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. Author manuscript; available in PMC: 2016 Jan 20.
Published in final edited form as: J Am Coll Cardiol. 2015 Jan 20;65(2):133–147. doi: 10.1016/j.jacc.2014.09.086

FIGURE 2. NS Loss Causes Altered Morphology During Senescence.

FIGURE 2

(A) Phase contrast microscopy of young (YCPC) and old (OCPC) mouse CPCs, and FhCPC and AhCPC (lines FH-05 and H10-004) stained with senescence associated beta-galactosidase (SA-β-gal). Arrows indicate blue, senescent cells. (B) Graph indicates senescent cell counts. (C) Phase contrast images of SA-β-gal+ binucleate (2 nuclei, binuc.) and polynucleate (>2 nuclei, polynuc.) cells; and percentage of multinucleate YCPC/OCPCs are shown. (D) NS declines in flat, senescent OCPCs (arrows). Confocal microscopy of OCPCs stained with Phalloidin (Phal, grey), NS (red), and Sytox blue (SB, blue) (left). Phase contrast image of the same cells treated with SA-β-gal (right). (E) Immunoblot and densitometric analyses showing NS knockdown in YCPCs (N = 12). (F) Phase contrast (top panel, Line H10-005) and electron micrographs (bottom panel, Line H11-020) of AhCPCs with NS knockdown demonstrating flattened, SA-β-gal+ cells (arrows). (G–H) Senescent (G) and binucleated cell count (H) (N>200 cells). Scale bar: 100µm (A,C,F); 50 µm (D).

*p < 0.05; **p < 0.01; ***p < 0.001 (relative to each corresponding group in YCPCs/AhCPC+Lt-sh-control in C)

Lt-sh = Lentivirus harboring short hairpin ribonucleic acid; Other abbreviations as in Figure 1.