Extracellular GalSph and GlcSph enter the lysosomes of MCF7 cells.

(A, B) Quantitites of HexSph (GalSph and GlcSph) in cell lysates (cells) and cell culture medium (media) of MCF7 cells treated with 45 μM GalSph (A) or 32 μM GlcSph (B) for indicated times were analyzed by shotgun lipidomics. (C) The molar percentages (mol%) of HexSph in MCF7 cells treated with 45 μM GalSph or 32 μM GlcSph for indicated times were analyzed as in (A). (D-G) The molar percentages of HexSph (D), BMP/PG (E), CL (F), and PC (G) lipid classes in total cell lysates (cells) and in purified lysosomes of MCF7 cells treated as indicated for 3 hours were analyzed as in (A). Error bars, SD of three independent experiments. *, P P P P < 0.0001 comparing the treated values to the first measurement at 0.02 hours (A, B) or to the control appropriate sample (D-G) and analyzed by unpaired t-test (A, B, E-G) or two-way ANOVA followed by Sidak’s multiple comparisons test (D). BMP, bis(monoacylglycero)phosphate; PG, phosphatidylglycerol; CL, cardiolipin; PC, phosphatidylcholine.