Human Platelet Lysate-Derived Nanofibrils as Building Blocks to Produce Free-Standing Membranes for Cell Self-Aggregation

abstract

Amyloid-like fibrils are garnering keen interest in biotechnology as supramolecular nanofunctional units to be used as biomimetic platforms to control cell behavior. Recent insights into fibril functionality have highlighted their importance in tissue structure, mechanical properties, and improved cell adhesion, emphasizing the need for scalable and high-kinetics fibril synthesis. In this study, we present the instantaneous and bulk formation of amyloid-like nanofibrils from human platelet lysate (PL) using the ionic liquid cholinium tosylate as a fibrillating agent. The instant fibrillation of PL proteins upon supramolecular protein-ionic liquid interactions was confirmed from the protein conformational transition toward cross-beta-sheet-rich structures. These nanofibrils were utilized as building blocks for the formation of thin and flexible free-standing membranes via solvent casting to support cell self-aggregation. These PL-derived fibril membranes reveal a nanotopographically rough surface and high stability over 14 days under cell culture conditions. The culture of mesenchymal stem cells or tumor cells on the top of the membrane demonstrated that cells are able to adhere and self-organize in a three-dimensional (3D) spheroid-like microtissue while tightly folding the fibril membrane. Results suggest that nanofibril membrane incorporation in cell aggregates can improve cell viability and metabolic activity, recreating native tissues' organization. Altogether, these PL-derived nanofibril membranes are suitable bioactive platforms to generate 3D cell-guided microtissues, which can be explored as bottom-up strategies to faithfully emulate native tissues in a fully human microenvironment.

keywords

FIBRILLATION; VINCULIN; PEPTIDE; DISEASE; MATRIX; MODEL

subject category

Chemistry; Science & Technology - Other Topics; Materials Science

authors

Monteiro, C; Gomes, MC; Bharmoria, P; Freire, MG; Coutinho, JAP; Custódio, CA; Mano, JF

Groups

G4 - Renewable Materials and Circular Economy
G5 - Biomimetic, Biological and Living Materials

Projects

CICECO - Aveiro Institute of Materials (UIDB/50011/2020)

CICECO - Aveiro Institute of Materials (UIDP/50011/2020)

Associated Laboratory CICECO-Aveiro Institute of Materials (LA/P/0006/2020)

Full human-based multi-scale constructs with jammed regenerative pockets for bone engineering. (REBORN)

Human based bioinks to engineer physiologically relevant tissues (HumanINK)

Unlocking the scientific excellence and innovation capacity of the University of Aveiro in supramolecular multicomponent biomaterials for enabling advanced biomaterials for healthcare (SUPRALIFE)

acknowledgements

This work was developed within the scope of the project CICECO-Aveiro Institute of Materials, UIDB/50011/2020 (DOI 10.54499/UIDB/50011/2020), UIDP/50011/2020 (DOI 10.54499/UIDP/50011/2020) and LA/P/0006/2020 (DOI 10.54499/LA/P/0006/2020), financed by national funds through the FCT/MCTES (PIDDAC). The authors would like to acknowledge the European Research Council for the Advanced Grant Agreement number H2020-ERC-AdG-883370 for the project REBORN and for the Proof-of-Concept Grant Agreement number ERC-2022-PoC-101082210 for the project HumanINK. This work was funded by the European Union's Horizon Europe research and innovation programme under the grant agreement No. 101079482 ("SUPRALIFE"). This work was also supported by the Foundation for Science and Technology through the individual contract 2020.01647.CEECIND of C.A.C. and the doctoral grant SFRH/BD/144640/2019 of C.F.M. P.B. acknowledges La-Caixa junior research leadership-postdoctoral program grant (ID: 100010434, fellowship code: LCF/BQ/P122/11910023) for financial support.