Technical NoteA biodegradable nanofiber scaffold by electrospinning and its potential for bone tissue engineering
Introduction
Engineering living tissue for reconstructive surgery requires an appropriate cell source, optimal culture conditions and a biodegradable scaffold as the basic elements. While specialized cells remain an important source, stem cells have emerged as a promising new alternative. Recent advances in stem cell biology have shown that mesenchymal stem cells (MSCs) can differentiate into cells of mesenchymal tissues such as bone, cartilage, muscle, tendon, ligament and fat, and are expected to play an important role in the repair of skeletal defects [1], [2].
It is also a well-accepted paradigm that the culture conditions affect the quality of the engineered tissue, and the development of bioreactors to provide optimal culture conditions remains an active field of research [3]. Recently, a rotational oxygen-permeable bioreactor system (ROBS) has been developed in our laboratory to provide optimal oxygen tension and mechanical stresses to cell-polymer constructs in culture. Osteoblasts derived from MSCs of neonatal rats were cultured on poly(dl-lactide-co-glycolide) (PLGA) foams, and mineralization as well as three-dimensional bone formation was observed [4].
Regarding the scaffold, it is generally agreed that a highly porous microstructure with interconnected pores and a large surface area is conducive to tissue ingrowth. For bone regeneration, pore sizes between 100 and 350 μm and porosities of more than 90% are preferred, and a variety of different scaffold fabrication techniques has been reported [5].
This study assesses the potential of electrostatic fiber spinning, or electrospinning, as an alternative scaffold fabrication technique to engineer bone in vitro using ROBS. Electrospinning produces highly porous non-woven fabrics consisting of ultrafine fibers. A wide variety of polymers have been electrospun and several applications have been proposed in recent years based on the small fiber diameters and high porosities, e.g. Refs. [6], [7]. Poly (ε-caprolactone) PCL was chosen as a model polymer due to its lack of toxicity, low cost and slow degradation [8]. MSCs were harvested from the bone marrow of neonatal rats and seeded on electrospun (PCL) scaffolds. The cell-polymer constructs were cultured with osteogenic supplements under dynamic culture conditions for 4 weeks. Cell behavior was observed using scanning electron microscopy (SEM), histological and immunohistochemical analyses.
Section snippets
Materials
PCL with an average molecular weight of 80,000 (80 kDa) was obtained from Aldrich (Milwaukee, WI), and dissolved in chloroform under gentle stirring to obtain a 10 wt% solution. All solvents were analytical grade and used as received from Sigma (St. Louis, MO).
Fabrication of polymer scaffolds
The polymer solution was delivered at a constant flow rate (Q=0.1 ml/min, Harvard Apparatus PHD 2000 syringe pump, Holliston, MA) to a metal capillary (1.6 mm OD, 1 mm ID, 50 mm length, Cole-Parmer, Vernon Hills, IL) connected to a high-voltage
Scanning electron microscopy
An SEM micrograph of electrospun PCL is shown in Fig. 2a. The three-dimensional fibrous mesh consists of fibers with diameters ranging from 20 nm to 5 μm. Most of the fiber diameters are less than 1 μm, and the average diameter is 400 nm (±200 nm). The standard deviation is denoted in parentheses. In addition to the broad fiber diameter distribution, occasional extreme outliers with diameters of up to 5 μm were observed. The fibers often had non-uniform diameters, i.e., the diameter varied along an
Discussion
This study focuses on the electrospinning process for the production of a nanofiber scaffold and assesses the cell behavior as an indicator for the potential for bone tissue engineering. Unlike conventional fiber spinning processes that produce fibers with diameters in the micron range, electrospinning is capable of producing fibers in the nanometer diameter range, or nanofibers, that are typically deposited in the form on non-woven fabrics. The large diameter reduction from a millimeter-scale
Conclusion
For clinical applications, especially the regeneration of specific size defects, structural integrity and sufficient cell penetration are two key requirements. This study has shown that electrospun PCL scaffolds provide an environment that supports mineralized tissue formation and may be a suitable candidate for the treatment of bone defects. The shapeability of electrospun PCL scaffolds may also prove useful in clinical applications. Current work is in progress to determine the in vivo
Acknowledgments
This study was supported by the Center for Integration of Medicine and Innovative Technology (CIMIT).
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These authors contributed equally.