Periostin is predominantly expressed in collagen-rich fibrous connective tissues that are subjected to constant mechanical stresses including: heart valves, tendons, perichondrium, cornea, and the periodontal ligament (PDL). Based on these data we hypothesize that periostin can regulate collagen I fibrillogenesis and thereby affect the biomechanical properties of connective tissues. Immunoprecipitation and immunogold transmission electron microscopy experiments demonstrate that periostin is capable of directly interacting with collagen I. To analyze the potential role of periostin in collagen I fibrillogenesis, gene targeted mice were generated. Transmission electron microscopy and morphometric analyses demonstrated reduced collagen fibril diameters in skin dermis of periostin knockout mice, an indication of aberrant collagen I fibrillogenesis. In addition, differential scanning calorimetry (DSC) demonstrated a lower collagen denaturing temperature in periostin knockout mice, reflecting a reduced level of collagen cross-linking. Functional biomechanical properties of periostin null skin specimens and atrioventricular (AV) valve explant experiments provided direct evidence of the role that periostin plays in regulating the viscoelastic properties of connective tissues. Collectively, these data demonstrate for the first time that periostin can regulate collagen I fibrillogenesis and thereby serves as an important mediator of the biomechanical properties of fibrous connective tissues.

Polymer nanofibers exhibit properties that make them a favorable material for the development of tissue engineering scaffolds, filtration devices, sensors, and high strength lightweight materials. Electrospinning is a versatile method commonly used to manufacture polymer nanofibers. Collection of electrospun nanofibers across two parallel plates is a technique useful for creating nanofiber structures because it allows for the collection of linearly oriented individual nanofiber arrays and these arrays can be easily transferred to other substrates or structures. It is of importance to have some understanding of the capabilities of this collection method, such as the maximum length of fibers that can be collected across two parallel plates. The effect of different electrospinning parameters on maximum fiber length, average fiber diameter, diameter uniformity, and fiber quality was explored. It was shown that relatively long continuous polycaprolactone (PCL) nanofibers with average diameters from approximately 350 nm to 1 µm could be collected across parallel plates at lengths up to 35-50 cm. Experimental results lead to the hypothesis that even longer continuous nanofibers over 50 cm could be collected if the size of the parallel plates were increased. Extending the maximum fiber length that can be collected across parallel plates could expand the applications of electrospinning. Polymer solution concentration, plate size, and applied voltage were all shown to have varying effects on maximum fiber length, fiber diameter, and fiber uniformity.

Three dimensional multicellular aggregate, also referred to as cell spheroid or microtissue, is an indispensable tool for in vitro evaluating antitumor activity and drug efficacy. Compared with classical cellular monolayer, multicellular tumor spheroid (MCTS) offers a more rational platform to predict in vivo drug efficacy and toxicity. Nevertheless, traditional processing methods such as plastic dish culture with nonadhesive surfaces are regularly time-consuming, laborious and difficult to provide uniform-sized spheroids, thus causing poor reproducibility of experimental data and impeding high-throughput drug screening. In order to provide a robust and effective platform for in vitro drug evaluation, we present an agarose scaffold prepared with the template containing uniform-sized micro-wells in commercially available cell culture plates. The agarose scaffold allows for good adjustment of MCTS size and large-scale production of MCTS. Transparent agarose scaffold also allows for monitoring of spheroid formation under an optical microscopy. The formation of MCTS from MCF-7 cells was prepared using different-size-well templates and systematically investigated in terms of spheroid growth curve, circularity, and cell viability. The doxorubicin cytotoxicity against MCF-7 spheroid and MCF-7 monolayer cells was compared. The drug penetration behavior, cell cycle distribution, cell apoptosis, and gene expression were also evaluated in MCF-7 spheroid. The findings of this study indicate that, compared with cellular monolayer, MCTS provides a valuable platform for the assessment of therapeutic candidates in an in vivo-mimic microenvironment, and thus has great potential for use in drug discovery and tumor biology research.

