Kapton® mit zell-selektiven Oberflächenbeschichtungen für verbesserte Elektroden-Nerv-Interaktionen

Einleitung: Hinsichtlich der Effektivität der Elektroden-Nerv-Interaktion spielt das Trägermaterial auditorischer Implantate eine besondere Rolle: Dessen Oberflächenbeschaffenheit sollte das Bindegewebswachstum hemmen. Hierbei stellen beschichtete Polyimide das Material der Zukunft zur Elektrodenumhüllung dar. Eine Strategie zur Oberflächenmodifikation von Polyimiden besteht in der photochemischen Anbindung geeigneter Polymere. Methoden: Für die Beschichtung wurden photochemisch reaktive DMAA-, DEAA- und MTA-enthaltende Copolymere sowie ein photoaktives Chitosanderivat verwendet. Das Polyimid Kapton® wurde mit den Polymeren via Rotationsbeschichtung oder Aufsprühen und anschließender Bestrahlung mit UV-Licht beschichtet. Der Nachweis der Polymerbeschichtungen erfolgte über ATR-IR-Spektroskopie, Kontaktwinkel-messungen, XPS und AFM. Die Zellselektivität wurde durch Kultivierung der lentiviral modifizierten murinen Fibroblasten-Zelllinie NIH3T3 untersucht. Die Zellviabilität der Fibroblasten wurde unter Verwendung eines Redox-Farbstoffs Resazurin quantitativ bestimmt. Die Morphologie der Zellen wurde fluoreszenzmikroskopisch untersucht. Ergebnisse: Die Synthese der photochemisch reaktiven Polymere sowie die Beschichtung des Kapton® erwies sich als erfolgreich. Alle Polymere wirkten im Vergleich zu unbehandeltem Kapton® inhibierend auf das Fibroblastenwachstum. Die Beschichtung mit dem Chitosanderivat und dem DMAA-enthaltenden Copolymer wirkte sogar antiadhäsiv. Schlussfolgerungen: Chitosanderivat und das DMAA-Copolymer stellen potentiell klinisch relevante Beschichtungen dar, die die Bindegewebsbildung effektiv hemmen und damit die Impedanzen reduzieren. Der Effekt der Polymerbeschichtungen auf die Viabilität von neuronalen Zellen ist Gegenstand laufender Untersuchungen.Unterstützt durch: SFB599, Teilprojekt D2Der Erstautor gibt keinen Interessenkonflikt an

Polymer Coatings of Cochlear Implant Electrode Surface - An Option for Improving Electrode-Nerve-Interface by Blocking Fibroblast Overgrowth.

Overgrowth of connective tissue and scar formation induced by the electrode array insertion increase the impedance and, thus, diminish the interactions between neural probes as like cochlear implants (CI) and the target tissue. Therefore, it is of great clinical interest to modify the carrier material of the electrodes to improve the electrode nerve interface for selective cell adhesion. On one side connective tissue growth needs to be reduced to avoid electrode array encapsulation, on the other side the carrier material should not compromise the interaction with neuronal cells. The present in vitro-study qualitatively and quantitatively characterises the interaction of fibroblasts, glial cells and spiral ganglion neurons (SGN) with ultrathin poly(N,N-dimethylacrylamide) (PDMAA), poly(2-ethyloxazoline) (PEtOx) and poly([2-methacryloyloxy)ethyl]trimethylammoniumchlorid) (PMTA) films immobilised onto glass surfaces using a photoreactive anchor layer. The layer thickness and hydrophilicity of the polymer films were characterised by ellipsometric and water contact angle measurement. Moreover the topography of the surfaces was investigated using atomic force microscopy (AFM). The neuronal and non-neuronal cells were dissociated from spiral ganglions of postnatal rats and cultivated for 48 h on top of the polymer coatings. Immunocytochemical staining of neuronal and intermediary filaments revealed that glial cells predominantly attached on PMTA films, but not on PDMAA and PEtOx monolayers. Hereby, strong survival rates and neurite outgrowth were only found on PMTA, whereas PDMAA and PEtOx coatings significantly reduced the SG neuron survival and neuritogenesis. As also shown by scanning electron microscopy (SEM) SGN strongly survived and retained their differentiated phenotype only on PMTA. In conclusion, survival and neuritogenesis of SGN may be associated with the extent of the glial cell growth. Since PMTA was the only of the polar polymers used in this study bearing a cationic charge, it can be assumed that this charge favours adhesion of both glial cells and SG neurons glial cells and SGN

Scanning electron microsopy of the adherent SGN following cultivation on polymer monolayers and ornithine/laminin coated glass plates.

<p>Two days after seeding all cell types of the SG were distributed on ornithine/laminin coated glass plates showing characteristic morphologies (positive control, <b>A</b>) as well on PMTA layers (<b>B</b>), whereas on PEtOx films (<b>C</b>) flat cells with broad lamellopodia were mainly observed. In contrast, only few cells were adherent on the PDMAA coated surfaces (<b>D</b>). Tiny spherical spots with different sizes in the range of less than 1 μm were visible on both PEtOx (<b>C</b>) and PDMAA coatings (<b>D</b>). They indicate either adsorption of ECM molecules or residual cell bodies or extensions, which were putatively torn off due to lacking mechanical stability. Furthermore, flattened pale spots could be exclusively observed on PEtOx coatings (<b>C</b>) indicating attempts of the cells to form adhesion points due to areas without polymer coating.</p

Water contact angle (WCA) and layer thickness determined by ellipsometry.

<p>Water contact angle (WCA) and layer thickness determined by ellipsometry.</p

Determination of the number of fibroblasts and glial cells cultivated on polymer coated and uncoated glass plates following immunostaining in relation to the total cell number.

<p>Cell count of the fibroblasts and glial cells presented in SG following positive staining with anti-vimentin and anti-p75 antibodies in relation to the total number of DAPI stained cells. The cells were seeded on glass plates with polymer layers PDMAA, PMTA and PEtOx as well on glass plates with ornithine/laminin coating assigned as positive control (PosCtrl). Each data point is presented as mean and SEM of the total cell number (<b>A</b>) and percentage of the number of fibroblasts and glial cells in relation to the total cell number counted in n = 2 (PEtOx), n = 4 (PosCtrl.) and n = 5 (PDMAA, PMTA) fluorescence images (<b>B</b>).</p

Primary antibodies used in this study.

<p>Listing of the primary for immunostaining of cell specific antigens in the SGN, fibroblasts and glial cells.</p