Nongenomic regulation of chondrocyte membrane fluidity by 1,25-(OH) 2D 3 and 24,25-(OH) 2D 3 is dependent on cell maturation

Abstract

1,25-(OH) 2D 3 and 24,25-(OH) 2D 3 regulate rat costochondral chondrocyte cultures in a metabolite-specific manner; 1,25-(OH) 2D 3 targets primarily growth zone cells (GC) and 24,25-(OH) 2D 3 targets primarily resting zone cells (RC). Some of the effects are nongenomic, since incubation of isolated membrane fractions with the metabolites results in regulation of enzyme activities comparable to that seen in culture. This study examined whether changes in membrane fluidity might be one mechanism involved in the nongenomic regulatory pathway. Chondrocyte cultures were incubated with the vitamin D metabolites and changes in plasma membrane fluidity monitored using the fluorophore, TMA-DPH, which is specific for membranes exposed to external fluids. Isolated matrix vesicles were also incubated directly with the metabolites and anisotropy of the membrane, as well as alkaline phosphatase-specific activity, determined. 1,25-(OH) 2D 3 caused a rapid and constant increase in alkaline phosphatase-specific activity in GC matrix vesicles; 24,25-(OH) 2D 3 caused an increase in RC matrix vesicle enzyme activity that was both dose- and time-dependent. Matrix vesicles produced by GC had a lower degree of fluidity than their parent plasma membranes or RC plasma membranes and matrix vesicles. Fluidity of the GC membrane fractions was increased by 1,25-(OH) 2D 3 in a dose- and time-dependent manner. 1,25-(OH) 2D 3 had no effect on the fluidity of the RC membranes. 24,25-(OH) 2D 3 caused a decrease in fluidity in GC at later time points. This metabolite caused an increase in fluidity of RC plasma membranes that returned to normal levels by 6 h; however, the increase induced in the matrix vesicles remained elevated throughout the 24-h experimental period. The results demonstrate that vitamin D metabolites can alter membrane fluidity in a cell-, metabolite-, and membrane-specific manner. For GC matrix vesicles, 1,25-(OH) 2D 3-dependent increased fluidity correlates with increased alkaline phosphatase activity. While 24,25-(OH) 2D 3 increases enzyme activity, the relation of this increase to changes in fluidity is less clear, supporting previous observations concerning differential specificity of response to the two metabolites in growth plate chondrocytes.