Cellular and Functional Characterization of Microparticles in Sepsis-Associated Coagulopathy

Background: Disseminated intravascular coagulation (DIC) is a complex pathology characterized by microvascular thrombosis, dysregulated fibrinolysis, endothelial activation, and endothelial disruption. It is the endpoint of the coagulopathy commonly associated with sepsis and is clinically associated with multi-organ failure and/or significant bleeding. Inflammatory processes mediated by chemokines and cytokines including soluble CD40 ligand (sCD40L), interleukin 6 (IL-6), vascular endothelial growth factor (VEGF), endocan, and tissue factor play an important role in DIC pathogenesis; and in turn, dysregulation of coagulation factors effects further inflammation. Cellular damage and apoptotic processes result in the generation of microparticles, a diverse population of small, cell-derived phospholipid vesicles displaying characteristic surface proteins. Microparticles are associated with inflammation and coagulation in sepsis; however, the precise role they play remains unclear.

Methods: Plasma samples from 75 patients with confirmed DIC were included in this study. 50 plasma samples from normal healthy individuals were obtained from a commercial source (George King Biomedical, Overland Park, KS).  Endocan, VEGF, sCD40L, and IL-6 were measured using commercially available sandwich ELISA assays (Lunginnov, Lillie, France/R&D Systems, Minneapolis, MN). Microparticles, and microparticles expressing tissue factor were measured using commercially available functional assays (Hyphen Biomedical, Paris, France).

Results: All inflammatory biomarkers were significantly elevated in DIC MP-TF (DIC- 1.85±1.3 pg/ml; N-0.39.6±0.2 pg/ml, p<0.001) ,  sCD40L (DIC-618±571 pg/ml; N-177.6±221.4 pg/ml, p<0.00) VEGF (DIC->740 pg/ml; N-32.6±50.9 pg/ml, p=0.007),  endocan (DIC-10.6 ng/ml±10.9; N-1.81±0.36 ng/ml, p<0.001) and IL-6 (DIC-4968±5719 pg/ml; N-59.2±118.1 pg/ml, p<0.001).  Microparticles were elevated compared to normal (p<0.001) and correlated significantly with sCD40L (p=0.0005) and VEGF (0.039). sCD40L and VEGF also correlated significantly (p=0.001)

Discussion: The correlation between microparticles, sCD40L and VEGF found in these patients strongly supports the widespread activation of platelets in DIC. It is possible that this process contributes to the thrombocytopenia often seen in sepsis. In addition there was an elevation in both endocan and VEGF suggesting that, while endocan may have an effect on the level of VEGF in vivo, platelets appear to be the major source. Interestingly, microparticles specifically carrying tissue factor did not show correlation with any of the other biomarkers. This may be because these microparticles may be shed from multiple sources, particularly both activated monocytes and endothelial cells, and this may have been a confounding factor. Thus, these results provide further evidence of the widespread inflammatory milieu present in patients with DIC, particularly elucidating possible cross-talk between platelets and endothelium.  Profiling of these biomarkers may be helpful in the risk stratification of patients with sepsis associated coagulopathy and following their clinical progress.