*Research Center of Occupational Medicine, Third Hospital of Peking University, Beijing, China †Department of Medicine, University of Colorado Denver, Aurora, Colorado ‡Department of Medicine, Denver Health Medical Center, Denver, Colorado.
Shock. 2017 Dec;48(6):666-673. doi: 10.1097/SHK.0000000000000907.
Extracellular histones are cationic damage-associated molecular pattern molecules capable of directly inducing cellular injury via charge-mediated interactions with plasma membranes. Accordingly, histones released into the plasma during critical illness are known to contribute to the onset and propagation of lung injury. Vascular injury (with consequent degradation of the endothelial glycocalyx) simultaneously releases anionic heparan sulfate fragments (hexa- to octasaccharides in size) into the plasma. It is unknown whether this endogenous release of heparan sulfate fragments modulates charge-dependent histone cytotoxicity, or if exogenous heparan sulfate fragments could therapeutically attenuate histone-induced lung injury. Using isothermic calorimetry, we found that extracellular histones only bind to heparan sulfate fragments ≥ 10 saccharides in size, suggesting that glycocalyx-derived heparan sulfate hexa/octasaccharides are incapable of intercepting/neutralizing circulating histones. However, we found that even heparan sulfate fragments incapable of histone binding (e.g., tetrasaccharides) attenuated histone-induced lung injury in vivo, suggesting a direct, size-independent protective effect of heparan sulfate. We found that histones had no effect on human neutrophils ex vivo but exerted toll-like receptor-independent cytotoxicity on human pulmonary microvascular endothelial cells in vitro. This cytotoxicity could be prevented by either the addition of negatively charged (i.e., highly sulfated) heparan sulfate tetrasaccharides (incapable of binding histones) or decasaccharides (capable of binding histones). Taken together, our findings suggest that heparan sulfate oligosaccharides may directly exert pulmonary endothelial-protective effects that attenuate histone-mediated lung injury.
细胞外组蛋白是带正电荷的损伤相关分子模式分子，能够通过与质膜的电荷介导相互作用直接诱导细胞损伤。因此，在危重病期间释放到血浆中的组蛋白被认为有助于肺损伤的发生和发展。血管损伤（随之而来的内皮糖萼降解）同时将带负电荷的肝素硫酸片段（大小为六到八糖）释放到血浆中。目前尚不清楚这种内源性肝素硫酸片段的释放是否调节依赖电荷的组蛋白细胞毒性，或者外源性肝素硫酸片段是否可以治疗性地减轻组蛋白诱导的肺损伤。使用等温量热法，我们发现细胞外组蛋白仅与大小≥ 10 个糖的肝素硫酸片段结合，这表明糖萼衍生的肝素硫酸六糖/八糖不能拦截/中和循环中的组蛋白。然而，我们发现即使不能与组蛋白结合的肝素硫酸片段（例如四糖）也能减轻体内组蛋白诱导的肺损伤，这表明肝素硫酸具有直接的、与大小无关的保护作用。我们发现组蛋白对体外人中性粒细胞没有影响，但在体外对人肺微血管内皮细胞发挥 TLR 非依赖性细胞毒性。这种细胞毒性可以通过添加带负电荷（即高度硫酸化）的肝素硫酸四糖（不能结合组蛋白）或十糖（能够结合组蛋白）来预防。综上所述，我们的研究结果表明肝素硫酸寡糖可能直接发挥肺内皮保护作用，减轻组蛋白介导的肺损伤。
