CXCR1 Mediates Dynamic Changes in the Vascular Niche and Promotes Hematopoietic Stem and Progenitor Cell Function

The vascular niche is an important regulator of hematopoietic stem and progenitor cell (HSPC) function during development and in response to non-physiologic stress. The zebrafish caudal hematopoietic territory (CHT) is a vascular niche that serves as the primary site of hematopoiesis from 36 hours post fertilization (hpf) to 6 days post fertilization (dpf). We have recently identified CXCL8/CXCR1 signaling as a positive regulator of HSPC colonization of the zebrafish caudal hematopoietic territory (CHT) during late embryogenesis. This observation raised the question whether CXCR1 signaling might induce dynamic changes in the CHT that favor HSPC colonization. CXCR1 was expressed at high levels in endothelial cells using a kdrl(VEGFR2):CXCR1;kdrl:mCherry double transgenic line. The CHT was imaged by fluorescence confocal microscopy, reconstructed in 3 dimensions and the volume measured using digital image analysis software. Overexpression of CXCR1 within the endothelial cells of these animals increased the volume of the CHT by 28% (p=0.02). To understand how CXCR1 affects the dynamics of niche development, we globally overexpressed CXCR1 beginning at 36 hpf using a heat shock induction system and performed time lapse confocal microscopy from 52 to 72 hpf. This revealed that overexpression of CXCR1 consistently increased the CHT volume from 53 to 72 hpf compared to control (21% increase at 72 hpf, p=0.004). To understand whether CXCR1 acted directly on the vascular niche or indirectly via secreted factors or circulating cells, we created parabiotic zebrafish by fusing kdrl:mCherry embryos at 4 hpf. One half of each parabiotic animal was modified by DNA microinjection to globally overexpress CXCR1 or GFP as a control via heat shock induction at 36 and 48 hpf. The volume of the CHT was measured in each half of each parabiotic animal at 72 hpf. In control parabiotics overexpressing GFP, there was no difference in CHT volume between modified and unmodified sides of the organism. However, in parabiotics overexpressing CXCR1, the CHT of the modified side was 27% larger compared with the unmodified side (p=0.012), consistent with our previous results and suggesting that CXCR1 acts directly on the niche in this system. We then asked whether this volume change could affect HSPC engraftment. Parabiotic animals were created using Runx1:mCherry embryos that carry an HSPC-specific reporter transgene as "donors" and WT embryos as "recipients". The recipient niche was modified as before to overexpress CXCR1 or GFP as a control. At 72 hpf there was no difference in HSPC colonization of donor and recipient niches when the recipient niche expressed GFP. However, when the recipient niche expressed CXCR1, there was a significant increase in HSPC colonization of the recipient niche compared to the donor niche (11.4+/-2.4 vs 19.8+/-3.5 HSPCs per CHT, p=0.02). Taken together, these results identify a novel role for CXCL8/CXCR1 signaling in HSPC biology and they provide a new example of how innate immune signaling pathways are important for interactions between stem and progenitor cells and the niche. Administration of CXCL8 to hematopoietic stem cell transplant recipients may therefore improve HSPC engraftment and clinical outcomes in patients who are being treated for hematologic malignancies.