Primer and interviews: The dynamic stem cell niche

Abstract

A stem cell niche is a microenvironment that supports self-renewal of a population of stem cells, and their production of differentiated cells. While the definition evokes images of a stem cell Shangri-La—where a serene stem cell pool nestles within a niche that shelters and sustains it—the reality is much more tumultuous. Niches are subject to an ever-changing maelstrom of environmental factors, the ravages of old age, and the sly tactics of disease. Presented here is a basic overview of the different ways in which stem cell niches respond to local and systemic environments, and their impact on stem cell behavior. The primer culminates with a discussion of the topic with stem cell and niche biologists D. Leanne Jones, Ph.D., and Tudorita Tumbar, Ph.D. Developmental Dynamics 240:737–743, 2011. © 2011 Wiley-Liss, Inc.

The stem cell niche exists to ensure there is an adequate stem cell pool at the ready to maintain homeostasis, repair, and regeneration of a tissue or organ. The niche performs these duties by maintaining self-renewal of stem cells during homeostasis, and by activating their proliferation in response to injury. Only upon their departure from the protective niche microenvironment can stem cells initiate differentiation. The mission of the niche is conserved despite the fact that its composition varies widely between species, and among different organs and tissues within a single species.

There are two classes of stem cell niches. Stromal niches are localized to discrete anatomical locations and contain specialized niche support cells that maintain long-term stem cell activity. Classic examples include somatic hub cells and cap cells in the Drosophila testis and ovary, respectively. Each of these niche cell types reside in close proximity to, and supports maintenance of, their respective germline stem cell (GSC) pool. By contrast, stem cells regulated by epithelial niches lack specialized support cells, instead often contacting basement membrane and their own mature progeny. This is the case among Drosophila intestinal stem cells (ISCs), which are localized along the basement membrane, adjacent to differentiated enterocytes that mobilize ISC activation. Similarly, mammalian skeletal muscle stem cells (satellite cells) are wedged adjacent to mature myofibers that control their proliferation, and under basal lamina that surrounds them. Within both classes, niches are often complex, with regulatory cues emanating from more than one niche cell type.

The challenge for any niche is to maintain an adequate stem cell pool, even when the host organism is under duress. Depicted here are six different “personas” that niches adopt, depending upon conditions within local and larger, systemic environments. The basis for each persona is illustrated with examples from the literature; however, the reader is referred to excellent reviews for detailed descriptions of niche-stem cell interactions in various models (Jones and Wagers, 2008; Morrison and Spradling, 2008; Voog and Jones, 2010). Whereas it is clear that all stem cell niches are dynamic, it remains to be determined whether every niche exhibits characteristics from all of the personas depicted below.

THE MULTIFACETED NICHE

Nurturer

In this defining role, the niche builds a stem cell sanctuary with cell adhesion and signaling molecules. Niches secrete soluble factors, including growth factors and cytokines, that coax stem cells to self-renew and refrain from differentiating. Cell adhesion and extracellular matrix molecules anchor stem cells to the niche, and their juxtaposition facilitates communication. For example in the Drosophila ovary, somatic niche cap cells are tightly bound to GSCs by means of adherens junctions containing DE-cadherin, and Armadillo, a β-catenin homolog. GSCs are thus positioned to receive cap cell secreted, short-range BMP ligands, Decapentaplegic (Dpp) and Glass bottom boat (Gbb), which suppress the GSC differentiation promoting gene, bag of marbles (bam). Genetic disruption of BMP signaling, DE-cadherin, or Armadillo, results in GSC loss (Song et al., 2002, 2004). In fact, niche signaling and adhesion are required for maintenance of various stem cell populations in female and male Drosophila gonad (Kawase et al., 2004; Voog et al., 2008; Leatherman and Dinardo, 2010).

