Polyelectrolyte multilayers of poly (l-lysine) and hyaluronic acid on nanostructured surfaces affect stem cell response.

Laser interference lithography (LIL) and the layer-by-layer (LbL) technique are combined here for the first time to design a system with variable nanotopographies and surface viscoelasticity to regulate cell behavior. LIL is used to generate hexagonally arranged nanostructures of gold with different periodicity. In contrast, LBL is used to assemble a multilayer system of poly-l-lysine and hyaluronic acid on top of the nanostructures. Moreover, the viscoelastic properties of that system are controlled by chemical cross-linking. We show that the topography designed with LIL is still present after multilayer deposition and that the formation of the multilayer system renders the surfaces hydrophilic, which is opposite to the hydrophobic nature of pristine nanostructures. The heterogenic system is applied to study the effect on adhesion and differentiation of human adipose-derived stem cells (hADSC). We show that hADSC spreading is increasing with cross-linking degree on flat multilayers, while it is decreasing on nanostructures modified with multilayers. In addition, early effects on signal transduction processes are seen. Finally, hADSC differentiation into chondrogenic and osteogenic lineages is superior to adipogenic lineages on nanostructures modified with multilayers. Hence, the presented system offers great potential to guide stem cell differentiation on surfaces of implants and tissue engineering scaffolds.

Cross-linking multilayers of poly-l-lysine and hyaluronic acid: Effect on mesenchymal stem cell behavior.

BACKGROUND: Cells possess a specialized machinery through which they can sense physical as well as chemical alterations in their surrounding microenvironment that affect their cellular behavior. AIM: In this study, we aim to establish a polyelectrolyte multilayer system of 24 layers of poly-l-lysine and hyaluronic acid to control stem cell response after chemical cross-linking. METHODS AND RESULTS: The multilayer build-up process is monitored using different methods, which show that the studied polyelectrolyte multilayer system grows exponentially following the islands and islets theory. Successful chemical cross-linking is monitored by an increased zeta potential toward negative magnitude and an extraordinary growth in thickness. Human adipose-derived stem cells are used here and a relationship between cross-linking degree and cell spreading is shown as cells seeded on higher cross-linked polyelectrolyte multilayer show enhanced spreading. Furthermore, cells that fail to establish focal adhesions on native and low cross-linked polyelectrolyte multilayer films do not proliferate to a high extent in comparison to cells seeded on highly cross-linked polyelectrolyte multilayer, which also show an increased metabolic activity. Moreover, this study shows the relation between cross-linking degree and human adipose-derived stem cell lineage commitment. Histological staining reveals that highly cross-linked polyelectrolyte multilayers support osteogenic differentiation, whereas less cross-linked and native polyelectrolyte multilayers support adipogenic differentiation in the absence of any specific inducers. CONCLUSION: Owing to the precise control of polyelectrolyte multilayer properties such as potential, wettability, and viscoelasticity, the system presented here offers great potential for guided stem cell differentiation in regenerative medicine, especially in combination with materials exhibiting a defined surface topography.

Introduction of Laser Interference Lithography to Make Nanopatterned Surfaces for Fundamental Studies on Stem Cell Response.

The extracellular matrix (ECM) is a nanostructured environment that provides chemical, mechanical, and topographical stimuli for various cellular functions. Here, we introduce the application of laser interference lithography (LIL) to generate hexagonally arranged gold nanostructures of three different dimensions on silicon to study the effect of feature dimensions on human adipose-derived stem cells (hADSC) in terms of adhesion, growth, and differentiation. Self-assembled monolayers (SAM) were used to passivate the background silicon surface with a long-chain polyethylene glycol (PEG), whereas the gold nanostructures were activated with mercaptoundecanoic acid (MUDA) to direct protein adsorption and cell adhesive structures to them, only. It was possible to show that the size and distance of the nanostructures affected the spreading of hADSC with a decrease of cell size with the increase of feature dimensions, which corresponded also to the expression of focal adhesions and presence of the small GTPase RhoA. Effects of these early events, related to outside-in signal transduction, were visible by an enhanced cell growth on smaller feature dimensions and distinct effects on cell differentiation. Because of the precise control of chemical and topographical cues, the presented system offers great potential to study effects of material topography on stem cell behavior, which may pave the way for applications in tailoring surfaces of implants and tissue engineering scaffolds.