Abstract
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.
Highlights
Serum calcium levels in healthy adults remain stable because the amount of calcium entering the extracellular compartment matches the amount excreted by the kidneys
In order to confirm the molecular appearance of calcium sensing receptor (CaSR) mRNA and protein, Reverse transcription- polymerase chain reaction (RT-PCR) and Western blot analysis were performed on the mouse mesangial cell lines (MMC)
The results obtained in the current study demonstrate for the first time that the extracellular calcium sensing receptor (CaSR) is expressed in the renal glomerular mesangial cells, and CaSRs play an important role in cell proliferation
Summary
Serum calcium levels in healthy adults remain stable because the amount of calcium entering the extracellular compartment matches the amount excreted by the kidneys. This balance is mainly achieved by the calcium-sensing receptor (CaSR) that is located on the surface of the various types of cells (Houillier et al, 2003). Extracellular calcium is essential for a number of vital processes, including bone mineralization, blood coagulation, regulation of enzymatic activity and the modulation of the permeability and excitability of the plasma membranes For these reasons, the calcium concentration in the extracellular fluids is under strict control by a complex homeostatic system that includes the bones, kidney, intestines and the parathyroid and thyroid glands (Brown, 1991; Shin et al, 2003, Jung et al, 2005). The CaSR belongs to the type III family of G-proteincoupled receptors that comprises the metabotropic glutamate receptor and the putative vomeronasal organ receptors (Brown et al, 1993)
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