The vastness and complexity of mammalian organs have made it challenging to perform experiments, such as in vivo genetic manipulation, that definitively test mechanisms regulating niche/stem cell interactions. However, a mixture of in vivo and in vitro experiments suggest that adhesion and signaling molecules are also important for mammalian stem cell maintenance. Transcriptional profiling of bulge stem cells in the hair follicle (HF) implicates the use of signal transduction pathway, cell adhesion, and other molecules for signaling and responding to their environment (Tumbar et al., 2004). In addition, co-culture experiments demonstrate that within the neural subventricular zone (SVZ) vascular niche, endothelial cells employ the Notch signaling pathway to stimulate self-renewal of neural stem cells (NSCs; Shen et al., 2004). Furthermore, when adhesion of SVZ NSC lineage cells to endothelial cells are disrupted in vivo by infusing mouse brains with integrin receptor blocking antibodies, proliferation of NSC lineage cells are altered (Shen et al., 2008). In both the SVZ and HF, multiple cell types are believed to contribute to the niche microenvironment (Riquelme et al., 2008). At present, it remains unclear how multiple niche components, and their cognizant signaling and adhesion molecules, work together to control stem cell behavior.

Collaborator

Stem cells are not merely sponges that soak up the good will of the niche. Evidence for a give-and-take relationship has been demonstrated in the mouse HF and Drosophila ovary. When HF bulge stem cells are expanded in culture and then grafted back into nude mice, they generate new HFs, including a new bulge niche (Blanpain et al., 2004). This finding suggests that these stem cells can organize their own niche, which in turn maintains the stem cell population. The intricate relationship between stem cell and niche is defined in more detail in the Drosophila ovary, where niche formation and maintenance is dependent upon stem cell expression of Delta, ligand for the Notch receptor. Upon Delta overexpression, niche cell numbers increase, and in a feedback mechanism, the expanded niche supports an enlarged GSC pool. Loss of Notch signaling, on the other hand, reduces both GSC and niche cell numbers (Ward et al., 2006; Song et al., 2007). The latter studies suggest that when lines of communication between stem cells and niche are broken, both cell types suffer.

Commander

When stem cell reinforcements are needed for tissue repair or regeneration, the niche is transformed from resting place to command center. In an effort to replace lost differentiated cells, the niche instructs stem cells to proliferate. Stem cells differentiate when they migrate from the niche, divide asymmetrically, and/or alter spindle orientation (Voog and Jones, 2010, see also “A Conversation With the Experts” below). In some instances, proliferation signals come directly from mature, injured tissue that double as an epithelial niche. For example, injured myofibers activate Notch and injured Drosophila intestinal enterocytes activate JAK-STAT induced proliferation in their stem cell cohorts (Conboy and Rando, 2002; Jiang et al., 2009). In vertebrates, proliferation signals also come from endothelial cells that line blood vessels. During liver regeneration, hepatocyte proliferation is induced by angiocrine factors, in this case Wnt2 and hepatocyte growth factor (HGF), that are expressed by liver sinusoidal endothelial cells (LSECs; Ding et al., 2010). The positioning of endothelial cells within vasculature suggests that they may be sensitive to blood-borne factors released upon injury (Spiegel et al., 2008). Strategic localization may help niches to mount an effective, rapid response.

Sentinel

Often interfaced between stem cells and the greater host environment, niches are also poised to alert stem cells to larger, systemic changes. Starvation conditions, for instance, provoke metabolic changes that demand immediate action on many fronts, including stem cell activity. When Drosophila are subject to protein starvation, levels of circulating insulin-like peptides (dILPs) drop, a change directly sensed by ovarian niche cap cells. In response, the niche coordinates a scaling back of the GSC pool by reducing Notch signaling and DE-cadherin expression, presumably in an effort to conserve resources (Hsu and Drummond-Barbosa, 2009).

