Journal: Journal of Innate Immunity
Article Title: Unique Francisella Phosphatidylethanolamine Acts as a Potent Anti-Inflammatory Lipid
Figure Lengend Snippet: Phosphatidylethanolamine (PE) and phosphatidylcholine (PC) isolated from SchuS4 impair inflammatory responses in human dendritic cells (hDC). Total lipid from SchuS4 was fractionated into 3 classes (fractions I–III [FI–III]) using aminopropyl silica solid-phase extraction, and these were tested for their ability to inhibit cytokine production in hDC. a Each fraction (20 µg/mL) or crude lipid (2 µg/mL) was added to hDC (3–4 wells/group) and incubated overnight. Control samples were treated with EtOH diluted similarly to lipids. hDC were then stimulated with E. coli LPS (5 ng/mL), and supernatants were collected the next day to assess TNF-α and IL-12p40. A–C indicate individual donors. Data were analyzed using one-way-ANOVA followed by Tukey's multiple-comparisons analysis. Error bars represent SD. b Each fraction was analyzed for lipid content by thin-layer chromatography. The indicated lipid fractions were spotted onto a silica plate and developed with Chl: MeOH:H 2 O (7: 3: 025). Lipid standards (Std), consisting of lyso-PC (a), PC (b), lyso-PE (c), and PE (d) were also added to the plate. Bands were visualized using primuline and UV illumination. The arrow designated as (1) indicates the presence of PE and (2) designates PC in FIII. c FIII was further fractionated by silica solid-phase extraction into 2 fractions, enriched for either PE (silica fraction [SF]1) or PC (SF2). SF1, SF2 (at the indicated concentrations), and crude lipid (10 µg/mL) were incubated with hDC overnight, the cells were then stimulated with E. coli LPS (5 ng/mL), and supernatants collected the next day to assess TNF-α and IL-12p40. Mock, mock-treated control. A–C indicate individual donors. Error bars represent SD.
Article Snippet: Primuline, ninhydrin, copper sulfate, isopropanol, acetone, hexane, ethyl acetate, and triglyceride standards were from Sigma-Aldrich, chloroform (Chl) from Macron (Center Valley, PA, USA), and methanol (MeOH) from Burdick Jackson (Muskegon, MI, USA).
Techniques: Isolation, Incubation, Thin Layer Chromatography