Figure 6.

MEDEP-E14 cells exhibit high S1P import activity.A, total RNA prepared from HUVECs and MEDEP-E14 cells was subjected to quantitative real-time RT-PCR using specific primers for SPNS2, MFSD2B, and GAPDH. Values presented are means ± SD and indicate quantities relative to that of GAPDH from three independent reactions. B and C, HUVECs and MEDEP-E14 cells were labeled with 0.2 μCi [³²P]Pi or 0.2 μCi [³²P]S1P for 3 h in the presence of 1% bovine serum albumin. Lipids were extracted and separated by TLC, followed by detection (B) and quantification (C) using an imaging analyzer BAS2500. Values represent the means ± SD of the radioactivity of [³²P]PE relative to that in [³²P]Pi-labeled HUVECs from three independent experiments (∗∗p < 0.01; Student's t test). D, total RNA prepared from HUVECs and MEDEP-E14 cells was subjected to quantitative real-time RT-PCR using specific primers for PLPP1–3 and GAPDH. Values presented are means ± SD and indicate quantities relative to that of GAPDH from three independent reactions. HUVECs, human umbilical vein endothelial cells; MEDEP, MEDEP-E14; MFSD2B, major facilitator superfamily domain containing 2B; PE, phosphatidylethanolamine; Pi, orthophosphoric acid; PLPP, phospholipid phosphatase; SM, sphingomyelin; S1P, sphingosine-1-phosphate; SPNS, sphingolipid transporter; #unidentified lipid.