Within the course of the project, the team first focused on designing and completing the building blocks (engineered cells, biomaterials and 3D printing set-ups), that are needed to create (bioprint) the pancreatic tissue constructs to be used as drug testing platforms and implantable grafts. In particular, we have developed two new synthetic biology-based strategies that enable the activation of cell responses upon exposure to light stimuli. As a proof-of-concept of the functionality of these networks, production and release of insulin in cell lines was demonstrated. In addition, we developed a robust method to generate pancreatic islets from iPSC using a precise array of soluble factors. These stem cell-derived islets are rich in beta-, alpha- and delta-like cells, and respond to glucose stimuli with proportional insulin secretion, and are already in use in our cultures involving bioprinted constructs. In parallel, several gelatin-based hydrogels formulations have been developed, and a base design that allows the formation of 3D clusters of engineered beta-like cells, to preserve the identity and function of iPSC-derived pancreatic islets as well as the formation of interconnected capillary networks from a co-culture of endothelial HUVEC cells and mesenchymal stromal cells has been defined. The material can be shaped via volumetric printing, and their functionality via the embedding of ECM matrix components. To sculpt these cell-laden materials, a new volumetric, tomographic 3D printing technology has been developed, enabling the rapid fabrication of centimeter scale constructs in less than 30 seconds. Several strategies have been developed to ensure the encapsulation of high cell densities contextually to a high shape fidelity and printing resolution.
Combining this knowledge and technologies, we built a fully functioning set-up for the sterile perfusion of geometrically complex, centimetre scale constructs printed from hydrogels displaying low mechanical properties was designed and tested. Perfusion culture of endothelial cells seeded in bioprinted channels was achieved. Bioprinting of iPSC derived islet organoids (obtained by chemical differentiation) is possible, culture up to 28 days, with nearly 100% viability has been performed. This system allows for simulating systemic flow, and to subject the islets to different type of metabolic and chemical challenges, such as fluctuations of glucose concentrations, as well as to the addition of anti-diabetic drugs (i.e. GLP-1 analogs) and toxic compounds. The system is a promising platform for advanced in vitro drug testing to reduce animal experimentation. In addition, we also investigated the therapeutic potential of both iPSC islets and of cell lines designed to act as beta cells. The engineered cells were able to restore glicemic control in diabetic mice.
Moreover, the dissemination framework for ENLIGHT was established in the first six months of the project. This included the website, logo, and social media platforms, along with a data management plan. The ENLIGHT partners were active in dissemination and communication efforts, including press releases by international media outlets, the development of project videos, presentations at virtual, global scientific conferences, and the publishing of peer-reviewed publications.
