OBJECTIVE: To explore the genetic basis for a child featuring idiopathic epilepsy and autism. METHODS: Peripheral blood samples of the child and his parents were collected with informed consent for the extraction of genome DNA. Whole exome sequencing was carried out for the family trio. Candidate variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The proband was found to harbor a heterozygous nonsense c.3025C>T (p.Arg1009Ter) variant in exon 7 of the CASR gene exon 7, which may produce a truncated protein. Based on the guidelines of the American College of Medical Genetics and Genomics, the variant was predicted to be deleterious and classified as possibly pathogenic (PVS1+PM2). CONCLUSION The c.3025C>T (p.Arg1009Ter) variant of the CASR gene probably underlay the disease in this child.
Autistic Disorder ; Child ; Epilepsy/genetics* ; Exons ; Heterozygote ; Humans ; Receptors, Calcium-Sensing/genetics* ; Whole Exome Sequencing

The study was designed to investigate the effects and mechanism of a calcium-sensing receptor (CaSR) polymorphism at E942K on the proliferation of gastric cancer cells. Single nucleotide polymorphisms (SNPs) were detected between gastric cancers group and normal controls group by DNA sequence analysis. The cell model was constructed by transfection of E942K mutant plasmid and wild-type (WT) plasmid into SGC-7901 and HEK-293 cells. The effect of E942K mutation on cell proliferation ability was detected by CCK8 and cell clone formation experiments. The effect of E942K mutation on calcium signaling was detected by calcium imaging. Western blot experiments were used to detect changes in phosphorylation levels of key proteins ERK1/2 and β-catenin in downstream signaling pathways after E942K mutation. The results showed that the mutation rate of E942K in gastric cancer group was significantly higher than that in normal control group (P < 0.05). CCK8 and cell clone formation experiments showed that E942K mutation significantly improved the proliferation ability of SGC-7901 gastric cancer cells and HEK-293 cells. E942K mutation enhanced calcium signaling in SGC-7901 and HEK-293 cells. E942K mutation enhanced ERK1/2 phosphorylation without affecting β-catenin phosphorylation. The results suggest that E942K mutation in CaSR may ultimately promote the proliferation of gastric cancer cells by enhancing intracellular calcium signaling and ERK1/2 phosphorylation. These results have potential clinical implications for the diagnosis and targeted therapy of gastric cancer.
Calcium ; Cell Proliferation ; HEK293 Cells ; Humans ; MAP Kinase Signaling System ; Mutation ; Receptors, Calcium-Sensing ; genetics ; Stomach Neoplasms ; genetics

OBJECTIVETo investigate the effect of interleukin-6 (IL-6) and calcium sensing receptor (CASR) gene polymorphisms on bone mass accrual in Chinese adolescent girls.

METHODSA total of 228 premenarche Chinese girls (9-11.5 years) were recruited for a 2-year study. Bone mineral densities (BMD) and Bone mineral contents (BMC) in the total body, total left hip including femoral neck, trochanter, intertrochanter and Ward's triangle area, and lumbar spine (L1-L4) were measured by DEXA. The -634C/G polymorphism of IL-6 gene was detected by PCR-restriction fragment length polymorphism and A986S polymorphism of CASR gene was detected by allele-special mutagenically separated amplication-PCR.

RESULTSThere were 176 available subjects when the 2-year study was completed. The -634C/G polymorphism of IL-6 gene and A986S polymorphism of CASR gene were significantly associated with bone mass accrual after adjusting the potential confounding factors. Girls with CC genotype of IL-6 -634G/C gene had higher percentage of increase in BMD of total body (P= 0.027) and femoral trochanter (P= 0.028) than those with CG+ GG genotypes. Girls with AA genotype of CASR gene had greater percentage of increase in L1-L4 lumbar spine BMC (P= 0.022) and Ward's triangle area BMD (P= 0.049) than their AS+ SS counterparts. Subjects with G allele of IL-6 gene in combination with S allele of CASR gene had less increase in BMC of femoral neck and L1-L4 lumbar spine.

CONCLUSIONAdolescent girls carrying the G allele of IL-6 gene at -634C/G and S allele of CASR gene at A986S could potentially be the risk population of lower bone mass accrual.

