Human in-vitro microphysiological systems integrating polymeric nanofilms and high-resolution 3D printing

Microphysiological systems (MPSs) and or organs-on-chip have revolutionized in vitro modelling, drug discovery and toxicology by enabling the recreation of fundamental human physiological mechanisms in vitro in a dynamic and controlled environment. A design approach involving single or multiple cell types confined in fluidic compartments connected by a microporous membrane is essential for replicating tissue and cell characteristics. However, current platforms lack essential components of human organs due to manufacturing constraints. A first aspect is related to the complexity to replicate the endothelial basement membranes (BMs), due to substrate thickness, stiffness and poor permeability. The second one is the difficulty to precisely pattern cells within MPSs due to the complexity of integrating protein template-based coating with standard manufacturing processes. Within this thesis, a 200-nm porous Poly(D,L-lactic acid) (PDLLA) nanofilm is proposed as an alternative to commercially available membranes for recapitulating the biomechanical properties, porosity and thickness of the BM in an endothelialbarrier- on-chip. PDLLA nanofilms demonstrate enhanced elasticity and controlled pore diameters, facilitating cellular compartmentalisation and molecular exchange. Their integration into a double-layer microfluidic device enabled the formation of a tight endothelium and the co-culture of human endothelial cells and astrocytes, towards the development of an endothelial barrier-on-chip. In addition, aerosol-jet printing (AJP) of poly(3,4-ethylenedioxythiophene):polystyrene sulphonate (PEDOT:PSS), a conductive polymer blend, is proposed as an alternative micro-patterning approach for endothelial cell alignment within a microfluidic chamber. AJP allows the precise patterning of 50-µm wide lines of PEDOT: PSS onto a microfluidic chamber. Human endothelial cells seeded in the chamber preferably attached to the PEDOT:PSS pattern and retained pattern-induced alignment and elongation over 7 days of culture exclusively under capillary flow. These ultrathin polymeric films and high-resolution 3D printing protocol, combined with the conductive properties of the transparent PEDOT:PSS, open unlimited opportunities in in vitro modelling, in particular for fine reconstruction of organized multicellular structures and for in situ sensing, which are needed to improve reproducibility and utility of organs-on-a-chip.