Research Paper Advance Articles

Figure 4. LKU4 negatively regulates H₂O₂-induced adipocyte senescence through NDN upregulation. (A) Reporter gene analysis using p21-promoter–Luc in HEK293T cells. (B–G) γH2AX, p53, p21, and NDN protein expression (B), SA-β-gal staining (scale bars, 200 μm) (C), RT-qPCR analysis of SASP genes (D) and mitochondrial function-associated genes (E), ROS levels (F), mtDNA copy number and CS activity (G) in primary adipocytes. Primary adipocytes differentiated from SVF cells were transfected with expression plasmids and NDN siRNA for 12 h, followed by treatment with 100 μM H₂O₂, 50 μM sirtinol, and LKU4–CM for another 24 h, as indicated, before analysis. (H) Oxygen consumption rate (OCR) analysis using a Seahorse XFe analyzer in 3T3-L1 adipocytes overexpressing the indicated expression plasmids in the absence or presence of LKU4–CM. (I) Intracellular TG levels in primary adipocytes. Differentiated primary adipocytes were transfected with expression plasmids and NDN siRNA and then treated with 100 μM H₂O₂, 50 μM sirtinol, and LKU4–CM, as indicated.