCaR cells.

Taken together, the above data support that the tight control of PTH secretion and

synthesis by Ca may in part be explained by the fact that high Ca enhances the

inhibitory action of vitamin D on parathyroid glands by augmenting VDR


Besides 1,25(OH)2D3 and Ca, several other factors have been reported to regulate

VDR expression and/or function. Calcimimetics, such as R-568 and cinacalcet HCl,

are allosteric modulators of the CaR, acting by increasing the sensitivity of the parathyroid gland to extracellular Ca. Thus, it was hypothesized that administration of

the calcimimetic R-568 may result in increased VDR expression in parathyroid tissue. In in vitro studies with whole rat parathyroid glands, the calcimimetic elicited

an increase in VDR mRNA similar to the maximum increase detected with 1.5 mM

Ca [32]. Treatment with R-568 also increased VDR protein in normal rat parathyroid glands and, interestingly, in human parathyroid glands with diffuse but not

nodular hyperplasia. These results support the convenience of using vitamin

D-calcimimetic combinations in the clinical settings. Any increase in VDR would

facilitate the inhibitory feedback of vitamin D on the parathyroid glands and would

assist in optimization of the positive action of the pharmacological administration

of 1,25(OH)2D3 or other vitamin D analogs, which in turn may reduce unwanted

side effects as high CaxP products and vascular calcifications.

It is now well-known that FGF23 decreases PTH secretion and PTH mRNA [2].

Furthermore, in normal rat parathyroid glands in vitro, addition of FGF23 to the

low-Ca medium increased parathyroid VDR and CaR mRNA and protein expression to levels similar to those observed with a high-Ca concentration [17]. The

FGF23-dependent changes in VDR and CaR expression were paralleled by the activation of the ERK1/2 activation. FGF23 also increased VDR and CaR expression

and activated ERK1/2 in the parathyroid glands of normal rats in vivo. This upregulation of CaR and VDR may represent another mechanism whereby FGF23 reduces

parathyroid function.

Tài liệu bạn tìm kiếm đã sẵn sàng tải về

3 The Calcium-Sensing Receptor and Its Implication in the Control of PTH Secretion

Tải bản đầy đủ ngay(0 tr)