INNOVATIONS FROM THE HORIZON 2020 EUROPEAN SCREENED PROJECT: IRON-DOPED HYDROXYAPATITE NANOPARTICLES (FEHA NPS) FOR MAGNETIC GUIDANCE OF RAT THYROCYTES ONTO THYROID LOBE-LIKE 3D COLLAGEN SCAFFOLDS

The Horizon 2020 European SCREENED Project (#825745,https://www.unipr.it/notizie/interferenti-endocrini-luniversita-di-parma-vincitrice-di-un-finanziamento-europeo-horizon) is focussed on developing 3D in vitro assays based on 3D organoids engineered with mammalian thyroid cells, for screening the effect of low doses Endocrine Disruptors (EDs). Exploiting cellular internalization of CNR-ISTEC fabricated, biocompatible and bioresorbable iron-doped hydroxyapatite nanoparticles (FeHA NPs), we used the certified adult rat thyrocyte cell line FRTL5 (CLS n.500407) whose thyroid-specific proteomic profile is fully comparable to that of adult male rat primary thyrocytes. FIRTL5 monolayers were challenged for 6 hs with FeHA NPs; commercially available SPION FLUIDMag NPs were used as control. Following uptake of both NPs types, cell viability was assessed at 6, 24 and 48 hs whereas evaluation of internalized NPs was immediately achieved by ICP analysis. Using both NPs, FIRTL5 cells remained viable in extremely high percentage. Similar, intracellular accumulation of NPs resulted close to maximal saturation with both types of NPs. For magnetic guidance, both unmagnetized and magnetized FIRTL5 cells were grown inside an ad hoc engineered culture chamber having at its center a 3D cylindric and porous collagen scaffold mimicking a decellularized rat thyroid lobe, harboring at its core a 3x3 mm static magnet generating a magnetic field of 320 mT. Light fluorescence, immunocytochemistry, and ultrastructural (TEM, SEM) microscopies revealed that magnetized cells densely accumulated onto the 3D scaffold, colonizing the empty follicular-like cavities. Collectively, we succeeded in setting an innovative 3D bioartificial thyroid lobe-like organoid based on thyroid epithelial cells magnetized with FeHA NPs, and their guidance onto a 3D collagen scaffold surrogating a decellularized 3D thyroid lobe matrix, thus providing a new assay model for in vitro testing low doses of EDs.