Department of Medical Microbiology and Immunology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United States.
Department of Foundational Medical Studies, Oakland University William Beaumont School of Medicine, Rochester, MI, United States.
Front Immunol. 2020 Jul 22;11:1460. doi: 10.3389/fimmu.2020.01460. eCollection 2020.
The complement system alternative pathway (AP) can be activated excessively in inflammatory diseases, particularly when there is defective complement regulation. For instance, deficiency in complement regulators CD55 and CD59, leads to paroxysmal nocturnal hemoglobinuria (PNH), whereas Factor H mutations predispose to atypical hemolytic uremic syndrome (aHUS), both causing severe thrombohemolysis. Despite eculizumab being the treatment for these diseases, benefits vary considerably among patients. Understanding the molecular mechanisms involved in complement regulation is essential for developing new treatments. Properdin, the positive AP regulator, is essential for complement amplification by stabilizing enzymatic convertases. In this study, the role of properdin in red blood cell (RBC) lysis and endothelial cell opsonization in these AP-mediated diseases was addressed by developing assays using PNH patient RBCs and human primary endothelial cells, where the effects of inhibiting properdin, using novel monoclonal antibodies (MoAbs) that we generated and characterized, were compared to other complement inhibitors. In models of PNH, properdin inhibition prevented hemolysis of patient PNH type II and III RBCs more than inhibition of Factor B, C3, and C5 (>17-fold, or >81-fold, or >12-fold lower molar IC values, respectively). When tested in an aHUS hemolysis model, the anti-properdin MoAbs had 11-fold, and 86-fold lower molar IC values than inhibition of Factor B, or C3, respectively ( < 0.0001). When comparing target/inhibitor ratios in all hemolysis assays, inhibiting properdin was at least as efficient as the other complement inhibitors in most cases. In addition, using endothelial cell assays, the data indicate a critical novel role for properdin in promoting complement activation on human endothelial cells exposed to heme (a hemolysis by-product) and rH19-20 (to inhibit Factor H cell-surface protection), as occurs in aHUS. Inhibition of properdin or C3 in this system significantly reduced C3 fragment deposition by 75%. Altogether, the data indicate properdin is key in promoting RBC lysis and complement activation on human endothelial cells, contributing to the understanding of PNH and aHUS pathogenesis. Further studies to determine therapeutic values of inhibiting properdin in complement-mediated diseases, in particular those that are characterized by AP dysregulation, are warranted.
补体系统替代途径 (AP) 在炎症性疾病中可能过度激活，特别是在补体调节缺陷时。例如，补体调节因子 CD55 和 CD59 的缺乏导致阵发性夜间血红蛋白尿症 (PNH)，而因子 H 突变易导致非典型溶血尿毒症综合征 (aHUS)，两者均导致严重的血栓性溶血性疾病。尽管依库珠单抗是这些疾病的治疗方法，但患者之间的获益差异很大。了解补体调节涉及的分子机制对于开发新的治疗方法至关重要。备解素是 AP 的正向调节因子，通过稳定酶转化物对补体进行放大是必不可少的。在这项研究中，通过开发使用 PNH 患者 RBC 和人原代内皮细胞的测定方法，研究了备解素在这些 AP 介导的疾病中对 RBC 裂解和内皮细胞调理作用的作用，其中比较了使用我们生成和表征的新型单克隆抗体 (MoAb) 抑制备解素的效果与其他补体抑制剂的效果。在 PNH 模型中，与抑制因子 B、C3 和 C5 相比（摩尔 IC 值分别降低 17 倍、81 倍和 12 倍以上），抑制备解素可更有效地防止 II 型和 III 型 PNH 患者 RBC 的溶血（>17 倍，或>81 倍，或>12 倍）。在 aHUS 溶血模型中进行测试时，抗备解素 MoAb 的摩尔 IC 值比抑制因子 B 或 C3 分别低 11 倍和 86 倍（<0.0001）。在比较所有溶血测定中的靶标/抑制剂比值时，在大多数情况下，抑制备解素的效率与其他补体抑制剂相当。此外，通过内皮细胞测定，数据表明备解素在促进暴露于血红素（溶血的副产物）和 rH19-20（抑制因子 H 细胞表面保护）的人内皮细胞上补体激活方面具有关键的新作用，这在 aHUS 中发生。在该系统中抑制备解素或 C3 可使 C3 片段沉积减少 75%。总的来说，数据表明备解素是促进人内皮细胞上 RBC 裂解和补体激活的关键因素，有助于理解 PNH 和 aHUS 的发病机制。有必要进一步研究抑制补体介导疾病中备解素的治疗价值，特别是那些以 AP 失调为特征的疾病。
