A novel microphysiological system with integrated biomimetic scaffolds as an advanced in vitro model of the endocrine pancreas

Globally, rates of impaired glucose tolerance and diabetes have exploded, with half a billion people now living with diabetes. Research focused on unraveling the mechanisms behind diabetes to develop more effective therapeutic approaches relies mainly on rodents and conventional cell culture models. However, both models have significant drawbacks, which in turn limit the transferability of insights gained from them. The ethical constraints and shortcomings of animal models, the inadequacies of existing cell-based models, and the need to study disease under controlled conditions are giving rise to a field at the interface of tissue engineering and (patho)physiology that is focused on the development of advanced in vitro models. These are defined as experimental systems that contain living human cells and mimic tissue- and organ-level physiology in vitro by exploiting advances in tissue engineering and microfabrication. With this in mind, I propose an approach that goes beyond the state of the art of current models of the endocrine pancreas by using advanced biomedical techniques (e.g., 3D bioprinting, microphysiological systems). The newly developed approach is based on a microphysiological platform that will be optimized for the integration of biomimetic scaffolds with incorporated islets of Langerhans. The advanced model combines for the first time 3D printed biomimetic scaffolds incubated under the dynamic environmental conditions of a microphysiological system (MPS). To achieve the project goal, I will first develop novel hydrogel-based bioinks and functionalize them with pancreas-specific extracellular matrix (ECM) and basement membrane building blocks. The bioinks will be compatible with additive manufacturing techniques to fabricate scaffolds that serve as ECM surrogates and mimic the characteristics and interactions of the pancreatic microenvironment. Both the bioinks and the scaffolds will be systematically developed and characterized with respect to their structural, mechanical, and biochemical properties. On this basis, the optimal scaffold will be carefully selected in terms of its composition, while in parallel, the environmental parameters in the developed MPS will be defined and modified to support optimal islet viability and functionality. In this way, the scaffolds will meet structural, mechanical, and biochemical requirements, while the MPS will provide dynamic microenvironmental conditions and constant diffusion of key soluble molecules, leading to the emergence of complex biochemical gradients necessary for endocrine function. I will perform complex functional analyzes (i) measurement of reactive oxygen and nitrogen species production, (ii) quantification of proinflammatory chemokines by multiplex ELISA, (iii) detailed morphological analyzes by nanocomputed tomography and transmission electron microscopy, (iv) immunocytochemical phenotypic studies, (v) measurement of glucose-dependent insulin secretion, and (vi) confocal imaging of calcium dynamics on ""artificial tissue slices"" to validate that islets in the proposed in vitro model retain their native-like morphology, endocrine phenotype, viability, and secretory functionality over a significantly longer period than existing in vitro models. The proposed approach will lead to the development of one of the most advanced in vitro models of the endocrine pancreas while overcoming the limitations associated with animals and both acute tissue slices and isolated islets, which are currently the most representative in vitro models of the endocrine pancreas. By reducing the need for in vivo testing, I also address the ethical concerns highlighted in the ""3R"" principles for ethical research. Such a model will allow more complex experiments on human pancreatic islets under controlled conditions to unravel the mechanisms of diseases associated with pancreatic islet dysfunction and lead to more effective or even new models for diagnosis, prevention, and therapy.