Journal: Journal of Lipid Research

Article Title: Aerobic glycolysis drives differentiation of unilocular adipocytes

doi: 10.1016/j.jlr.2026.101023

Figure Lengend Snippet: HUVAS adipocytes are characterized by AMPK activation and high fatty acid uptake. A, B: Capillary Western blot analysis of p-AMPK T172 , total AMPK, p-HSL S660 , p-HSL S563 (shown with 2 contrasts), total HSL, and CD36 as well as quantification of the p-AMPK/AMPK and p-HSL/HSL ratios after normalization to total protein levels (Fig. S6A, S7A) across HUVAS, 2D, and 3D cultures from 3 technical replicates of the same individual on d30. C, D, isoprenaline-induced (C, shown as induced/basal) and basal (D) lipolysis measured by glycerol release across cultures from 8 technical replicates of the same individual. Similar results were obtained from cells from >three different cell donors. E, F: Quantification of the uptake of a BODIPY-labelled long-chain fatty acid tracer (BODIPY-FA) across HUVAS, 3D, and 2D cultures (E), and in HUVAS and 3D cultures in the absence or presence of the CD36 inhibitor SMS121 or vehicle (F). N ≥ 5 throughout; values in (F) are expressed as relative to the mean of HUVAS control to combine data from several experiments and cell donors. G–H: Quantification of CD36 mRNA expression expressed as normalized counts (G) and protein from capillary Western blot analysis (H) after normalization to total protein levels ( A) from 3 technical replicates of the same individual on d30. I–K: Effects of chronic treatment of HUVAS with CD36 inhibitor SMS121 d20-d30 shown as representative confocal images at d30 (I), adipocyte lipid droplet diameter (J), and unilocularity (K). n = 40 cells per spheroid and 2 spheroids per individual. Scale bars: 50 μm.

Article Snippet: Immunoblotting was performed using the following primary antibodies against: PLIN1 (1:10, PA5-81240, Fisher Scientific), CIDEC (1:10, PA1-46128, Invitrogen), UCP1 (1:50, MAB6158, R&D Systems), PPARγ (1:50, 2435S, Cell Signaling), FAS (1:50, 3180, BioTechne), ATGL (1:50, 2138, Cell Signaling), Total OXPHOS cocktail (1:50, Ab110411 , Abcam), TOMM20 (1:50, NBP1-81556, BioTechne), PTP1B (1:50, Ab244207 , Abcam), HIF1α (1:100, NBP2-75977, Novus), P-AMPK (Thr172, 1:10, 2535, Cell Signaling), total AMPK (1:50, 2532, Cell Signaling), P-HSL (Ser660, 1:10, 45,804, Cell Signaling), P-HSL (Ser563, 1:10, 4139, Cell Signaling), total HSL (1:50, 4107, Cell Signaling), and CD36 (1:50, AF1955, R&D Systems).

Techniques: Activation Assay, Western Blot, Control, Expressing