In vertebrates, niches may be alerted to systemic changes through connections with vasculature (see “Commander” above), and to the nervous system. Hematopoietic stem progenitor cells (HSPCs) continuously migrate between the bone marrow and bloodstream, a mechanism for homeostasis. Partly based on the observation that the bone marrow niche microenvironment bears substantial innervation by the sympathetic nervous system, it was hypothesized that HSPCs and their niche are regulated by the nervous system (Spiegel et al., 2008). Supporting this idea, noradrenergic signals from the sympathetic nervous system mediate osteoblastic production of CXCL12/Stromal Derived Factor-1 (SDF-1), a factor that attracts HSPCs to the bone marrow niche and supports HSPC maintenance (Katayama et al., 2006; Sugiyama et al., 2006). The finding is provocative considering that the sympathetic nervous system helps to coordinate the body's response to stress. Further study of the role of extrinsic signals in regulating niche and stem cells will likely reveal intriguing connections between stress or disease states, and homeostasis, regeneration and repair dynamics.

Senior Citizen

As with most living things, the niche weakens and deteriorates with age. Over time, the niche atrophies and exhibits reduced expression of cell adhesion molecules and stem cell self-renewal factors (Conboy et al., 2003; Boyle et al., 2007; Bouab et al., 2011). Stem cells also undergo deleterious changes, including a decline in proliferation rate (Pan et al., 2007). These changes contribute to age-related declines in tissue maintenance, repair, and regeneration experienced by animals across the kingdom.

Although aging niche and stem cells undergo cell autonomous changes, paracrine factors also indirectly govern the cells' aging phenotype. As with flies fed a protein-poor diet, diminishing levels of dILPs also lead to degenerating GSC and niche activity in the aging Drosophila ovary (Hsu and Drummond-Barbosa, 2009). Interestingly, these deficiencies can be overcome by genetically activating insulin or downstream Notch signaling in the niche, suggesting that stem cells actually retain substantial activity in aging mammals. Rodent muscle in aged mice display weakened Notch signaling, resulting in diminished satellite cell activity and repair and regenerative potential. However, perfusion of in vivo aged muscle with serum from younger mice leads to recovery of Notch signaling and rejuvination of satellite cell activity (Conboy et al., 2005). Experiments like these demonstrate that systemic factors have potent control over stem cell and niche activity during aging.

Villain

Many types of cancers derive from misregulation of stem cells. Therefore, it's easy to imagine that under abnormal conditions, the niche's power over stem cell behavior could be used toward ill effect. Described here are five experimentally supported hypotheses depicting how niche-related mechanisms drive tumorigenesis and metastasis. Cancer exploits a niche's “collaborator” function in the first scenario, while the remaining four describe variations in which the “nurturer” and “commander” roles are abused: (1) In a feedback mechanism, cancer cells stimulate tumor-associated-stroma to secrete niche-like signals that signal cancer cell proliferation, thereby promoting metastasis (Karnoub et al., 2007); (2) Cancer cells hone to endogenous stem cell niches, where niche signals induce their proliferation (Sheng et al., 2009); (3) A premetastatic niche refers to a non-niche microenvironment that gains a niche-like status when migratory tumor cells take residence, and endogenous signals from the microenvironment induce proliferation (Kaplan et al., 2005); (4) An ectopic niche forms when cells abnormally express niche-like signals that can support an ectopic stem cell expansion (Song et al., 2007); and (5) A latent niche refers to existing niche-like microenvironments that can support the inappropriate proliferation, self-renewal, or survival of wayward stem, or other competent cells. Unlike in other scenarios, these cells are inappropriately nudged into the microenvironment during aberrant development or injury-induced mechanisms (McGovern et al., 2009). The sobering implication of this latter model is that it illustrates how tumorigenesis can initiate in a previously cancer-free host by genetically normal stem and niche cells. These hypotheses demonstrate how the dynamic nature of the stem cell niche makes it a malleable platform from which cancer can carve a foothold.

A CONVERSATION WITH THE EXPERTS

Featured here is an interview with Leanne Jones, Ph.D., and Tudorita Tumbar, Ph.D. (Fig. 1), who discuss current topics in stem cell/niche research and the future of the field.

Figure 1. (L) Leanne Jones, Ph.D., Assistant Professor, Salk Institute for Biological Studies. (R) Tudorita Tumbar, Assistant Professor, Department of Molecular Biology and Genetics, Cornell University.

Developmental Dynamics: What is your lab's research focus?

