No abstract

Electrospinning has become a widely implemented technique for the generation of nonwoven mats that are useful in tissue engineering and filter applications. The overriding factor that has contributed to the popularity of this method is the ease with which fibers with submicron diameters can be produced. Fibers on that size scale are comparable to protein filaments that are observed in the extracellular matrix. The apparatus and procedures for conducting electrospinning experiments are ostensibly simple. While it is rarely reported in the literature on this topic, any experience with this method of fiber spinning reveals substantial ambiguities in how the process can be controlled to generate reproducible results. The simplicity of the procedure belies the complexity of the physical processes that determine the electrospinning process dynamics. In this article, three process domains and the physical domain of charge interaction are identified as important in electrospinning: (a) creation of charge carriers, (b) charge transport, (c) residual charge. The initial event that enables electrospinning is the generation of region of excess charge in the fluid that is to be electrospun. The electrostatic forces that develop on this region of charged fluid in the presence of a high potential result in the ejection of a fluid jet that solidifies into the resulting fiber. The transport of charge from the charge solution to the grounded collection device produces some of the current which is observed. That transport can occur by the fluid jet and through the atmosphere surrounding the electrospinning apparatus. Charges that are created in the fluid that are not dissipated remain in the solidified fiber as residual charges. The physics of each of these domains in the electrospinning process is summarized in terms of the current understanding, and possible sources of ambiguity in the implementation of this technique are indicated. Directions for future research to further articulate the behavior of the electrospinning process are suggested.

The in vitro generation of a three-dimensional (3-D) myocardial tissue-like construct employing cells, biomaterials, and biomolecules is a promising strategy in cardiac tissue regeneration, drug testing, and tissue engineering applications. Despite significant progress in this field, current cardiac tissue models are not yet able to stably maintain functional characteristics of cardiomyocytes for long-term culture and therapeutic purposes. The objective of this study was to fabricate bioactive 3-D chitosan nanofiber scaffolds using an electrospinning technique and exploring its potential for long-term cardiac function in the 3-D co-culture model. Chitosan is a natural polysaccharide biomaterial that is biocompatible, biodegradable, non-toxic, and cost effective. Electrospun chitosan was utilized to provide structural scaffolding characterized by scale and architectural resemblance to the extracellular matrix (ECM) in vivo. The chitosan fibers were coated with fibronectin via adsorption in order to enhance cellular adhesion to the fibers and migration into the interfibrous milieu. Ventricular cardiomyocytes were harvested from neonatal rats and studied in various culture conditions (i.e., mono- and co-cultures) for their viability and function. Cellular morphology and functionality were examined using immunofluorescent staining for alpha-sarcomeric actin (SM-actin) and gap junction protein, Connexin-43 (Cx43). Scanning electron microscopy (SEM) and light microscopy were used to investigate cellular morphology, spatial organization, and contractions. Calcium indicator was used to monitor calcium ion flux of beating cardiomyocytes. The results demonstrate that the chitosan nanofibers retained their cylindrical morphology in long-term cell cultures and exhibited good cellular attachment and spreading in the presence of adhesion molecule, fibronectin. Cardiomyocyte mono-cultures resulted in loss of cardiomyocyte polarity and islands of non-coherent contractions. However, the cardiomyocyte-fibroblast co-cultures resulted in polarized cardiomyocyte morphology and retained their morphology and function for long-term culture. The Cx43 expression in the fibroblast co-culture was higher than the cardiomyocytes mono-culture and endothelial cells co-culture. In addition, fibroblast co-cultures demonstrated synchronized contractions involving large tissue-like cellular networks. To our knowledge, this is the first attempt to test chitosan nanofiber scaffolds as a 3-D cardiac co-culture model. Our results demonstrate that chitosan nanofibers can serve as a potential scaffold that can retain cardiac structure and function. These studies will provide useful information to develop a strategy that allows us to generate engineered 3-D cardiac tissue constructs using biocompatible and biodegradable chitosan nanofiber scaffolds for many tissue engineering applications.

Electrospinning is a widely used processing method to form fibrous tissue engineering scaffolds that mimic the structural features of the native extracellular matrix. Electrospun fibers made of collagen have been sought because it is a natural structural protein that supports cell attachment and growth. Yet, conventional solvents used to electrospin collagen can result in the loss of hydrolytic stability and fiber morphology of the scaffold. This study evaluated the effect of commonly used synthetic and natural crosslinking agents, genipin, glutaraldehyde, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), and EDC with N-hydroxysulfosuccinimide (EDC-NHS), on electrospun collagen. Crosslinked collagen scaffolds were assessed for structural integrity in an in vitro immersion study for up to 3 months. Their cytocompatibility was evaluated by human mesenchymal stem cell morphology and proliferation. Our results showed that dimensional stability and cytocompatibility of crosslinked electrospun collagen scaffolds are dependent on the type of crosslinking agent used. Collagen scaffolds treated with EDC and EDC-NHS were structurally stable and retained fiber structure for up to 3 months and were cytocompatible. Therefore, EDC and EDC-NHS are favorable crosslinking agents for electrospun collagen that can be utilized in tissue engineering applications.