High extracellular calcium attenuates adipogenesis in 3T3-L1 preadipocytes

Abstract

We studied the effect of extracellular Ca 2+ concentration ([Ca 2+] e) on adipocyte differentiation. Preadipocytes exposed to continuous [Ca 2+] e higher than 2.5 mmol/l accumulated little or no cytoplasmic lipid compared to controls in 1.8 mmol/l [Ca 2+] 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 [Ca 2+] e inhibited expression of peroxisome proliferator-activated receptor γ, CCAAT/enhancer binding protein α, and steroid regulatory binding element protein. High [Ca 2+] 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 [Ca 2+] e. Treatment of 3T3-L1 cells with high [Ca 2+] e did not significantly affect cell number or viability and did not trigger apoptosis. Levels of intracellular Ca +2 remained unchanged in various [Ca 2+] e. Treatment of 3T3-L1 with pertussis toxin (PTX) partially restored lipid accumulation and increased differentiation markers in cells treated with 5 mmol/l [Ca 2+] e. ‘Classical’ parathyroid cell Ca 2+ sensing receptors (CaSR) were not detected either by RT-PCR or by Western blotting. These results suggest that continuos exposure to high [Ca 2+] e inhibits preadipocyte differentiation and that this may involve a G-protein-coupled mechanism mediated by a novel Ca 2+ sensor or receptor.