Leanne Jones: My lab is primarily interested in the mechanisms regulating stem cell behavior, in particular the role of the local microenvironment (or niche), and how the relationship between stem cells and their environment changes during aging and disease progression.

Drosophila is emerging as an ideal system for studying the relationship between stem cell activity, tissue homeostasis, and longevity. In addition to established protocols for measuring lifespan, adult flies have several tissues that are maintained by resident stem cells. Genetic tools that allow inducible tissue and temporal specific gene expression and the ability to generate clones of stem cells that can be analyzed in vivo provide a distinct advantage in defining the effects of aging on stem cell behavior.

In an initial study to determine how aging affects stem cell behavior in the Drosophila testis, we found that the number and activity of germline stem cells decreases. However, one of the most striking observations we made was the decrease in the expression of a key self-renewal factor from adjacent niche support cells. So, one project in the lab is to investigate the mechanism by which this age-related decrease in self-renewal factors occurs. Behavior of stem cells in the Drosophila intestine also change with age and in response to chronic stress; therefore, we're also comparing and contrasting how somatic and germline stem cells respond to different types of environmental changes.

Tudorita Tumbar: Our research aims at understanding the molecular mechanisms that govern cell fate decisions in tissue stem cells. We use the mouse skin and hair follicle as a model system, and focus our experiments in vivo, trying to capture stem cell behavior in an unperturbed niche, in the absence of cell transplantation. While I was a postdoctoral fellow in Dr. Elaine Fuchs' laboratory, I invented a new system to target and specifically label cells with infrequent divisions based on histone H2B-GFP retention. Using this system we have characterized a putative population of hair follicle stem cells. Later on, in my laboratory at Cornell University, we were able to demonstrate in vivo at the single cell level that stem cells in the hair follicle are infrequently dividing and they behave as a population in choosing their fate.

Our working model is that some stem cells choose to differentiate without self-renewal, while other cells replenish the pool by creating more stem cells. We are currently asking how transcription factors that regulate self-renewal and differentiation in hair follicle stem cells interplay with epigenetic modifications to control tissue stem cell quiescence and cell fate decisions. Whenever possible, we try to link our findings in the mice with human disease. For example, we found that Runx1, a transcription factor essential for timely hair follicle stem cell activation from quiescence plays a role in skin squamous cell carcinoma.

Dev Dyn: What initially provoked your interest in this field?

LJ: My background is in cancer biology, so I'm fascinated by how a cell “decides” to keep proliferating or to initiate a differentiation program. Each time a stem cell divides, the daughter cells make the decision to self-renew (proliferate) or initiate differentiation; therefore, stem cells seemed to be the perfect system in which to study this question.

TT: There were two aspects that attracted me to stem cell research: first was the promise for therapy of currently un-curable diseases; second was the promise of a rich basic science field with potential for new cell and developmental biology mechanisms that would be responsible for determining unique stem cell properties and behavior.

Dev Dyn: Which papers have most impacted your research?

LJ: (1) This paper (Conboy et al., 2003) begins to tease apart the role of the systemic/local environments in contributing to the aging of tissues maintained by stem cells.

(2) Leatherman and Dinardo (2008) reinterprets data that I published as a postdoc. The results from our lab and the Matunis lab in 2001 indicated that JAK-STAT signaling acted autonomously in germ cells to regulate germline stem cell (GSC) proliferation. However, this study demonstrated that signaling from the somatic cells was sufficient to specify GSC proliferation in a nonautonomous manner. This study reminds me that (1) even when you think you know what is happening, you probably don't understand everything and (2) you should always be challenging the current model as you discover new genes and develop new tools.

(3) These back-to-back articles (Micchelli and Perrimon, 2006; Ohlstein and Spradling, 2006) first report of intestinal stem cells in flies. These papers have allowed my lab to take what we are doing in the germ line and directly compare and contrast those findings with another stem cell system. Because the two tissues are quite different, it allows us to explore both germline specific and stem cell generalizable phenomena.

