Functional heterogeneity of the bone marrow vascular niche

Abstract

Sinusoidal endothelial cells (SECs) comprise the platform where trafficking into and out of the BM occurs and where hematopoietic stem and progenitor cells (HSPC) harbor and receive cues for self-renewal, survival, and differentiation. Therefore, SECs are referred to as a bone marrow vascular niche (BMVN). Hematopoietic regeneration has been shown to occur only with concurrent angiogenic regeneration. However, there are still not sufficient means to identify and isolate SECs, therefore the "niche endothelial cell" remains incompletely characterized. VEGF-receptor-3 (VEGFR3) is expressed exclusively by the SECs, while Sca1 and Tie2 are only expressed on the VEGFR3(-) arteriolar endothelium. We previously demonstrated the importance of vascular recovery in hematopoietic regeneration from myelosuppression due to cytotoxic agents or whole-body irradiation. Therefore to establish the functional importance of SECs, the mechanisms underlying BMVN regeneration were examined utilizing a 5-fluorouracil (5-FU) myelosuppression model of vascular damage. Injection of antibodies against murine VEGFR-1 and -2 had no significant effect on hemangiogenic recovery. However, when soluble VEGFR-1, a decoy receptor for VEGF-A and PlGF, was injected after 5-FU, both angiogenic remodeling and regeneration of megakaryopoiesis were delayed. In conclusion, we show that the bone marrow vasculature comprises heterogeneous compartments. SECs are distinguished from arterioles by unique immunophenotypes. Regeneration of damaged SECs is the rate-limiting step in hematopoietic regeneration from myelosuppressive therapy. Novel, high-efficiency VEGF-binding drugs in combination with chemotherapeutic agents may lead to cases of prolonged cytopenia.

Figure 1

BM SECs are VEGFR3⁺. WT C57BL/6 mice were stained with anti–pan endothelial cell antigen (clone MECA-32) and anti-VEGFR-3 (clone AFL4). Note that SECs are VEGFR3⁺ while MECA32⁺ arterioles are VEGFR3⁻ (black arrows)

Figure 2

Blockade of VEGF signaling in BM leads to vascular disruption and regional impairment of recovery after myelosuppression. C57Bl/6 mice were treated with 250 mg/kg 5-FU followed by administration of adenoviral vectors encoding for soluble VEGFR (AdsVEGFR) or vehicle at 24 h after 5-FU. Femurs were harvested at Day 10 and then stained for H&E, VEGFR-3, or TSP-1 as described in text. (A) H&E and VEGFR-3 immunostaining of metaphyseal region in control versus. AdsVEGFR group (red and green arrows, respectively). Note reduction in vascular integrity of VEGFR3⁺ SECs in treated animals. 200×. (B) H&E and anti-TSP immunohistochemistry of diaphyseal region in treated versus control. Note reduction in thrombopoiesis in treated group as measured by TSP immunoreactivity (yellow arrows). 200×.