title: Extracellular calcium-sensing receptor mediated signalling is involved in human vascular smooth muscle cell proliferation and apoptosis.
abstract: Calcium-sensing receptor (CaSR) plays key role in vascular calcification in patients with chronic kidney disease (CKD). We investigated the role of CaSR in regulating smooth muscle cell (SMC) proliferation and apoptosis. Incubation with 300 microM neomycin (CaSR agonist) resulted in 7.5-fold (p<0.05) increase in ERK1,2 phosphorylation. It was reduced (p<0.01) by 10 microM PD98059 (MEK1 inhibitor), indicating that CaSR agonist-induced effects were mediated via MEK1/ERK1,2 pathway. ERK1,2 phosphorylation was abolished by 5 microM U73122 (PLC inhibitor), indicating that PLC signalling was crucial for MEK1/ERK1,2 activation. Confirming PLC activation, inositol triphosphate (IP3) production was increased by neomycin/gentamycin (p<0.05) and reduced by U73122. To confirm that ERK1,2 and PLC signalling were mediated via CaSR, Human Aortic SMC (HAoSMC) were transfected with CaSR siRNA. CaSR knockdown resulted in lower ERK1,2 neomycin response and IP3 production (p<0.01). Neomycin increased HAoSMC proliferation >3-fold, which was reduced in CaSR knockdown cells (p<0.01) and further inhibited by PD98059 and U73122 (p<0.05). Apoptosis was not affected by neomycin treatment. U73122 produced 3.5-fold increase in HAoSMC apoptosis, which was further increased by CaSR knockdown (5-fold, p<0.05). In conclusion, stimulation of CaSR leads to activation of MEK1/ERK1,2 and PLC pathways and up-regulation of cell proliferation. CaSR-mediated PLC activation is important for SMC survival and protection against apoptosis.
authors.affiliation: The Clinical Sciences Research Institute, Warwick Medical School, Coventry, UK. g.molostvov@warwick.ac.uk
authors.forename: Guerman
authors.initials: G
authors.lastname: Molostvov
authors.forename: Simon
authors.initials: S
authors.lastname: Fletcher
authors.forename: Rosemary
authors.initials: R
authors.lastname: Bland
authors.forename: Daniel
authors.initials: D
authors.lastname: Zehnder
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D004247:DNA / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004963:Estrenes / Q000494:pharmacology; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000201:enzymology; D032389:Myocytes, Smooth Muscle / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009355:Neomycin / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011419:Propidium / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D044169:Receptors, Calcium-Sensing / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection
chemical_list: D004963:Estrenes; D011760:Pyrrolidinones; D034741:RNA, Small Interfering; D044169:Receptors, Calcium-Sensing; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D011419:Propidium; D015544:Inositol 1,4,5-Trisphosphate; D004247:DNA; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D009355:Neomycin