(4) Two papers from the Kenyon lab (Hsin and Kenyon, 1999; Arantes-Oliveira et al., 2002) reveal that the germ line, specifically the proliferating germline stem cells, are a “pro-aging” tissue in worms whereas the somatic component of the gonad promotes longevity. How the germ line and/or reproduction influences aging and longevity is a fascinating question to me.

TT: (1) Thomson et al. (1998) was crucial for my career path. I read it at a time when I was completing my graduate studies in large-scale chromatin and looking for a postdoctoral position. I was impressed by the possibilities that this study opened up and by the stem cell field in general. I was irreversibly fascinated with cell fate acquisition and the complexity of cell intrinsic and cell extrinsic pathways that magically combine to create three-dimensional tissues.

(2) Work from Lavker's lab gave me the theoretical and experimental basis needed to design my own experimental system that allowed me, and later on other people in different areas of stem cell biology, to isolate and characterize a most quiescent population of tissue stem cells.

(3) This paper (Robinett et al., 1996) was published from my graduate study laboratory on work that was done before my joining this research team. Although it did not directly impact my current research, it did shape my thinking and approach in the most profound manner. This study inspired me to think out of the box and dare to try something different. I came up with a novel H2B-GFP pulse-chase system to mark cells in mouse tissues in vivo based on proliferation history and to quantify their divisions.

Dev Dyn: How has the study of stem cell niches in invertebrates informed vertebrate stem cell research? What are some important differences?

LJ: Stem cells are rare, and for a long time, specific markers and tools were lacking to identify stem cells in mammalian systems. In worms and flies, stem cells could be easily identified based on lineage-tracing experiments and specific markers, allowing them to be studied in vivo in the context of the tissues they sustain. Therefore, scientists using these systems were able to identify signaling pathways and niche components to establish paradigms for how stem cell behavior could be regulated that provided a platform for the scientists working in more complex stem cell systems.

An important difference stems from recent data indicating that one of the primary factors influencing stem cell behavior in mammalian systems is the vasculature (especially hematopoietic system, neural stem cells, and spermatogonial stem cells). Flies don't have blood or blood vessels, so this is one major difference. However, the tissues that serve similar functions, hemolymph and the trachea, respectively, could serve important stem cell support roles in flies.

TT: It is very interesting to see how work in invertebrates translates to vertebrates. Model systems such as Drosophila neuroblast and germ line, or C. elegans germ line have been very useful. Based on evidence at hand, it appears that the mouse nervous system and the muscle stem cells seem to follow a simple model of asymmetric cell fate decisions, similar with that proposed in Drosophila. However, recent work from my group (Zhang et al., 2009) and other groups (Clayton et al., 2007; Snippert et al., 2010) suggests that the long-lived progenitors (stem cells) in some tissues do not self-renew and differentiate by the well-accepted asymmetric cell fate decisions model. Instead the behavior of these stem cells can be described by a population deterministic model similar with that proposed by Judith Kimble for the C. elegans germ line, in which symmetric or unidirectional decisions might be the norm.

It seems important to keep an open mind and make as few assumptions as one can when starting to use a new system that has not been characterized in the enormous detail of the Drosophila. Major differences that stem cells face when in a mouse versus a fly is the life span, and the size of the tissues that they need to maintain. Although the evidence we have is slim, my impression is that in hair follicles, the short-lived progenitors located in the matrix might behave similarly with the stem cells of Drosophila. They would self-renew and differentiate continuously by asymmetric divisions, as long as they last, which is a couple of weeks, a life span closely approaching that of a fly. In contrast the hair follicle stem cells seems to act as a reservoir of senior (long-lived) citizens from which periodically some of them are killed to differentiate and maintain the tissue while the others compensate for the loss by symmetric expansion. This periodicity becomes obscured in rapidly regenerative tissues such as epidermis and intestine, where the stem cell behavior could be modeled as a stochastic and continuous process of stem cell loss and proliferation. An important aspect of this matter is that different vertebrate tissues and stem cells might behave very differently, a concept that is not yet engrained in the field as it is difficult to resist the tendency to generalize models that we often take for granted.

