A matrix micropatterning platform for cell localization and stem cell fate determination

Abstract

To study the role of cell-ECM interactions, microscale approaches provide the potential to perform high throughput assessment of ECM microenvironments on cellular function and phenotype. Using a microscale direct writing (MDW) technique, we characterized the generation of multicomponent ECM microarrays for cellular micropatterning, localization, and stem cell fate determination. ECMs and other biomolecules of various geometries and sizes were printed onto epoxide-modified glass substrates for evaluation of cell attachment by human endothelial cells. The endothelial cells displayed strong preferential attachment to the ECM-patterned regions and aligned their cytoskeleton along the direction of the micropatterns. We next generated ECM microarrays that contained one or more ECM compositions (namely gelatin, collagen IV, and fibronectin) and then cultured murine embryonic stem cell (ESCs) on the microarrays. The ESCs selectively attached to the micropatterned features and expressed markers associated with a pluripotent phenotype, such as E-cadherin and alkaline phosphatase, when maintained in growth media containing leukemia inhibitory factor. In the presence of soluble factors retinoic acid and bone morphogenetic protein-4, the ESCs differentiated towards ectodermal lineage on the ECM microarray with differential ECM effects. The ESCs cultured on gelatin showed significantly higher levels of pan cytokeratin expression, when compared cells cultured on collagen IV or fibronectin, suggesting that gelatin preferentially promotes ectodermal differentiation. In summary, our results demonstrate that MDW is a versatile approach to print ECMs of diverse geometries and compositions onto surfaces, and it is amenable to the generation of multicomponent ECM microarrays for stem cell fate determination.

Three-dimensional structural niches engineered via two-photon laser polymerization promote stem cell homing

“…The results of the present work challenge the current approach to synthetic niche engineering, which is based on building a closed container in which the cells are confined and stimulated to differentiate [3][4][5]. We introduce the new perspective of building an open 3-D microenvironment, structurally interacting with individual cells, to guide their spontaneous homing and proliferation.…”

“…Cell spreading on these surfaces may induce an artificial cell polarity perturbing stem cell differentiation [2]. A step forward in this regard is the high-throughput microwell arrays used to probe stem cell fate under the effects of combinatorial factors [3][4][5]. The microwells in these systems confine cell microaggregates, in order to maintain them well localized and physically separated for the assays.…”

Age associated communication between cells and matrix: a potential impact on stem cell-based tissue regeneration strategies

“…96 Uncertainty exists regarding age-related changes in collagen synthesis by MSCs. Sun and colleagues found no difference in total protein content of ECM laid by young or old murine BMSCs, but did report that proteins weighing approximately 140 and 40 kDa were significantly less abundant in old ECM than in young.…”

Cardiac optogenetics

“…Guided by computational predictive tools, the combined application of biochemical, spatial, mechanical, and optogenetic-mediated electrical stimuli will provide for potentially powerful tools for high spatiotemporal control and interrogation of stem cell differentiation and synchronization [20][21][22][23]. However, like traditional electrical stimulation, optical stimulation could also lead to inhomogeneous activation and inhibition; factors such as viral transduction efficiency, geometric arrangement of the cells with respect to the light source, and delivery of light through surrounding tissue and blood will need to be optimized to maximize activation and inhibition fidelity.…”