Figure 4.

Follow-up of acute senescence in human and mouse cells.

Human MSCs and mouse fibroblasts were treated with peroxide hydrogen to induce senescence. We collected cells and performed immunocytochemistry to evaluate RB1 and RB2/P130 expression 3 and 24 hours following stressor treatment. The two time points were indicated as early and late phase of senescence.

(A) Fluorescence photomicrographs show the merging of cells stained with anti-RPS6 (green), anti–Ki-67 (red), and RB1 (blue) in human MSCs during early and late senescence. Light microscopy pictures show the same fields stained to detect senescence-associated beta-galactosidase. In early senescence, the arrow indicates a senescent cell that is Ki67(+) and RB1(+). In late senescence, the arrows indicate a senescent cell that is negative for Ki67 and RB1. RPS6 stands for 40S ribosomal protein S6 and was used to detect cells.

(B) Photomicrographs show the merging of cells stained with anti-RPS6 (green), anti–Ki-67 (red), and RB2 (blue) in human MSCs during early and late senescence. Light microscopy pictures show the same fields stained to detect senescence-associated beta-galactosidase. In early phase, the arrow indicates a senescent cell that is Ki67(+) and RB2(−). In late senescence, the arrows indicate a senescent cell that is Ki67(−) and RB2 (+).

(C and D) Experiments carried out in mouse fibroblasts. The panels show the same staining we reported in panels A and B, respectively. In panel C (early), the arrow indicates a senescent cell that is positive for Ki67 and RB1. In late phase, the cells become Ki67(−) but remained RB1 positive (see arrow).

In panel D (early), the cells are Ki67 (+) and RB2 (−). In the late phase, cells were Ki67(−) and became RB2 (+) (See the arrows).