1 Department of Biomedical Engineering, University of Memphis , Memphis, Tennessee.
2 Department of Microbiology, Immunology and Biochemistry, University of Tennessee Health Science Center , Memphis, Tennessee.
Tissue Eng Part A. 2017 Oct;23(19-20):1054-1063. doi: 10.1089/ten.TEA.2016.0452. Epub 2017 Feb 7.
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.
越来越多的证据表明，中性粒细胞是植入生物材料的第一反应者，通过分泌因子和通过 NETosis 释放中性粒细胞细胞外陷阱 (NETs)，为募集的免疫细胞启动微环境。在这项研究中，我们研究了电纺模板结构和组成在调节 NETosis 中的作用。使用小直径 (0.25-0.35 μm) 和大直径 (1.0-2.00 μm) 纤维制造了聚二恶烷酮 (PDO)、I 型胶原蛋白 (COL) 和混合 PDO-COL 模板 (PC) 的电纺模板。体外用人中性粒细胞评价了 3 小时和 24 小时的中性粒细胞-模板相互作用，并用体内 (大鼠皮下模型) 研究了 PDO 模板 1 天和 7 天。通过荧光显微镜和 On-cell Western 测定定量测定了模板结合的 NETs。体外结果表明，较大的纤维直径降低了 PDO 模板上的 NETosis，而 COL 的掺入则独立于纤维直径减弱了 NETosis。体内结果同样表明，在第 1 天，大直径 PDO 模板上的 NETs 程度较低，导致模板在第 7 天的组织整合程度较差。相比之下，在体内 24 小时大量 NETs 覆盖的小直径 PDO 模板，在第 7 天被胶囊样组织包围。这些初步的体内结果验证了体外模型，并表明 NETosis 可能是一种重要的生理反应，也是固有免疫反应对模板的关键预处理事件。总之，这些结果表明，在设计能够促进原位再生的模板时，对中性粒细胞与生物材料的急性对抗进行特征描述是非常重要的。