Alleles ; Bone Density ; genetics ; Child ; Female ; Genotype ; Humans ; Interleukin-6 ; genetics ; Polymerase Chain Reaction ; Polymorphism, Genetic ; genetics ; Receptors, Calcium-Sensing ; genetics

OBJECTIVE: To explore the genetic basis for a child with neonatal severe hyperparathyroidism. METHODS: Genomic DNA was extracted from peripheral blood samples from the patient and her parents. Whole exome sequencing was carried out to screen potential mutations. Suspected mutation was verified by Sanger sequencing. RESULTS: The proband was found to carry compound heterozygous variants c.179G>A (p.Cys60Tyr) and c.1525G>A (p.Gly509Arg) of the CaSR gene. The c.179G>A variant was derived from her mother and was unreported previously. The c.1525G>A variant was derived from her father and known to be pathogenic. CONCLUSION The compound heterozygous variants of c.179G>A and c.1525G>A of the CaSR gene probably underlie the disease in the patient. The results of genetic testing has enabled diagnosis and genetic counseling for her family.
Female ; Genetic Counseling ; Genetic Testing ; Humans ; Hyperparathyroidism/genetics* ; Infant, Newborn ; Infant, Newborn, Diseases/genetics* ; Mutation ; Pedigree ; Receptors, Calcium-Sensing/genetics* ; Whole Exome Sequencing

OBJECTIVETo investigate the relationship between calcium-sensing receptor protein (CaSR) expression and rat cardiomyocyte apoptosis and related signal transduction pathways.

METHODSThe CaSR, BCl2, Caspase3 protein and ERK1/2 phosphorylation or non-phosphorylation were detected by Western blot. Cardiomyocyte apoptosis was detected by flow cytometry and immunofluorescence.

RESULTSCaSR protein was detected in rat cardiac tissue and CaSR activator gadolinium (GdCl3) induced cardiomyocyte apoptosis and increased ERK1/2 phosphorylation and expression of BCl2 and activated Caspase3. The selective mitogen-activated protein kinase (MAPK) inhibitor PD98059 abolished gadolinium -induced ERK1/2 activation and BCl2 expression, further increased the activation of Caspase3 and cardiomyocyte apoptosis.

CONCLUSIONOur results demonstrate the CaSR existence in cardiomyocytes and CaSR activation by gadolinium can induce myocyte apoptosis by activating Caspase3 and tyrosine protein kinase pathway.

Animals ; Apoptosis ; Caspase 3 ; metabolism ; Female ; Male ; Myocardium ; metabolism ; Myocytes, Cardiac ; metabolism ; RNA, Messenger ; biosynthesis ; Rats ; Rats, Wistar ; Receptors, Calcium-Sensing ; biosynthesis ; genetics ; Signal Transduction

Adult ; Aged ; Female ; Humans ; Hyperparathyroidism, Secondary ; metabolism ; pathology ; Immunohistochemistry ; Male ; Middle Aged ; Parathyroid Glands ; chemistry ; RNA, Messenger ; analysis ; Receptors, Calcium-Sensing ; analysis ; genetics ; Uremia ; metabolism

OBJECTIVETo study the effect of calcium-sensing receptor (CaSR) agonists and antagonists on the expression of CaSR in neonatal mice with persistent pulmonary hypertension (PPHN), and to clarify the role of CaSR in neonatal mice with PPHN.

METHODSForty-nine neonatal mice were randomly divided into four groups: control (n=10), hypoxia (PPHN; n=11), agonist (n=13), and antagonist (n=15). The mice in the PPHN, agonist, and antagonist groups were exposed to an oxygen concentration of 12%, and those in the control group were exposed to the air. The mice in the agonist and antagonist groups were intraperitoneally injected with gadolinium chloride (16 mg/kg) and NPS2390 (1 mg/kg) respectively once daily. Those in the PPHN and the control groups were given normal saline daily. All the mice were treated for 14 consecutive days. Hematoxylin and eosin staining and immunohistochemistry were used to observe the changes in pulmonary vessels. Laser confocal microscopy was used to observe the site of CaSR expression and measure its content in lung tissues. qRT-PCR and Western blot were used to measure the mRNA and protein expression of CaSR in lung tissues.

RESULTSCompared with the control group, the PPHN group had significant increases in the pulmonary small artery wall thickness and the ratio of right to left ventricular wall thickness (P<0.05), which suggested that the model was successfully prepared. Compared with the control group, the PPHN group had a significant increase in the mRNA and protein expression of CaSR (P<0.05), and the agonist group had a significantly greater increase (P<0.05); the antagonist group had a significant reduction in the mRNA and protein expression of CaSR (P<0.05).

CONCLUSIONSCaSR may play an important role in the development of PPHN induced by hypoxia in neonatal mice.

