Selenoprotein P attenuates oxidative senescence and ferroptosis-associated lipid peroxidation in dental pulp stem cells through a FOXM1-dependent program

Figure 5 Forkhead box protein M1 is required for the anti-senescence, antioxidant, and ferroptosis-associated protective effects of selenoprotein P/selenite co-treatment. A: Quantitative real-time polymerase chain reaction analysis of forkhead box protein M1 (FOXM1) knockdown efficiency in dental pulp stem cells transfected with two independent small interfering RNAs (siF1 and siF2); B: Western blot analysis of FOXM1 protein after transfection with siF1 or siF2; C: Densitometric quantification of FOXM1 protein; siF1 showed higher knockdown efficiency and was used for subsequent experiments; D: Representative SA-β-gal staining images showing that FOXM1 knockdown (Trt + siF1) weakens the anti-senescence effect of Trt. Scale bars = 50 μm; E: Quantification of SA-β-gal-positive cells; F: Western blot analysis of senescence markers (P53, P21, and P16) and the proliferation marker proliferating cell nuclear antigen; G: Densitometric quantification of proteins shown in panel F; H: CCK-8 assay showing that FOXM1 depletion compromises the proliferative rescue induced by Trt; I: Reduced glutathione levels; J: Oxidized glutathione levels; K: Glutathione/oxidized glutathione ratio; L: Western blot analysis of TFRC and glutathione peroxidase 4; M: Densitometric quantification of TFRC and glutathione peroxidase 4; N: Representative FerroOrange staining images showing re-accumulation of labile Fe²⁺ in the Trt + siF1 group. Scale bars = 20 μm; O: Quantification of FerroOrange fluorescence intensity; P: Quantitative real-time polymerase chain reaction analysis of downstream targets (MYBL2, SESN3, and CDC20). Data are presented as mean ± SD. ^aP < 0.05. FOXM1: Forkhead box protein M1; DPSC: Dental pulp stem cell; PCNA: Proliferating cell nuclear antigen; GSH: Glutathione; GSSG: Oxidized glutathione; GPX4: Glutathione peroxidase 4.