Vitamin C Treatment Rescues Prelamin A-Induced Premature Senescence of Subchondral Bone Mesenchymal Stem Cells

Figure 3

Prelamin A accumulation triggered SCB-MSCs and increased DNA damage, apoptosis resistance, and cell cycle arrest. Representative confocal images of (a) γ-H2AX staining and (b) quantification of γ-H2AX foci. Blue: DAPI; red: γ-H2AX. Scale bar: 25 μm. At least 100 nuclei were analyzed per sample. was used for statistical significance. (c) Representative confocal images of 53BP1 in MSC/GFP and MSC/PLA. Blue: DAPI; red: 53BP1. Scale bar: 50 μm. (d) Remarkable increase of GADD45A mRNA level (). (e) Comparison of mRNA expression levels of Shelterin complex (POT1, RAP1A, TERF1, TERF2, TPP1, and TINF2) (, ). (f) The apoptotic cells were decreased in SCB-MSC overexpressing prelamin A using Annexin V/7AAD. (g) Quantitative RT-PCR showed the downregulation of caspase 3 and the significant elevation in the mRNA level of Bcl2 (, ). (h) Flow cytometry analysis showed plunged S phase in the MSC/PLA, compared with the MSC/GFP (). (i) Protein levels of p21 and p53 significantly increased in prelamin A-overexpressed SCB-MSCs. (j) Quantitative RT-PCR verified the upregulation of p53 and p21 in MSC/PLA (). (k) The downregulated CDK2 and CCNE in the MSC/PLA in mRNA level demonstrated the block in G1/S checkpoint (, ).

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