Fig. 7
The addition of α-KG rescues the phenotypes induced by ILF2 knockdown. (A-C) Proliferation rate status of MDA-MB-231 cells was assessed through CCK-8, colony formation assays, and EdU assays, n = 3. scale bar: 50μm. D Cell cycle analysis was conducted via flow cytometry, involving propidium iodide (PI) staining on MDA-MB-231 cells, n = 3. E Apoptosis detection was performed using a flow cytometry assay. Annexin V and propidium iodide (PI) staining were employed on MDA-MB-231 cells, n = 3. F Left, oxygen consumption rate (OCR) on addition of oligomycin (Oligo), fluorocarbonyl cyanide phenylhydrazone (FCCP) and rotenone plus antimycin A (R&A) (n = 4). Right, basal respiration, ATP-coupled respiration and maximal respiration (n= 4). G-H Relative lactate level (G) and relative ATP level (H) in MDA-MB-231 cells with ILF2 knockdown and α-KG supplementation were shown, n = 3. (I) Relative lactate level (left) and relative ATP level (right) in MDA-MB-231 cells with ILF2 knockdown and α-KG supplementation were shown, n = 3. J-N In vivo studies involved BALB/c athymic nude mice subcutaneously injected with MDA-MB-231 cells, n = 5. Images of xenograft s were captured using a digital camera (J). Tumor weight was measured on day 50 (K). Tumor volume was measured at ten-day intervals, calculated as volume = length × (width)2/2 (L). Immunohistochemical images showing Ki67 and PCNA staining in control, shILF2-, α-KG-, and shILF2 combined plus α-KG-treated groups. scale bar: 100μm (M). Immunofluorescence images showcased TUNEL staining in control, shILF2-, α-KG-, and shILF2 combined plus α-KG-treated groups. scale bar: 60μm (N). Statistical analyses are depicted in bar graphs. Data are presented as mean ± SD. Significance levels are denoted as * for p<0.05, ** for p<0.01, and *** for p<0.001, as determined by the t-test