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Functionalization of electrospun fibers of poly(ε-caprolactone) with star shaped NCO-poly(ethylene glycol)-stat-poly(propylene glycol) for neuronal cell guidance

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Abstract

Microfibers produced with electrospinning have recently been used in tissue engineering. In the development of artificial implants for nerve regeneration they are of particular interest as guidance structures for cell migration and axonal growth. Using electrospinning we produced parallel-orientated biocompatible fibers in the submicron range consisting of poly(ε-caprolactone) (PCL) and star shaped NCO-poly(ethylene glycol)-stat-poly(propylene glycol) (sPEG). Addition of the bioactive peptide sequence glycine-arginine-glycine-aspartate-serine (GRGDS) or the extracellular matrix protein fibronectin to the electrospinning solution resulted in functionalized fibers. Surface characteristics and biological properties of functionalized and non-functionalised fibers were investigated. Polymer solutions and electrospinning process parameters were varied to obtain high quality orientated fibers. A polymer mixture containing high molecular weight PCL, PCL-diol, and sPEG permitted a chemical reaction between hydroxyl groups of the diol and isocyanante groups of the sPEG. Surface analysis demonstrated that sPEG at the fiber surface minimized protein adhesion. In vitro experiments using dorsal root ganglia explants showed that the cell repellent property of pure PCL/sPEG fibers was overcome by functionalization either with GRGDS peptide or fibronectin. In this way cell migration and axonal outgrowth along fibers were significantly increased. Thus, functionalized electrospun PCL/sPEG fibers, while preventing non-specific protein adsorption, are a suitable substrate for biological and medical applications.

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Abbreviations

BSA:

Bovine serum albumin

DAPI:

4′,6-Diamidino-2-phenylindole dihydrochloride

DCM:

Dichloromethane

DIV:

Days in vitro

DMEM:

Dulbecco’s modified eagle medium

DMSO:

Dimethyl sulfoxide

DRG:

Dorsal root ganglia

ECM:

Extracellular matrix

FCS:

Fetal calf serum

GRGDS:

One letter code of the peptide sequence GlyArgGlyAspSer

Mw :

Molecular weight

NF200:

Neurofilament 200 kDa

NGS:

Normal goat serum

NMP:

N-Methylpyrrilidone

PBS:

Phosphate buffer saline

PCL:

Poly(ε-caprolactone)

PCL-ol:

PCL diol

PEG:

Poly(ethylene glycol)

PNS:

Peripheral nervous system

S100:

Protein antibody, marker for Schwann cells

SC:

Schwann cells

SEM:

Scanning electron microscopy

sPEG:

Star shaped NCO-poly(ethylene glycol)-stat-poly(propylene glycol)

THF:

Tetrahydrofuran

TBS-T:

Tris-buffered saline with 1% Triton-X 100

wt%:

Weight percent

XPS:

X-ray photoelectron spectroscopy

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Acknowledgments

We thank Marie Pradella for help with evaluating the cell experiments. Kristin Michael and Bernd Hoffmann kindly allowed us to use their equipment for making composite photographs of DRG explants. This work was supported by a grant from the Interdisciplinary Centre for Clinical Research “BIOMAT” within the faculty of Medicine at Aachen University (RWTH) (TV B111) and by DFG-Graduiertenkolleg 1035 “Biointerface”. Julia Bockelmann was supported by a Marie-Curie EST grant from the EU (EURON).

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Authors and Affiliations

  1. Institute of Technical and Macromolecular Chemistry and DWI e.V., RWTH Aachen University, Pauwelsstr. 8, 52056, Aachen, Germany

    Kristina Klinkhammer, Chariklia Simitzis, Dirk Grafahrend, Jürgen Groll, Martin Möller & Doris Klee

  2. Institute of Biology II, RWTH Aachen University, Mies-van-der-Rohe-Str. 15, 52074, Aachen, Germany

    Julia Bockelmann & Jörg Mey

  3. EURON Graduate School of Neuroscience, Maastricht, Netherlands

    Julia Bockelmann & Jörg Mey

  4. Institute of Neuropathology, Medical Faculty, RWTH Aachen University, Pauwelsstr. 30, 52074, Aachen, Germany

    Gary A. Brook

  5. IZKF-BIOMAT, Aachen University Hospital, RWTH Aachen University, Pauwelsstr. 30, 52074, Aachen, Germany

    Gary A. Brook

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  1. Kristina Klinkhammer
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  2. Julia Bockelmann
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  9. Doris Klee
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Correspondence to Doris Klee.

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Klinkhammer, K., Bockelmann, J., Simitzis, C. et al. Functionalization of electrospun fibers of poly(ε-caprolactone) with star shaped NCO-poly(ethylene glycol)-stat-poly(propylene glycol) for neuronal cell guidance. J Mater Sci: Mater Med 21, 2637–2651 (2010). https://doi.org/10.1007/s10856-010-4112-7

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