A Small-Sized Population of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Shows High Stemness Properties and Therapeutic Benefit

Figure 5

EGFR knockdown induces senescence and results in a loss of differentiation capability. Small-size populations of mesenchymal stem cells (MSCs) were transfected with scramble siRNA (si Con) or EGFR siRNA (si EGFR). (a) Morphological differences were observed by microscopy for adhesion (upper panel) or single cells (lower panel,  μm). (b) Cell sizes of the three populations were assessed during expansion. The expression levels were normalized to those observed in naïve cells, defined as 1-fold expression (, ; , ). (c) Cells were cultured under regular conditions and population doubling (PD) was assessed until cell growth ceased (, ; , ). (d) The cells were stained with SA β-gal, and the activity was measured by counting positively-stained cells at P9 or P12 (, ; ,  μm). (e) The expression of senescence-associated proteins was measured by immunoblotting analysis and β-actin was used as a control at P12. The expression levels were normalized to those observed in naïve cells, defined as 1-fold expression (means ± SD, ; ). (f) After 2 weeks of osteogenic induction, the level of ALP was determined by staining ( μm).

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