All ETDs from UAB

Advisory Committee Chair

Jillian R Richter

Advisory Committee Members

Palaniappan Sethu

Brant Wagener

Document Type

Thesis

Date of Award

2018

Degree Name by School

Master of Science in Biomedical Engineering (MSBME) School of Engineering

Abstract

Trauma is the leading cause of death for Americans between the ages of 1 and 44. Implementation of massive transfusion protocols has improved early mortality due to traumatic hemorrhage; however, long-term complication rates associated with hemorrhagic shock remain high due to the development of multiple organ failure. In particular, resuscitation with large volumes of stored red blood cells (RBC) introduces high concentrations of free heme, which contributes to the development of acute lung injury (ALI) after trauma. Angiopoietin-2 (Agpt-2) is a mediator of trauma-related lung injury, but the effect of RBC storage-derived free heme on Agpt-2 regulation is unknown. The goal of this study was to determine (1) the effects of free heme formed during RBC storage on pulmonary microvascular endothelial cell (PMEC) function and Agpt-2 release and (2) the effects of PMEC-derived Agpt-2 on barrier function and inflammation of alveolar epithelial cells (AEC). To test this, RBC were collected from mice and stored for 14 days, during which time free heme levels doubled. PMEC barrier function was measured following treatment with fresh or stored RBC supernatant. The effects of hemin and Agpt-2 were determined by blocking with hemopexin (Hpx) and an Agpt-2 neutralizing antibody. PMEC barrier damage caused by stored RBC supernatant treatment was significantly attenuated with the addition of Hpx but not with the addition of an Agpt-2 neutralizing antibody, suggesting that PMEC barrier dysfunction caused by free heme is not mediated by Agpt-2. It is unclear whether Agpt-2 release from PMEC is mediated by heme following RBC supernatant treatment due to high variability and high background of samples via ELISA. Recombinant Agpt-2 treatment significantly upregulated inflammation in AEC; however, AEC barrier function was not affected by conditioned media from RBC-treated PMEC with low concentrations of Agpt-2, suggesting Agpt-2 is a mediator of AEC inflammation but effects on barrier function are unclear. More data are needed to determine the effects of stored blood transfusions on Agpt-2 release and the role of Agpt-2 in dysfunction of the lung-capillary barrier to develop therapies aimed at improving patient outcomes following trauma.

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