HSC migrate in several circumstances: (a) HSC are thought to migrate through the blood to seed new sites of hematopoiesis during embryonic and fetal development (for review see reference 2
); (b) HSC migrate continuously between BM and blood in normal adult animals (3
); (c) HSC migrate in large numbers from BM to blood after the treatment of animals with cytotoxic agents such as Cy and/or cytokines such as G-CSF (23
); and (d) injected HSC migrate efficiently to hematopoietic tissues after transplantation in experimental animals, or after clinical transplantation in humans (28
). Because HSC have now been shown to also have the capacity to give rise to nonhematopoietic tissues such as liver, migrating HSC may represent a source of pluripotent cells that are constantly available for the repair of damaged organs. Here, we report the first comprehensive study to address the chemotactic responsiveness of HSC to chemokines.
The role of chemokines in hematopoiesis includes guiding progenitor cells to microanatomical sites in BM or thymus for proper maturation (29
). However, the roles of chemokines in the migrations of HSC during embryonic and fetal development, the maintenance of adult HSC niches, and the physiological flux of HSC between BM and blood in adults are unclear. Here we showed that in normal adult animals, BM-derived LT-HSC and ST-HSC displayed a sharply restricted responsiveness pattern to chemokines; they only migrated appreciably in response to SDF-1α. Consistent with this, HSC express mRNA for CXCR4, the only known receptor for SDF-1α. The SDF-1α–CXCR4 interaction is important in hematopoietic development, and may have a role in engraftment of BM by HSC. Among other defects, mice lacking either SDF-1α or CXCR4 lack lymphomyeloid hematopoiesis in fetal BM (30
). The antibody blockade of CXCR4 prevented engraftment of adult SCID mouse BM by CD34+
-enriched human cord blood cells (32
Our data suggest that HSC, in contrast to other leukocyte subsets, only respond to SDF-1α. However, the possibility exists that HSC may respond to other chemokines or chemoattractants that we have not tested in this study. Although HSC express mRNA for CCR3 and CCR9 (lack of antibodies against murine chemokine receptors precluded determination of whether CCR3 or CCR9 proteins were present on the cell surface of HSC), they did not migrate in response to eotaxin or RANTES, ligands for CCR3, or in response to TECK, the ligand for CCR9. These data indicate that receptor mRNA expression alone does not adequately predict chemotactic responsiveness, as has been observed in other studies (24
). Knockout mice that do not express functional CCR3 have been prepared and no hematopoietic defects have been observed (Gerard, C., personal communication). Similarly, preliminary analyses of CCR9 knockout mice do not reveal notable defects in the development of most lymphoid compartments (Wurbel, M.A., and Malissen, B., personal communication [35
]). Lack of migration by HSC to eotaxin, RANTES, and TECK is also consistent with the absence of essential roles for CCR3 and CCR9 in hematopoiesis.
Current models of leukocyte trafficking hold that chemotactic responsiveness plays a critical role in the homing of cells to particular microenvironments and in positioning cells within these microenvironments (36
). Most leukocyte subsets express multiple chemokine and chemoattractant receptors, and migrate in response to several chemokines (37
). The possession of multiple chemokine and chemoattractant receptors has been shown to allow cells to maneuver in a stepwise fashion through spatial arrays of chemokine and chemoattractant gradients (39
). This raises the question of the significance of the extremely selective responsiveness pattern to chemokines exhibited by HSC.
BM has the challenging task of promoting the growth of several cell lineages. The presumption that various lineages have unique requirements for maturation has led to the notion that BM is partitioned functionally into specialized microenvironments, or “niches.” Although HSC niches have not been characterized, evidence of their existence comes from the observation that cloned stromal lines are heterogeneous in their ability to maintain hematopoiesis in vitro (40
). A second observation supporting the niche hypothesis is that transplanted HSC, when injected intravenously as discrete boluses, do not engraft BM of syngeneic recipient animals unless the dose of HSC is extremely large (44
), or the recipient's BM is injured by radiation/cytotoxic drugs.
Given the unique requirement of HSC niches in supporting both the maintenance of multipotency and differentiation, it is tempting to speculate that the restricted chemotactic responsiveness of HSC we observed might be important in localizing HSC within their niches, at least when they are actively participating in hematopoiesis. Possession by HSC of multiple functional chemokine receptors in a complex environment such as BM would be undesirable as it could lead to the inappropriate migration of HSC out of the HSC niche. Ma et al. (49
) recently proposed that CXCR4 is required for the retention of B lineage and granulocytic precursors in the fetal liver and BM. The possibility that CXCR4 might similarly promote the retention of HSC in their BM niches remains to be fully tested, but is supported by the observation that the administration to mice of an SDF analogue that downmodulates CXCR4 resulted in a greater than 30-fold increase in the number of circulating Thy-1.1lo
In addition to promoting the maintenance of HSC in specific microanatomical sites within BM, CXCR4 might allow efficient homing back to BM by those HSC that are released into the bloodstream. Using a cross-circulation model, we have found that HSC continuously flux between BM and blood under physiological conditions. HSC released from the BM into the circulation recolonize BM in other locations (5
). Therefore, the circulating pool of HSC must have a mechanism for the efficient rehoming to BM. The SDF-1α–CXCR4 interaction appears to play a key role in the homing of transplanted human hematopoietic progenitors to BM of NOD/SCID mice (32
), and SDF-1α promotes integrin-mediated arrest on vascular endothelium of circulating CD34+
). The SDF-1α–CXCR4 interaction may have a similar role in the rehoming to BM of murine HSC released physiologically into the circulation or injected intravenously for transplantation.
Aiuti et al. (16
) raised the possibility of a direct role for SDF-1α in hematopoietic progenitor mobilization. In a study of human BM and MPB CD34+
cells, fewer MPB CD34+
cells responded to SDF-1α than CD34+
cells isolated from BM, and it was suggested that the reduction in SDF-1α responsiveness might be part of the mechanism of progenitor mobilization. But only a fraction of CD34+
cells are HSC (17
). We found that mouse HSC from MPB, or from BM or spleens of Cy/G-CSF–treated animals, were equally responsive to SDF-1α as HSC from BM of untreated control animals. These data do not support a role for changes in SDF-1α responsiveness in cytokine-induced HSC mobilization. The disparity between the results of the current study and those of Aiuti et al. (16
) might be due to the differences between progenitors and HSC, or it may reflect species- or mobilization protocol–specific effects. In preliminary experiments, the migration of mobilized HSC to the following chemokine panel was also tested: JE, eotaxin, thymus- and activation-regulated chemokine, RANTES, MIP-1β, MIP-3α, MIP-3β, I-309, KC, IL-8, MIG, and SDF-1α. Similar to untreated BM, mobilized HSC migrated only to SDF-1α and did not respond to the other chemokines or to G-CSF (unpublished data).
In conclusion, we report that HSC migrate to SDF-1α but not to chemokines signaling through other known chemokine receptors. To our knowledge, this is the first report of a leukocyte subset that responds to a single chemokine, which makes HSC highly specialized in this regard. It remains to be determined whether SDF-1α–CXCR4 interactions play a role in HSC localization within BM, or in the rehoming to BM by HSC that are released physiologically into the circulation. SDF-1α responsiveness is preserved in HSC isolated from BM, blood, and spleens of Cy/G-CSF–mobilized mice.