Fabppm

Routine Western Blotting procedures were used to detect the total expression of fatty acid transport proteins: FATP2, FATP5 (Santa Cruz Biotechnology, USA), FAT/CD36, FABPpm (Abcam, UK), ABCA1 (Thermo Scientific, USA), MTP (Santa Cruz Biotechnology, USA), ACBP and L-FABP (Abcam, UK) as well as the proteins directly involved in lipogenesis (FAS; Cell Signaling, USA), oxidation pathway (CPT 1; Santa Cruz Biotechnology, USA) and lipid metabolism (PPARα, LXR, SREBP1c, pAMPK; Cell Signaling, USA) as previously described in details by Konstantynowicz-Nowicka et al. [17 ]. All the antibodies used in our procedures were monoclonal except for FATP2 and FAT/CD36. Cell lysates were separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose membranes. After blocking with 5% nonfat dry milk, the membranes were immunoblotted with primary antibodies of interest and incubated with secondary antibodies labeled with horseradish peroxidase (HRP). Obtained protein bands were quantified densitometrically using a ChemiDoc visualization system (Bio Rad, Warsaw, Poland). Equal protein loading was controlled by Ponceau S staining. The expression of all the proteins was standardized to the GAPDH (Santa Cruz Biotechnology, USA) expression and the control was set as 100%.