Institut für Immunologie und Transfusionsmedizin and.
Zentrum für Innovationskompetenz-Humorale Immunreaktionen bei kardiovaskulären Erkrankungen, Universität Greifswald, Greifswald, Germany.
Blood. 2019 Feb 28;133(9):978-989. doi: 10.1182/blood-2018-05-850370. Epub 2018 Dec 20.
Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating anti-platelet factor 4 (PF4)/heparin antibodies. Platelet activation assays that use "washed" platelets are more sensitive for detecting HIT antibodies than platelet-rich plasma (PRP)-based assays. Moreover, heparin-exposed patients vary considerably with respect to the risk of PF4/heparin immunization and, among antibody-positive patients, the risk of subsequent "breakthrough" of clinical HIT with manifestation of thrombocytopenia. We used washed platelets and PRP, standard laboratory HIT tests, and physicochemical methods to identify a plasma factor interfering with PF4/heparin complexes and anti-PF4/heparin antibody-platelet interaction, thus explaining differences in functional assays. To investigate a modulating risk for PF4/heparin immunization and breakthrough of HIT, we also tested 89 plasmas from 2 serosurveillance trials. Fibronectin levels were measured in 4 patient groups exhibiting different degrees of heparin-dependent immunization and expression of HIT. The heat-labile plasma protein, fibronectin, inhibited PF4 binding to platelets in a dose-dependent fashion, particularly in washed (vs PRP) systems. Fibronectin also inhibited PF4/heparin binding to platelets, anti-PF4/heparin antibody binding to PF4/heparin complexes, and anti-PF4/heparin antibody-induced platelet activation as a result of PF4/heparin complex disruption. In addition, plasma fibronectin levels increased progressively among the following 4 patient groups: enzyme-linked immunosorbent assay (ELISA)+/serotonin-release assay (SRA)+/HIT+ < ELISA+/SRA+/HIT- ∼ ELISA+/SRA-/HIT- < ELISA-/SRA-/HIT-. Altogether, these findings suggest that fibronectin interferes with PF4/heparin complex formation and anti-PF4/heparin antibody-induced platelet activation. Reduced fibronectin levels in washed platelet assays help to explain the greater sensitivity of washed platelet (vs PRP) assays for HIT. More importantly, lower plasma fibronectin levels could represent a risk factor for PF4/heparin immunization and clinical breakthrough of HIT.
肝素诱导的血小板减少症（HIT）是由血小板激活的抗血小板因子 4（PF4）/肝素抗体引起的。与基于富含血小板血浆（PRP）的检测相比，使用“洗涤”血小板的血小板激活检测对检测 HIT 抗体更敏感。此外，肝素暴露的患者在 PF4/肝素免疫的风险方面存在很大差异，并且在抗体阳性的患者中，随后发生伴有血小板减少症的临床 HIT 突破的风险也存在差异。我们使用洗涤血小板和 PRP、标准实验室 HIT 检测以及物理化学方法来鉴定一种干扰 PF4/肝素复合物和抗 PF4/肝素抗体-血小板相互作用的血浆因子，从而解释了功能检测中的差异。为了研究 PF4/肝素免疫和 HIT 突破的调节风险，我们还测试了来自 2 项血清学监测试验的 89 个血浆样本。在表现出不同程度肝素依赖性免疫和 HIT 表达的 4 个患者组中，测量了纤维连接蛋白水平。热不稳定血浆蛋白纤维连接蛋白以剂量依赖的方式抑制 PF4 与血小板的结合，特别是在洗涤（与 PRP 相比）系统中。纤维连接蛋白还抑制 PF4/肝素与血小板、抗 PF4/肝素抗体与 PF4/肝素复合物的结合，以及抗 PF4/肝素抗体诱导的血小板激活，这是由于 PF4/肝素复合物的破坏。此外，在以下 4 个患者组中，血浆纤维连接蛋白水平逐渐增加：酶联免疫吸附测定（ELISA）+/血清素释放测定（SRA）+/HIT+<ELISA+/SRA+/HIT-∼ELISA+/SRA-/HIT-<ELISA-/SRA-/HIT-。总之，这些发现表明纤维连接蛋白干扰 PF4/肝素复合物的形成和抗 PF4/肝素抗体诱导的血小板激活。在洗涤血小板检测中纤维连接蛋白水平降低有助于解释洗涤血小板（与 PRP 相比）检测对 HIT 的更高敏感性。更重要的是，较低的血浆纤维连接蛋白水平可能是 PF4/肝素免疫和 HIT 临床突破的危险因素。
