Mass production of nanofibrous extracellular matrix with controlled 3D morphology for large-scale soft-tissue regeneration

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Alamein, M., Stephens, S., Liu, Q., Skabo, S., & Warnke, P.H. (2013). Mass production of nanofibrous extracellular matrix with controlled 3D morphology for large-scale soft-tissue regeneration. Tissue Engineering Part C: Methods, 19(6), 458-472

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Copyright © Mary Ann Liebert, 2013

2013 HERDC Submission. FoR code: 100404;110504




Aim: Biomaterials that mimic the nanofibrous architecture of the natural extracellular matrix (ECM) are in the focus for stem cell hosting or delivery in tissue engineering of multilayered soft tissues such as skin, mucosa, or retina. Synthetic nanofibers for such ECM are usually produced by single-syringe electrospinning with only one needle-jet at very low production rates of 0.005–0.008 g·min−1. The aim of this study was to utilize a novel industrial needle-free multijet electrospinning device with the potential for mass production of nanofibrous ECM (NF-ECM) exhibiting a controlled three-dimensional (3D) morphology for large-scale applications such as large area skin regeneration in patients with burns.

Methods: The novel NanoSpider™ NS200, an industrial apparatus originally designed for electrospinning of nanofibrous textile meshes, was used to fabricate 3D NF-ECMs of the following synthetic and natural biopolymers: collagen, gelatin, poly(caprolactone) (PCL), and poly(l-lactide-co-glycolide) (PLGA). Different concentrations of Gelatin polymer solution were electrospun under varying processing conditions, namely speed of spinning electrode rotation (u) and electric field intensity (E) by altering applied voltage (v) or the distance between electrodes (h) to achieve homogeneous desirable 3D morphology. Nanofiber diameters were assessed by scanning electron microscopy (SEM). Biocompatibility was tested by WST-1 (water-soluble tetrazolium salt) proliferation assay of seeded human mesenchymal stem cells (HMSCs). Biological performance of HMSCs on 3D PLGA NF-ECM was compared to two-dimensional (2D) PLGA film controls via SEM and confocal microscopy. Western blotting addressed the expression of surface adhesion proteins; focal adhesion kinase (FAK), phosphorylated FAK (pY397), α-tubulin, paxillin, vinculin. and integrin subunits; α5, αv, and β1 proteins.

Results: Large-scale mass production of NF-ECM membranes with a highly homogenous nanofiber morphology and 3D architecture could be produced with an extremely high production rate of 0.394±0.013 g·min−1·m−1 when compared to standard procedures. This was achieved by electrospinning a 20% wt/v gelatin solution, in an electric field intensity of 0.381 kV·mm−1. The nanofibers possessed diameters of around 180±40 nm with 28% deviation. HSMCs proliferation was significantly improved on NF-ECMs derived from collagen, gelatin, and PLGA when compared to PCL or flat coverglass controls (p

Conclusion: Needle-free multijet electrospinning can be used to mass produce artificial ECMs with intrinsic biocompatibility and desirable integration of stem cells for large-scale applications.

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