Cell-laden Hydrogel Microfibers Research Articles

Upregulation of biochemical and biophysical properties of cell-laden microfiber, silk-hyaluronic acid composite

Cell-laden filament-like hydrogels are advantageous for many applications including drug screening, tissue engineering, and regenerative medicine. However, most of the designed filament vehicles hold weak mechanical properties, which hinder their applications in specific tissue engineering. We present a binary hybrid silk and hyaluronic acid hydrogel microfiber generated through a microfluidic system to encapsulate cells with superior mechanical properties and biocompatibility. Cell-laden hydrogel microfibers were continuously produced through coaxial double orifice microfluidic device and horseradish peroxidase mediated crosslinking, which conjugated introduce phenolic moieties in the backbone of silk fibroin and HA derivatives (Silk-Ph and HA-Ph, respectively). The iterative hybrid Silk-Ph + HA-Ph fibers were fabricated in tunable size distribution between 195 and 680 μm through control of outer flow velocity. Tensile strength and maximum stain of prepared Silk-Ph + HA-Ph sample upregulated more than three times higher than the single HA-Ph sample, which demonstrated significant impacts of synthesized silk derivative in hydrogel fiber composition. The proteolytic degradation of microfibers manipulated by hyaluronidase and collagenase treatment. Encapsulation process and crosslinking did not insert any harmful effect on cell viability (> 90%) and the cells maintained their growth ability after encapsulation process. Cellular filament-like tissue fabricated from proliferation of cells in Silk-Ph + HA-Ph microfiber.

Mesenchymal Stem Cell-Laden Hydrogel Microfibers for Promoting Nerve Fiber Regeneration in Long-Distance Spinal Cord Transection Injury

Mesenchymal stem cell (MSC)-based regenerative medicine is widely considered as a promising approach for repairing tissue and re-establishing function in spinal cord injury (SCI). However, low survival rate, uncontrollable migration, and differentiation of stem cells after implantation represent major challenges toward the clinical deployment of this approach. In this study, we fabricated three-dimensional MSC-laden microfibers via electrospinning in a rotating cell culture to mimic nerve tissue, control stem cell behavior, and promote integration with the host tissue. The hierarchically aligned fibrin hydrogel was used as the MSC carrier though a rotating method and the aligned fiber structure induced the MSC-aligned adhesion on the surface of the hydrogel to form microscale cell fibers. The MSC-laden microfiber implantation enhanced the donor MSC neural differentiation, encouraged the migration of host neurons into the injury gap and significantly promoted nerve fiber regeneration across the injury site. Abundant GAP-43- and NF-positive nerve fibers were observed to regenerate in the caudal, rostral, and middle sites of the injury position 8 weeks after the surgery. The NF fiber density reached to 29 ± 6 per 0.25 mm2 at the middle site, 82 ± 13 per 0.25 mm2 at the adjacent caudal site, and 70 ± 23 at the adjacent rostral site. Similarly, motor axons labeled with 5-hydroxytryptamine were significantly regenerated in the injury gap, which was 122 ± 22 at the middle injury site that was beneficial for motor function recovery. Most remarkably, the transplantation of MSC-laden microfibers significantly improved electrophysiological expression and re-established limb motor function. These findings highlight the combination of MSCs with microhydrogel fibers, the use of which may become a promising method for MSC implantation and SCI repair.

Cell fibers promote proliferation of co-cultured cells on a dish

This paper describes a co-culture method using cell fiber technology. Cell fibers are cell-laden hydrogel microfibers, in which cells are cultured three-dimensionally and allowed to reach more mature state than the conventional two-dimensional cell culture. Cells in the cell fibers are encapsulated by alginate shell. Only cellular secretome is released into the surrounding environment through the shell while the cells were retained by the fiber. With their high handleability and retrievability, we propose to use the cell fibers for co-culture to ensure steady supply of cellular secretome. We cultured mouse C2C12 myoblasts with mouse 3T3 fibroblasts encapsulated in the cell fibers for two days. The number of C2C12 cells increased proportionally to the number of co-cultured 3T3 fibers, suggesting that the secretome of 3T3 fibers promoted survival and proliferation of C2C12 cells. We believe that cell fiber technology is a useful tool for co-culturing cells, and it will contribute to both basic cell biology and tissue engineering with its unique features.

Microfluidics-Based Fabrication of Cell-Laden Hydrogel Microfibers for Potential Applications in Tissue Engineering.

Fibrous hydrogel scaffolds have recently attracted increasing attention for tissue engineering applications. While a number of approaches have been proposed for fabricating microfibers, it remains difficult for current methods to produce materials that meet the essential requirements of being simple, flexible and bio-friendly. It is especially challenging to prepare cell-laden microfibers which have different structures to meet the needs of various applications using a simple device. In this study, we developed a facile two-flow microfluidic system, through which cell-laden hydrogel microfibers with various structures could be easily prepared in one step. Aiming to meet different tissue engineering needs, several types of microfibers with different structures, including single-layer, double-layer and hollow microfibers, have been prepared using an alginate-methacrylated gelatin composite hydrogel by merely changing the inner and outer fluids. Cell-laden single-layer microfibers were obtained by subsequently seeding mouse embryonic osteoblast precursor cells (MC3T3-E1) cells on the surface of the as-prepared microfibers. Cell-laden double-layer and hollow microfibers were prepared by directly encapsulating MC3T3-E1 cells or human umbilical vein endothelial cells (HUVECs) in the cores of microfibers upon their fabrication. Prominent proliferation of cells happened in all cell-laden single-layer, double-layer and hollow microfibers, implying potential applications for them in tissue engineering.

Development of a perfusable 3D liver cell cultivation system via bundling-up assembly of cell-laden microfibers

Although the reconstruction of functional 3D liver tissue models invitro presents numerous challenges, it is in great demand for drug development, regenerative medicine, and physiological studies. Here we propose a new approach to perform perfusion cultivation of liver cells by assembling cell-laden hydrogel microfibers. HepG2 cells were densely packed into the core of sandwich-type anisotropic microfibers, which were produced using microfluidic devices. The obtained microfibers were bundled up and packed into a perfusion chamber, and perfusion cultivation was performed. We evaluated cell viability and functions, and also monitored the oxygen consumption. Furthermore, fibers covered with vascular endothelial cells were united during the perfusion culture, to form vascular network-like conduits between fibers. The presented technique can structurally mimic the hepatic lobule invivo and could prove to be a useful model for various biomedical research applications.

Improvement in the Mechanical Properties of Cell-Laden Hydrogel Microfibers Using Interpenetrating Polymer Networks.

Microencapsulation of cells is a promising technique in biomedical applications such as cell therapy. Recently, cell-laden hydrogel microfibers have been proposed as another shape microcapsule instead of microbeads; however, these are brittle with little stretching capability. This paper describes a cell-laden hydrogel microfiber that showed enhanced mechanical properties and handleability by using a double-network (DN) hydrogel consisting of alginate and polyacrylamide. The DN hydrogel microfiber supported approximately 6-fold higher strain and exhibited 10-fold higher tensile strength than the conventional alginate form. The DN hydrogel microfiber could also encapsulate pancreatic β cells while maintaining cell viability and function. The in vivo functionality of the DN hydrogel microfiber was demonstrated by transplanting 3D assemblies of the microfibers into the intraperitoneal or subcutaneous space of diabetic mice, which successfully decreased their blood glucose levels. Thus, cell-laden DN hydrogel microfibers may represent a promising material for various biomedical applications.