Noncanonical Wnt Signaling Maintains Hematopoietic Stem Cells in Different Zones

Abstract 639

Hematopoietic stem cells (HSCs) are maintained in a balance between quiescent state and proliferating state. While proliferating HSCs are critical for supporting the routine blood production, quiescent HSCs are essential for long term maintenance and can also be activated to replenish lost proliferating or active HSCs. How the different states of HSCs are regulated is a fundamental question. Accumulated evidence supports a model that quiescent HSCs are located in the endosteal zone and active HSCs are in the perivascular zone. The underlying signaling to regulate the quiescence and activation in different niches remains largely unknown. To address this question, we have analyzed the expression profile of Wnt receptors, Frizzleds, in HSCs. We found that noncanonical Wnt signaling via receptor Frizzled8 (Fz8) and co-receptor Flamingo presents in and functionally maintains quiescent HSCs in the endosteal zone (Sugimura, et al., Cell 2012). However, it has not been clear whether and how active HSCs in the perivascular zone are regulated by Wnt signaling.

Recently, we detected that another noncanonical Wnt receptor, Frizzled5 (Fz5), is expressed in metabolically active (indicated by Mitotracker) HSCs and also in Nestin-GFP⁺ mesenchymal stem cells (MSCs) in the perivascular zone of central marrow. Fz5 expresses neither in H2B-GFP label-retaining quiescent HSCs nor in endosteal cells and nor in sinusoidal cells as well. Using an Mx1-Cre:Fz5 knockout mouse model, we found a 60% decrease of HSCs isolated from central marrow, but no change in the number of HSCs isolated from endosteum. Functionally, hematopoietic reconstitution was not affected in the primary transplantation, but was substantially decreased (by 80%) in the secondary transplantation compared to the control.

This indicates that Fz5 maintains HSCs in the perivascular zone. To examine the role of Fz5 in Nestin⁺ MSCs for HSC maintenance, we examined the Nestin-Cre:Fz5 model. We observed a large loss of CD49^hiHSCs that were reported to represent intermediate (IT) HSCs. We further found a correlation of the quiescent vs. active HSCs respectively to long term (LT) HSCs vs. IT-HSCs with the latter population sensitive to 5FU treatment. Mechanistically we observed that Fz5 inactivation also led to a loss of Cdc42 polarity in the HSCs residing in the perivascular niche. The results suggest that Fz5-mediated noncanonical Wnt signaling regulates polarity of active HSCs in the perivascular zones. Future study is required to see whether the Fz5-Cdc42 mediated polarity in HSCs is associated with symmetric vs. asymmetric division.

We propose that noncanonical Wnt signaling maintains quiescent and active HSCs reside respectively in the endosteal zone and the perivascular zone. In these zones, Fz8 and Fz5 are differentially expressed and mediate noncanonical Wnt signaling for HSC maintenance in the endosteal niche and to regulate active HSC action in the perivascular niche.