Background: Since sphingosine-1-phosphate (Sph-1-P) plays an important role as an extracellular mediator through interaction with specific cell surface receptors, especially in the area of vascular biology and immunology/haematology, determination of its plasma concentration may become important from the clinical viewpoint. Thus, we attempted to develop a method of measuring the plasma Sph-1-P concentration for use in the clinical laboratory setting.
Methods: After two-step lipid extraction, Sph-1-P was coupled with o-phthaldialdehyde, and the resultant fluorescent derivative was separated by high-performance liquid chromatography. C17-Sph-1-P was used as the internal standard, instead of dihydrosphingosine-1-phosphate, which had been used previously for the same purpose but was actually detected in plasma.
Results: Our procedures for preparing the plasma samples and assay Sph-1-P were found to be satisfactory for clinical laboratory testing. The plasma Sph-1-P concentrations were significantly higher in men (413.1 +/- 52.0 nmol/L; mean +/- SD) than in women (352.4 +/- 39.7 nmol/L). Unexpectedly, strong positive correlations were found between the plasma Sph-1-P concentration and red blood cell (RBC)-related parameters, rather than platelet-related parameters.
Conclusions: Our present study confirmed the possibility of the clinical introduction of plasma Sph-1-P measurement, and in addition, suggested that RBCs may be involved in the regulation of plasma Sph-1-P concentrations.