Suppressing upregulation of fibrinogen after polytrauma mitigates thrombosis in mice

Abstract

Background: Polytrauma results in systemic inflammation and increased circulating fibrinogen, which increases the risk of microvascular and macrovascular thrombosis that contributes to secondary organ damage and venous thromboembolism (VTE). There are no clinically approved agents to prevent hyperfibrinogenemia after polytrauma. We hypothesized that preventing the increase in fibrinogen levels after polytrauma would suppress thrombosis.

Methods: Small-interfering ribonucleic acid (siRNA) against fibrinogen was encapsulated in lipid nanoparticles (siFibrinogen). Mice underwent a model of polytrauma and were then given varying doses of siFibrinogen, control siRNA, or no treatment. Fibrinogen was measured for 1 week via enxyme-linked immunosorbent assay (ELISA). To model postinjury VTE, the inferior vena cava was ligated 2 days after polytrauma in a portion of the mice. Thrombus weight was measured 48 hours after the inferior vena cava was ligated.

Results: Treatment with siFibrinogen prevented hyperfibrinogenemia after trauma without exacerbating the hypofibrinogenemic state that occurs in the acute injury period (1 hour). In treated groups, fibrinogen was significantly lower from 6 hours postinjury through the 7-day monitoring period. Maximal fibrinogen reduction was observed at 72 hours. Here, mice that received 2.0 mg/kg of siFibrinogen had 1% of normal values relative to untreated mice, and mice that received 1.0 or 0.5 mg/kg had 4%. Mice treated with siFibrinogen that underwent the postinjury VTE model had significantly reduced thrombus weight compared with control siRNA-treated animals. More notably, among all siFibrinogen treated mice, 12 of 18 were completely protected from thrombosis, compared with 0 of 9 displaying protection in the control group.

Conclusion: The rise of fibrinogen and the size of thrombi after polytrauma can be mitigated via the administration of siRNA against fibrinogen. siFibrinogen represents a promising novel target for VTE prophylaxis posttrauma.

Keywords: Fibrinogen; mice; polytrauma; siRNA; thrombosis.

Figure 1.. siFibrinogen mitigates the rise of fibrinogen after polytrauma in mice.

Mice (n=3 /group/timepoint) were given a single dose of varying concentrations of siFibrinogen. (A) Schematic of polytrauma model. (B) Time-course of fibrinogen levels up to 1 week after polytrauma relative to baseline. Data expressed as mean ± SEM and analyzed by 1-way ANOVA. * P < 0.05; @ P < 0.01; & P < 0.001, # P < 0.0001, ns = no significant difference.

Figure 2.. Suppressing excess fibrinogen production after polytrauma decreases thrombosis risk in mice.

After polytrauma, mice (n=5–9/ group) were given a single dose of varying concentrations of siFibrinogen or siLuciferase after polytrauma, then underwent IVC ligation to stimulate venous thrombosis. (A) Representative image of in situ thrombus in a mouse given 2.0 mg/kg siFibrinogen. (B) Representative image of in situ thrombus in a mouse given 2.0 mg/kg siLuciferase. Blue arrows point to IVC. (C-D) Fibrinogen concentration (C) and thrombus weight (D) at the time of euthanasia and clot harvest. Data expressed as mean ± SEM and analyzed by 1-way ANOVA. ** P < 0.01; **** P < 0.0001.