Dev Dyn: It has been shown that the niche and stem cells respond to changes in the larger systemic environment caused, for example, by aging or diet. What is your view of the role of the niche in interfacing with systemic cues?

TT: These are very exciting news, and it will be interesting to see how many tissues show similar responses. The data will allow us to better dissect what are stem cells intrinsic or specific characteristics and what is due to the environment. Right now the balance seems to shift toward a determinant role of the environment, although I predict that this will change in the future as we might become better at uncovering unique stem cell intrinsic properties. In that respect, the embryonic stem cells lead the way, and we have seen that these cells have special cell cycle properties and genome organization at the level of chromatin structure and epigenetic modifications. We only have a dim glimpse of these processes in tissue stem cells, mainly because it is much more difficult to do biochemistry with small populations of sorted tissue stem cells. It will also be interesting to see what these results really mean in the context of unperturbed tissues in higher organisms, when shifting the assays from in vitro and injury models such as transplantation, to a more natural setting. The in vivo data we have from Drosophila seem to be promising.

LJ: Ultimately, stem cell behavior is regulated by the integration of extrinsic cues with intrinsic factors. We are learning more and more about how changes in the environment lead to changes in stem cell behavior; however, our work has clearly demonstrated that simply manipulating the environment is not always sufficient to reverse or override intrinsic changes. Therefore, the identification and understanding of intrinsic, molecular responses to extrinsic changes, such as those that occur during aging or disease progression, will ultimately allow us to take the next step to develop strategies to manipulate stem cell behavior to counter negative or enhance positive influences from the environment.

Dev Dyn: Would you predict that systemic cues only impact niche/stem cell dynamics under extreme conditions, or do you expect that they are in constant flux throughout the lifetime of an animal?

LJ: Tissue homeostasis requires a precise balance between stem and progenitor cells, and the local microenvironment plays a major role in ensuring this balance. Therefore, there is constant communication between niche support cells and stem cells. With regards to how systemic signals could impact this communication, several labs have studied how signaling by means of the insulin/IGF pathway regulates stem cell behavior in the gonads and the intestine. Daniela Drummond-Barbosa's group, who work in the female germ line, has done some of the most elegant work in this area. As the levels of circulating Drosophila insulin-like peptides (dILPs) clearly fluctuate in response to nutritional conditions and with age, I would predict that systemic cues are constantly influencing local stem cell-niche cell interactions.

TT: The recent data from my laboratory and from other laboratories suggest that stem cells in mouse tissues such as hair follicle, epidermis, intestine, and testis, maintain their pool by a population deterministic model. In these models some stem cells are lost by differentiation or cell death while others that remain in the niche seem to subsequently compensate the loss by what appears to be a symmetric expansion. In light of these and other data, the field is now reconsidering asymmetric cell fate decisions as an inherent property of tissue stem cells during normal homeostasis. The stem cell loss could be modeled as a stochastic process in rapidly regenerative tissues with a constant need for cells. In the hair follicle, we showed that stem cell loss is tightly coupled with the initiation of a regenerative growth cycle, and is likely dependent upon both the localization of cells within their niche, and localization near the differentiating signals. I do think the flux of stem cells is continuous, or at least periodic, and that it is programmed into the normal tissue maintenance capacity.

Dev Dyn: Tudorita, the bulge niche is somewhat ill-defined. What do you think contributes to the bulge niche microenvironment?

TT: Clustering stem cells themselves seem to secrete molecules known to control cell proliferation, survival, and differentiation. It seems conceivable that these molecules create a protective “cloud” around the bulge, thus contributing to the “niche.” It is possible, although unlikely based on the evidence at hand, that some of these quiescent bulge cells are dedicated niche cells, or that some other cells in this rich bulge environment perform this function. It is also possible, and in my mind most likely, that there is a combination of several cell populations, including dermal cells, clustering of bulge cells themselves, blood vessels and nerves, which are rich in this zone, that all contribute to the creation of a specialized stem cell environment. This makes a challenging problem for tissue reconstruction in vitro, and speaks of the complexity of three-dimensional vertebrate tissues and their intimate connection with their environment.

