Customizable, engineered substrates for rapid screening of cellular cues

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 μ m 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.

Potential sealing and repair of human FM defects after trauma with peptide amphiphiles and Cx43 antisense

“…33 This could be achieved with ligands such as RGDs to improve actin cytoskeletal dynamics and mechanotransduction of cellular cues and synthesis of structural proteins such as elastin, collagen or fibronectin that will improve mechanical resilience, functionality and matrix viscosity. [34][35][36] Under the current experimental conditions, the peptide nanofibrous network is too soft and viscous to facilitate F-actin nuclear contraction and surface cell mechanics due to the limited surface stiffness of the material to support the typical purse-string contraction mechanism, as previously described. 17 In contrast, recent self-assembling membrane systems were designed to promote protein conformational changes that guide assembly to form soft biomaterials or are triggered by fibronectin to form nano sized networks.…”

Multifunctional Biomaterials: Combining Material Modification Strategies for Engineering of Cell-Contacting Surfaces

“…248 Furthermore, high throughput platforms and multiwell arrays enable rapid screening of different cellular stimuli, such as surface exposed topography or rigidity cues, simultaneously or isolated to assess their influence on cell behavior. 249 Research on biomaterials for tissue engineering is poised to grow in the future 250 , as personalized regenerative medicine approaches using stem cells are moving towards clinical applications 251 .…”

Nanotopography controls single-molecule mobility to determine overall cell fate

“…Cells were allowed to differentiate in response to the nanopatterns without biochemical inducers of osteogenesis. Polystyrene patterned with the same nanopit arrays was used to study differentiation, to ensure the isolated effects of each nanopattern without confounding paracrine effects 57 . Cells were seeded onto patterned polystyrene at 5000 cells/cm 2 and allowed to grow for either 2 days or 28 days.…”