Skeletal Stem Cell PDGFR-beta Signaling as a New Regulator of the HSC Niche

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PROJECT SUMMARY/ABSTRACT Platelet-derived growth factor (PDGF) and its corresponding receptors (PDGFRα and PDGFRβ) regulate organogenesis and tissue homeostasis. Abnormal receptor signaling has been implicated in developmental syndromes and adult diseases such as fibrosis, cardiovascular disease, and cancer. Therefore, PDGF receptor activation and its downstream signaling have been proposed as potential therapeutic targets. Recent reports have implicated PDGFRβ signaling in regulating bone marrow (BM) microenvironment and hematopoietic stem cell (HSC) maintenance. Although skeletal stem cells (SSCs) are known to express PDGFRβ, a regulatory role of PDGFRβ in HSC niche is unknown. Our goal is to determine the roles of PDGFRβ in HSC niche maintenance and SSC function. Our central hypothesis is that PDGFRβ signaling in SSCs regulates the size of the HSC niche and blood-forming functions of the BM. With our expertise in PDGF signaling and novel genetic approaches, this project will address the central hypothesis with the two following aims: 1) Characterize the impact of SSC PDGFRβ signaling on the HSC pool maintenance and myeloproliferative neoplasia (MPN) development and 2) Define the functional consequences on SSCs/stromal lineages with altered PDGFRβ signaling. In Aim 1, tamoxifen-inducible Gli1 Cre recombinase mouse line (Gli1-CreER) will be used to conditionally induce a gain-of-function PDGFRβ knock-in allele or to inactivate the endogenous Pdgfrb gene in Gli1+ SSCs. Experiments using WT and mutants will focus on the evaluation of functional changes in BM and blood with altered PDGFRβ in SSCs. We will transplant BM from PDGFRβ mutants to irradiated wild type (WT) mice (or vice versa) to determine the extent at which blood forming cells acquire permanent functional alterations from the PDGFRβ mutant environment. Finally, we will determine whether altered PDGFRβ- regulated stromal environment alters MPN progression using BM transplantation with JAK2-V617F mutant BM cells. In Aim 2, in order to characterize cellular changes and functionality in HSC niche-supportive SSCs and stromal lineages in PDGFRβ mutants, we will use in vitro SSC assays to evaluate functional changes in SSC stemness, differentiation, and cytokine production. Finally, SSCs sorted from BM of PDGFRβ mutants or controls will be transplanted into WT kidney capsules to determine the impact of SSC PDGFRβ signaling on the modulation of BM formation and HSC colonization in vivo. The results of these projects will generate new knowledge of PDGFRβ signaling-mediated stroma-to-HSC niche crosstalk and facilitate the development of potential therapeutic targets on BM and hematopoietic diseases.