Decreased expression of calcium-sensing receptor messenger ribonucleic acids in parathyroid adenomas

The cloning of the calcium sensing receptor (CaR) confirmed that parathyroid cells monitor extracellular calcium concentration ([Ca2+]ext) via a receptor-type mechanism. This lead to the hypothesis that abnormalities in the expression and/or function of the CaR could explain the biochemical abnormalities in primary hyperparathyroidism (PHPT). Cultured cells from parathyroid adenomas of patients operated for PHPT were used to monitor real-time changes in intracellular calcium concentration ([Ca2+]i) as measured by fluorescent microscopy using the Fura-2/AM dye. We found that CaR agonists trigger release of intracellular calcium pools and such responses are amplified by increasing the affinity of IP3 receptors. Using confocal microscopy to monitor membrane trafficking in living parathyroid cells labelled with the fluorescent dye FM1-43, we found that a decrease in [Ca2+]i rather than an absolute change in [Ca2+]ext is the main stimulus for exocytosis from human parathyroid cells. These data suggest that, in PHPT, a defective signalling mechanism from the CaR allows cells from parathyroid adenomas to maintain low [Ca2+]i with uninhibited PTH secretion in the face of hypercalcaemia. Over longer periods of time, CaR controls parathyroid proliferation via changes in tyrosine phosphorylation. We found that multiple proteins of molecular weight 20–65 kDa are phosphorylated within 10–60 min in response to CaR agonists. Further work demonstrated that high [Ca2+]i stimulates the expression of bcl-2 oncoprotein in cultured human parathyroid cells and that, in parathyroid adenomas, predominant expression of bcl-2 rather than bax oncoprotein might prevent apoptosis and explain the slow growth rate of these tumours. More recently, it became apparent that CaR stimulates cell proliferation in several cell types not involved in calcium homeostasis. Using archived histological material from 65 patients who died with metastatic breast cancer, we identified CaR expression predominantly in tumours from patients who developed bone rather than visceral metastases (35 of 49 versus 7 of 16; P < 0.01, chi-squared test). These data suggest that CaR expression has the potential to become a new biological marker predicting the risk of bone metastases in patients with breast cancer. A prospective study should investigate if patients with CaR-positive tumours are more likely to develop bone metastases and whether they could benefit more from prophylactic treatment with bisphosphonates or the newly developed CaR antagonists.

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Methylation Status of the CpG Islands in Vitamin D and Calcium-Sensing Receptor Gene Promoters Does Not Explain the Reduced Gene Expressions in Parathyroid Adenomas

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jc.2013-1699 ◽

2013 ◽

Vol 98 (10) ◽

pp. E1631-E1635 ◽

Cited By ~ 23

Author(s):

Shweta Varshney ◽

Sanjay Kumar Bhadada ◽

Naresh Sachdeva ◽

Ashutosh Kumar Arya ◽

Uma Nahar Saikia ◽

...

Keyword(s):

Vitamin D ◽

Receptor Gene ◽

Methylation Status ◽

Cpg Islands ◽

Gene Promoters ◽

Calcium Sensing Receptor ◽

Gene Expressions ◽

Calcium Sensing ◽

Parathyroid Adenomas ◽

Calcium Sensing Receptor Gene

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Calcium-sensing receptor expression and signalling in human parathyroid adenomas and primary hyperplasia

Clinical Endocrinology ◽

10.1046/j.1365-2265.2000.00933.x ◽

2000 ◽

Vol 52 (3) ◽

pp. 339-348 ◽

Cited By ~ 62

Author(s):

Sabrina Corbetta ◽

Giovanna Mantovani ◽

Andrea Lania ◽

Stefano Borgato ◽

Leonardo Vicentini ◽

...

Keyword(s):

Receptor Expression ◽

Calcium Sensing Receptor ◽

Calcium Sensing ◽

Parathyroid Adenomas

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Osteocalcin may participate to the bone-parathyroid crosstalk through activation of the calcium-sensing receptor in human parathyroid adenomas

Endocrine Abstracts ◽

10.1530/endoabs.70.aep143 ◽

2020 ◽

Author(s):

Chiara Verdelli ◽

Tavanti Giulia Stefania ◽

Riccardo Maggiore ◽

Gilberto Mari ◽

Veronica Sansoni ◽

...

Keyword(s):

Calcium Sensing Receptor ◽

Calcium Sensing ◽

Parathyroid Adenomas

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Decrease in vitamin D receptor and calcium-sensing receptor in highly proliferative parathyroid adenomas

Acta Endocrinologica ◽

10.1530/eje.0.1480403 ◽

2003 ◽

pp. 403-411 ◽

Cited By ~ 56

Author(s):

S Yano ◽

T Sugimoto ◽

T Tsukamoto ◽

K Chihara ◽

A Kobayashi ◽

...

