Impaired Hemostasis Caused By Deficit of Tissue Factor-Triggered Blood Coagulation Leads to Alveolar Hemorrhages and Increased Mortality in Influenza A Infection

Objective: Viral infections lead to activation of coagulation which enhances inflammatory responses. For instance, influenza A is associated with activation of coagulation and the increased risk for thrombotic events, such as myocardial infarction. Here we investigated the role of the coagulation cascade and platelets in influenza A virus (IAV) infection in mice.

Methods: To investigate the role of the coagulation cascade in influenza infection, we examined the effect of either low levels of tissue factor or wild-type mice that were administered with 4 μg/mL of warfarin via drinking water. To study the role of platelets in influenza infection, we examined protease-activated receptor 4 (PAR-4, the main functional receptor on mouse platelets) deficient mice or wild-type mice treated with 0.15 mg/mL of clopidogrel or 0.4 mg/mL of aspirin via drinking water. All mice were infected intranasally with mouse adapted influenza A H1N1/PR8 (IAV). Changes in body weights and survival were analyzed up to 14 days after infection. In addition, bronchoalveolar lavage fluid (BALF) or lungs were collected and analyzed 7 or 14 days after virus challenge.

Results: LowTF mice exhibited higher mortality rate and increased loss of body weights compared to littermate control mice after IAV infection (P<0.01). At day 7, BALF of LowTF was red and had increased red blood cell numbers and hemoglobin levels compared to infected control mice (P<0.001). Thrombin-antithrombin complex (TAT) levels in the BALF of infected LowTF mice were lower than that of control mice. In addition, lung histology revealed that LowTF mice had extensive hemorrhages. Similarly, mice treated with warfarin and infected with IAV showed higher mortality compared with the warfarin-treated mice without infection or IAV-infected mice without drugs. Warfarin treated mice also had red BALF, higher red blood cell number and hemoglobin levels in the BALF, and lung bleeding. PAR-4 deficient mice exhibited visible lung hemorrhages with increased hemoglobin levels in the BAL (P<0.01) and increased mortality compared to controls (P<0.05), although the bleeding was less than LowTF mice. Treatment of mice with clopidogrel or aspirin did not increased lung bleeding or mortality.

Conclusion: Pulmonary hemostasis requires both TF-mediated activation of blood coagulation and thrombin activation of platelets during IAVinfection. Use of antithrombotics during IAV infection may increase the risk of lung bleeding.