Research Laboratory, Nordland Hospital, Bodø, Norway.
Faculty of Health Sciences, K. G. Jebsen TREC, UiT - The Arctic University of Norway, Tromsø, Norway.
J Thromb Haemost. 2018 May;16(5):905-918. doi: 10.1111/jth.13979. Epub 2018 Mar 23.
Essentials Complement, Toll-like receptors and coagulation cross-talk in the process of thromboinflammation. This is explored in a unique human whole-blood model of S. aureus bacteremia. Coagulation is here shown as a downstream event of C5a-induced tissue factor (TF) production. Combined inhibition of C5 and CD14 efficiently attenuated TF and coagulation.
Background There is extensive cross-talk between the complement system, the Toll-like receptors (TLRs), and hemostasis. Consumptive coagulopathy is a hallmark of sepsis, and is often mediated through increased tissue factor (TF) expression. Objectives To study the relative roles of complement, TLRs and TF in Staphylococcus aureus-induced coagulation. Methods Lepirudin-anticoagulated human whole blood was incubated with the three S. aureus strains Cowan, Wood, and Newman. C3 was inhibited with compstatin, C5 with eculizumab, C5a receptor 1 (C5aR1) and activated factor XII with peptide inhibitors, CD14, TLR2 and TF with neutralizing antibodies, and TLR4 with eritoran. Complement activation was measured by ELISA. Coagulation was measured according to prothrombin fragment 1 + 2 (PTF ) determined with ELISA, and TF mRNA, monocyte surface expression and functional activity were measured with quantitative PCR, flow cytometry, and ELISA, respectively. Results All three strains generated substantial and statistically significant amounts of C5a, terminal complement complex, PTF , and TF mRNA, and showed substantial TF surface expression on monocytes and TF functional activity. Inhibition of C5 cleavage most efficiently and significantly inhibited all six markers in strains Cowan and Wood, and five markers in Newman. The effect of complement inhibition was shown to be completely dependent on C5aR1. The C5 blocking effect was equally potentiated when combined with blocking of CD14 or TLR2, but not TLR4. TF blocking significantly reduced PTF levels to baseline levels. Conclusions S. aureus-induced coagulation in human whole blood was mainly attributable to C5a-induced mRNA upregulation, monocyte TF expression, and plasma TF activity, thus underscoring complement as a key player in S. aureus-induced coagulation.
目的 研究补体系统、Toll 样受体（TLR）和止血之间的广泛相互作用。消耗性凝血病是败血症的标志，通常通过增加组织因子（TF）表达来介导。方法 用三种金黄色葡萄球菌菌株 Cowan、Wood 和 Newman 孵育 lepirudin 抗凝的人全血。用 compstatin 抑制 C3，用 eculizumab 抑制 C5，用肽抑制剂抑制 C5a 受体 1（C5aR1）和激活的 XII 因子，用中和抗体抑制 CD14、TLR2 和 TF，用 eritoran 抑制 TLR4。通过 ELISA 测量补体激活。根据 ELISA 测定的凝血酶原片段 1+2（PTF）测量凝血，并用定量 PCR、流式细胞术和 ELISA 分别测量 TF mRNA、单核细胞表面表达和功能活性。结果 三种菌株均产生大量且具有统计学意义的 C5a、末端补体复合物、PTF 和 TF mRNA，并在单核细胞上显示出大量 TF 表面表达和 TF 功能活性。C5 切割的抑制最有效地和显著地抑制了 Cowan 和 Wood 两种菌株的所有六种标志物，以及 Newman 菌株的五种标志物。补体抑制的作用完全依赖于 C5aR1。当与阻断 CD14 或 TLR2 联合使用时，C5 阻断作用同样增强，但与 TLR4 无关。TF 阻断显著降低 PTF 水平至基线水平。结论 金黄色葡萄球菌诱导的人全血凝血主要归因于 C5a 诱导的 mRNA 上调、单核细胞 TF 表达和血浆 TF 活性，从而突出了补体在金黄色葡萄球菌诱导的凝血中的关键作用。
