New tools and methodologies for engineering the human gut microbiota

The human gut microbiota is widely considered as a metabolic organ hidden within our bodies. One of the challenges in the research is to recreate an in vitro model of the human gut microbiota to study how the microorganisms regulate different metabolic pathways within human health and disease. For this reason, the aim of my PhD thesis is to investigate various methods to recreate a model of the gut microbiota in vitro and connect it to eukaryotic cell cultures. To do this, various tools from tissue engineering were used. First, a scaffold was made to create a 3D structure that could not only promote adhesion and proliferation of the microorganisms but also maintain a constant level of microbial composition. Next, the effect of a dynamic culture on the human gut microbiota was studied by evaluating the reaction of the microorganisms to different levels of shear stress. In this case, a controlled shear stress culture chamber was designed and manufactured to perform the tests, which showed several benefits when switching between static and dynamic culture conditions. Finally, several culture chambers and devices were designed and manufactured for dynamic culture of the human gut microbiota. A micro dialyzer was fabricated to connect the microorganisms compartment to the eukaryotic cells, thus avoiding possible infections and cytotoxic effects from the metabolites produced by the microorganisms. In particular, the effects of this co-culture were examined on intestine and bone models to investigate the crosstalk in the microbiota-gut-bone axis. All these tools and methods could be considered as the building blocks to recreate more complex models that can be used to study how microorganisms interact with eukaryotic cells in vitro analyzing the effects of diets, drugs, probiotics, and prebiotics.