Simultaneous Sessions I: Hematopoiesis, stem cells and microenvironment
The success of clinical hematopoietic stem cell (HSC) transplants is highly dependent on the ability of HSC to home to bone marrow (BM). Although it is appreciated that hematopoietic recovery post-transplant is correlated to infused HSC number, greater understanding of homing mechanisms should allow us to identify improved transplant strategies that are not solely reliant on number but also HSC quality. Mobilized blood is the most common source of HSC for transplant and the gold standard mobilization agent, granulocyte-colony stimulating factor (G-CSF), is known to impair the homing ability of HSC. G-CSF-mobilization of HSC is known to be associated with reduced expression of key adhesion molecules and we hypothesize this to include junctional adhesion molecule A (JAM-A), a transmembrane protein enriched on long-term HSC. Currently, the specific role of JAM-A in HSC mobilization, homing and stem cell maintenance is not known.
To assess the role of junctional adhesion molecule A (JAM-A) in mobilization, homing and engraftment of HSC as well as stem cell maintenance in BM. In addition, we aim to identify a G-CSF independent mobilization strategy that does not reduce expression of key adhesion receptors, thus providing a mobilized product with enhanced homing capacity.
Anti-JAM-A antibody treatment of HSC followed by transplant into either C57BL/6 or NSG mice were used to assess the role of JAM-A in homing and engraftment of murine or human HSC, respectively. Mice were injected with anti-JAM-A blocking antibody for 3 consecutive days to determine whether prolonged inhibition of JAM-A in vivo leads to mobilization of HSC or alteration of HSC maintenance in BM. Mice were mobilized with G-CSF or a small molecule combination of the α4β1/α9β1 integrin antagonist “BOP” and the CXCR4 antagonist “AMD3100” and blood stem cells were comparatively assessed using 1) using single cell RNAseq 2) flow cytometric analysis of key adhesion molecules, including JAM-A, CXCR4, α4β1 and α9β1 and 3) homing and long-term transplantation.
JAM-A has multi-faceted roles in HSC regulation during steady state and following stem cell mobilization. Specifically, JAM-A is critical for HSC homing and engraftment post-transplant and contributes to the preferential maintenance of HSC quiescence in the endosteal niche (region closely associated with bone/BM interface). Furthermore, G-CSF dependent HSC mobilization significantly reduces JAM-A expression on HSC as a result of JAM-A cleavage by tumor necrosis factor α-converting enzyme, which is upregulated by G-CSF administration. In addition, HSC mobilized with G-CSF have significantly impaired homing potential, which is attributed to lower levels of JAM-A, CXCR4 as well as α4β1 and α9β1 integrins compared to cells mobilized using the small molecule combination of the α4β1/α9β1 integrin antagonist BOP and the CXCR4 antagonist AMD3100. Consequently, we show that a single injection of BOP plus AMD3100 for 1 hour was capable of mobilizing HSC and progenitors with significantly enhanced long-term multi-lineage engraftment potential compared to a 4-day G-CSF approach.
Our results identify JAM-A as a critical regulator of stem and progenitor trafficking to and maintenance in BM and suggests the use of G-CSF independent mobilization strategies (e.g. BOP plus AMD3100) that do not abrogate JAM-A expression or function should result in improved clinical stem cell transplants.