COMPUTATIONAL AND BIOLOGICAL VALIDATION OF A NEW NICHOID SCAFFOLD OPTIMIZED FOR HUMAN MESENCHYMAL STEM CELLS

INTRODUCTION: Adipose-derived mesenchymal stem cells (hADMSCs) represent a promising source for cell therapies: they can be harvested directly from the patient with moderately invasive procedures and re-implanted without eliciting immune response. In particular, the physiologic niche microenvironment of stem cells, is characterized by a unique combination of biophysical, biochemical and biomechanical properties and nowadays it is a challenging field of research. Biomaterials tend to mimic the stem cells niches recreating the physiological system. In this context, the micro-scaffold “Nichoid” has shown to induce pluripotency in stem cells thanks to its peculiar 3D structure inducing isotropic mechanical stimuli (1,2). The aim of this study is to fabricate and validate a new geometry of the Nichoid optimized for the expansion of hADMSCs and investigate the mechanotransduction process of cells in different scaffold pore configurations, using a combined computational and experimental approach. METHODS: Nichoids were fabricated by an innovative two-photon polymerization technique using a biocompatible photoresist (3). To investigate the biomechanical stimuli of cells, we developed a computational model of the structural interaction between cells and the scaffold pore, using COMSOL Multiphysics. We predicted the Von Mises stresses on the scaffold structure and the maximal displacement of cells adhering to the scaffold. Then, we compared these mechanical parameters with several biomolecular and bioinformatics analyses performed on hADMSCs expanded on Nichoid scaffolds, as RNA-seq, Real Time RT-PCR and immunofluorescence assays. RESULTS: We compared different pores dimension (10x10um, 20x20um and 30x30um) and we identified the most suitable structure for hADMSCs (Fig.A). Our hypothesis is based on how displacements and deformation of the scaffold influence the isotropy of the cells (Fig.B). This phenomenon it is strictly connected to a roundish nucleus which corresponds to higher expression of stemness genes (Fig.C). Indeed, the cubic conformation of the scaffold correlated to increased pluripotency, verified by the up-regulation of the stem cell gene OCT4. DISCUSSION: The cubic Nichoid recreated the ex-vivo physiological environment optimized for hADMSCs. Moreover, cells expanded in this new configuration of the scaffold could be easily collected in view of an autologous transplantation. Therefore the presented biomechanical and biological validation represents a great promise for the Nichoid’s application in regenerative medicine, for example in the field of neurodegenerative diseases as Parkinson’s disease and Spinal Cord Injury. ACKNOWLEDGEMENTS: ERC projects NICHOID G.A. 646990, NICHOIDS G.A. 754467, and MOAB G.A. 825159; FET-OPEN project IN2SIGHT G.A. 964481; ESA project NICHOID-ET G.A. 4000133244/20/NL/GLC; NC3Rs projects MOAB, G.A. NC/C01903/1 and NC/C019201/1; MIUR-FARE project BEYOND, G.A. R16ZNN2R9K; FISR-COVID project LUNGCHILDMSC G.A. FISR2020IP_02959; Fondazione Giordano Dell’Amore, Cariplo Factory, Politecnico di Milano, Fondazione Bassetti and Fondazione Triulza (S2P project EGGS&BEACON) and Fondazione Romeo ed Enrica Invernizzi.