Microrna Mediated Regulation of Hematopoietic Stem Cell Aging

Aging hematopoietic stem cells (HSCs) exhibit numerous functional alterations including reduced capacity for self-renewal, myeloid-biased differentiation, and reduced production of mature lymphocytes and red blood cells. Interventions such as calorie restriction (CR) and rapamycin (Rapa) treatment have been shown to increase lifespan and to delay the onset of age-related diseases, and some studies have demonstrated that they may improve HSC function through poorly defined mechanisms. We and others have shown that microRNAs (miRNAs) are potent cell-intrinsic regulators of HSC self-renewal and lineage specification and also contribute to age-related disorders such as acute myeloid leukemia (AML) and the myelodysplastic syndromes (MDS). We hypothesized that miRNAs may underlie the recovery of HSC function observed in anti-aging mouse models, and thus we characterized miRNA expression profiles from HSCs (Lin-c-Kit+Sca-1+CD34-CD150+) from young mice (12-16 weeks old), old mice (20-22 months), and old mice that had been treated with anti-aging interventions. Evaluation of HSCs from CR and Rapa treated old mice revealed numerous changes consistent with inhibition/reversal of age-related HSC changes including a 5-fold reduction in HSC frequency (p=0.04), 2-fold increase in erythroid progenitors (pro-erythroblasts, p=0.04), 2.5 fold increase in common lymphoid progenitors (CLP; Lin-c-Kit+Sca-1+CD127+FLK2+, p=0.05), as well as 3.5-fold increase in peripheral blood B cells (p=0.002), 2.2 fold decrease in platelets (p=0.01), and increased red blood cells (p=0.04). These changes were associated with statistically significant increases in the percentage of HSCs in S/M/G2 (p=0.045), and undergoing apoptosis (p=0.05). Using a TaqMan-based qPCR expression profiling method evaluating 750 miRNAs, we found that old HSCs exhibited altered expression of 91 miRNAs compared to young (FDR <0.1, P <0.05). Moreover, HSCs from both CR and Rapa treated old mice exhibited expression of 60 miRNAs at levels similar to young, normal HSCs. miR-125b, a miRNA we and others previously showed to positively regulate HSC self-renewal, was reduced 2.2-fold in old mice, and its expression was restored in CR and Rapa treated HSCs. Lentivirally mediated expression of miR-125b in old HSCs increased their long-term reconstitution capacity 8.1-fold compared to control old HSCs based on donor chimerism levels at 16 weeks post-transplantation, resulting in chimerism levels similar to mice transplanted with young HSCs expressing miR-125b. The improved HSC engraftment capacity of old HSCs transduced with miR-125b was accompanied by statistically significant increases in the frequencies of lymphoid biased HSCs (Lin-c-Kit+Sca-1+CD34-CD150neg-low), megakaryocyte-erythroid progenitors (MEPs), CLPs, and peripheral blood B- and T-cells, compared to old HSCs transduced with control lentivirus (p<0.05 for all indicated cell types). While enforced expression of high levels of miR-125b in mouse HSPCs has been reported to induce myeloid leukemias, there was no evidence of a hematologic malignancy in mice transplanted with miR-125b transduced old HSCs up to 6 months post-transplantation. Overall, these results demonstrate that functional HSC aging phenotypes can be that inhibited/reversed by anti-aging interventions, that miR-125b regulates HSC aging, and that anti-aging interventions may exert their positive effects on HSC function by regulating miR-125b expression.