The potential of stem cells in clinics and as a diagnostic tool is still largely unmet, partially due to a lack of in vitro models that efficiently mimic the in vivo stem cell microenvironment—or niche—and thus would allow reproducible propagation of stem cells or their controlled differentiation in vitro. The current methodological challenges in studying and manipulating stem cells have spurred intense development and application of microfabrication and micropatterning technologies in stem cell biology. These approaches can be readily used to dissect the complex molecular interplay of stem cells and their niche and study single-cell behavior in high-throughput. Increased merging of microfabrication with advanced biomaterials technologies may ultimately result in functional artificial niches capable of recapitulating extrinsic stem cell regulation in vitro and on a single-cell level.

Introduction

Stem cells are defined by their unique capability to self-renew and produce differentiated progeny, which makes them extremely interesting cellular sources for clinical tissue engineering and for in vitro drug discovery applications. However, the clinical and pharmaceutical application of stem cells is still hampered by a lack of tractable stem cell culture techniques. In particular, many adult stem cell types cannot easily be maintained in culture without compromising multipotentiality, and the directed differentiation of embryonic or induced pluripotent stem (iPS) cells and their assembly into functional tissues is still highly challenging, if not impossible (1).

In vivo, stem cell self-renewal and differentiation are tightly controlled by a complex niche that physically hosts the stem cells in an anatomically well-defined location within a tissue (Figure 1A). The key function of the niche is the perpetual maintenance of a pool of slowly dividing stem cells. Stem cells in the niche are surrounded by support (or niche) cells and are exposed to additional extrinsic signaling cues originating from interactions with the extracellular matrix (ECM) as well as various soluble stimuli. The spatially and temporally controlled presentation of these stimuli is assumed to instruct stem cell behavior by balancing the number of quiescent and cycling stem cells. Cell divisions can result in two daughter cells with the same or disparate fates (Figure 1B). Asymmetric self-renewal division (asymmetric with respect to the function of the two daughter cells), resulting in homeostatic conditions, could either be induced by an asymmetric distribution of cell-intrinsic, fate-determining proteins or by exposing the two equal daughter cells to different local microenvironments (2). Symmetric self-renewal divisions would result in the expansion of the stem cell pool at the population level (2,3,4).

Figure 1. Microfabricated platforms to study the interaction of stem cells with their niche. (Click to enlarge)

While classical biological methodologies—ranging from high-throughput gene expression analyses or fluorescence-activated cell sorting (FACS) to in vivo experiments—have significantly increased our understanding of the phenotypic stem cell makeup and the genetic mechanisms that control stem cell behavior, they are not ideally suited to elucidate the extrinsic mechanisms of stem cell regulation. In vivo experiments on stem cell niches are often hindered by low accessibility (e.g., niches in the bone marrow, brain, and muscle) and by the difficulties to specifically manipulate niches genetically. On the other hand, commonly used in vitro approaches lack the means to recapitulate the spatial and temporal niche signaling, and are built on materials with biophysical properties that do not mimic stem cell niches. Furthermore, many classical in vitro approaches are based on population scale cell analyses, which neglect the fact that stem cell populations are not homogeneous, while FACS analyses miss the dynamics and genealogical relationships in behaviors of large numbers of single cells.

These shortcomings have spurred the development and application of new generations of cell culture platforms building on microfabrication technologies as well as advanced biomaterials approaches. Microfabrication is a generic term describing the construction of miniaturized structures—ranging from a few to hundreds of microns—that can be fabricated via numerous techniques such as photolithography, soft lithography, and microfluidics (5,6). The use of these techniques in stem cell biology is often motivated by the low reagent consumption, high throughput, and shorter analysis times (7). These technologies promise to enable specialized applications and processes not imaginable on a larger scale.

In this review, we discuss emerging applications of such miniaturized platforms in stem cell biology. Since the rapidly growing body of literature on micron-scale systems for cell culture applications is already too large to be discussed comprehensively, we will mainly focus on those methods that have been applied in stem cell biology, as well as a few examples that have not yet been interfaced with stem cell biology but appear particularly promising in this context. We distinguish between three main types of applications: (i) high-throughput platforms to screen for many different niche factors and their combinations (Figure 1C); (ii) artificial niche models to recapitulate key aspects of natural niches in vitro, including spatially heterogeneous substrates, gradients, and 3-D microenvironments (Figure 1D); and (iii) high-throughput single-cell handling techniques to assess the heterogeneity and dynamics of stem cells and their progeny (Figure 1E). Notably, since many types of stem cells are sensitive to the biophysical characteristics of their niche, we emphasize approaches that incorporate advanced biomaterials into micron-scale platforms. We strongly believe that some of the discussed approaches will improve the validity of many in vitro stem cell experiments and will ultimately reveal novel biological insights not discoverable using conventional methodologies.