Dev Dyn: Are there new techniques that you anticipate will help define niches and their properties?

TT: We need to identify the native niche components in vivo, in the unperturbed tissues, and be able to generate and characterize large amounts of mutants in higher organisms. What we would really benefit from is new imaging methods capable of generating large amounts (high throughput) of data characterizing and identifying different cell types and their arrangements within unprocessed tissues. Fluorescent immunolabeling techniques, the current workhorse of quantitative imaging technology, require extensive labeling steps coupled with a need to examine a multitude of markers that make meaningful high-throughput screening of phenotypes in complex vertebrate tissues a nearly impossible task. Especially promising is quantitative microscopy taking advantage of the natural interactions of transparent samples with light, mainly phase and polarization. To take advantage of these data we will need systematic and automated methods for storing, categorizing (understanding), and accessing it. The quantitative imaging data should generate feature-based searchable databases usable in direct quantitative comparisons across experiments performed by different investigators and in different laboratories. We need to be able to do “big science” by building synergies among smaller scale projects. Together, the aforementioned technologies will allow us to rapidly sift through vast amounts of data to identify subtle cell and tissue organization phenotypes, which would otherwise go unnoticed, although in fact they might be the most crucial for understanding stem cell and niche behavior.

LJ: I completely agree that being able to visualize stem cell activity in real time in vivo is an essential tool that will provide us with invaluable information about how stem cells behave during normal tissue homeostasis in wild type animals, in mutants, or in models of human disease. This information will allow us to build models to predict how transplanted stem cells may respond to different environments in the course of regenerative medicine. In addition, this information will help bioengineers generate niches, or even tissues, in vitro that can be used to maintain and/or expand stem cells for therapeutic uses.

Dev Dyn: What are some exciting ideas that are emerging in the field? What important questions remain to be answered?

TT: An important question is whether fate switches from stem to progenitor cells are reversible in normal homeostasis of adult vertebrate tissues. We have seen that just a couple of the right transcription factors can reverse the fate of a differentiated cell to embryonic stem cells. Does de-differentiation happen naturally during normal homeostasis and what is the extent of this phenomenon? When stem cell daughters undergo symmetric fate decisions to become stem cells and compensate for stem cell loss, is that a result of a symmetric or asymmetric stem cell division? The latter would be coupled with the ability of the more differentiated stem cell daughter to reverse back to the stem cell state when needed. Because divisions are rare within vertebrate stem cell pools, this has been difficult to monitor directly. In the Drosophila germ line there is evidence that such reversibility exists, although the reverted cells were deemed less potent and their accumulation in the stem cell pool was posed as the cause of tissue aging. It will be interesting to see how this model applies to vertebrates and how is it reconciled with the notion that stem cell pools can be rejuvenated by systemic cues.

LJ: I also look forward to seeing how universal “de-differentiation” within tissues will be as a mechanism to maintain stem cells, either as a basic mechanism underlying tissue homeostasis or in response to tissue damage.

In addition, I think that the role of local O₂ levels, including ROS, in regulating stem cell behavior is quite interesting. This observation is tied to the fact that several stem cells reside within relatively hypoxic niches and that the vasculature is proving to be an integral component of the local niche in many systems.

One of the many fundamental questions that we are trying to tackle is how aging affects stem cells and the niche. Many patients that we hope to be able to treat with regenerative medicine and stem cell-based therapies will be older individuals. Data from our lab and others indicate that the niche also ages suggests it will likely be insufficient to supply these patients with “young, healthy” stem cells. Therefore, we must understand how the niche ages and consider ways to rejuvenate the niche or co-transplant new niche cells along with the stem cells. Furthermore, if we hope to recapitulate aging-related disease models in vitro, in the course of reprogramming patient fibroblasts, it will be essential to understand how the affected cell types age in normal individuals.

Acknowledgements

Many thanks to D. Leanne Jones and Tudorita Tumbar for generously sharing their expertise, insights, and time. The author regrets that due to space constraints she was unable to reference many excellent papers.