Production of functionalised micro and nanostructured polymer surfaces to trigger mesenchymal stem cell differentiation

MARTÍNEZ, E.; RÍOS-MONDRAGÓN, I.; PLA-ROCA, M.; RODRÍGUEZ SEGUÍ, S.A.; ENGEL, E.; MILLS, C. A.; SISQUELLA, X.; PLANELL, J.A.; SAMITIER, J.

Barcelona

Congreso; 13th European Congress on Biotechnology (ECB13); 2007

In vivo, mammalian cells interact with one another triggering diverse intracellular processes that control cell development. Similarly, the surrounding environment, constituting the extracellular matrix (ECM) and soluble factors, causes cells to adapt, reprogramming their intracellular  apparatus. Cells can sense topological, chemical and physical cues within the surrounding milieu. Indeed, mammalian cells respond to nano/microscale features on artificial surfaces. Hence, artificial bio-functionalised substrates with nano and micropatterns mightmake cells develop through different pathways, depending on the substrate design, in a non-invasive approach; specifically controlling processes such as cell adhesion, survival, proliferation, migration and differentiation. The principle relies on genetic reprogramming via intracellular signalling pathways, due to specific reactions through customized nanostructured surfaces in contact with cell surface receptors. The ?CellPROM? European  integrated project, involving 27 collaborating partners (both industrial and academic) from 12 European countries, entails the production of these nanostructured surfaces for controlling cellular characteristics. Substrate topology and biochemistry can be highlycontrollable factors. An understanding of these aspects will help in creating functional engineered tissues, as well as in the design of many implantable medical devices. It is not hard to envisage nanoscale devices with the ability to stimulate stem cells to produce partially or completely differentiated cells for regenerative therapies. The main objective of our work is to apply novel micro and nanofabrication techniques and surface modification strategies to generate well-defined topographical and biochemical cues for cell culture. To achieve this goal, nanoembossing has been used to impart micro and  nanostructures into polymer substrates, and their surface properties modified by micro-contact printing, nanoplotting and dip-pen nanolithography of ECMproteins, which are attached covalently to the surface. The modified surfaces are then used to study their influence on cell adhesion, morphology, proliferation and differentiation. Nanoembossing is achieved using nanoimprint lithography apparatus in biocompatible polymers such as poly(methyl methacrylate) (PMMA). Moulds for the nanoembossing are produced using conventional photolithography techniques combined with dry etching processes (micro) and focussed ion beam lithography or e-beam lithography (nano). Micro/nanopatterning of biochemical factors has beenundertaken by a variety of techniques, including micro-contact printing using PDMS stamps, nanolitre drop delivery (nanoplotter) and dip pen nanolithography, performed using a dedicatedAFM instrument. Results show that surface functionalisation with adhesion proteins such as fibronectin can be used to selectively attach and confine cells at specific surface locations. When micro and nanopatterned, fibronectin can also alter cell morphology, cytoskeletal organisation and stress level. Surface topography is instrumental in cell guidance and alignment processes, but it alsogreatly affects cell morphology. The study of both topographical and biochemical patterns is therefore particularly relevant when examining cell differentiation.