Two anatomical niches for hematopoietic stem cells (HSCs) have been reported in the bone marrow, but a distinct function for each of these niches has remained unclear. A new role in stem cell proliferation has now been identified for the adhesion molecule E-selectin expressed by bone marrow endothelial cells at the vascular niche (pages 1651–1657).
A prerequisite for HSC function is the ability to travel through the bloodstream and home to the bone marrow. These circulating HSCs develop low-affinity contacts with the cell adhesion molecules E- and P-selectin on the endothelium in a process called rolling that precedes firm adhesion via ICAM-1 and VECAM-1 (ref. 1). Upon egress from the vasculature, HSCs reside in, or adjacent to, specialized microenvironments in the bone marrow2,3. The endothelial or perivascular niche is adjacent to bone marrow endothelial sinuses, and its perivascular stromal cells produce the HSC chemoattractant CXCL12, whereas the endothelium provides the HSC survival and proliferation-inducing factor c-Kit ligand (also known as stem cell factor)2,3 (Fig. 1). The majority of HSCs are quiescent for prolonged periods, and only a minority undergoes asymmetric division to produce one HSC and one non–self-renewing, transit-amplifying progenitor cell4.
In this issue of Nature Medicine, Winkler et al.5 describe a new role for E-selectin expressed exclusively by endothelial cells of the vascular niche, showing that this molecule is a key regulator of HSC proliferation. Mice deficient in either the E- or P-selectin gene (Sele−/− or Selp−/− mice) show mild phenotypes, but mice lacking both selectins have extreme leukocytosis, marked extramedullary hematopoiesis and a lack of leukocyte rolling and die of opportunistic bacterial infections due to low leukocyte extravasation at sites of inflammation6. Winkler et al.5 have now demonstrated increased HSC quiescence and self-renewal potential in Sele−/− mice and in wild-type mice given an E-selectin antagonist, GMI-1070.
The authors determined HSC proliferation by measuring BrdU incorporation into DNA during S phase. In wild-type mice, BrdU labeled 50% of HSCs in 3.3 d, whereas it took 9.5 d to label HSCs to the same extent in Sele−/− mice. BrdU has some HSC toxicity that could trigger their proliferation, leading to an underestimate of the number of quiescent cells4. Nevertheless, Winkler et al.5 found that this HSC proliferation was suppressed in the absence of E-selectin. The authors then used three assays to determine HSC cell cycle status, confirming that loss of E-selectin increases HSC quiescence. Using a long-term competitive repopulation assay, they showed that sixfold more HSCs survived treatment with hydroxyurea, an S-phase–specific cytotoxic drug, in Sele−/− compared to wild-type mice. This slower cell cycling and higher frequency of quiescent HSCs when E-selectin is absent was unexpected, because, at least in the periphery, P- and E-selectins are largely redundant in function1. The authors detected E-selectin only in CD31+ endothelial cells, and the expression of E-selectin was 16-fold higher in the endothelium of vasculature near the interface with the endosteal region compared to the central sinusoidal vasculature (Fig. 1). There was a marked increase in E-selectin–expressing endothelial cells in this endosteal region during recovery from irradiation. Despite differences in HSC proliferation between E-selectin–deficient and wild-type mice, there was no difference in their total femoral HSC numbers. Using reciprocal HSC transplantation, Winkler et al.5 showed that the proliferative effect was not cell autonomous but was due to E-selectin binding to receptors on HSCs.
High-dose or repeated rounds of chemotherapy or irradiation can eventually exhaust the self-renewal capacity of the HSC pool, leading to prolonged bone marrow suppression and life-threatening neutropenia in people being treated for cancer. Therefore, Winkler et al.5 investigated the role of E selectin in protection from chemotherapy- or irradiation-induced HSC damage. In Sele−/− mice, more HSCs survived after a single injection of the antimetabolite 5-flurouracil compared to in wild-type mice. With consecutive rounds of cytotoxic treatment, the Sele−/− mice were clearly more resistant and survived a median of 82 d longer than wild-type mice. Similarly, the Sele−/− mice showed greater HSC survival after treatment with the alkylating agent cyclophosphamide than wild-type mice. The Sele−/− mice also showed accelerated recovery of peripheral leukocytes after nonlethal irradiation compared with wild-type mice. These studies support the authors' conclusion that E-selectin has an unexpected additional role as a crucial component of the endothelium of the vascular niche that induces HSC proliferation, self-renewal and chemo- and radiosensitivity.
