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Although functional association between Wnt signaling and bone homeostasis has been well described through genetic ablation of Wntless (Wls), the mechanisms of how osteoblastic Wls regulates the fate of bone marrow stromal cells (BMSCs) and hematopoietic stem cells (HSCs) in relation to age are not yet understood. Here, we generated Col2.3-Cre;Wls fl/fl mice that were free from premature lethality and investigated age-related impacts of osteoblastic Wls deficiency on hematopoiesis, BM microenvironment, and maintenance of BMSCs (also known as BM-derived mesenchymal stem/stromal cells) and HSCs. Ablation of osteoblastic Wls deteriorated BM microenvironment and bone mass accrual along with age-independent effects on functions of BMSCs. Osteoblastic Wls deletion impaired HSC repopulation and progeny with skewing toward myeloid lineage cells only at old stage. As proven by hallmarks of stem cell senescence, osteoblastic Wls ablation differentially induced senescence of BMSCs and HSCs in relation to age without alteration in their BM frequency. Our findings support that deletion of Wls in Col2.3-expressing cells induces senescence of BMSCs and impairs BM microenvironment in age-independent manner. Overall, long-term deterioration in BM microenvironment contributes to age-related HSC senescence with impaired progeny and hematopoiesis, which also suggests possible roles of osteoblastic Wls on the maintenance of BM HSCs.

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Nuclear factor (erythroid-derived 2)-like 2 (Nrf2)/heme oxygenase-1 (HO-1) signal is known to play important roles in controlling bone homeostasis. This study examined how oxidative stress affects the mineralization of embryonic stem (ES) cells by exposing them to glucose oxidase (GO), which continuously generates H2O2 at low concentrations. The roles of Nrf2/HO-1 and mitogen-activated protein kinases on osteogenesis in GO-exposed ES cells were also investigated. GO treatment at relatively low concentrations did not change the viability of ES cells, whereas it enhanced osteogenic differentiation and mineralization in the cells. GO treatment (1 mU/ml) augmented the induction of runt-related transcription factor 2 (Runx2), Nrf2, and HO-1 in ES cells. GO-mediated acceleration of Runx2 expression and mineralization was inhibited either by Nrf2 knockdown or by treating with 5 μM PD98059, an inhibitor of phospho-extracellular signal-regulated kinase (p-ERK). The GO-stimulated mineralization was also suppressed by treating the cells with reduced glutathione or catalase, but not by superoxide dismutase or N-acetyl-cysteine. Collectively, our results demonstrate that a mild oxidative stress activates Nrf2/HO-1 signaling and an ERK-mediated pathway, and facilitates the mineralization of ES cells with a corresponding increase in Runx2.

Total body irradiation (TBI) serves as an effectively curative therapy for cancer patients and adversely causes long-term residual bone marrow (BM) injury with premature senescence of hematopoietic stem cells (HSCs), which is mediated by increased production of reactive oxygen species (ROS). In the present study, we investigated how the exposure time of TBI in a mouse model affects HSCs and whether the treatment of caffeic acid (CA), a known dietary phenolic antioxidant, has a radioprotective effect. Single (S)-TBI at a sublethal dose (5 Gy) caused relatively higher induction of mitochondrial ROS and senescence-related factors in HSCs than those in hematopoietic progenitor cells (HPCs) and Lineage-Sca-1 + c-Kit + (LSK) cells, as well as reduced clonogenic formation and donor cell-derived reconstituting capacity. Repetitive double (D)-TBI (two months after the S-TBI at a dose of 5Gy) further weakened HSPC function via mitochondrial ROS accumulation and senescence-associated β-galactosidase (SA-β-gal) activity. Oral administration of CA (20 mg/kg) five times before and once immediately after TBI ameliorated ROS generation and TBI-induced HSC senescence and its radioprotective effect was long lasting in S-TBI mice but not in D-TBI mice. Further, supplementation of CA also induced apoptotic cell death of colon cancer cells. Collectively, these findings indicate that CA has a dual effect, ameliorating HSC senescence-accompanied long-term BM injury in S-TBI mice and stimulating apoptotic cell death of colon cancer cells.