Abstract
This study presents novel biocompatible Polydimethylsiloxane (PDMS)-based micromechanical tweezers (μTweezers) capable of the stiffness characterization and manipulation of hydrogel-based organoids. The system showed great potential for complementing established mechanical characterization methods such as Atomic Force Microscopy (AFM), parallel plate compression (PPC), and nanoindentation, while significantly reducing the volume of valuable hydrogels used for testing. We achieved a volume reduction of ~0.22 μl/sample using the μTweezers vs. ~157 μl/sample using the PPC, while targeting high-throughput measurement of widely adopted micro-mesoscale (a few hundred μm-1500 μm) 3D cell cultures. The μTweezers applied and measured nano-millinewton forces through cantilever’ deflection with high linearity and tunability for different applications; the assembly is compatible with typical inverted optical microscopes and fit on standard tissue culture Petri dishes, allowing mechanical compression characterization of arrayed 3D hydrogel-based organoids in a high throughput manner. The average achievable output per group was 40 tests per hour, where 20 organoids and 20 reference images in one 35 mm petri dish were tested, illustrating efficient productivity to match the increasing demand on 3D organoids’ applications. The changes in stiffness of collagen I hydrogel organoids in four conditions were measured, with ovarian cancer cells (SKOV3) or without (control). The Young’s modulus of the control group (Control—day 0, E = 407± 146, n = 4) measured by PPC was used as a reference modulus, where the relative elastic compressive modulus of the other groups based on the stiffness measurements was also calculated (control-day 0, E = 407 Pa), (SKOV3-day 0, E = 318 Pa), (control-day 5, E = 528 Pa), and (SKOV3-day 5, E = 376 Pa). The SKOV3-embedded hydrogel-based organoids had more shrinkage and lowered moduli on day 0 and day 5 than controls, consistently, while SKOV3 embedded organoids increased in stiffness in a similar trend to the collagen I control from day 0 to day 5. The proposed method can contribute to the biomedical, biochemical, and regenerative engineering fields, where bulk mechanical characterization is of interest. The μTweezers will also provide attractive design and application concepts to soft membrane-micro 3D robotics, sensors, and actuators.
Highlights
Based on the design dimensions and the elastic modulus E = 2.46 MPa we found in our previous study [28], the calculated PDMS cantilever spring constant was (0.0765 N/m), and COMSOL stationary solid mechanics’ simulation resulted in a spring constant (0.08 N/m)
The stiffness of the hydrogel-based organoids was measured for four groups using the micro tweezers, namely, Control at day 0, ovarian cancer cells SKOV3 at day 0, Control at day 5, and SKOV3 at day 5 (Fig 8)
A common approach is the use of contact mechanics models
Summary
Studies have reported increased drug resistance in 3D cultures compared to 2D monolayer models, indicating that 3D models better represent in vivo conditions [3, 4]. The use of 3D models has shown great potential in studying different types of cancer, developing a better understanding of the 3D environmental cellular cues and signals [2], supporting the existing therapeutic approaches, and creating novel targeted precision medicine approaches [5]. The substrate or hydrogel matrix has a significant effect on the status of the cultured cells. Many have investigated substrate stiffness as the environmental parameters on the cell morphology (acini, rounded, protrusions, or invasive). Different epithelial cancer cell lines have shown various morphological cellular behaviors in response to substrate stiffness [9]
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