Inhibition of early plasmin activation following thermal injury attenuates systemic inflammatory markers and inflammation within injured muscle

Abstract

Introduction Burn injuries drive systemic inflammation, increasing the risk of organ dysfunction, and impeding the repair of injured tissues. Traumatic injuries provoke excess activation of plasmin, the protease responsible for degrading fibrin clots, causing bleeding complications. Although burn injuries rarely carry bleeding risks, excess plasmin activity can drive inflammatory signaling and vascular permeability, suggesting plasmin contributes to burn-induced inflammation. Therefore, we hypothesized that excess plasmin activation provoked by burn injury exacerbates inflammation at the site of tissue injury and systemically. Methods A validated, murine burn model in combination with muscle injury was employed to assess the study hypothesis. Sham animals received only calf muscle injuries. Mice expressing an NF-κB-inducible luciferase reporter were used to quantify inflammatory signaling by bioluminescence in tissues. To inhibit plasmin activity, mice received plasminogen antisense oligonucleotide (Plg ASO, IONIS Pharmaceuticals) before injury, or antifibrinolytic drug tranexamic acid (TXA) administered for 12 hours post-burn. Controls received control (Ctl) ASO or saline. Plasmin activation was measured by circulating complexes of plasmin and its inhibitor α2-antiplasmin (PAP). Inflammation was measured serially by plasma interleukin-6 (IL-6), neutrophil to lymphocyte ratio (NLR), and NF-κB activity in the injured muscle. Results Burn injury resulted in a 3-5 fold increase in plasmin activity measured by PAP(P<0.05) compared with sham animals and provoked a significant increase in plasma IL-6(P<0.005) and NLR(P<0.05) within 6-12 hours post-injury, at levels consistent with transient systemic inflammation. In mice without burns, muscle injury alone provoked NF-κB activity from 1-3 days post-injury with resolution by 5 days. With the addition of the burn injury, NF-κB activity within the injured calf muscle was significantly elevated between 1-3 days post-injury (P<0.01) and prolonged in the injured muscle (Figure 1). When plasmin was reduced with Plg ASO and TXA treatments, plasma IL-6 and NLR levels were reduced within 24 hours post-burn (P<0.05), and NF-κB activity within muscle was significantly reduced between 1-3 days post-burn (P<0.05)(Figure 1). Conclusion Burns provoke systemic and augment local inflammation in a muscle injury remote to the burn site. Inhibition of early plasmin activation after burn reduces systemic inflammatory markers and local inflammation within injured tissues. Significance Burn injuries incite significant global inflammation, increasing the risk of life-threatening complications, and chronic comorbidities. Because plasma drives inflammation after acute injury, it may be a therapeutic target to reduce tissue inflammation. However, plasmin is essential for fibrinolysis and tissue repair, and the beneficial effects of inhibiting plasmin following burn are likely time-dependent.