Figure 2.

Overexpression of Spns2 does not alter cellular levels of S1P and dihydro-S1P or other sphingolipids. HEK 293 cells transfected with control or Spns2 were washed, and culture medium was replaced with medium containing phosphatase inhibitor. After 2 h, lipids were extracted from cells, and dihydrosphingosine, dihydro-S1P, sphingosine, and S1P (A), different chain length species of ceramide (B), monohexosylceramide (C), and sphingomyelin (D) were determined by LC-ESI-MS/MS. Numbers indicate chain length, followed by the number of double bonds in the fatty acid. Data are averages of triplicate determinations and are expressed as picomoles of lipid per million cells. Similar results were found in two additional experiments.