3176 – MAC-1 MARKS A QUIESCENT AND FUNCTIONALLY SUPERIOR HSC SUBSET DURING REGENERATION

Abstract

Hematopoietic stem cells are capable of sustaining a lifelong production of mature blood cells as well as rapid hematopoietic regeneration upon acute stress. Mouse hematopoietic stem cells (HSCs) at steady state have been extensively defined both molecularly and functionally. When exposed to regenerative stress, however, HSCs undergo several immunophenotypical changes that limit the possibilities of isolating them at high purity. As a consequence, actively regenerating HSCs have remained less defined, and it is of great interest to identify markers that specifically label activated HSCs to gain further knowledge about their molecular and functional properties. Macrophage-1 antigen (Mac-1), a cell surface marker typically expressed by myeloid cells, has been shown to be upregulated on HSCs in response to the cytotoxic drug 5-fluorouracil (5FU). Here, we assessed the expression of Mac-1 on HSCs during regeneration following transplantation and observed a transient but distinct increase in Mac-1 expression during the early reconstitution phase 4 weeks after transplantation. Remarkably, from serial transplantation experiments, we found that the functional reconstitution potential was highly enriched in this Mac-1+ portion of the HSC pool. A similar correlation between Mac-1 expression and functional HSC activity was seen following 5-FU treatment. Moreover, in contrast to previous reports, we found that Mac-1 expression inversely correlates with cell cycling, and global transcriptome analysis showed that regenerating Mac-1+ HSCs share molecular features with stem cells with low mitotic history and low historic progeny output. Taken together, our results suggest that Mac-1 expression marks predominantly quiescent and functionally superior HSCs during early regeneration. Hematopoietic stem cells are capable of sustaining a lifelong production of mature blood cells as well as rapid hematopoietic regeneration upon acute stress. Mouse hematopoietic stem cells (HSCs) at steady state have been extensively defined both molecularly and functionally. When exposed to regenerative stress, however, HSCs undergo several immunophenotypical changes that limit the possibilities of isolating them at high purity. As a consequence, actively regenerating HSCs have remained less defined, and it is of great interest to identify markers that specifically label activated HSCs to gain further knowledge about their molecular and functional properties. Macrophage-1 antigen (Mac-1), a cell surface marker typically expressed by myeloid cells, has been shown to be upregulated on HSCs in response to the cytotoxic drug 5-fluorouracil (5FU). Here, we assessed the expression of Mac-1 on HSCs during regeneration following transplantation and observed a transient but distinct increase in Mac-1 expression during the early reconstitution phase 4 weeks after transplantation. Remarkably, from serial transplantation experiments, we found that the functional reconstitution potential was highly enriched in this Mac-1+ portion of the HSC pool. A similar correlation between Mac-1 expression and functional HSC activity was seen following 5-FU treatment. Moreover, in contrast to previous reports, we found that Mac-1 expression inversely correlates with cell cycling, and global transcriptome analysis showed that regenerating Mac-1+ HSCs share molecular features with stem cells with low mitotic history and low historic progeny output. Taken together, our results suggest that Mac-1 expression marks predominantly quiescent and functionally superior HSCs during early regeneration.