Fig. 3: α-KG induced oxidative stress by promoting ROS production and ATP reduction in DHL.

A–C OCI-LY1 and OCI-LY10 cells were treated with 15 mM DM-αKG for 24 h and performed RNA-seq (3 biological replicates for each group). Cells treated with 0.1% DMSO were used as control. Bubble plot profiled the most significantly differentially expressed GO terms (A). Contributing genes were displayed by a string diagram (B). GSEA plot of the NADH metabolic process (C, NES = 1.5, p < 0.05). D, E OCI-LY1 and OCI-LY10 cells were treated with different agents for 24 h: (1) 0.1% DMSO; (2) 15 mM DM-αKG; (3) 1 μM Fer-1 pre-treatment followed by 15 mM DM-αKG. DCFDA assay was performed to detect intracellular ROS levels, represented by flow cytometry plots (D) and relative fluorescence quantification (E). F, G OCI-LY1 and OCI-LY10 cells were treated with 0.1% DMSO or 15 mM DM-αKG for 24 h. Reagents containing D-luciferin and firefly luciferase were used to determine ATP levels, represented by the luminescence intensity (F). Immunoblot analysis revealed the protein levels of PCG-1α, NRF-1, and ERRα in DMSO- and DM-αKG-treated cells. Wild-type cells without drug treatment were used as the positive control (Blank) (G). All data were presented as means ± SD of 3 independent experiments. Immunoblot images in (G) were the representation of 3 independent experiments. The data were analyzed by two-tailed t test in (E) and (F). ^*P < 0.05, ^**P < 0.01, ^***P < 0.001.