Promoting Cell Migration and Neurite Extension along Uniaxially Aligned Nanofibers with Biomacromolecular Particles in a Density Gradient.

A simple method based upon masked electrospray is reported for directly generating both unidirectional and bidirectional density gradients of biomacromolecular particles on uniaxially aligned nanofibers. The method has been successfully applied to different types of biomacromolecules, including collagen and a mixture of collagen and fibronectin or laminin, to suit different types of applications. Collagen particles in a unidirectional or bidirectional gradient are able to promote the linear migration of bone marrow stem cells or NIH-3T3 fibroblasts along the direction of increasing particle density. In the case of particles made of a mixture of collagen and fibronectin, their deposition in a bidirectional gradient promotes the migration of Schwann cells from two opposite sides toward the center, matching the scenario in peripheral nerve repair. As for a mixture of collagen and laminin, the particles in a unidirectional gradient promote the extension of neurites from embryonic chick dorsal root ganglion in the direction of increasing particle density. Taken together, the scaffolds featuring a combination of uniaxially aligned nanofibers and biomacromolecular particles in density gradient can be applied to a range of biological studies and biomedical applications.

An engineered macroencapsulation membrane releasing FTY720 to precondition pancreatic islet transplantation.

Macroencapsulation is a powerful approach to increase the efficiency of extrahepatic pancreatic islet transplant. FTY720, a small molecule that activates signaling through sphingosine-1-phosphate receptors, is immunomodulatory and pro-angiogenic upon sustained delivery from biomaterials. While FTY720 (fingolimod, Gilenya) has been explored for organ transplantation, in the present work the effect of locally released FTY720 from novel nanofiber-based macroencapsulation membranes is explored for islet transplantation. We screened islet viability during culture with FTY720 and various biodegradable polymers. Islet viability is significantly reduced by the addition of high doses (≥500 ng/mL) of soluble FTY720. Among the polymers screened, islets have the highest viability when cultured with poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV). Therefore, PHBV was blended with polycaprolactone (PCL) for mechanical stability and electrospun into nanofibers. Islets had no detectable function ex vivo following 5 days or 12 h of subcutaneous implantation within our engineered device. Subsequently, we explored a preconditioning scheme in which islets are transplanted 2 weeks after FTY720-loaded nanofibers are implanted. This allows FTY720 to orchestrate a local regenerative milieu while preventing premature transplantation into avascular sites that contain high concentrations of FTY720. These results provide a foundation and motivation for further investigation into the use of FTY720 in preconditioning sites for efficacious islet transplantation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 555-568, 2018.

Effect of connective tissue growth factor delivered via porous sutures on the proliferative stage of intrasynovial tendon repair.

Recent growth factor, cell, and scaffold-based experimental interventions for intrasynovial flexor tendon repair have demonstrated therapeutic potential in rodent models. However, these approaches have not achieved consistent functional improvements in large animal trials due to deleterious inflammatory reactions to delivery materials and insufficient induction of targeted biological healing responses. In this study, we achieved porous suture-based sustained delivery of connective tissue growth factor (CTGF) into flexor tendons in a clinically relevant canine model. Repairs with CTGF-laden sutures were mechanically competent and did not show any evidence of adhesions or other negative inflammatory reactions based on histology, gene expression, or proteomics analyses at 14 days following repair. CTGF-laden sutures induced local cellular infiltration and a significant biological response immediately adjacent to the suture, including histological signs of angiogenesis and collagen deposition. There were no evident widespread biological effects throughout the tendon substance. There were significant differences in gene expression of the macrophage marker CD163 and anti-apoptotic factor BCL2L1; however, these differences were not corroborated by proteomics analysis. In summary, this study provided encouraging evidence of sustained delivery of biologically active CTGF from porous sutures without signs of a negative inflammatory reaction. With the development of a safe and effective method for generating a positive local biological response, future studies can explore additional methods for enhancing intrasynovial tendon repair. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2052-2063, 2018.

A General Strategy for Generating Gradients of Bioactive Proteins on Electrospun Nanofiber Mats by Masking with Bovine Serum Albumin.

Electrospun nanofibers are widely used in tissue engineering owing to their capability to mimic the structures and architectures of various types of extracellular matrices. However, it has been difficult to incorporate a biochemical cue into the physical cue provided by the nanofibers. Here we report a simple and versatile method for generating gradients of bioactive proteins on nanofiber mats. We establish that the adsorption of bovine serum albumin (BSA) onto nanofibers is a time- and concentration-dependent process. By linearly increasing the volume of BSA solution introduced into a container, a gradient in BSA is readily generated across the length of a vertically oriented strip of nanofibers. Next, the bare regions uncovered by BSA can be filled with the bioactive protein of interest. In demonstrating the potential application, we examine the outgrowth of neurites from dorsal root ganglion (DRG) isolated from chick embryos and then seeded on aligned polycaprolactone nanofibers covered by nerve growth factor (NGF) with a uniform coverage or in a gradient. In the case of uniform coverage, the neurites extending from DRG show essentially the same length on either side of the DRG cell mass. For the sample with a gradient in NGF, the neurites extending along the gradient (i.e., increase of NGF concentration) were significantly longer than the neurites extending against the gradient.

Spatiotemporal oxygen sensing using dual emissive boron dye-polylactide nanofibers.

Oxygenation in tissue scaffolds continues to be a limiting factor in regenerative medicine despite efforts to induce neovascularization or to use oxygen-generating materials. Unfortunately, many established methods to measure oxygen concentration, such as using electrodes, require mechanical disturbance of the tissue structure. To address the need for scaffold-based oxygen concentration monitoring, a single-component, self-referenced oxygen sensor was made into nanofibers. Electrospinning process parameters were tuned to produce a biomaterial scaffold with specific morphological features. The ratio of an oxygen sensitive phosphorescence signal to an oxygen insensitive fluorescence signal was calculated at each image pixel to determine an oxygenation value. A single component boron dye-polymer conjugate was chosen for additional investigation due to improved resistance to degradation in aqueous media compared to a boron dye polymer blend. Standardization curves show that in fully supplemented media, the fibers are responsive to dissolved oxygen concentrations less than 15 ppm. Spatial (millimeters) and temporal (minutes) ratiometric gradients were observed in vitro radiating outward from the center of a dense adherent cell grouping on scaffolds. Sensor activation in ischemia and cell transplant models in vivo show oxygenation decreases on the scale of minutes. The nanofiber construct offers a robust approach to biomaterial scaffold oxygen sensing.