Abstract
BackgroundNeutrophils are the first responders in wound healing after injury that mediate pro- and anti-inflammatory activities i.a. through the formation of extracellular traps (NETs). However, excessive NETs presence in wound tissue can cause local hyperinflammation and -coagulation resulting in delayed wound healing. To improve wound healing, we aimed to examine the role of NETs and DNase1 on primary and secondary wound healing.MethodsThe study included 93 C57BL/6 mice, with 3 different genotypes: wildtype, Pad4-, and DNase1-Knockout (KO). Pad4-KO mice show limited NETs formation, while DNase1-KO mice cannot disintegrate them. All 3 genotypes were included in (1) a laparotomy group and (2) a thermal injury group. Animals in both groups either received DNase1 or a vehicle i.p. post wound induction and wound assessment and euthanasia were conducted. Laparotomy and burn scars were assessed using the stony brook scar evaluation scale and modified Yeong scale respectively. Tissue was analyzed histologically using H&E staining. Ly6g, Collagen I and III, SMA, and Fibrinogen were visualized and neutrophils activation (NE, MPO) and NETs (H3cit) formation assessed.ResultsAll animals survived with no complications. DNase1 treatment led to a significantly improved scar appearance in both groups, which was also seen in Pad4-KO mice. In the laparotomy group DNase1 improved collagen deposition and fibrin concentration was significantly reduced by DNase1 treatment. Markers of neutrophil activation were significantly reduced in the treatment and Pad4-KO group. In the thermal injury group wound closure time was significantly reduced after DNase1 treatment and in the Pad4-KO group. Even though inflammation remained high in the thermal injury model over time, neutrophil activation and NETs formation were significantly reduced by DNase1 treatment compared to controls.DiscussionPrimary and secondary intention wound healing is improved by targeting NETs through DNase1 treatment or genetic KO, as assessed by wound closure time and scar appearances. Additionally, wound stability was not affected by DNASE treatment. The results suggest that overall wound healing is accelerated and DNase1 appears to be a promising option to reduce scar formation; which should be evaluated in humans.
Highlights
Neutrophils are the first responders in wound healing after injury that mediate pro- and anti-inflammatory activities i.a. through the formation of extracellular traps (NETs)
Effects of Anti-neutrophils form extracellular traps (NETs) Treatment on Wound Healing Treatment with DNase1 led to a significantly improved Stony Brook Scar Evaluation Scale at 21 days post wound induction compared to controls that received a vehicle
A similar effect occurred in the Pad4-KO mice, which scored significantly better in the scar scale in comparison to wild type mice treated with the vehicle
Summary
Neutrophils are the first responders in wound healing after injury that mediate pro- and anti-inflammatory activities i.a. through the formation of extracellular traps (NETs). After an injury our innate immune system reacts within hours and days through an array of mechanisms which can exhibit both pro- and anti-inflammatory activities [1]. Neutrophils are the most abundant cell type in the circulatory system, are regarded as the first line of defense in the innate immune system, and constitute the main leukocytes involved in the early phase of wound healing [2, 3]. Neutrophiles can reverse transmigration and reenter the circulatory system after shifting their phenotype towards a proinflammatory state with a longer life span of about 5.4 days, leading to severe systemic inflammation [5]
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