Keyword(s):

Vitamin D ◽

Vitamin D Receptor ◽

Serum Levels ◽

Clinical Severity ◽

Regression Analyses ◽

Calcium Sensing Receptor ◽

Calcium Sensing ◽

Strong Positive Relationship ◽

Parathyroid Adenomas ◽

Single Regression

OBJECTIVE: A significant decrease in vitamin D receptor (VDR) and calcium-sensing receptor (CaSR) protein expression has been demonstrated recently in parathyroid (PT) adenomas. In this study, we investigated the relationships between the proliferative activity of parathyroid glands (PTGs) and the expression of VDR as well as CaSR, and compared it with the clinical severity in patients with primary hyperparathyroidism (1 degrees HPT). DESIGN: Seven patients with 1 degrees HPT were included in this study. Four patients with thyroid carcinoma served as controls. METHODS: Immunohistochemical staining was performed on serial sections of PTGs with specific antibodies against CaSR, VDR, and Ki67. Areas examined in each section were selected at random in relation to the gland size. The number of Ki67-positive cells was expressed as a labeling index (LI; positive cells per 1000 PT cells). The expression of CaSR and VDR was semi-quantitatively analyzed based on the intensity of staining. After averages of the scores from all areas were calculated, CaSR and VDR scores, and Ki67 LI were assigned to each gland for use in statistical analyses. RESULTS: In PT adenomas, scores of VDR and CaSR were markedly lower than in normal PTGs (P<0.01), while the proportion of Ki67-positive cells in PT adenomas was significantly higher than in normal PTGs (P<0.01). Single regression analyses revealed that Ki67 LI was positively correlated with serum levels of intact parathyroid hormone and Ca, and PTG weight (R=0.70, P<0.05, R=0.78, P<0.01 and R=0.84, P<0.05 respectively). Ki67 LI was negatively correlated with CaSR and VDR scores (R=-0.78, P<0.01 and R=-0.72, P<0.05 respectively). Moreover, there was a strong positive relationship between CaSR and VDR expression (R=0.95, P<0.001). CONCLUSIONS: Marked decreases in VDR and CaSR expression could, at least in part, be responsible for the high proliferation of PT cells and the pathological progression of 1 degree HPT.

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Reduced Calcium Sensing Receptor (CaSR) Expression Is Epigenetically Deregulated in Parathyroid Adenomas

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/clinem/dgaa419 ◽

2020 ◽

Vol 105 (9) ◽

pp. 3015-3024

Author(s):

Priyanka Singh ◽

Sanjay Kumar Bhadada ◽

Divya Dahiya ◽

Ashutosh Kumar Arya ◽

Uma Nahar Saikia ◽

...

Keyword(s):

Protein Expression ◽

Parathyroid Adenoma ◽

Histone Modifications ◽

Chromatin Immunoprecipitation ◽

Methylation Status ◽

Sequencing Analysis ◽

Calcium Sensing Receptor ◽

Calcium Sensing ◽

Bisulphite Sequencing ◽

Parathyroid Adenomas

Abstract Aim Reduced calcium sensing receptor (CaSR) expression has been implicated in parathyroid tumorigenesis, but the underlying mechanism remains elusive. Accordingly, we aimed to explore the epigenetic changes (DNA methylation and histone modifications) involved in CaSR regulation in sporadic parathyroid adenomas and correlate epigenetic state with disease indices. Experimental Design Forty sporadic parathyroid adenomas and 10 control parathyroid tissues were studied. Real-time quantitative PCR (qPCR) for mRNA and immunohistochemistry for protein expression of CaSR were performed. The methylation status of the CaSR promoter 2 was determined by bisulphite sequencing analysis of sodium bisulphite-converted DNA. To determine the role of histone modifications in the CaSR regulation, chromatin immunoprecipitation-qPCR assay was performed. Results Real-time qPCR revealed reduced CaSR mRNA expression with a fold reduction of 0.12 (P < 0.0001) in parathyroid adenomas. Immunohistochemistry revealed reduced protein expression of CaSR in 90% (36/40) of adenomas. The promoter 2 region of CaSR displayed significant hypermethylation in 45% (18/40) of the adenomas compared with the controls (6.7%; 1 of 10) (P < 0.002). Bisulphite sequencing analysis revealed maximum methylated CpG at glial cell missing 2 binding site on the CaSR promoter 2 compared to other CpG sites. The methylation status of CaSR correlated directly with plasma intact parathyroid hormone levels in patients with parathyroid adenoma. With chromatin immunoprecipitation-qPCR analysis, H3K9me3 levels showed increased enrichment by 10-fold in adenomas and correlated with CaSR-mRNA expression (r = 0.61; P < 0.003). Treatment with 5-aza-2′deoxycytidine restored the expression of CaSR in a parathyroid cell line. Conclusion Our data suggest that hypermethylation and increased H3K9me3 of the CaSR promoter 2 are involved in silencing CaSR expression in sporadic parathyroid adenoma.

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