Characterization of the effects of BTK inhibition and monocyte-produced IL-8 on the hematopoietic stem cell niche

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The Ohio State University

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Hematopoietic stem and progenitor cells (HSPCs) are multipotent stem cells that give rise to all blood cell lineages. During early zebrafish development, HSPCs interact closely with endothelial cells in an endothelial niche known as the Caudal Hematopoietic Tissue (CHT). How these interactions influence HSPC fate and clonality is not well-understood, however. The chemokine Interleukin-8 (IL-8) may alter HSPC fate by extending the time that HSPCs reside within the CHT. IL-8 is produced by a number of cell types, including monocytes. Using a zebrafish model, we aim to characterize the impact of enforced expression of IL-8 by macrophages on HSPC interactions with the endothelial niche. HSPCs colonize the CHT beginning approximately 32 hours post fertilization before migrating to the kidney marrow at about 6 days post fertilization where they reside for the life of the zebrafish. While in the CHT, HSPCs can be directly imaged by fluorescence microscopy using a fluorescent reporter driven by the HSPC-specific runx1 + 23 enhancer element (runx1+23:GFP). We have generated a transgenic zebrafish line, which ectopically expressesIL-8 in macrophages using the macrophage-specific promoter mpeg1.1 (mpeg1.1:cxcl8 2A mCherry). We hypothesize that the F1 offspring of these lines will show that IL-8 expression by macrophages will increase the residency time of HSPCs in the CHT compared to a non-expressing control group. We are also treating developing fish with Ibrutinib, a therapeutic protein kinase inhibitor that inhibits the IL-8 PI3K/Akt signaling pathway. We hypothesize that HSPCs in treated fish will be fewer in number and will reside in the endothelial niche for less time than non-treated controls. Understanding these interactions will shed light on alterations of HSPC cell fate due to IL-8 expression and provide potential therapeutic targets for patients with hematopoietic disorders



stem cell niche, stem cell fate, genetics, zebrafish, hematology