Fig. 1

The design, property, senescence alleviation, and age-related phenotype improvement of SeHed. a The construction of SeHed. b The top ten active transcription factors upregulating pro-senescent genes in oxidative stressed-cellular senescence model predicted by TFactS software. NF-κB transcription factor family, including NF-κB (p52), RELA (p65), and REL (cRel) are remarked as red sign. c (left) The construction of luciferase reporter plasmid (pGL3B-NF8p). The NF8p (binding core sequence: NGGGGACTTTCCN), containing eight repeated core sequences, was cloned into the pGL3B plasmid. (right) The dual-luciferase assay was performed in HEK293T cells co-transfected with pRT-L plus pGL3B-NF8p plasmid and treated with H₂O₂ (100 μM), TNFα (10 ng/ml), and QNZ (0.5 μM) was applied for 8 h. d The dual-luciferase assay was performed in HEK293T cells co-transfected with pRT-L plus pGL3B-NF8p plasmid and treated with H₂O₂ (100 μM), TNFα (10 ng/ml), and NAC (2 mM) was applied for 8 h. e Representative of western blots for HA (mCat), p65, and p-p65 were shown. NIH3T3 cells were infected with lentivirus for expressing the SeHed (pNF8p-HA-mCAT-puro). Then, cells were treated with H₂O₂ (100 μM) or TNFα (10 ng/ml) for different times. f Representative of western blots for mCat, p65, and p-p65 in SeHed-expressed NIH3T3 cells treated with H₂O₂ (100 μM), NAC (2 mM) or QNZ (0.5 μM) for 4 and 8 h. g Cellular ROS and mtROS levels were monitored in H₂O₂ or ADR induced-cellular senescence models. The cellular ROS and mtROS levels were labeled by DCFH-DA and mitoSOX, respectively, and quantified by fluorescence microplate reader in Mock- and SeHed-expressed NIH3T3 cells. h Representative images of SA-β-gal staining and the ratio of SA-β-gal positive cells were shown. The NIH3T3 and MRC-5 cells were infected with lentivirus expressing the Mock or SeHed, respectively. Then, these cells were incubated with H₂O₂, ADR or Rot to induce cellular senescence. i, j Mice were injected with two doses of AAV9-SeHed virus, respectively (2.5 × 10¹¹ GC/mice and 5 × 10¹¹ GC/mice). four weeks later, all mice were intraperitoneally injected with LPS (5 mg/kg) for 10 h. (n = 3/group). i Relative mCat mRNA expression in the liver was analyzed by qRT-PCR assay. j Representative images of western blots for mCat in the liver were shown. k The MDA level in serum and liver were shown. (left) Mice were divided into Saline, D-gal (100 mg/kg/day) and SeHed (AAV9-SeHed, 5 × 10¹¹ GC/mice, plus D-gal (100 mg/kg/day)) groups. (right) Mice were divided into Standard diet (SD), High-fat diet (HFD) and SeHed (AAV9-SeHed (5 × 10¹¹ GC/mice) plus HFD) groups (n = 5–6/group). l Representative images of SA-β-Gal staining of epididymal fat tissue. m Representative images of western blots for p16, p65, p-p65, and mCat in the liver. 1, 2, 3 represent mice num. n (up) Representative images of myotubes formation in Mock-, CMV-mCAT (CMV)- and SeHed-expressed mouse primary myoblast cells cultured in DMEM medium with 2% HS (Horse Serum) for 3 days. (down) Representative images of myotubes formation in Mock-, CMV- and SeHed-expressed H9C2 cells cultured in DMEM medium with 2% HS (Horse Serum) and 10 μM retinoic acid for 6 days. Scale bars, 50 μm. o Representative images of western blots for Myogenin and mCat in Mock-, CMV- and SeHed-expressed H9C2 cells cultured in a medium with 2% HS plus 10 μM retinoic acid for 6 days. p Schematic overview of SeHed system in aging and diseases. All data were shown as mean ± SD, *P < 0.05, **P < 0.01, ***P < 0.001; NS no significance