"Three Dimentional brain in vitro models via electrofluidodynamics" - 3D NEUROGLIA - 3st Year report of the project

An accurate control of physical and chemical interactions between cells and extracellular matrix at the multiscale level is mandatory to understand the biological phenomena involving in neurogenesis and neurodegeneration. In this context, 3D neuroglia aims to design nano-structured biomaterials and nanocomposites including peculiar topological/chemical/physical features, controlled at micro-, sub micro- and nano- scale level, to validate their use as 3D models for mimic and/or in vitro supporting the functionalities of the natural extracellular matrix (ECM). For this purpose, electrofluidodynamic processes were variously investigated. During the first year of the project, electrospinning was selected to fabricate Poly ?-caprolactone (- GEL) and Poly ?-caprolactone/Gelatin fibres (+Gel), to form 3D instructive platforms and we have demonstrated their ability to variously affect the growth of astrocytes and neurons, as a function of their chemical and morphological/topological cues. During the second year of the project, more complex fibrous platforms based on the integration of intrinsically conductive polymers - i.e., polyaniline PANi - were also optimized to investigate the astrocyte response through electrical stimulation, in addition to biophysical and biochemical cues, just present in the previously tested models. During the third year of the project, in vitro studies on these platforms were further assessed for long-term in vitro neurophysiological investigations to validate the use PANi nano-needles synthesized in our labs and integrated into electrospun fibres as bio conductive platforms able to improve astrocytes interface, under controlling their bio-electrical properties. Meanwhile, the fabrication of micro-fabricated platforms was proposed to design symmetric 3D cultures able to model key biological processes, more efficaciously than asymmetric models based on electrospun fibres. For this purpose, electro fluid dynamic processes were properly customized to manipulate biopolymers under the application of high voltages electric fields to form microgel systems. During the 3rd year, the process was optimized for the fabrication of sodium alginate micro-gels with different physical and mechanical properties. At this preliminary stage, in vitro studies were performed to validate their use for in vitro studies with human mesenchymal stem cells (hMSCs), This screening allowed to investigate the effect of physical properties of microgels on the biological response of hMSCs cultured at the interface. In the view of these biological results, in the last year, it is planned to investigate the interaction of microgels with neurospheres as innovative 3D in vivo like models for brain.