3D printing of compliant materials for industrially scalable muscle tissue modelling devices

In vitro tissue models are engineered biological structures that aim to replicate physiologically relevant microanatomy and function. Devices supporting the culture and analysis of artificial tissues have the potential to facilitate drug research, which is hindered by the poor predictability of efficacy and toxicity of drug compounds provided by two-dimensional cell culture assays and animal studies. As one of the leading causes of drug candidate rejection is cardiac or musculoskeletal toxicity, the development of engineered muscle tissues may bring about better treatment options for various conditions.

Previous work in the research group demonstrated the use-case of compliant 3D printed hydrogel pillars with elastic properties as micromechanical cues in muscle tissue culture in a laboratory-scale format. To enable widespread adoption of muscle models in drug development, tissue culture devices should ideally be produced in microtiter plate (MTP) layout to integrate into the workflow of the pharmaceutical industry. However, producing arrays of microfeatures over an extended area generally comes with challenges relating to loss of feature detail and non-uniform feature quality over the substrate area, affecting the readout of the contractility assay. To aid the upscaling process to MTP format, the thesis describes factors that influence the 3D printing outcome along with strategies to alleviate these challenges. Furthermore, an evaluation framework to assess 3D printer performance is established.

Although the previously developed hydrogel tissue modelling device has a proven track record of maintaining engineered muscle tissues and enabling contractile force readout, its dehydration leads to loss of structural integrity. The requirement of hydration imposes significant restrictions on post-processing, long-term storage and transportation of the device. Therefore, a compliant non-hydrated alternative was envisioned as the next generation tissue culture platform. A commercially available 3D printable elastomer was found to have suitable processability and mechanical properties as a substitute for the hydrogel based on bending stiffness measurements with a custom-built functional quality control setup. Development of a non-hydrated analogue of the hydrogel resin was also attempted without success. To enable active stretching of engineered muscle tissues, non-contact actuation of micropillars with magnetic fillers was explored but proved unfeasible due to the low success rate of printing and the aggregation of filler particles.

Lastly, strategies to reduce the cytotoxicity of 3D printed tissue modelling devices were studied in-depth. Biocompatibility treatments relying on removal of toxic compounds from the polymer network were investigated, along with the application of a thin coating to isolate the device from the tissue and the culture medium. The devices were shown to be compatible with electron beam irradiation as an industrially scalable sterilization method. The biocompatibility and contractility readout of platforms with thin coating surpassed the performance of most other toxicity mitigation methods in a 3-week long culture with stem cell-derived human cardiomyocytes. A further advantage of the thin layer coating is the limited absorption of test compounds by the elastomer. However, there is a trade-off between micropillar compliance and impermeability.

Overall, this work provides insight into converting muscle tissue modelling to an upscaled format, as well as demonstrates various biocompatibility treatment approaches of non-hydrated platforms with a focus on process scalability. The findings can contribute to the commercialization and widespread usage of muscle tissue culture devices in drug development, disease modelling and personalized medicine.