Amino- and carboxy-functionalized nano- and microstructured surfaces for evaluating the impact of non-biological stimuli
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Amino- and carboxy-functionalized nano- and microstructured surfaces for evaluating the impact of non-biological stimuli on adhesion, proliferation and differentiation of primary skin-cells Petra J. Kluger2, Marc Panas 2, Lena Schober2, Günter E. M.Tovar1,2, Heike Mertsching2, Kirsten Borchers2
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Institute for Interfacial Engineering, University of Stuttgart, Fraunhofer-Institute for Interfacial Engineering and Biotechnology Nobelstr. 12, 70569 Stuttgart, Germany
ABSTRACT To gain basic insight into the impact of non-biological features on cells’ behaviour, primary skin-cells (keratinocytes and fibroblasts) were cultured on amine-functionalized or carboxyfunctionalized planar, nano- or microstructured surfaces. Sintered layers of silica nano- or microparticles were used to fabricate structures in the range of naturally occurring structuresizes. Organo-chemical functionalization was achieved using organo-functional silanes. Primary skin cells isolated from human foreskin were cultivated on these interfaces. Keratinocytes showed significantly better adhesion and proliferation on amino-functionalized surfaces than on carboxy-functionalized surfaces. On amino-functional surfaces decreasing proliferation was detected from planar to microstructured surfaces. Fibroblasts tended to proliferate stronger on carboxylated surfaces. Actin and vinculin, a protein involved in the formation of focal adhesions, were equally expressed on all surfaces, proofing intact cell-substrate contacts. INTRODUCTION Tissue Engineering is an interdisciplinary research field with the goal to manufacture in vitro tissues and organs [1, 2]. A crucial factor is the functional long-term cultivation of primary cells. Therefore, scaffolds are needed, featuring structurally and functionally tailored surfaces for the isolation and cultivation of primary cells in early differentiation stages [3, 4]. However, the number of studies, which examine cell behaviour of primary cells, is still negligible. In addition only few systematic studies investigated different structural effects on cell adhesion, proliferation and differentiation [5, 6]. Often structural stimuli compete with biomolecule-coatings like fibronectin or adhesion sequences like Arg-Gly-Asp (RGD) [7]. As these biomolecules mediate cell adhesion to the ECM via cellular adhesion receptors, the structural signals may be overcompensated and cannot be distinguished from the biochemical stimuli. Insight into the cells’ preferences concerning structure and the impact of organo-chemical functionality of the substrate helps to built up improved cell culture systems as well as optimized biomaterials and tailored surfaces for implants without the use of cost-intensive biological components.
In this study, we evaluated the behaviour of primary human keratinocytes and fibroblasts on planar, nano- and microstructures surfaces, which were functionalized by amine-functional and carboxy-functional groups. The globular geometries were generated using silica nanoparticles or microparticles (mean
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