Infusion of Cultured Endothelial Progenitor Cells Boosts Hematopoietic Recovery and Increases the Expression of LepR⁺ Cells after Irradiation

The hemopoietic system's function of producing blood cells and immune cells during thorough life relies on the Maintainance and differentiation of hematopoietic stem cells (HSCs). HSCs activities were tightly and finely tuned by a specialized microenvironment called "niche" in the bone marrow. However, during the routine treatment of hematopoietic diseases, such as chemotherapy and radiotherapy, the niche was inevitably impaired which is one of the essential causes of hematopoietic suppression. Our previous research results showed that infusion of cultured endothelial progenitor cells (EPCs) could boost hematopoietic recovery after HSCT. And the latent mechanisms could be that EPCs can differentiate into endothelial cells (ECs) in vivo. However, the roles of EPCs play in the bone marrow niche and the specific mechanism remain unknown. Here we designed further to investigate the effect and mechanism of EPCs in boosting bone marrow microenvironment recovery after total body irradiation (TBI). This may offer a new cue for therapy of hematopoietic suppression after the treatment of malignant hematological diseases.

Bone marrow-derived Primary EPCs isolation and culture were described as before. Briefly, Bone marrow cells from Balb/c or C57BL/6J mice aged 6-8 weeks were cultured with EGM-2 medium which was changed in 36 h, the adherent cells formed colonies identified as elongated sprouting cells radiating from a central core of round cells and identified as VEGFR2⁺; CD133⁺; CD144⁺; SCA1⁺ by FACS on days 4-6. To assess the effects of the EPCs in boosting hematopoiesis, Balb/c mice underwent TBI with a lethal dose and were injected withEPCs or the same volume of PBS via tail vein in 4-6 hours. The survival of the EPC group was elevated ( survival: 8/9 vs 0/13; p＜0.001). C57BL/6J mice underwent TBI with a sublethal dose and were injected with the cultured EPCs isolated from C57BL/6-Tg(CAG-EGFP) mice or the same volume of PBS via tail vein in 4-6 hours. Results showed that EPCs boost the recovery of hematopoietic cells in the bone marrow and peripheral blood cells; Immunofluence (IF) staining showed that Infused EPCs with GFP fluorescence were home to bone marrow and differentiated into ECs which were marked with endomucin (EMCN); Moreover, assessments of the alteration of bone marrow niche using H&E staining revealed that infusion of EPCs reduced the accumulation of fat and using multi-analyte flow assay showed that mice infused with EPCs express a higher level of M-CSF and TGF-β in plasma while the level of SCF and IL-6 both in plasma and bone marrow supernatant were lower compared to TBI. Furthermore, to assess the change of sinusoidal niche, we used IF staining and found that the sinusoids were dilated and perisinusoidal stromal cells identified as LepR⁺ were massively lost on the 7^th day after TBI while the dilated sinusoidal lumens were narrowed (11.05±1.789 vs 20.86±4.573，p=0.0258) and LepR⁺ cells were upregulated on the 21^st day after the infusion of EPCs which indicated that the sinusoidal niche has a better recovery compared to TBI.

In Conclusion, EPCs could differentiate into endothelial cells in vivo boosting the recovery of the hematopoietic system. And the potential mechanism is that EPCs repair the bone marrow sinusoidal niche damaged by irradiation.

No relevant conflicts of interest to declare.