Hematopoietic stem cell regeneration through paracrine regulation of the Wnt5a/Prox1 signaling axis
Qiqi Lin, Limei Wu, Srinivas Chatla, Fabliha A. Chowdhury, Neha Atale, Jonathan Joseph, Wei Du
Qiqi Lin, Limei Wu, Srinivas Chatla, Fabliha A. Chowdhury, Neha Atale, Jonathan Joseph, Wei Du
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Research Article Cell biology Hematology

Hematopoietic stem cell regeneration through paracrine regulation of the Wnt5a/Prox1 signaling axis

Abstract

The crosstalk between the BM microenvironment (niche) and hematopoietic stem cells (HSCs) is critical for HSC regeneration. Here, we show that in mice, deletion of the Fanconi anemia (FA) genes Fanca and Fancc dampened HSC regeneration through direct effects on HSCs and indirect effects on BM niche cells. FA HSCs showed persistent upregulation of the Wnt target Prox1 in response to total body irradiation (TBI). Accordingly, lineage-specific deletion of Prox1 improved long-term repopulation of the irradiated FA HSCs. Forced expression of Prox1 in WT HSCs mimicked the defective repopulation phenotype of FA HSCs. WT mice but not FA mice showed significant induction by TBI of BM stromal Wnt5a protein. Mechanistically, FA proteins regulated stromal Wnt5a expression, possibly through modulating the Wnt5a transcription activator Pax2. Wnt5a treatment of irradiated FA mice enhanced HSC regeneration. Conversely, Wnt5a neutralization inhibited HSC regeneration after TBI. Wnt5a secreted by LepR+CXCL12+ BM stromal cells inhibited β-catenin accumulation, thereby repressing Prox1 transcription in irradiated HSCs. The detrimental effect of deregulated Wnt5a/Prox1 signaling on HSC regeneration was also observed in patients with FA and aged mice. Irradiation induced upregulation of Prox1 in the HSCs of aged mice, and deletion of Prox1 in aged HSCs improved HSC regeneration. Treatment of aged mice with Wnt5a enhanced hematopoietic repopulation. Collectively, these findings identified the paracrine Wnt5a/Prox1 signaling axis as a regulator of HSC regeneration under conditions of injury and aging.

Authors

Qiqi Lin, Limei Wu, Srinivas Chatla, Fabliha A. Chowdhury, Neha Atale, Jonathan Joseph, Wei Du

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Figure 5

LepR+ niche cell–derived Wnt5a inhibits β-catenin accumulation and represses Prox1 expression in irradiated HSPCs.

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LepR+ niche cell–derived Wnt5a inhibits β-catenin accumulation and repre...
(A) Wnt5a neutralization increases β-catenin accumulation in cocultured HSPCs. LSK cells and LepR+ cells were subjected to 300 cGy irradiation and cocultured with anti-Wnt5a (2 μg/mL) or control IgG for 5 days. Levels of β-catenin in suspension cells were determined. IgG: n = 6; α-Wnt5a: n = 8. (B) Wnt5a treatment reduces β-catenin accumulation in cocultured HSPCs. WT LSK cells and LepR+ cells were subjected to 300 cGy irradiation and cocultured with rWnt5a (100 ng/mL) or vehicle (saline) for 5 days. IgG: n = 6; α-Wnt5a: n = 8. (C) Deletion of Ctnnb1 abrogates the effect of Wnt5a neutralization on Prox1 expression in cocultured HSPCs. LSK cells from Ctnnb1fl/fl or Ctnnb1fl/flVav1-Cre mice were cocultured with WT LepR+ cells and anti-Wnt5a (2 μg/mL) or control IgG for 5 days after 300 cGy irradiation. Prox1 expression was determined. Ctnnb1fl/flVav1-Cre, IgG: n = 9; others: n = 6. (D) Deletion of Ctnnb1 abolishes the effect of rWnt5a. LSK cells were cocultured with LepR+ cells in the presence of rWnt5a (100 ng/mL) or vehicle for 5 days after 300 cGy irradiation. Ctnnb1fl/fl, vehicle: n = 8; Ctnnb1fl/fl, rWnt5a: n = 9; Ctnnb1fl/flVav1-Cre, vehicle: n = 6; Ctnnb1fl/flVav1-Cre, rWnt5a: n = 7. (E) Ctnbb1 deletion abrogates the dampening effect of Wnt5a neutralization to 1000 progeny cells from cocultures were transplanted into BoyJ recipients. Ctnnb1fl/flVav1-Cre, α-Wnt5a: n = 12; others, n = 10. (F) Ctnbb1 deletion abolishes the promoting effect of rWnt5a to 1000 progeny cells from cocultures of were transplanted into BoyJ recipients. Ctnnb1fl/fl, vehicle; Ctnnb1fl/flVav1-Cre, rWnt5a: n = 10; Ctnnb1fl/fl, rWnt5a; Ctnnb1fl/flVav1-Cre, vehicle: n = 12. Statistics were performed in the indicated groups: 2-tailed, paired t test (parametric). *P < 0.05; **P < 0.01.