Merck
  • Home
  • Search Results
  • Calcium sensing receptor activation by calcimimetic R-568 in human amniotic fluid mesenchymal stem cells: correlation with osteogenic differentiation.

Calcium sensing receptor activation by calcimimetic R-568 in human amniotic fluid mesenchymal stem cells: correlation with osteogenic differentiation.

Stem cells and development (2014-07-19)
Caterina Pipino, Pamela Di Tomo, Domitilla Mandatori, Eleonora Cianci, Paola Lanuti, Meritxell B Cutrona, Letizia Penolazzi, Laura Pierdomenico, Elisabetta Lambertini, Ivana Antonucci, Vittorio Sirolli, Mario Bonomini, Mario Romano, Roberta Piva, Marco Marchisio, Assunta Pandolfi
ABSTRACT

Human amniotic fluid mesenchymal stem cells (hAFMSCs) are promising for therapeutic applications in bone damage. Calcium sensing receptor (CaSR), a G protein-coupled receptor, plays a physiological role in the regulation of bone metabolism. Thus, the bone CaSR could be targeted by calcimimetic agonists, which may be potentially helpful in treating bone diseases. The aim of our study was to characterize CaSR expression in hAFMSCs and to assess the activity of calcimimetic R-568 during in vitro osteogenesis. Using western blotting, immunofluorescence, and flow cytometry, we consistently observed constitutive CaSR in osteo-differentiating hAFMSCs. Notably, both R-568 and calcium significantly enhanced hAFMSC osteogenic differentiation after exposure to osteogenic medium. To provide further evidence of the involvement of CaSR in osteogenesis, we correlated its expression with that of established osteogenic markers, that is, alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2), and osteopontin (OPN), and novel, not yet completely defined regulators of osteogenesis. Among these are β-catenin and Slug, which are mediators of Wnt signaling, and nuclear factor of activated T cells c1 (NFATc1), which plays a critical role in calcium/calcineurin signaling. Taken together, our results demonstrate that CaSR is expressed in hAFMSCs, positively correlates with osteogenic markers, and is activated by R-568. Notably, downregulation of CaSR by RNA interference supports the conclusion that CaSR activation plays a central role in hAFMSC osteogenesis. Thus, this study provides significant information on the mechanisms of hAFMSC osteogenesis, which could provide additional molecular basis for the use of calcimimetics in bone regenerative medicine.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Sodium chloride, 99.999% trace metals basis
Sigma-Aldrich
Sodium chloride-35Cl, 99 atom % 35Cl
Sigma-Aldrich
Sodium chloride, random crystals, optical grade, 99.9% trace metals basis
Sigma-Aldrich
Tris(tert-butoxy)silanol, 99.999%
Sigma-Aldrich
Sodium chloride, AnhydroBeads, −10 mesh, 99.99% trace metals basis
Sigma-Aldrich
Sodium chloride, AnhydroBeads, −10 mesh, 99.999% trace metals basis
Sigma-Aldrich
Tris(tert-butoxy)silanol, packaged for use in deposition systems
Sigma-Aldrich
Sodium chloride solution, 0.85%
Sigma-Aldrich
Sodium chloride, tested according to Ph. Eur.
Sigma-Aldrich
Sodium chloride, BioUltra, for molecular biology, ≥99.5% (AT)
Supelco
Hydrogen chloride – ethanol solution, ~1.25 M HCl, for GC derivatization, LiChropur
Supelco
Sodium chloride, certified reference material for titrimetry, certified by BAM, ≥99.5%
Supelco
Hydrogen chloride – 2-propanol solution, ~1.25 M HCl (T), for GC derivatization, LiChropur
Supelco
Calcium standard for AAS, analytical standard, 1.000 g/L Ca+2 in hydrochloric acid, traceable to BAM
Sigma-Aldrich
Sodium chloride solution, BioUltra, for molecular biology, ~5 M in H2O
Sigma-Aldrich
Hydrochloric acid solution, ~6 M in H2O, for amino acid analysis
Sigma-Aldrich
Sodium azide, BioUltra, ≥99.5% (T)
Sigma-Aldrich
Sodium azide, purum p.a., ≥99.0% (T)
Sigma-Aldrich
Sodium azide, ReagentPlus®, ≥99.5%
Sigma-Aldrich
Sodium chloride solution, 5 M in H2O, BioReagent, for molecular biology, suitable for cell culture
Sigma-Aldrich
Sodium azide, BioXtra
Sigma-Aldrich
7-Aminoactinomycin D, ~97% (HPLC), powder
Sigma-Aldrich
Hydrochloric acid, 36.5-38.0%, BioReagent, for molecular biology
Sigma-Aldrich
Sodium chloride solution, 5 M
Supelco
Hydrochloric acid solution, volumetric, 0.1 M HCl (0.1N), endotoxin free
Sigma-Aldrich
Hydrochloric acid solution, 1.0 N, BioReagent, suitable for cell culture
Sigma-Aldrich
Sodium chloride solution, 0.9% in water, BioXtra, suitable for cell culture
Sigma-Aldrich
Sodium chloride, meets analytical specification of Ph. Eur., BP, USP, 99.0-100.5%
Sigma-Aldrich
Sodium chloride, for molecular biology, DNase, RNase, and protease, none detected, ≥99% (titration)
Sigma-Aldrich
Sodium chloride, BioReagent, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99%