Fibrin-derived Peptide Bβ15-42 and Earthworm Fibrinolytic Enzyme A Inhibit Atherosclerosis and Inflammation via the VE-cadherin Pathway

Elevated fibrinogen increases risk for cardiovascular and cerebrovascular diseases. The biological effects of the fibrin-derived peptide Bβ15–42 are different from fibrin and promote vascular and anti-inflammatory effects. Fragments of fibrinogen cleavage by earthworm fibrinolytic enzyme A (EFEa) are structurally similar to Bβ15–42. Therefore, we evaluated if Bβ15–42 and EFEa have anti-atherosclerotic effects. To investigate the anti-atherosclerotic effect of Bβ15–42 and EFEa, we used New Zealand rabbits fed a high cholesterol diet to promote atherosclerosis. Human umbilical vascular endothelial cells with high expression of VE-cadherin were used to determine the mechanism of action of Bβ15–42 and EFEa to inhibit the deleterious effect of fibrin. Both Bβ15–42 and EFEa significantly reduced atherosclerosis in rabbits fed a high cholesterol diet. Vascular fibrinogen deposition and inflammatory cell aggregation was significantly reduced in rabbits treated with Bβ15–42 or EFEa. In addition, Bβ15–42 and EFEa stabilized the structure of endothelial cells and decreased inflammatory cell migration. Bβ15–42 and EFEa protected endothelial cell function by reducing the effect of fibrinogen in the VE-cadherin pathway. Therefore, the fibrin-derived Bβ15–42 peptide exhibited anti-atherosclerotic effects and reduced vascular fibrinogen deposition and inflammatory cell aggregation in the aorta. Furthermore, EFEa has similar Bβ15-42-like anti-atherosclerotic effects.

Platelet-derived TLT-1 is a prognostic indicator in ALI/ARDS and prevents tissue damage in the lungs in a mouse model

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) affect >200 000 individuals yearly with a 40% mortality rate. Although platelets are implicated in the progression of ALI/ARDS, their exact role remains undefined. Triggering receptor expressed in myeloid cells (TREM)–like transcript 1 (TLT-1) is found on platelets, binds fibrinogen, and mediates clot formation. We hypothesized that platelets use TLT-1 to manage the progression of ALI/ARDS. Here we retrospectively measure plasma levels of soluble TLT-1 (sTLT-1) from the ARDS Network clinical trial and show that patients whose sTLT-1 levels were >1200 pg/mL had nearly twice the mortality risk as those with <1200 pg/mL (P < .001). After correcting for confounding factors such as creatinine levels, Acute Physiology And Chronic Health Evaluation III scores, age, platelet counts, and ventilation volume, sTLT-1 remains significant, suggesting that sTLT-1 is an independent prognostic factor (P < .0001). These data point to a role for TLT-1 during the progression of ALI/ARDS. We use a murine lipopolysaccharide-induced ALI model and demonstrate increased alveolar bleeding, aberrant neutrophil transmigration and accumulation associated with decreased fibrinogen deposition, and increased pulmonary tissue damage in the absence of TLT-1. The loss of TLT-1 resulted in an increased proportion of platelet-neutrophil conjugates (43.73 ± 24.75% vs 8.92 ± 2.4% in wild-type mice), which correlated with increased neutrophil death. Infusion of sTLT-1 restores normal fibrinogen deposition and reduces pulmonary hemorrhage by 40% (P ≤ .001) and tissue damage by 25% (P ≤ .001) in vivo. Our findings suggest that TLT-1 uses fibrinogen to govern the transition between inflammation and hemostasis and facilitate controlled leukocyte transmigration during the progression of ARDS.

9. Investigating the relationship between Fibrinogen deposition and myelomonocytic cell infiltration in EAE Spinal Cord

Multiple sclerosis and its animal model, experimental autoimmune encephalitis (EAE), are primarily characterized by inflammatory degeneration. It has also been shown that deposition of fibrinogen from the blood in spinal cord tissue increased in MS and EAE. Fibrinogen is a substrate molecule found in the blood for a receptor called CD11 expressed on the surface of both brain and blood immune cells. Using a technique to light up specific tissue (called immunofluorescence), we demonstrate that there is a positive correlation between levels of fibrinogen deposition in spinal cord and the progression of clinical symptoms. Contrary to previous research, our past work indicated that brain immune cells play a limited role in diseaseprogression, while immune cells in the blood are critical. Thus an alternative interpretation of these results may be that blocking fibrinogen deposition within the spinal cord during EAE reduces the infiltration of blood immune cells into the spinal cord. Investigating this experimentally, however, is complicated by the inability to distinguish blood and brain resident immune cells. To circumvent this, we use parabiosisirradiation-separation models to investigate the replacement of circulating immune cells, which leads to the dynamic assessment of blood immune cell infiltration in spinal cord. We have used fibrinogen deposits during EAE, thereby successfully identifying a pathological component, and a immunofluorescence tovisualize fluorescent blood cells within spinal cord and their relationship to therapeutic target for multiple sclerosis.