High extracellular calcium attenuates adipogenesis in 3T3-L1 preadipocytes

We studied the effect of extracellular Ca2+ concentration ([Ca2+](e)) on adipocyte differentiation. Preadipocyte.s exposed to continuous [Ca2+], higher than 2.5 mmol/l accumulated little or no cytoplasmic lipid compared to controls in 1.8 mmol/l [Ca2+](e). Differentiation was monitored by Oil Red O staining of cytoplasmic lipid and triglyceride assay of accumulated lipid, by RT-PCR analysis of adipogenic markers, and by the activity of glycerol-3-phosphate dehydrogenase (GPDH). Elevated [Ca2+](e) inhibited expression of peroxisome proliterator-activated receptor gamma, CCAAT/enhancer binding protein alpha and steroid regulatory binding element protein. High [Ca2+](e) significantly inhibited differentiation marker expression including adipocyte fatty acid binding protein, and GPDH. The decrease in Pref-1 expression that accompanied differentiation also was prevented by high [Ca2+](e). Treatment of 3T3-L1 cells with high [Ca2+](e) did not significantly affect cell number or viability and did not trigger apoptosis. Levels of intracellular Ca+2 remained unchanged in various [Ca2+](e) Treatment of 3T3-L1 with pertussis toxin (PTX) partially restored lipid accumulation and increased differentiation markers in cells treated with 5 mmol/l [Ca2+](e). 'Classical' parathyroid cell Ca2+ sensing receptors (CaSR) were not detected either by RT-PCR or by Western blotting. These results suggest that continuos exposure to high [Ca2+](e) inhibits preadipocyte differentiation and that this may involve a G-protein-coupled mechanism mediated by a novel Ca2+ sensor or receptor. (C) 2004 Elsevier Inc. All rights reserved.