Designing Drug-eluting Regeneration Templates to Down-regulate Neutrophil Extracellular Traps

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The proposal aims to examine the ability of N-α-benzoyl-N5-(2-chloro-1-iminoethyl)-L-ornithine amide (Cl-amidine) eluting electrospun templates in regulating the level of neutrophil extracellular traps (NETs) which have been shown to be induced in acute interacting neutrophils. The templates will be electrospun such that the resulting architectures vary in fiber diameter, pore size, drug loading, and surface chemistry to create diverse microenvironments to allow examination of the role of each variable in regulating the NET formation and subsequent host response. The overriding hypothesis is that the degree of NETosis (or the level of template preconditioning NETs) can be down regulated by the inclusion of Cl-amidine impregnation and/or adsorption onto the electrospun fibers and that the eluted drug functioning to minimize NET formation will lead to a desired, accelerated marginal tissue healing and enhanced tissue integration of the templates. More specifically, Aim 1 will define the role of the varied surface- area-to-volume ratios, degradation rates, surface chemistries, and Cl-amidine loading by fiber impregnation during electrospinning and/or template post-processing surface adsorption, characterize the in vitro release profiles, and finally determine the in vitro functionality in inhibiting NETosis. Aim 2 will then determine the Cl-amidine loaded PDO templates capacity to down- regulate the degrees of NETosis in vivo and determine the tissue healing/integration outcomes associated with the Cl-amidine down-regulated degree of NETosis in a rat subcutaneous model. It is anticipated that increasing template fiber diameters/pore sizes and altering surface chemistry with a significant burst drug release capacity will invoke a minimal degree of NETs, favoring tissue regeneration. We anticipate that this study will begin to break new ground regarding neutrophil- template interaction, and more critically, provide evidence for additional template-derived functionality (Cl-amidine elution) in down-regulating a potential initiator of chronic inflammation and impaired tissue healing. This functionality is potentially a critical missing component that may provide a means for tissue engineers to more efficiently harness the innate immune response (acute template preconditioning) that will lead to the enhancement of in situ regeneration, or more precisely, minimize failure derived by chronic inflammation and fibrosis.