Tanon 5500

The cultured cells and mouse aortas were lysed by the RIPA buffer containing 0.1 mmol/L phenylmethanesulfonyl fluoride (Beyotime). The cytoplasmic and nuclear proteins were extracted by the corresponding protein extraction kits (Beyotime). The protein concentration was quantified by a BCA Protein Assay Kit. The protein samples were subjected to SDS-PAGE and then transferred to polyvinylidene difluoride membranes (Millipore, Billerica, MA, USA). After blockade in 5% skim milk for 1 h at room temperature, the membranes were incubated with primary antibodies against asprosin (FNab09797, 1:1000), LXRα (ab176323, 1:2000), PCSK9 (ab95478, 1:1000), CD36 (ab133625, 1:500), SR-A (AF1797, 1:1000), Elk-1 (ab11847, 1:500), p-Elk-1 (Ser383; ab34270, 1:300), p-Elk-1 (Ser389; ab28818, 1:300), p-Elk-1 (Thr417; ab28817, 1:300), p38 (ab223619, 1:500), p-p38 (ab32557, 1:1000), JNK (ab112501, 1:500), p-JNK (ab124956, 1:1000), ERK1/2 (ab17942, 1:1000), p-ERK1/2 (ab223500, 1:400), ABCA1 (ab7360, 1:200), ABCG1 (ab52617, 1:1000), histone H3 (ab176842, 1:3000) or β-actin (ab115777, 1:3000) overnight at 4 °C. Subsequently, the membranes were incubated and reacted with HRP-conjugated secondary antibodies (1:3000) for 2 h at room temperature. The immunoreactive bands were visualized using the BeyoECL Plus kit (Beyotime) on Tanon 5500 (Shanghai, China). β-actin was used as an internal control.

The tissues and cultured cells were lysed by RIPA buffer (Beyotime) containing 0.1 mmol/L phenylmethylsulfonyl fluoride. Protein extracts were quantified and then subjected to SDS-PAGE, followed by immunoblotting with mouse monoclonal antibody against ABCA1 (ab18180, 1:500, Abcam, Cambridge, MA, USA), rabbit monoclonal antibody against ABCG1 (ab52617, 1:1000, Abcam), rabbit monoclonal antibody against CD36 (ab133625, 1:500, Abcam), mouse polyclonal antibody against SR-A (AF1797, 1:1000, R&D Systems, MN, USA), rabbit monoclonal antibody against LXRα (ab176323, 1:500, Abcam), rabbit polyclonal antibody against HDAC1 (ab53091, 1:1000, Abcam), rabbit monoclonal antibody against HDAC3 (ab32369, 1:1000, Abcam), mouse monoclonal antibody against HDAC5 (ab50001, 1:300, Abcam), mouse monoclonal antibody against HAT-1 (sc-390562, 1:500, Santa Cruz, TX, USA), or rabbit monoclonal antibody against β-actin (ab115777, 1:1000, Abcam). After rinsing with PBS-T, the membranes were incubated with HRP-labeled secondary antibodies (1:3000, Beyotime). The proteins were visualized using Tanon 5500 (Shanghai, China) and BeyoECL Plus (Beyotime), and β-actin was used as an internal control.