Evaluation of preosteoblast behavior on biomimetic intersected nanotube frameworks for tissue engineering applications

Details

Supervisors

Demoustier-Champagne, Sophie ; Dupont-Gillain, Christine C.

Faculty

Ecole polytechnique de Louvain

Degree label

Master [120] : ingénieur civil biomédical

Abstract

Extensive research in the past few decades has shed light on the role of the extracellular matrix (ECM) in the control of cell functions. By providing a complex combination of physical, chemical and nanotopographical cues to the cells, the ECM influences cell behaviors, such as adhesion, growth or differentiation. These findings have paved the way for the elaboration of new tissue engineering scaffolds and biointerfaces mimicking the ECM by presenting similar cues. In this context, this master thesis focuses on the response of mouse MC3T3-E1 preosteoblastic cells seeded on nanostructured biointerfaces mimicking the bone ECM. The studied biointerfaces are self-standing hard-templated frameworks, made of intersected rigid polypyrrole nanotubes and functionalized with a biomimetic multilayer coating of collagen (COL) and hyaluronan (HA) through Layer-by-Layer assembly. The nanotubular polypyrrole framework imitates the porous and nanofibrous structure of the bone native ECM and provides mechanical strength while the biomimetic coating, made of constituents present in bone native ECM, offers biomolecular cues to the cells. The elaboration process and the material characterization of the biointerfaces had already been completed in previous research works. The objective of this study was, therefore, to test cell behavior on the designed biointerfaces. The specific effects of the nanostructure and biomimetic (HA/COL)-coating of the biointerfaces were investigated by studying the response of MC3T3-E1 cells on both coated and non-coated polypyrrole nanotube frameworks. Cell adhesion and proliferation after 24 hours of culture on the biointerfaces were assessed by a fluorometric DNA quantification assay and epifluorescence microscopy. The ability to induce osteoblastic differentiation was evaluated after 7, 14 and 21 days, by colorimetric quantification of alkaline phosphatase activity, an early marker of osteoblastic differentiation. 24-h adhesion and proliferation results showed that the polypyrrole nanotube frameworks, both bare and coated, enhanced cell adhesion compared to bare glass substrates. The polypyrrole nanotube frameworks also promoted cell spreading and filopodia formation. The diameter of the polypyrrole nanotubes was found to influence cell adhesion, as frameworks of smaller diameter displayed increased cell attachment. In the absence of fetal bovine serum (FBS), the effect of the biomimetic (HA/COL)-coating was found to depend on the nanostructure of the underlying substrate. It enhanced cell adhesion on coated frameworks of larger diameter nanotubes but slightly attenuated it on frameworks of smaller diameter nanotubes. In the presence of FBS, the effect of the biomimetic coating was masked by the adsorption of a mixture of proteins naturally present in the serum. Osteoblastic differentiation was found to be significantly enhanced on both bare and coated biointerfaces, respectively only after 21 days and after both 14 and 21 days, compared to bare and coated glass substrates. This study provides a first evaluation of the adhesion, proliferation and differentiation of preosteoblastic cells on newly developed ECM-like biointerfaces. The results reveal the promising potential of such biointerfaces for the control of osteoblastic differentiation. The developed biointerfaces offer a platform for the delivery of signals to cells, with potential applications in cell biology, biomaterials science and tissue engineering.