Poster O18 - Fabricated micropillar constructs for inferring forces generated from annular tissue explants

Introduction: Understanding the mechanical forces generated by cells and tissues of complex 3D geometries on their substrates and environment is important for investigating many biological processes such as tissue morphogenesis, wound healing, and collective cell migration. Current methods for quantifying forces generated by tissues on their environment such as Traction Force Microscopy (TFM), support mainly planar tissue geometries. These methods are insufficient for investigating the contractile forces generated by complex three-dimensional tissues and tissue explants because they measure interactions of tissues resting on a substrate and are unable to measure forces generated by cells and tissues resting above the substrate. We have developed a novel method to infer the forces generated by contractile toroidal tissue explants through the fabrication of a novel micropillar construct.

Materials and

Methods: We designed a novel micropillar construct to quantify the forces generated by toroidal tissue geometries. Each pillar is 900µm in height, with a semicircular cross-section 50µm in radius. Each construct contains two pillars separated by 375µm. The final pillar dimensions are small enough to be threaded through toroidal shaped explant tissue with an inner diameter of 475µm (Figure 1A).

The pillars were fabricated using a DLP bioprinter and a hydrogel bioink we synthesized. The bioink is formulated with polyethylene glycol diacrylate (PEGDA, 12.5v%), four-arm polyethylene glycol acrylate (PEG-Ac, 5w/v% ), and a 20mM solution of lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) photoinitiator.

Toroidal explants of contractile tissue were prepared from Xenopus laevis embryos, and the micropillar structure was threaded through the explant of initial inner diameter of approximately 500µm. The pillar structures were imaged continuously for 60 minutes using fluorescent stereo zoom microscopy (Figure 1B). To infer forces generated by the explant, we derived a constitutive relation of exerted force and beam end deflection taking into account pillar material stiffness and pillar geometry.

Results, Conclusions, and Discussions: Results

Our printed micropillar bioink material was determined to have a Young’s modulus of approximately 14.2kPa by dynamic mechanical analysis. Our micropillar constructs are sensitive to sub-uN forces and have been measured to deflect 4µm with application of 0.25µN of force using micro-scale tension-compression testing. Micropillar structures can be threaded through toroidal tissue explants to infer forces generated by the contractile tissue as shown in Figure 1B.

Discussion and Conclusions

We have created a novel biomaterial construct and associated method to infer the contractile forces generated by multi-dimensional tissues and explants of toroidal geometry. Our structure and associated modeling framework exhibits remarkable sensitivity, allowing for the detection and analysis of forces as low as 0.25µN. Our approach allows for robust analysis of 3-dimensional toroidal tissue explant contractility. Quantifying the contractile force generated by tissues of complex geometries may be applicable for investigating the mechanics of many biological systems such as embryology, wound healing, and muscle physiology.

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