The mechanism behind E-cadherin–mediated regulation of HSC proliferation remains obscure. Further investigation into the E-selectin ligands expressed on HSCs and their function upon E-selectin binding are required. Several E-selectin ligands have been identified on hematopoietic cells and on leukemia cells and a variety of solid tumor types1. Although diverse proteins seem to function as E-selectin ligands, the core species of these ligands are decorated with sialofucosylated carbohydrates such as sialyl Lewis X and sialidated glycosphingolipids (which have recently been recognized as E-selectin ligands)1. Winkler et al.5 found that treatment of mouse HSCs with 1-phenyl-2-palmitoyl-3-morpholino-1-propanol (PPMP), a potent inhibitor of the glucosylceramide synthetase involved in the synthesis of glycosphingolipids, reduced their E-selectin binding.
The authors excluded most of the canonical E-selectin ligands in their search for one that was expressed in the endothelial niche and that specifically suppressed HSC proliferation. In an attempt to characterize noncanonical E-selectin receptors, they undertook 'pull-down' proteomic studies with E-selectin–coated magnetic beads but could not detect any of the candidate E-selectin ligands CD43, CD65, CD147 or death receptor 3 (also known as TNFRSF25) in these pull-downs. However, they did identify the canonical E-selectin ligand 1 (ESL-1), a part of the E-selectin receptor-ligand signaling complex1,7, in these assays. The authors do not propose a mechanism to account for the HSC proliferation that may result from binding of ESL-1 to E-selectin. It will be of interest to investigate this further in light of a study showing that ESL-1 regulates transforming growth factor-β (TGF-β)8. TGF-β is one of the most potent inhibitors of HSC proliferation, and neutralization of TGF-β releases HSCs from quiescence (reviewed in refs. 9,10). TGF-β downmodulates receptors for a number of cytokines known to stimulate HSC proliferation10. It is synthesized as an inactive precursor (proTGF-β) that undergoes a cleavage and maturation process in the Golgi apparatus in a process involving latent TGF-β–binding protein 3 (LTBP3)8. Gain of ESL-1 in the Golgi region, or knockdown of LTBP3, decreases TGF-β signaling8. The mechanism involves ESL-1 binding to proTGF-β, resulting in inhibition of furin-mediated maturation of proTGF-βl, which leads to decreased extracellular amounts of bioactive TGF-β8. Esl1−/− mice show increased mature TGF-β, increased intraosseous TGF-β signaling and decreased proliferation of chondrocytes with development of chondroplasia8. The link between ESL-1 and TGF-β may explain the authors' observation on E-selectin stimulation of HSC proliferation5, and additional studies in this area using the Esl1−/− mouse model should be informative in this respect.
The authors suggest that the aberrant glycosylation and expression of de novo E-selectin ligands is characteristic of many tumor cells and could aid their homing and engraftment into the bone marrow. Their findings imply that interaction of tumor cells with endothelium expressing E-selectin might stimulate tumor growth and proliferation, as metastasis frequently involves endothelial E-selectin binding to tumor E-selectin ligands, which, in turn, may induce tumor cell proliferation. They also point out that chemotherapy-induced myelosuppression remains a clinical problem and that reducing this dose-limiting side effect would translate to less infection, less need for support with blood products and less need for hospitalization of patients treated for cancer with chemotherapy. The use of an E-selectin antagonist such as GMI-1070 might also be of therapeutic value in preventing metastasis from primary breast or prostate tumor sites to the blood marrow and would inhibit tumor cell and HSC proliferation. This would protect HSCs from damage by cell-cycle–specific chemotherapy. However, on the negative side, such a strategy could equally promote tumor stem cell dormancy and protection from chemotherapy that targets cycling tumor cells, a possibility that will need to be investigated.
Zarbock, A., Ley, K., McEver, R.P. & Hidalgo, A. Blood 118, 6743–6751 (2011).
Ding, L., Saunders, T.L., Enikolopov, G. & Morrison, S.J. Nature 481, 457–462 (2012).
Méndez-Ferrer, S. et al. Nature 466, 829–834 (2010).
Wilson, A. et al. Cell 135, 1118–1129 (2008).
Winkler, I.G. et al. Nat. Med. 18, 1651–1657 (2012).
Frenette, P.S., Mayadas, T.N., Rayburn, H., Hynes, R.O. & Wagner, D.D. Cell 84, 563–574 (1996).
Miyaoka, Y. et al. Biochem. J. 440, 33–41 (2011).
Yang, T. et al. J. Clin. Invest. 120, 2474–2485 (2010).
Blank, U. & Karlsson, S. Leukemia 25, 1379–1388 (2011).
Fortunel, N.O., Hatzfeld, A. & Hatzfeld, J.A. Blood 96, 2022–2036 (2000).
Declaration: M.A.S.M. holds patents on granulocyte colony–stimulating factor [AU: with which patent office?] and receives royalties from Amgen.
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