The development of high-throughput screening approaches for stem cell engineering

Abstract

It has become increasingly clear that both soluble factors, such as growth factors, and insoluble factors, including the surfaces on which cells grow, can have controlling effects on stem cell behavior and differentiation. While much progress has been made in biomaterial design and application, the rational design of biomaterial cues to direct stem cell behavior and differentiation remains challenging. Recent advances in automated, high-throughput methods for synthesizing and screening combinatorial biomaterial libraries and cellular microenvironments promise to accelerate the discovery of factors that control stem cell behavior. Specific examples include miniaturized, automated, combinatorial material synthesis and extracellular matrix screening methods as well microarrayed methods for creating local microenvironments of soluble factors, such as small molecules, siRNA, and other signaling molecules.

Figure 1

A schematic diagram of high-throughput approaches in biomaterials research (The image is kindly provided by Dr. Ellenberg, and reproduced with permission from [18]).

Figure 2

Biomaterial microarray design. (a) Monomers used for microarray synthesis. (b) Monomers were mixed at a 70:30 ratio pairwise in all possible combinations with the exception of monomer 17, which was substituted with * to increase polymer hydrophilicity. To facilitate analysis, all 24 polymers composed of 70% of a particular monomer were printed as a 6 × 4 group on the array, as highlighted by the red and yellow boxes. Three blocks of 576 polymers were printed on each slide, with a center to center spacing of 740 μm. (c) Printed polymer array imaged by Arrayworx reader. Blocks composed of 70% monomer 1 and 70% monomer 6 are highlighted in red and yellow, respectively. (d) Differential interference contrast light microscopy of typical polymer spot overlaid with a few fluorescent cells (red) (reproduced with permission from [20^••]).

Figure 3

A montage of representative images of PLLA morphology from AFM data (top panels, field of view in each image is 20 μm) and corresponding cell count from fluorescent microscopy (bottom panels, field of view in each image is 1500 μm) (reproduced with permission from [35]).

Figure 4

Cell shape-dependent control of focal adhesion formation on micropatterned adhesive islands of different size coated with fibronectin. (A) Diagram of an assortment of square adhesive islands with sides ranging from 10 to 50 μm in length that were micropatterned onto substrates using microcontact printing. (B) An immunofluorescence micrograph showing staining for adsorbed fibronectin selectively limited to the square islands. (C) A differential interference contrast micrograph of bovine capillary endothelial cells cultured on different sized, square fibronectin islands. (D) Fluorescent confocal micrographs of individual, vinculin-labeled cells cultured on square islands of different sizes (lengths of sides are indicated). (E) Quantitation of total vinculin and total phosphotyrosine labeling per cell, for cells cultured on different sized squares. Over 30 cells per condition were averaged; error bars indicate standard error of the mean (The image is kindly provided by Dr. Ingber, and reproduced with permission from [43]).