Animals ; Hypoxia ; complications ; Lung ; pathology ; Mice ; Myocardium ; pathology ; Persistent Fetal Circulation Syndrome ; etiology ; pathology ; Pulmonary Artery ; pathology ; RNA, Messenger ; analysis ; Receptors, Calcium-Sensing ; analysis ; genetics ; physiology

Calcium Channels, L-Type ; genetics ; metabolism ; Esophageal Achalasia ; genetics ; metabolism ; Esophagus ; metabolism ; Humans ; Nitric Oxide Synthase ; metabolism ; Nitric Oxide Synthase Type I ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Real-Time Polymerase Chain Reaction ; Receptors, Calcium-Sensing ; genetics ; metabolism

Isolated hypoparathyroidism (IH) shows heterogeneous phenotypes and can be caused by defects in a variety of genes. The goal of our study was to determine the clinical features and to analyze gene mutations in a large cohort of Korean patients with sporadic or familial IH. We recruited 23 patients. They showed a broad range of onset age and various values of biochemical data. Whole exome sequencing was performed on two affected cases and one unaffected individual in a family. All coding exons and exon-intron borders of GCMB, CASR, and prepro-PTH were sequenced using PCR-amplified DNA. In one family who underwent the whole exome sequencing analysis, approximately 300 single nucleotide changes emerged as candidates for genetic alteration. Among them, we identified a functional mutation in exon 2 of GCMB (C106R) in two affected cases. Besides, heterozygous gain-of-function mutations in the CASR gene were found in other subjects; D410E and P221L. We also found one single nucleotide polymorphism (SNP) in the prepro-PTH gene, five SNPs in the CASR gene, and four SNPs in the GCMB gene. The current study represents a variety of biochemical phenotypes in IH patients with the molecular genetic diagnosis of IH.
Adult ; Aged ; Asian Continental Ancestry Group/*genetics ; Cohort Studies ; Heterozygote ; Humans ; Hypoparathyroidism/diagnosis/*genetics/pathology ; Middle Aged ; Nuclear Proteins/*genetics ; Parathyroid Hormone/*genetics ; Phenotype ; Polymorphism, Single Nucleotide ; Receptors, Calcium-Sensing/*genetics ; Registries ; Republic of Korea ; Transcription Factors/*genetics ; Young Adult

The extracellular calcium sensing receptor (CaSR) belongs to the type III family of G-protein-coupled receptors, a family that comprises the metabotropic glutamate receptor and the putative vomeronasal organ receptors. The CaSR plays an important role for calcium homeostasis in parathyroid cells, kidney cells and other cells to directly 'sense' changes in the extracellular calcium ion concentration ((Ca2+)o). The mesangial cells are known to be involved in many pathologic sequences through the mediation of altered glomerular hemodynamics, cell proliferation, and matrix production. In this study, we examined the expression of the CaSR in the mouse mesangial cell lines (MMC, ATCC number CRL-1927). Reverse transcription- polymerase chain reaction (RT-PCR) was perform with CaSR-specific primers, and this was followed by nucleotide sequencing of the amplified product; this process identified the CaSR transcript in the MMCs. Moreover, CaSR protein was present in the MMCs as assessed by Western blot and immunocytochemical analysis using a polyclonal antibody specific for the CaSR. Functionally, (Ca2+)o induced the increment of the intracellular calcium concentration ((Ca2+)i) in a dose-dependent manner. This (Ca2+)i increment by (Ca2+)o was attenuated by the pretreatment with a phospholipase C inhibitor (U73122) and also by a pretreatment with a CaSR antagonist (NPS 2390). The similar results were also obtained in IP3 accumulation by (Ca2+)o. To investigate the physiological effect of the CaSR, the effect of the (Ca2+)o on cell proliferation was studied. The increased (Ca2+)o (up to 10 mM) produced a significant increase in the cell numbers. This mitogenic effect of (Ca2+)o was inhibited by the co-treatment with a CaSR antagonist. From these results, the (Ca2+)o-induced (Ca2+)i elevation in the MMC is coupled with the extracellular calcium sensing receptor. Furthermore, (Ca2+)o produces a mitogenic effect in MMCs.
Animals ; Calcium/metabolism ; Cell Line ; Cell Proliferation ; Inositol 1,4,5-Trisphosphate/metabolism ; Mesangial Cells/*cytology/*metabolism ; Mice ; RNA, Messenger/genetics/metabolism ; Receptors, Calcium-Sensing/genetics/*metabolism ; Research Support, Non-U.S. Gov't