Electrospun Template Architecture and Composition Regulate Neutrophil NETosis In Vitro and In Vivo

• Published in: Tissue engineering. Part A Vol. 23; no. 19-20; pp. 154 - 1063
• Main Authors: Fetz, Allison E., Neeli, Indira, Rodriguez, Isaac A., Radic, Marko Z., Bowlin, Gary L.
• Format: Journal Article
• Language: English
• Published: United States Mary Ann Liebert, Inc 01.10.2017
• Subjects: Angiogenesis; Animals; Biochemistry; Biomaterials; Biomedical engineering; Biomedical materials; Collagen; Collagen (type I); Electrospinning; Emergency medical services; Extracellular Traps - metabolism; Female; Fluorescence; Genotype & phenotype; Humans; Immune response; Immune system; Immunology; Inflammation; Innate immunity; Leukocytes (neutrophilic); Male; Microscopy; Neutrophils; Neutrophils - metabolism; Polydioxanone - chemistry; Preconditioning; Proteins; Rats, Sprague-Dawley; Special Focus Articles; Staining and Labeling; Tissue engineering; Tissue Engineering - methods; Tissue Scaffolds - chemistry; United States > US; Tennessee; tissue regeneration; NETosis; electrospinning; innate immune response; neutrophils; templates
• ISSN: 1937-3341; 1937-335X
• DOI: 10.1089/ten.tea.2016.0452

Mounting evidence indicates that neutrophils, first responders to an implanted biomaterial, prime the microenvironment for recruited immune cells by secreting factors and releasing neutrophil extracellular traps (NETs) through NETosis. In this study, we investigated the role of electrospun template architecture and composition in regulating NETosis. Electrospun polydioxanone (PDO), collagen type I (COL), and blended PDO-COL templates (PC) were fabricated with small-diameter (0.25–0.35 μm) and large-diameter (1.0–2.00 μm) fibers. Neutrophil–template interactions were evaluated in vitro for 3 and 24 h with human neutrophils, and the PDO templates were studied in vivo (rat subcutaneous model) for 1 and 7 days. Template-bound NETs were quantified by fluorescent microscopy and an On-cell Western assay. The in vitro results indicate that larger fiber diameters reduced NETosis on PDO templates, whereas the incorporation of COL attenuated NETosis independent of fiber diameter. The in vivo results similarly revealed a lower degree of NETs on large-diameter PDO templates at 1 day, resulting in marginal tissue integration of the templates at 7 days. In contrast, the small-diameter PDO templates, which were coated in a large amount of NETs at 24 h in vivo , were surrounded by capsule-like tissue at 7 days. These preliminary in vivo results validate the in vitro model and signify NETosis as a potentially significant physiological response and a critical preconditioning event for the innate immune response to templates. In conclusion, these results demonstrate the importance of characterizing the neutrophil's acute confrontation with biomaterials to engineer templates capable of promoting in situ regeneration.