Hierarchical polymer niches for enhanced cell attachment

Research context / Theoretical framework Controlling and understanding adhesion of cells on artificial surfaces remains as a critical topic in materials and life sciences. In this regard, combination of top-down (contact printing) and bottom-up approaches (ATRP polymerization + layer-by-layer adsorption of polyelectrolytes and proteins) appears as a promising strategy for the design and fabrication of cell-appealing interfaces. Interestingly, this methodology allows going from 2D to 3D-like hierarchical structures of hybrid content (niches) that influence a subsequent cell attachment on top, by better exposing the specific binding sites (RGD, IKVAV moieties) towards target membrane receptors (i.e. integrins, CD44). Complementary use of Atomic Force Microscopy (AFM), with a living cell as probe, together with Quartz Crystal Microbalance with Dissipation (QCM-D), will enable an early-stage analysis and quantification of these cell-substrate interactions on the nanoscale. Hypotheses/ Research questions / Objectives The main hypotheses of the project are the following: i) Combination of substrate-anchored polymer brushes and layer-by-layer deposited polyelectrolyte chains give rise to soft 3D niches for the enhanced adsorption of ECM proteins. The transformation of 2D interfaces into 3D-like architectures will, in turn, enhance cell attachment and proliferation of cells, with particular impact on both cell morphology and the number of cell-substrate connections formed; ii) The use of Contact-Printing techniques before the grafting-from of the brushes allows the fabrication of localized individual 3D attachment points. The localized presence of specific molecules will influence the cell-substrate affinity with final impact on cell morphology and the establishment of a different number of cell-surface contacts; iii) Single-Cell Probe Force Spectroscopy (SCPFS) technique is sensitive enough to identify early stage attachment events in cell-substrate contacts. The use of a living cell acting as indenting probe will determine events taking place on the nano- and microscale. Approach / Methods The following methods will be used to study substrate preparation and cell adhesive behaviour: Atomic force microscopy (AFM) in SCPFS mode, (confocal) fluorescence microscopy, quartz crystal microbalance with dissipation (QCMD), scanning electron microscopy (SEM), and cell culture protocols.