S. A. Olson1, B. K. Osborn1, M. E. Cotton1, J. D. Krocker1, J. C. Cardenas1 1University Of Texas Health Science Center At Houston, McGovern Medical School, Department Of Surgery, Center For Translational Injury Research, Houston, TX, USA
Introduction: Trauma-induced coagulopathy (TIC) is a condition associated with both early hemorrhage-related deaths and late-stage deaths due to multiple organ failure (MOF). While improvements to hemostatic resuscitation have significantly reduced hemorrhage-related deaths, the incidence of MOF among TIC patients remains high. Disruption of vascular endothelial cells (EC) is a unifying mechanism in the development of MOF following severe trauma, and hyperfibrinolysis, defined as accelerated clot breakdown, is a key driver of TIC. Fragment X is a fibrinogen degradation product released during hyperfibrinolysis. The goal of this project was to determine the effects of fragment X on EC expression of paracellular junctional proteins and barrier function.
Methods: Human pulmonary microvascular cells (HPMECs) were treated with increasing concentrations of fragment X (1, 10, and 100 μg/mL), and barrier function was monitored over 24 hours using the xCELLigence live-cell monitoring system. Quantitative PCR (qPCR) was performed to measure changes in EC expression of gene targets including both tight (occludin) and adherens (VE-cadherin) junctional proteins compared to vehicle-treated control. Immunofluorescent cytostaining was done to validate qPCR findings followed by visualization by fluorescent microscopy.
Results: Fragment X treatment resulted in significant increases in endothelial permeability over time (p<0.05, Figure 1A). There was a significant reduction in VE-cadherin mRNA expression in fragment X-treated HPMECs compared to control (p=0.0107, Figure 1B). Additionally, immunofluorescent cytostaining and subsequent visualization by fluorescent microscopy confirmed decreased levels of VE-cadherin following HMPEC fragment X treatments.
Conclusion: Fragment X induces EC hyperpermeability by reducing VE-cadherin expression. This suggests that disrupting EC-fragment X interaction can mitigate EC barrier disruption and organ edema, allowing us to next identify a targeted approach to prevent fragment X from engaging ECs following trauma, thus preventing MOF.
