Nutlin-3 treatment activates p53 and corrects the aberrant HSPC expansion of asrij KO mice. (A) Schematic showing protocol followed for Nutlin-3 treatment of mice and their subsequent analyses. Eight-month-old asrij floxed and KO mice were injected intraperitoneally with 20 mg/kg of Nutlin-3 and then analyzed for p53 activation by immunoblotting and HSPC frequencies by flow cytometry. (B) Immunoblot analysis of HSPCs shows increased wtp53 levels in Nutlin-3 treated mice compared with DMSO-treated controls in floxed and KO genotypes (n = 4 per genotype). Loading control: GAPDH. (C) Representative flow cytometry plots and graph show that Nutlin-3 reduces the increased HSPC frequencies observed in KO mice to near control levels (n = 6 per genotype). (D) Model representing mechanism of action of Asrij that regulates HSC quiescence. Blue solid line indicates protein interaction. Statistically significant differences in HSPC frequencies determined using ANOVA: single factor analysis are indicated. Error bars denote standard error of mean. *P < .05 and ***P < .001.