Serum from rats subjected to traumatic brain injury and hemorrhagic shock induces profound endothelial barrier dysfunction

Abstract

IntroductionTraumatic brain injury (TBI) has been associated with development of indirect acute respiratory distress syndrome (ARDS). However, the causative relationship between TBI and lung injury remains unclear. To explore potential mechanism linking TBI with development of ARDS, we characterized effects of serum factors released during TBI and hemorrhagic shock (HS) in a rat model on the pulmonary endothelial cell (EC) barrier dysfunction, a key feature of ARDS.MethodsThe adult male rat polytrauma model consisted of controlled cortical impact (CCI)–induced TBI followed by 30 minutes of hemorrhage shock (HS) induced by blood withdrawal. The HS phase was followed by a 1‐hour “prehospital” Hextend fluid resuscitation phase and then a 1‐hour “hospital phase” when shed blood was reinfused. 48 hours after the TBI, blood was drawn from right ventricle and serum samples were obtained. Serum was collected from the TBI or sham rats, and its effects on barrier properties of pulmonary endothelial cell culture were assessed by measurements of transendothelial electrical resistance (TER) using the electrical cell impedance sensor and visualization of fluorescent tracer penetration through EC monolayer using XPerT assay.ResultsWe found pronounced barrier‐disruptive effects of serum samples from animals exposed to controlled cortical impact (CCI) and HS, but not from sham operated rats. Rapid and sustained decrease in EC monolayer transendothelial resistance was observed within minutes after serum addition and remained below basal TER levels after incubation with CCI‐HS sera. Thrombin has been implicated in the early‐phase barrier disruptive activity of CCI‐HS serum, as both thrombin inhibitor and thrombin receptor antagonist attenuated the acute phase of serum‐induced TER decline, but did not affect TER decline at later time points. These findings suggest that thrombin elevation may play a role in early‐phase barrier disruptive activity of CCI‐HS serum. However, both the early and late phases of CCI‐HS‐induced EC permeability were inhibited by heparin. The late phase barrier disruptive effect of CCI‐HS was also prevented by serum preincubation with heparin‐sepharose. Pulmonary EC treated with serum from CCI‐HS rats for 3 hours have demonstrated significant decline in expression of EC junctional protein, VE‐Cadherin, and disassembly of peripheral EC adherens junction complexes monitored by immunostaining with VE‐cadherin antibody.ConclusionThese results suggest that exposure to CCI‐HS leads to production of humoral factors causing early and late‐phase barrier disruptive effects in lung vascular endothelium. While thrombin‐PAR1 signaling has been identified as mechanism of acute EC permeability increase by CCI‐HS serum, the factor(s) defining long‐term EC barrier disruption in CCI‐HS model remain to be determined.