Our area of interest is the movement / trafficking of hematopoietic stem and progenitor cells (HSPC) from the peripheral circulation into the marrow microenvironment. This process is incompletely understood, but it is crucial for not only native hematopoiesis, but also for successful engraftment after hematopoietic cell transplantation (HCT). We hypothesize that there are factors that positively affect HSPC migration as well as factors that negatively affect migration. The negative factors have not been described or studied to date. Our overarching goal and focus of this application is to study a new protein, DPT, which may impede HSPC trafficking to the marrow. Our prior investigations into the marrow nice factors that control HSPC migration after HCT were based in a zebrafish model (and the primary goal of my prior K08 award). Using this model, we determined that a specific protein, dermatopontin (DPT), a small protein that is both attached to the extracellular matrix and is also present in the plasma, is highly expressed after radiation. Furthermore, we found that DPT and was elevated in the circulation after radiation-induced damage. Adoptive transfer experiments utilizing recombinant DPT pretreated donor cells indicated that DPT had an inhibitory effect on HSPC migration to the marrow. We hypothesize that DPT interacts with HSPC and endothelial cells and acts as a barrier to HSPC migration to the marrow after transplant. We have 2 specific aims:
Specific Aim 1 : Determine the effect of extracellular DPT on cellular trafficking after HCT.
This aim will be accomplished through in vivo and in vitro experiments designed to confirm DPT's effects on cellular movement. Several experiments are designed to test DPT overexpression in an HCT system. The second aspect involves the creation of a dpt knockout zebrafish to study HSPC homing. Finally, we will attempt to generate preliminary data on DPT and HSPC homing in a murine system.
Specific Aim 2 : Determine the mechanism of DPT inhibition on HSPC trafficking. DPT binds to the surface of endothelial cells via integrins. Using an in vitro system of HSPC transendothelial migration, we will test whether DPT reduces HSPC-endothelial attachment in vitro and if it also cooperates with other ECM proteins such as fibrinogen or collagen. We have also recently developed novel methodology to evaluate HSPC-endothelial interactions in vivo using live fluorescent microscopy and will test the ability of DPT to alter these interactions. My K08-funded research allowed me to identify novel mediators of HSPC homing. The intention of this R03 is to expand on my research by investigating, in more detail, the role of a specific protein (DPT) in cellular homing and its mechanisms of action. Data on how DPT functions could help us select compounds or drugs to test if DPT inhibition can improve homing and engraftment in humans. Thus, this R03 is both a logical extension of my current K08 work on cellular homing, as well as an important next step to set up an R01 proposal.
Hematopoietic stem cell transplant is one of the most common cellular therapies and can cure a variety of leukemia's, lymphomas, hematologic and metabolic disorders. This application seeks to better understanding the engraftment process including the initial events that involve hematopoietic-endothelial cell interactions. This new knowledge may allow us to develop target strategies to improve the transplant process by accelerating engraftment and improving transplant efficacy; this would lead to decrease morbidity and morality in our patients.
| Kramer, Ashley C; Blake, Amanda L; Taisto, Mandy E et al. (2017) Dermatopontin in Bone Marrow Extracellular Matrix Regulates Adherence but Is Dispensable for Murine Hematopoietic Cell Maintenance. Stem Cell Reports 9:770-778 |
