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Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold.

ABSTRACT: An artificial three-dimensional (3D) culture system that mimics the tumor microenvironment in vitro is an essential tool for investigating the cross-talk between immune and cancer cells in tumors. In this study, we developed a 3D culture system using an electrospun poly(?-caprolactone) (PCL) nanofibrous scaffold (NFS). A hybrid NFS containing an uninterrupted network of nano- and submicron-scale fibers (400 nm to 2 µm) was generated by deposition onto a stainless steel mesh instead of an aluminum plate. The hybrid NFS contained multiplanar pores in a 3D structure. Surface-seeded mouse CT26 colon cancer cells and bone marrow-derived dendritic cells (BM-DCs) were able to infiltrate the hybrid NFS within several hours. BM-DCs cultured on PCL nanofibers showed a baseline inactive form, and lipopolysaccharide (LPS)-activated BM-DCs showed increased expression of CD86 and major histocompatibility complex Class II. Actin and phosphorylated FAK were enriched where unstimulated and LPS-stimulated BM-DCs contacted the fibers in the 3D hybrid NFS. When BM-DCs were cocultured with mitoxantrone-treated CT26 cells in a 3D hybrid NFS, BM-DCs sprouted cytoplasm to, migrated to, synapsed with, and engulfed mitoxantrone-treated CT26 cancer cells, which were similar to the naturally occurring cross-talk between these two types of cells. The 3D hybrid NFS developed here provides a 3D structure for coculture of cancer and immune cells.

SUBMITTER: Kim TE

PROVIDER: S-EPMC4780731 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold.

Kim Tae-Eon TE Kim Chang Gun CG Kim Jin Soo JS Jin Songwan S Yoon Sik S Bae Hae-Rahn HR Kim Jeong-Hwa JH Jeong Young Hun YH Kwak Jong-Young JY

International journal of nanomedicine 20160302

An artificial three-dimensional (3D) culture system that mimics the tumor microenvironment in vitro is an essential tool for investigating the cross-talk between immune and cancer cells in tumors. In this study, we developed a 3D culture system using an electrospun poly(ε-caprolactone) (PCL) nanofibrous scaffold (NFS). A hybrid NFS containing an uninterrupted network of nano- and submicron-scale fibers (400 nm to 2 µm) was generated by deposition onto a stainless steel mesh instead of an aluminu ...[more]

PMID: 27042051

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Project description:Tissue engineering strategies have emerged in response to the growing prevalence of chronic musculoskeletal conditions, with many of these regenerative methods currently being evaluated in translational animal models. Engineered replacements for fibrous tissues such as the meniscus, annulus fibrosus, tendons, and ligaments are subjected to challenging physiologic loads, and are difficult to track in vivo using standard techniques. The diagnosis and treatment of musculoskeletal conditions depends heavily on radiographic assessment, and a number of currently available implants utilize radiopaque markers to facilitate in vivo imaging. In this study, we developed a nanofibrous scaffold in which individual fibers included radiopaque nanoparticles. Inclusion of radiopaque particles increased the tensile modulus of the scaffold and imparted radiation attenuation within the range of cortical bone. When scaffolds were seeded with bovine mesenchymal stem cells in vitro, there was no change in cell proliferation and no evidence of promiscuous conversion to an osteogenic phenotype. Scaffolds were implanted ex vivo in a model of a meniscal tear in a bovine joint and in vivo in a model of total disc replacement in the rat coccygeal spine (tail), and were visualized via fluoroscopy and microcomputed tomography. In the disc replacement model, histological analysis at 4 weeks showed that the scaffold was biocompatible and supported the deposition of fibrous tissue in vivo. Nanofibrous scaffolds that include radiopaque nanoparticles provide a biocompatible template with sufficient radiopacity for in vivo visualization in both small and large animal models. This radiopacity may facilitate image-guided implantation and non-invasive long-term evaluation of scaffold location and performance.Statement of significanceThe healing capacity of fibrous musculoskeletal tissues is limited, and injury or degeneration of these tissues compromises the standard of living of millions in the US. Tissue engineering repair strategies for the intervertebral disc, meniscus, tendon and ligament have progressed from in vitro to in vivo evaluation using a variety of animal models, and the clinical application of these technologies is imminent. The composition of most scaffold materials however does not allow for visualization by methods available to clinicians (e.g., radiography), and thus it is not possible to assess their performance in situ. In this work, we describe a radiopaque nanofibrous scaffold that can be visualized radiographically in both small and large animal models and serve as a framework for the development of an engineered fibrous tissue.

Project description:Engineered tendon grafts offer a promising alternative for grafting during the reconstruction of complex tendon tears. The tissue-engineered tendon substitutes have the advantage of increased biosafety and the option to customize their biochemical and biophysical properties to promote tendon regeneration. In this study, we developed a novel centrifugal melt electrospinning (CME) technique, with the goal of optimizing the fabrication parameters to generate fibrous scaffolds for tendon tissue engineering. The effects of CME processing parameters, including rotational speed, voltage, and temperature, on fiber properties (i.e. orientation, mean diameter, and productivity) were systematically investigated. By using this solvent-free and environmentally friendly method, we fabricated both random and aligned poly (L-lactic acid) (PLLA) fibrous scaffolds with controllable mesh thickness. We also investigated and compared their morphology, surface hydrophilicity, and mechanical properties. We seeded human adipose derived mesenchymal stem cells (HADMSC) on various PLLA fibrous scaffolds and conditioned the constructs in tenogenic differentiation medium for up to 21 days, to investigate the effects of fiber alignment and scaffold thickness on cell behavior. Aligned fibrous scaffolds induced cell elongation and orientation through a contact guidance phenomenon and promoted HADMSC proliferation and differentiation towards tenocytes. At the early stage, thinner scaffolds were beneficial for HADMSC proliferation, but the scaffold thickness had no significant effects on cell proliferation for longer-term cell culture. We further co-seeded HADMSC and human umbilical vein endothelial cells (HUVEC) on aligned PLLA fibrous mats and determined how the vascularization affected HADMSC tenogenesis. We found that co-cultured HADMSC-HUVEC expressed more tendon-related markers on the aligned fibrous scaffold. The co-culture systems promoted in vitro HADMSC differentiation towards tenocytes. These aligned fibrous scaffolds fabricated by CME technique could potentially be utilized to repair and regenerate tendon defects and injuries with cell co-culture and controlled vascularization.

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Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold.

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Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold.

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