Abstract
Biophysical cues robustly direct cell responses and are thus important tools for in vitro and translational biomedical applications. High throughput platforms exploring substrates with varying physical properties are therefore valuable. However, currently existing platforms are limited in throughput, the biomaterials used, the capability to segregate between different cues and the assessment of dynamic responses. Here we present a multiwell array (3 × 8) made of a substrate engineered to present topography or rigidity cues welded to a bottomless plate with a 96-well format. Both the patterns on the engineered substrate and the well plate format can be easily customized, permitting systematic and efficient screening of biophysical cues. To demonstrate the broad range of possible biophysical cues examinable, we designed and tested three multiwell arrays to influence cardiomyocyte, chondrocyte and osteoblast function. Using the multiwell array, we were able to measure different cell functionalities using analytical modalities such as live microscopy, qPCR and immunofluorescence. We observed that grooves (5 μm in size) induced less variation in contractile function of cardiomyocytes. Compared to unpatterned plastic, nanopillars with 127 nm height, 100 nm diameter and 300 nm pitch enhanced matrix deposition, chondrogenic gene expression and chondrogenic maintenance. High aspect ratio pillars with an elastic shear modulus of 16 kPa mimicking the matrix found in early stages of bone development improved osteogenic gene expression compared to stiff plastic. We envisage that our bespoke multiwell array will accelerate the discovery of relevant biophysical cues through improved throughput and variety.
Highlights
Through its ability to regulate cell behavior, the cellular micro-environment plays a key role in health and disease [1,2,3,4]
A master stamp containing the patterns of interest are defined on silicon (figure 1(A)) or quartz (figure 1(B)) through standard fabrication techniques of electron beam lithography (EBL) and plasma etching
The mould inlay is normally prepared from a polymeric material to withstand high temperatures and high pressures required for high fidelity replication using injection moulding (figure 1(F)) [35]
Summary
Through its ability to regulate cell behavior, the cellular micro-environment plays a key role in health and disease [1,2,3,4]. Substrates that recapitulate substrate rigidity or surface topographical cues present in the cell environment have been shown in vitro to force cells to behave differently [12,13,14]. Even interaction of cells with artificial biophysical environments (i.e. topography or substrate rigidity not found in the natural cell niche) can powerfully change cell behavior by inducing cell signaling mechanisms through mechanotransduction [15,16,17,18]. Artificial biophysical environments have been shown to preferentially direct mesenchymal stem cell differentiation [19,20,21], alter endothelial cell functionality [22,23,24] and change in neurogenic subtype [14, 25]
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