Endothelial MAPK Activation Disrupts Hematopoiesis By Depleting Plasma SCGF

Adult hematopoietic stem cells (HSCs) are defined by their ability to undergo self-renewal and maintain the capacity to generate all of the mature hematopoietic cell types within the blood and immune system. Maintenance of the HSC is dependent upon its cell-intrinsic properties, as well as the extrinsic cues from the bone marrow (BM) microenvironment. Within the BM microenvironment there exists a diverse array of cellular hubs that comprise the HSC niche, including vascular endothelial cells (ECs), perivascular stromal cells, osteoblasts, sympathetic nerves, and cells part of the hematopoietic hierarchy such as macrophages, granulocytes, and megakaryocytes. Our group has demonstrated that BMECs are indispensable in supporting HSC self-renewal and differentiation into lineage-committed progeny in order to maintain balanced hematopoiesis. We have built upon this finding and have recently demonstrated that suppression of vascular inflammation via EC-specific inhibition of the canonical NF-kB signaling pathway resulted in significant enhancement of steady state and regenerative hematopoiesis, in part by interfering with MAPK signaling. Furthermore, we have shown in an in vitro setting that overexpression of MAPK in ECs results in differentiation of HSCs via the upregulation of pro-differentiation paracrine factors. Based on this data, we hypothesized that MAPK overexpression within ECs promotes severe defects in the hematopoietic system and we set out to develop strategies to overcome these deficiencies and enhance hematopoietic function.

To formally test if endothelial MAPK signaling impacts the ability of the BM endothelial niche to support functional hematopoiesis, we generated a mouse model in which the MAPK signaling pathway was selectively activated in ECs (CDH5-MAPK mice) and found that CDH5-MAPK mice manifested a profound decline in their HSC and progenitor potential resulting from loss of vascular integrity and increased inflammation in BMECs. Additionally, we found that CDH5-MAPK mice resulted in an increase in inflammation and a decrease in overall frequency in other critical BM niche cells, including Lepr+, Nestin+, and osteoblast cells. Suppression of the canonical NF-kB signaling pathway in ECs of CDH5-MAPK mice led to a complete restoration of their hematopoietic deficiencies and restored the proper function, inflammatory profile, and frequency in all BM niche cells. We analyzed expression of known pro-HSC factors, such as KitL and Cxcl12, and found no differences within BMECs or other BM niche cells. However, proteomic analysis of CDH5-MAPK mice revealed the deficiency of a novel, plasma-borne pro-hematopoietic factor, Stem Cell Growth Factor-a (SCGF), which was restored following EC-specific inhibition of NF-kB signaling. SCGF was recently described as a potent osteogenic factor than can rejuvenate bone homeostasis in aged mice. Analysis of bone health in CDH5-MAPK mice revealed a significant defect in bone homeostasis, suggesting a critical need in maintaining the proper cellular cross-talk between BMECs and other BM niche constituents. Infusion of recombinant SCGF into CDH5-MAPK mice led to a complete recovery of their functional HSC and progenitor defects, restoration of their vascular integrity and bone health, and suppression of inflammation within the microenvironment. Furthermore, we demonstrated that infusion of SCGF has potent therapeutic properties that enhance hematopoietic recovery and increase the functionality of the HSC following myelosuppressive insult in both control and CDH5-MAPK mice. Utilizing our genetic models, we were able to establish a role for SCGF in supporting the instructive capacity of the endothelial niche, and their interaction with other critical components of the HSC niche, to facilitate the maintenance and expansion of HSCs and improve their functional output following hematopoietic insults. We anticipate that future studies directed towards a thorough characterization of the core endothelial signaling networks will offer novel therapeutic approaches to accelerate the regeneration of the HSC pool by promoting vascular regeneration and enhancing the pro-HSC function of BMECs. These therapies could be utilized following myeloablation as well as to expand transplantable, repopulating HSCs ex vivo, thereby ameliorating life-threatening pancytopenia associated with chemo-irradiation regimens.