Antioxidant and antibacterial effects of human plasma-based scavenging protein cocktail on burn wound healing in vitro

In the United States, 400,000 or more fire or burn-related injuries are reported annually. Post-burn, there is a release of reactive oxygen species (ROS) as a result of hemolysis. Excess ROS causes oxidative stress that harms the surrounding healthy tissue and expands the wound area. Typically, plasma scavenger proteins such as haptoglobin, hemopexin, and transferrin help neutralize ROS by binding to free hemoglobin, heme, and iron, respectively. However, in severe burns, endogenous levels of scavenging proteins may not be sufficient to scavenge these hemolytic byproducts. Therefore, we evaluated the use of a human plasma-based protein cocktail containing haptoglobin, hemopexin, and transferrin as an exogenous source to reduce ROS-mediated injury. We show that human dermal fibroblasts injured by a sublethal dose of H 2 O 2 recover their ability to close a wound gap in an in vitro scratch assay. Furthermore, CellROX staining revealed a reduction in intracellular ROS levels following protein cocktail treatment. While immediate protein cocktail treatment post-H 2 O 2 significantly improved wound closure, this benefit was abolished if protein cocktail treatment was delayed by 1 or 4 hours. We also tested the ability of the protein cocktail to inhibit bacterial growth, and found that while it was not capable of doing so by itself, it potentiated E. Coli inhibition by the common antibiotic gentamicin. In conclusion, a protein cocktail containing haptoglobin, hemopexin, and transferrin is capable of both mitigating ROS-mediated injury and potentiating the inhibitory effect of antibiotics towards bacteria. This dual effect may find applications in the treatment of burn wounds to decrease burn wound expansion and infection. • ROS post-burn cause oxidative stress that harms the surrounding healthy tissue. • Endogenous scavenging proteins may not be sufficient to control the damage on time. • Human plasma-based protein cocktail (Hp-Hpx-Tf) reduced ROS-mediated injury. • Immediate cocktail treatment enhanced wound closure in the scratch assay. • Immediate cocktail treatment reduced the intracellular ROS levels. • Cocktail along with gentamicin enhanced bacterial inhibition under iron supplement.