Institute of Biomedical Engineering, Ecole Polytechnique, Montréal, Quebec, Canada H3C 3A7.
J Biomed Mater Res A. 2010 Jun 15;93(4):1429-41. doi: 10.1002/jbm.a.32638.
Chitosan is a polycationic and biocompatible polysaccharide composed of glucosamine and N-acetyl glucosamine that is chemotactic for neutrophils and stimulates wound repair through mechanisms that remain unclear. It was previously shown that chitosan depletes complement proteins from plasma, suggesting that chitosan activates complement. Complement activation leads to cleavage of C5 to produce C5a, a neutrophil chemotactic factor. Here, we tested the hypothesis that chitosan generates C5a in human whole blood, citrated plasma, and serum. C5a fragment appeared in coagulating whole blood, and mixtures of chitosan-glycerol phosphate/whole blood, in parallel with platelet and thrombin activation. However, in plasma and serum, thrombin and chitosan-GP failed to generate C5a, although native C3, C5, and factor B adsorbed noncovalently to insoluble chitosan particles incubated in citrated plasma, serum, EDTA-serum and methylamine-treated plasma. By surface plasmon resonance, pure C3 adsorbed to chitosan. The profile of serum factors associating with chitosan was consistent with a model in which anionic blood proteins with a pI lower than the pK(0) 6.78 of chitosan (the upper limit of chitosan pK(a)) associate electrostatically with cationic chitosan particles. Zymosan, a yeast ghost particle, activated complement in serum and citrated plasma, but not in EDTA-serum or methylamine plasma, to generate fluid-phase C5a, while C3b formed covalent cross-links with zymosan-associated proteins and became rapidly cleaved to iC3b, with factor Bb stably associated. These data demonstrate that chitosan is a nonreactive biomaterial that does not directly activate complement, and provide a novel basis for predicting anionic serum protein-chitosan interactions.
壳聚糖是一种带正电荷且具有生物相容性的多糖，由葡萄糖胺和 N-乙酰葡萄糖胺组成，对中性粒细胞具有趋化作用，并通过尚不清楚的机制刺激伤口修复。先前的研究表明壳聚糖会从血浆中耗尽补体蛋白，这表明壳聚糖会激活补体。补体激活导致 C5 裂解产生 C5a，这是一种中性粒细胞趋化因子。在这里，我们检验了壳聚糖在人全血、柠檬酸盐血浆和血清中产生 C5a 的假说。C5a 片段出现在凝固的全血中，并且壳聚糖-甘油磷酸/全血混合物与血小板和凝血酶的激活同时出现。然而，在血浆和血清中，尽管天然的 C3、C5 和因子 B 非共价吸附到在柠檬酸盐血浆、血清、EDTA 血清和甲胺处理的血浆中孵育的不溶性壳聚糖颗粒上，但凝血酶和壳聚糖-GP 未能产生 C5a。通过表面等离子体共振，纯 C3 吸附到壳聚糖上。与壳聚糖结合的血清因子的图谱与一种模型一致，该模型认为 pI 低于壳聚糖 pK(0)6.78（壳聚糖 pK(a)的上限）的带负电荷的血液蛋白通过静电与阳离子壳聚糖颗粒结合。酵母幽灵颗粒酵母聚糖在血清和柠檬酸盐血浆中激活补体，而不在 EDTA 血清或甲胺血浆中激活补体，以产生液相间 C5a，而 C3b 与酵母聚糖相关蛋白形成共价交联，并迅速裂解为 iC3b，因子 Bb 稳定结合。这些数据表明壳聚糖是一种非反应性生物材料，不会直接激活补体，并为预测带负电荷的血清蛋白-壳聚糖相互作用提供了新的依据。
