Figure 6.

(A) Western Blot analysis of phospho -ERK1/2^{thr202/tyr204} in Unpass- and Pass-ASC without or with exposure to α₅β₁-integrin-specific peptide agonist CRRETAWAC (5 μg/ml, 30 minutes) stim condition. GAPDH was used as internal protein loading control. A representative blot following signal detection is shown. Values for levels of phospho -ERK1/2^{thr202/tyr204} were normalized with respect to their corresponding GAPDH value and are expressed relative to levels in unstimulated Unpass-ASC *p < 0.05, **p < 0.01 (ANOVA with Dunnet’s multiple comparisons test, after data normalization by logarithmic transformation). (B) Number of colonies (expressed as % of the colonies in the Unpass-ASC condition) of Unpass-ASC alone or after stimulation of the α₅β₁-integrin pathway only once (boost condition) or for the whole duration of the culture (stim condition). *P < 0.05 (Kruskal-Wallis with Dunn multiple comparisons test); (C) Representative fields of histological sections of osteogenic constructs loaded with Unpass-ASC or Unpass-ASC stimulated with CRRETAWAC (stim condition) and stained with H&E. Size bar = 200 μm; (D) Quantification of bone tissue formation 12 weeks after implantation of constructs loaded with Unpass-ASC or Unpass-ASC stimulated with the CRRETAWAC. **p < 0.01 (t-test).