Effectiveness of Antioxidants in Treating Severely Injured Patients.

Hemorrhagic shock and ischemia-reperfusion (IR) injury present in military field and civilian emergency situations with several clinical and logistical challenges. Hemorrhagic shock is a leading cause of death in military combat and civilian trauma. It is the major cause of early death of injured soldiers accounting for about 50% of the deaths.

IR injury is one of the most common types of cell injuries that occur in a variety of clinical and military relevant field conditions, such as, coronary arterial disease, cardiopulmonary bypass, occlusive arterial disease, stroke, tourniquet application and re-plantation of amputated parts. In general, IR represents an acute inflammatory response after a restriction in blood supply event and subsequent restoration of blood flow.

It is well known that exposure of oxygen deprived tissues to oxygen during IR can lead to organ damage (e.g., the release of a tourniquet) because the sudden reintroduction of oxygen results in a burst of reactive oxygen/nitrogen species, that is, oxidative stress.

Oxidative stress, or an adverse biochemical event, is associated with tissues damaged produced by IR injury. Reducing oxidative stress associated with IR injury subsequent to hemorrhagic shock, tourniquet application, or other combat/civilian situations by using a clinical regimen of antioxidants, including a natural occurring body chemical, glutathione (GSH), is a reasonable science-based approach. Through early intervention with GSH or similar chemicals, we would have the potential of preventing injury morbidity and mortality.

Our objective is to determine the feasibility of using a precursor chemical for GSH in modulating the oxidative damage subsequent to IR injury. By preloading tissues with this GSH precursor chemical, we should be able to stimulate tissue to synthesize GSH which should prevent IR injury to such tissues.

Hemorrhagic shock is a condition of reduced blood delivery to vital organs with subsequent inadequate provision of oxygen and nutrients required for normal tissue and cellular function. Effective hemorrhage control and optimal resuscitation are required in the management of the severely injured patients; however, hemorrhagic shock/resuscitation can be viewed as a global oxygen deprivation/re-oxygenation injury.

For example, in hemorrhagic shock oxygen deprivation in various organs is followed by re-oxygenation during therapeutic intervention, i.e., ischemia-reperfusion (IR) injury. IR injury is one of the most common types of cell injuries that occur in a variety of clinical and military relevant field conditions, such as, coronary arterial disease, cardiopulmonary bypass, occlusive arterial disease, stroke, tourniquet application and re-plantation of amputated parts.

It is well known that exposure of oxygen deprived tissues to oxygen during reperfusion (eg., the release of a tourniquet) can lead to organ damage because the sudden reintroduction of oxygen results in a burst of reactive oxygen/nitrogen species (ROS/RNS). The step by step development of damage is associated with oxidative stress and commitment damages in macromolecules, such as; lipid, proteins, and nucleic acids, which subsequently lead to cytoskeletal damage, mitochondrial dysfunction and altered signal transduction.

Antioxidant depletion from within the damaged tissue, particularly, glutathione (GSH) results in disruption of cellular stability and programed cell death. Enzymatic and non-enzymatic antioxidants can protect tissue against oxidative damage; however, following ROS/RNS mediated tissue injury, such antioxidants may not afford adequate defense.

GSH is the most abundant antioxidant in a cellular system and often has been referred to as the first line of defense in preventing the deleterious effects of oxidative stress. When provided as a treatment, GSH and N-acetyl-cysteine (a GSH precursor) supplements have been reported to be protective in exercised-induced oxidative stress and gamma-glutamylcysteine ethyl ester is effective in increasing the nerve cell’s power plant levels of GSH. Likewise, GSH levels decrease in tissue injury and increasing the amount of GSH in the cell as a means to modulate oxidative brain injury has been a topic of interest.

Internal GSH plays an important protective role against ROS/RNS and administration of L-buthionine-(S,R)sulfoximine (BSO) an inhibitor of endogenous GSH, worsen experimental spinal cord injury. However, the use of exogenous GSH has not been totally successful because of difficulties in gut availability and the lack of translocation of GSH into tissue (such as the brain because of blood brain barrier).

GSH is a unique tripeptide compose of the amino acids; glutamate, cysteine, and glycine. Since the synthetic capacity of GSH in tissues subjected to oxygen deprivation has not been compromised, the use of substrates or GSH precursors is of current interest.

Gamma-glutamylcysteine (GGC) can be used as a GSH precursor, because the feedback inhibition of gamma-glutamylcysteine synthetase by GSH can be circumvented and this dipeptide can be taken up into cells and directly used as a substrate for GSH synthetase. Also, injection of GGC ester into tissues causes an increase in GSH levels.

This project will determine the feasibility of using a precursor chemical for glutathione in modulating the oxidative damage subsequent to ischemia-reperfusion injury.