Effect of chitosan and coagulation factors on the wound repair phenotype of bioengineered blood clots

Controlling the blood clot phenotype in a surgically prepared wound is an evolving concept in scaffold-guided tissue engineering. Here, we investigated the effect of added chitosan (80% or 95% Degree of Deacetylation, DDA) or coagulation factors (recombinant human Factor VIIa, Tissue Factor, thrombin) on inflammatory factors released by blood clots. We tested the hypothesis that 80% DDA chitosan specifically enhances leukotriene B4 (LTB4) production. Human or rabbit whole blood was combined with isotonic chitosan solutions, coagulation factors, or lipopolysaccharide, cultured in vitro at 37 °C, and after 4 hours the serum was assayed for LTB4 or inflammatory factors. Only 80% DDA chitosan clots produced around 15-fold more LTB4 over other clots including 95% DDA chitosan clots. All serum contained high levels of PDGF-BB and CXCL8. Normal clots produced very low type I cytokines compared to lipopolysaccharide clots, with even lower IL-6 and IL-12 and more CCL3/CCL4 produced by chitosan clots. Coagulation factors had no detectable effect on clot phenotype. Conclusion In blood clots from healthy individuals, 80% DDA chitosan has a unique influence of inducing more LTB4, a potent neutrophil chemoattractant, with similar production of PDGF-BB and CXCL8, and lower type I cytokines, compared to whole blood clots.