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Bacterial Infection-Mimicking Three-Dimensional Phagocytosis and Chemotaxis in Electrospun Poly(ε-caprolactone) Nanofibrous Membrane.

ABSTRACT: In this study, we developed a three-dimensional (3D) in vitro infection model to investigate the crosstalk between phagocytes and microbes in inflammation using a nanofibrous membrane (NM). Poly(ε-caprolactone) (PCL)-NMs (PCL-NMs) were generated via electrospinning of PCL in chloroform. Staphylococcus aureus and phagocytes were able to adhere to the nanofibers and phagocytes engulfed S. aureus in the PCL-NM. The migration of phagocytes to S. aureus was evaluated in a two-layer co-culture system using PCL-NM. Neutrophils, macrophages and dendritic cells (DCs) cultured in the upper PCL-NM layer migrated to the lower PCL-NM layer containing bacteria. DCs migrated to neutrophils that cultured with bacteria and then engulfed neutrophils in two-layer system. In addition, phagocytes in the upper PCL-NM layer migrated to bacteria-infected MLE-12 lung epithelial cells in the lower PCL-NM layer. S. aureus-infected MLE-12 cells stimulated the secretion of tumor necrosis factor-α and IL-1α in 3D culture conditions, but not in 2D culture conditions. Therefore, the PCL-NM-based 3D culture system with phagocytes and bacteria mimics the inflammatory response to microbes in vivo and is applicable to the biomimetic study of various microbe infections.

SUBMITTER: Lee SJ

PROVIDER: S-EPMC8399938 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Tissue engineering has become a promising therapeutic approach for bone regeneration. Nanofibrous scaffolds have attracted great interest mainly due to their structural similarity to natural extracellular matrix (ECM). Poly(lactide-co-?-caprolactone) (PLCL) has been successfully used in bone regeneration, but PLCL polymers are inert and lack natural cell recognition sites, and the surface of PLCL scaffold is hydrophobic. Silk fibroin (SF) is a kind of natural polymer with inherent bioactivity, and supports mesenchymal stem cell attachment, osteogenesis, and ECM deposition. Therefore, we fabricated hybrid nanofibrous scaffolds by adding different weight ratios of SF to PLCL in order to find a scaffold with improved properties for bone regeneration.Hybrid nanofibrous scaffolds were fabricated by blending different weight ratios of SF with PLCL. Human adipose-derived stem cells (hADSCs) were seeded on SF/PLCL nanofibrous scaffolds of various ratios for a systematic evaluation of cell adhesion, proliferation, cytotoxicity, and osteogenic differentiation; the efficacy of the composite of hADSCs and scaffolds in repairing critical-sized calvarial defects in rats was investigated.The SF/PLCL (50/50) scaffold exhibited favorable tensile strength, surface roughness, and hydrophilicity, which facilitated cell adhesion and proliferation. Moreover, the SF/PLCL (50/50) scaffold promoted the osteogenic differentiation of hADSCs by elevating the expression levels of osteogenic marker genes such as BSP, Ocn, Col1A1, and OPN and enhanced ECM mineralization. In vivo assays showed that SF/PLCL (50/50) scaffold improved the repair of the critical-sized calvarial defect in rats, resulting in increased bone volume, higher trabecular number, enhanced bone mineral density, and increased new bone areas, compared with the pure PLCL scaffold.The SF/PLCL (50/50) nanofibrous scaffold facilitated hADSC proliferation and osteogenic differentiation in vitro and further promoted new bone formation in vivo, suggesting that the SF/PLCL (50/50) nanofibrous scaffold holds great potential in bone tissue regeneration.

Project description:Recent studies have shown the potential of artificially synthesized conduits in the repair of peripheral nerve injury. Natural biopolymers have received much attention because of their biocompatibility. To investigate the effects of novel electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits (biopolymer nanofiber conduits) on the repair of peripheral nerve injury, we bridged 10-mm-long sciatic nerve defects with electrospun absorbable biopolymer nanofiber conduits, poly(ε-caprolactone) or silicone conduits in Sprague-Dawley rats. Rat neurologica1 function was weekly evaluated using sciatic function index within 8 weeks after repair. Eight weeks after repair, sciatic nerve myelin sheaths and axon morphology were observed by osmium tetroxide staining, hematoxylin-eosin staining, and transmission electron microscopy. S-100 (Schwann cell marker) and CD4 (inflammatory marker) immunoreactivities in sciatic nerve were detected by immunohistochemistry. In rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits, no serious inflammatory reactions were observed in rat hind limbs, the morphology of myelin sheaths in the injured sciatic nerve was close to normal. CD4 immunoreactivity was obviously weaker in rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits than in those subjected to repair with poly(ε-caprolactone) or silicone. Rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits tended to have greater sciatic nerve function recovery than those receiving poly(ε-caprolactone) or silicone repair. These results suggest that electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits have the potential of repairing sciatic nerve defects and exhibit good biocompatibility. All experimental procedures were approved by Institutional Animal Care and Use Committee of Taichung Veteran General Hospital, Taiwan, China (La-1031218) on October 2, 2014.

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Bacterial Infection-Mimicking Three-Dimensional Phagocytosis and Chemotaxis in Electrospun Poly(ε-caprolactone) Nanofibrous Membrane.

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