Extracellular matrix fibers (ECM) such as collagen, elastin, and keratin provide biological and physical support for cell attachment, proliferation, migration, differentiation and ultimately cell fate. Therefore, ECM fibers are an important component in tissue and organ development and regeneration. Meanwhile, polymer nanofibers could play the same critical role in tissue regeneration process. Fibrous structures can be fabricated from a variety of materials and methods with diameters ranging throughout the size scale where cells can sense individual fibers (several nanometers to several microns). Polymer nanofiber scaffolds can be designed in a way that predictably modulates a variety of important cell behaviors towards a desired overall function. The nanofibrous topography itself, independent of the fiber material, has demonstrated the potential to modulate cell behaviors desirable in tissue engineering such as: unidirectional alignment; increased viability, attachment, and ECM production; guided migration; and controlled differentiation. The versatility of polymer nanofibers for functionalization with biomolecules opens the door to vast opportunities for the design of tissue engineering scaffolds with even greater control over cell incorporation and function. Despite the promise of polymer nanofibers as tissue engineering scaffolds there have been few clinically relevant successes because no single fabrication technique currently combines control over structural arrangement, material composition, and biofunctionalization, while maintaining reasonable cost and yield. Promising strategies are currently being investigated to allow for the fabrication of optimal polymer nanofiber tissue engineering scaffolds with the goal of treating damaged and degenerated tissues in a clinical setting.

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At present there is no clinically effective treatment for injuries or pathological processes that disrupt the continuity of axons in the mature central nervous system. However, a number of studies suggest that a tremendous potential exists for developing biomaterial based therapies. In particular, biomaterials in the form of bridging substrates have been shown to support at least some level of axonal regeneration across the lesion site, but display a limited capacity for directing axons toward their targets. To improve the directionality and outgrowth rate of the axonal regeneration process, filaments and tubes appear promising, but the technology is far from optimized. As a step toward optimization, the influence of filament diameter and various extracellular matrix coatings on nerve regeneration was evaluated in this article using a dorsal root ganglion (DRG) explant model. An increasing pattern of alignment and outgrowth of neurites in the direction parallel the long axis of the packed filament bundles with decreasing filament diameters ranging from supracellular and beyond (500 to 100 mum), cellular (30 mum), down to subcellular size (5 mum) was observed. Such effects became most prominent on filament bundles with individual filament diameters in the range of cellular size and below (5 and 30 mum) where highly directional and robust neuronal outgrowth was achieved. In addition, laminin-coated filaments that approached the size of spinal axons support significantly longer regenerative outgrowth than similarly treated filaments of larger diameter, and exceed outgrowth distance on similarly sized filaments treated with fibronectin. These data suggested the feasibility of using a multifilament entubulation bridging device for supporting directional axonal regeneration.

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We previously demonstrated that coadministration of glial cell line-derived neurotrophic factor (GDNF) with grafts of Schwann cells (SCs) enhanced axonal regeneration and remyelination following spinal cord injury (SCI). However, the cellular target through which GDNF mediates such actions was unclear. Here, we report that GDNF enhanced both the number and caliber of regenerated axons in vivo and increased neurite outgrowth of dorsal root ganglion neurons (DRGN) in vitro, suggesting that GDNF has a direct effect on neurons. In SC-DRGN coculture, GDNF significantly increased the number of myelin sheaths produced by SCs. GDNF treatment had no effect on the proliferation of isolated SCs but enhanced the proliferation of SCs already in contact with axons. GDNF increased the expression of the 140 kDa neural cell adhesion molecule (NCAM) in isolated SCs but not their expression of the adhesion molecule L1 or the secretion of the neurotrophins NGF, NT3, or BDNF. Overall, these results support the hypothesis that GDNFenhanced axonal regeneration and SC myelination is mediated mainly through a direct effect of GDNF on neurons. They also suggest that the combination of GDNF administration and SC transplantation may represent an effective strategy to promote axonal regeneration and myelin formation after injury in the spinal cord.