Stowers Institute for Medical Research fellow Linheng Li has discovered a mechanism by which blood cell-making stem cells maintain a reservoir of cells that can replenish these stem cells under stressful conditions. His paper greatly advances our knowledge of these stem cells.
Li and his colleagues have identified two cell surface molecules that keep mouse hematopoietic stem cells from proliferating when they are not needed. Not all adult stem cells are created equally. Some are busy repairing damaged tissues while others held in reserve to replenish those stem cells that have worn themselves out healing other tissues. Blood cell-making stem cells or hematopoietic stem cells are no different. some are held in reserve while others are actively making new blood cells. What tells some to grow and others to lay low?
Li’s group found that two cell surface proteins known as Flamingo and Frizzled8 regulate hematopoietic stem cell (HSC) proliferation. The activity of these two proteins helps maintain a reserve pool of HSCs in mouse bone marrow. In doing so, they help control the delicate balance between long-term maintenance of stem cell populations in the bone marrow, and the requirements of ongoing tissue maintenance and regeneration.
Li explains: “HSCs daily produce billions of blood cells via a strict hierarchy of lineage-specific progenitors. Identifying the molecular signals that allow HSC populations to sustain this level of output over a lifetime is fundamental to understanding the development of different cell types, the nature of tumor formation, and the aging process. My hope is that these insights will help scientists progress towards new therapies for diseases of the blood.”
Currently, work in Li’s laboratory and other labs has provided a working model that predicts that HSCs consist of a population that only divides a few times a year and sit, essentially, in reserve. These reserve cells only become activated when they need to replace other active HSCs that have been damaged by the daily war-and-tear or in response to injury or disease that greatly depletes the blood supply (Li J. Exp Hematol. 2011;39(5):511-20 & Ezoe S, et al., Cell Cycle. 2004;3(3):314-8). Missing from this working model is how the reserve and active populations of HSCs are maintained and regulated.
Both of these HSC populations exist in specialized locations within the bone and these locations or “niches” provide external cues to regulate the proliferation of the respective HSC population. Approximately 90% of all HSCs frequently divide and are found in the central marrow near blood vessels and endothelial and perivascular cells. Reserve HSCs tend to be located in the spongy part of the bone (trabecular bone) at the end of long bones. Reserve HSCs are usually very intimately associated with immature versions of bone-making cells (pre-osteoblasts). In fact, the reserve HSCs tend to be in contact with the pre-osteoblasts and this contact is extremely important from a regulatory perspective.
In Li’s lab, graduate student Ryohichi Sugimura examined Flamingo and Frizzled8 on the surface of reserve HSCs. Both of these molecules participate in a signaling pathway known as the Wnt pathway, except that Wnt signaling normally occurs through a pathway known as the canonical Wnt pathway, but Wnt signaling can also occur though so-called non-canonical Wnt signaling pathways. Canonical Wnt signaling involves a secreted glycoprotein called the Wnt, which binds to a receptor. The receptor is a member of the Frizzled gene family and Frizzled binding to Wnt initiates a signaling cascade that culminates in the accumulation of a protein called beta-catenin in the nucleus. Beta-catenin associates with members of the TCF/LEF gene family to change gene expression in the cell. Noncanonical Wnt signaling occurs through changes in Calcium ion concentrations in the cell that elicit changes in cell behavior (see Rao TP, Kühl M Circ Res. 2010;106(12):1798-806).
Experiments in culture showed that reserve HSCs not only are in contact with pre-osteoblasts, but that the Flamingo protein accumulates at the interface between the HSC and the pre-osteoblast. Furthermore, Sugimura and his colleagues showed that Flamingo regulated the distribution of Frizzled8 in the HSC membrane. Other clues were also very curious. Members of the canonical Wnt signaling pathway in reserve HSCs were quite low, but components of the noncanonical Wnt signaling pathway were expressed at high levels in reserve HSCs.
Sugimura explained the significance of these data this way: “These observations indicated that the osteoblast niche provides a microenvironmentr in which non-canonical Wnt signaling prevails over canonical Wnt signaling under normal conditions. It also suggested that Flamingo and Frizzled8 may play a direct role in maintaining the pool of quiescent HSCs.”
Surgimura and his colleagues engineered mice that lacked the Flamingo and Frizzled8 proteins and they found that these mice had very few reserve HSCs. Additionally, HSC function was greatly reduced (>70%).
Treatment of normal mice with the drug 5-fluorouracil, which destroys dividing HSCs, confirmed the role of non-canonical Wnt signaling in the maintenance of reserve HSCs. Under these conditions, noncanonical Wnt signaling components decreased and canonical Wnt signaling components increased as the reserve HSCs transitioned to frequently-dividing HSCs.
Li concluded: “A better understanding of how the balance shifts between the two will provide the necessary mechanistic insight that allows us to reduce non-canonical Wnt-signaling and dial-up canonical Wnt signaling in order to activate quiescent HSCs during aging. But the knob will have to be turned carefully. If the balance shift too far in favor of canonical Wnt signaling, it may well increase the risk of leukemia.”