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

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

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

Familial benign hypocalciuric hypercalcemia (FBHH) is an autosomal dominant trait with high penetrance, clinically manifestating a relatively benign, lifelong, persistent hypercalcemia and hypocalciuria without hypercalcemic related complications. The calcium-sensing receptor (CaSR) plays an important role in the regulation of PTH secretion and calcium metabolism. Here we present a family with FBHH of an autosomal dominant inheritance. A heterozygous mutation of E297K (GAG -> AAG, exon 4) of CaSR gene was found in 3 family members. To our knowledge, it is the first confirmed case of FBHH with CaSR gene mutation in Korea.
Sequence Analysis, DNA ; Receptors, Calcium-Sensing/*genetics ; Pedigree ; Parathyroid Hormone/analogs & derivatives/genetics/metabolism ; *Mutation ; Metabolism, Inborn Errors/*genetics ; Male ; Korea ; Hypercalcemia/*genetics ; Humans ; Heterozygote ; Genes, Dominant ; Female ; Family Health ; Exons ; DNA Restriction Enzymes/metabolism ; DNA/metabolism ; Adult

Clinical studies reported hypomagnesaemia in long-term omeprazole usage that was probably due to intestinal Mg2+ wasting. Our previous report demonstrated the inhibitory effect of omeprazole on passive Mg2+ transport across Caco-2 monolayers. The present study aimed to identify the underlying mechanism of omeprazole suppression of passive Mg2+ absorption. By using Caco-2 monolayers, we demonstrated a potent inhibitory effect of omeprazole on passive Mg2+, but not Ca2+, transport across Caco-2 monolayers. Omeprazole shifted the %maximum passive Mg2+ transport-Mg2+ concentration curves to the right, and increased the half maximal effective concentration of those dose-response curves, indicating a lower Mg2+ affinity of the paracellular channel. By continually monitoring the apical pH, we showed that omeprazole suppressed apical acid accumulation. Neomycin and spermine had no effect on passive Mg2+ transport of either control or omeprazole treated monolayers, indicating that omeprazole suppressed passive Mg2+ transport in a calcium sensing receptor (CaSR)-independent manner. The results of western blot analysis showed that omeprazole significantly suppressed claudin (Cldn)-7 and -12, but not Cldn-2, expression in Caco-2 cells. By using apical solution of pH 5.5, 6.0, 6.5, and 7.0, we found that apical acidity markedly increased passive Mg2+ transport, Mg2+ affinity of the paracellular channel, and Cldn-7 and -12 expression in Caco-2 monolayers. Apical acidity abolished the inhibitory effect of omeprazole on passive Mg2+ transport and Cldn-7 and -12 expression. Our results provided the evidence for the regulation of intestinal passive Mg2+ absorption by luminal acidity-induced increase in Cldn-7 and -12 expression.
Absorption/drug effects ; Caco-2 Cells ; Calcium/metabolism ; Claudins/genetics/*metabolism ; Dose-Response Relationship, Drug ; Gene Expression/drug effects ; Humans ; Hydrogen-Ion Concentration ; Magnesium/*metabolism ; Omeprazole/*pharmacology ; Proton Pump Inhibitors/*pharmacology ; Receptors, Calcium-Sensing/metabolism