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. 2024 Apr 30;23(7):e14188. doi: 10.1111/acel.14188

FIGURE 2.

FIGURE 2

Doxycycline alleviates the cell senescence of Zmpste24 KO MEF and HGPS fibroblasts in vitro. (a) The q‐RT‐PCR analysis of mRNA expression of Il6, Il1b, p16 and p21 in ZKO MEF cells with or without DOX treatment. (b,c) The statistical analysis of abnormal nuclear membrane (b) and the 53BP1 foci number (c) in WT or ZKO MEF cells with or without DOX treatment. (d) The SA‐β‐Gal staining analysis of ZKO MEF cells at passage 8 with or without DOX treatment. Bar, 100 μm. (e) The q‐RT‐PCR analysis of mRNA expression of Il6, ILlb, p16 and p21 in HGPS skin fibroblasts with or without DOX treatment. (f) The western blotting analysis of lamin A/C, IL6, p16 and p21 protein expression in HGPS cells with or without DOX treatment. HGPS1, HGADFN122; HGPS2, HGADFN169. (g,h) The statistical analysis of abnormal nuclear membrane (g) and the γH2AX foci number (h) in normal human dermal fibroblasts (NHDFs) and HGPS skin fibroblasts with or without DOX treatment. (i) The SA‐β‐Gal staining analysis of HGPS fibroblasts at passage 29 with or without DOX treatment. Bar, 100 μm. (j) The FACS analysis of cell death (PI staining) in ZKO MEF cells and HGPS fibroblasts with or without DOX treatment. (k,l) The western blotting analysis of NAT10 and Ac‐Tubulin protein expression in WT and ZKO MEF cells (k), NHDFs and HGPS fibroblasts (l) with or without DOX treatment. (m) The representative figures of immunofluorescence (IF) staining with anti‐Ac‐α‐Tubulin antibody in ZKO MEF cells, HGPS fibroblasts and aorta tissue from ZKO mice treated with or without DOX. Bar, 50 μm. (n) The analysis of Ac‐Tubulin protein expression in HGPS cells transfected with control (Con) or NAT10 shRNAs, and treated with or without DOX. n.s., nonsignificant. * p < 0.05, ** p < 0.01, *** p < 0.001.