Sodium oleate

The isolated preadipocytes (SV cells) were seeded at a density of 1 × 10⁵ cells/cm² in DMEM/F12 (Gibco) with 10% fetal bovine serum (Gibco) at 5% CO₂. At 60% confluence, the cells were induced to differentiate by 160 μM sodium oleate (Sigma-Aldrich). The induced cells were harvested for qRT-PCR analysis every 12 or 24 h, continued for a total of 120 h.
For analysis of the effect of GATA3 overexpression on preadipocyte differentiation, ICP2 cells were seeded into 6-well plates at a density of 1 × 10⁵ cells/cm². At 60% confluence, the cells were transfected with the pCMV-Myc-GATA3 or pCMV-Myc (empty vector), using Invitrogen Lipofectamine^® 2000 (Invitrogen). At 24 h after transfection, 160 μM sodium oleate (Sigma-Aldrich) was added into the medium to induce preadipocyte differentiation. At 24 h after the induction of differentiation, the cells were stained by oil red O, or examined by qRT-PCR and western blot analysis. The c-Myc-GATA3 protein expression was detected by western blot. The adipogenic gene expressions of PPARγ, FABP4, and C/EBPα were analyzed by qRT-PCR. The sequences of primers are shown in Table 1.

Sodium palmitate and sodium oleate were obtained from Sigma‐Aldrich. Stock solution of 50 mM of each reagent was obtained by dissolution in H₂O at 70°C. Working solutions were made by conjugation of 2 mM palmitic acid or free fatty acid (FFA) mixture (Sodium palmitate: sodium oleate = 1:2) with 2% BSA‐MEM/F‐12.³⁶ Antibodies including anti‐LC3, anti‐Beclin‐1, anti‐ATG5 and anti‐β‐actin were obtained from Sigma‐Aldrich.
Mouse hepatocyte cell line AML‐12 was obtained from Shanghai Institutes for Biological Science, CAS. Cells were maintained in MEM/F12 medium supplemented with 10% foetal bovine serum, 10 μg/ml insulin, 5.5 μg/ml transferrin, 5 ng/ml selenium and 40 ng/ml dexamethasone. Culture medium containing 0.5% lipoprotein depleted foetal bovine serum (LPDS) was used when FFA was added.

Chemicals were obtained from Sigma Aldrich, including hexadecyltrimethylammonium bromide (CTAB, ≥98%), hydrogen tetrachloroaurate trihydrate (HAuCl₄·3H₂O), l-ascorbic acid, silver nitrate (AgNO₃), sodium borohydride (NaBH₄), hydrochloric acid (HCl, 37 wt% in water), 3-mercaptopropyltrimethoxysilane (MPTMS), sodium silicate solution, tetraethyl orthosilicate (TEOS), sodium hydroxide (NaOH), and potassium iodide (KI). Other chemicals obtained from additional suppliers were sodium oleate (NaOL, TCI), nitric acid (HNO₃, 70%, Merck), ethanol (analytical pure, Solveco) and iodine (I₂, VWR Chemicals).