Endothelial cell functions impaired by interferon in vitro: Insights into the molecular mechanism of thrombotic microangiopathy associated with interferon therapy
Endothelial cell functions impaired by interferon in vitro: Insights into the molecular mechanism of thrombotic microangiopathy associated with interferon therapy
复制标题
DOI：
10.1016/j.thromres.2018.01.039
复制
发表时间：
2018-03-01
影响因子：
7.5
通讯作者：
Wadhwa, Meenu
中科院分区：
文献类型：
--
作者：
Jia, Haiyan;Thelwell, Craig;Wadhwa, Meenu
简介：干扰素 (IFN)-α 和 IFN-ss 分别被批准用于治疗慢性丙型肝炎病毒感染和多发性硬化症，与影响肾功能的血栓性微血管病 (TMA) 有关。由于这种严重并发症的分子机制尚不清楚，我们旨在研究干扰素是否直接影响体外与血管生成和血液止血相关的内皮细胞功能，以及内皮细胞衍生的一氧化氮（NO）和前列环素的血管扩张剂。方法：通过BrdU掺入和alamarBlue测定法测量人脐静脉内皮细胞（HUVEC）的增殖和存活。在 HUVEC 和人真皮成纤维细胞的共培养物中评估血管生成。使用 ELISA 测量纤维蛋白溶解分子。分别使用荧光 NO 特异性探针和竞争性酶免疫测定法测量 NO 和前列环素。 结果：IFN-ss 1a 和 IFN-ss 1b 剂量依赖性地抑制 HUVEC 增殖，但 IFN-α 2a 和 IFN-α 2b 则不剂量依赖性抑制 HUVEC 增殖。一致地，IFN-ss 1a 和 IFN-ss 1b 也降低了 HUVEC 的存活率，但 IFN-α 却没有观察到这一点。然而，两种 IFN 亚型均抑制 VEGF 诱导的毛细血管样结构的发育，但 IFN-α 的作用不如 IFN-ss 有效。此外，两种 IFN 亚型均上调经处理的共培养物中干扰素诱导蛋白 10 的产生，同时抑制血管生成。此外，IFN-α 2a 和 IFN-ss 1a 减少了细胞内 NO 的产生，而 HUVEC 的前列环素释放不受 IFN 影响。重要的是，与未处理的细胞相比，IFN-ss 1a 和 IFN-ss 1b 处理的 HUVEC 均显示尿激酶型纤溶酶原激活剂释放显着减少，并且纤溶酶原激活剂抑制剂-1 的分泌量比组织型纤溶酶原激活剂高得多，表明纤溶活性降低。然而，IFN-α 在调节纤溶系统方面效果较差。结论：我们证明了 IFN 对血管生成和纤溶介导的内皮细胞功能产生有害影响，这可能会导致生理性内皮血栓抵抗力的丧失，并促进临床环境中血管并发症的发生。从机制上讲，我们的研究结果对于理解 IFN 疗法如何促进 TMA 的发展具有重要意义。
Introduction: Interferon (IFN)-alpha and IFN-ss approved for treatment of chronic hepatitis C viral infection and multiple sclerosis respectively have been linked to thrombotic microangiopathy (TMA) affecting renal function. Since the molecular mechanisms underlying this severe complication remain largely unclear, we aimed to investigate whether IFN affects directly in vitro endothelial cell functions associated with angiogenesis and blood haemostasis, as well as endothelial cell-derived vasodilators of nitric oxide (NO) and prostacyclin.Methods: Proliferation and survival of human umbilical vein endothelial cells (HUVECs) were measured by BrdU incorporation and alamarBlue assays. Angiogenesis was evaluated in co-cultures of HUVECs and human dermal fibroblasts. Fibrinolysis molecules were measured with ELISA. NO and prostacyclin were measured using a fluorescent NO-specific probe and a competitive enzyme immunoassay, respectively.Results: HUVEC proliferation was dose-dependently inhibited by IFN-ss 1a and IFN-ss 1b, but not by IFN-alpha 2a and IFN-alpha 2b. Consistently, IFN-ss 1a and IFN-ss 1b also reduced survival of HUVECs, but this again was not observed with IFN-alpha. However, both IFN subtypes inhibited VEGF-induced development of capillary-like structures, but the effect of IFN-alpha was less potent than IFN-ss. In addition, both IFN subtypes upregulated interferon inducible protein 10 production from treated co-cultures while suppressing angiogenesis. Furthermore, intracellular NO generation was reduced by IFN-alpha 2a and IFN-ss 1a, whereas prostacyclin release from HUVECs was not affected by IFN. Importantly, both IFN-ss 1a- and IFN-ss 1b-treated HUVECs showed a marked reduction in urokinase-type plasminogen activator release and a much greater secretion of plasminogen activator inhibitor-1 than tissue-type plasminogen activator compared with untreated cells, suggesting decreased fibrinolytic activity. IFN-alpha, however was less effective in modulating the fibrinolysis system.Conclusions: We demonstrate the detrimental effects of IFN on endothelial cell functions mediated with angiogenesis and fibrinolysis, which could potentially cause the loss of physiological endothelium thromboresistance and facilitate the development of vascular complications in a clinical setting. Mechanistically, our findings have implications for understanding how IFN therapy can foster the development of TMA.