To obtain conditioned media from BMMCs, 1 × 107 BMMCs were cultured in IL-3-containing BMMC complete medium for 2 days. Neutral lipids, phospholipids and free fatty acids were extracted from the conditioned media by solid-phase extraction using Sep-Pak C18 cartridges (Waters). Briefly, the conditioned medium was added to Sep-Pak columns and neural lipids were eluted with 10 ml of hexane. Free fatty acids were subsequently eluted with 10 ml of methyl formate. Phospholipids were finally eluted with 10 ml of methanol. The free fatty acid fraction was used as experiments.
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