Abstract：
BACKGROUND: Tafamidis is a molecular chaperone that stabilizes the transthyretin (TTR) homo-tetramer, preventing its dissociation and consequent deposition as amyloid fibrils in organ tissues. Tafamidis reduces mortality and the incidence of hospitalization for cardiovascular causes in patients with TTR amyloid (ATTR) cardiomyopathy. As ATTR cardiomyopathy is associated with a high risk of thromboembolic complications, we hypothesized that tafamidis may have a direct ancillary anti-thrombotic effect.
METHODS: Primary human aortic endothelial cells (HAECs) were treated with tafamidis at clinically relevant concentrations and with plasma of patients, before and after the initiation of treatment with tafamidis. The expression of TF was induced by incubation with Tumor Necrosis Factor α (TNFα). Intracellular expression of tissue factor (TF) was measured by western blot. TF activity was measured by a colorimetric assay. Gene expressions of TF were measured by quantitative polymerase chain reaction.
RESULTS: Treatment with tafamidis dose-dependently reduced the expression and activity of TNFα-induced TF. This effect was confirmed in cells treated with patients\' plasma. Signal Transducer and Activator of Transcription 3 (STAT3) phosphorylation was significantly inhibited by tafamidis. Incubation of HAECs with tafamidis and the STAT3 activator colivelin partially rescued the expression of TF.
CONCLUSIONS: Treatment with tafamidis lowers the thrombotic potential in human primary endothelial cells by reducing TF expression and activity. This previously unknown off-target effect may provide a novel mechanistic explanation for the lower number of thromboembolic complications in ATTR cardiomyopathy patients treated with tafamidis.
METHODS: Primary human aortic endothelial cells (HAECs) were treated with tafamidis at clinically relevant concentrations and with plasma of patients, before and after the initiation of treatment with tafamidis. The expression of TF was induced by incubation with Tumor Necrosis Factor α (TNFα). Intracellular expression of tissue factor (TF) was measured by western blot. TF activity was measured by a colorimetric assay. Gene expressions of TF were measured by quantitative polymerase chain reaction.
RESULTS: Treatment with tafamidis dose-dependently reduced the expression and activity of TNFα-induced TF. This effect was confirmed in cells treated with patients\' plasma. Signal Transducer and Activator of Transcription 3 (STAT3) phosphorylation was significantly inhibited by tafamidis. Incubation of HAECs with tafamidis and the STAT3 activator colivelin partially rescued the expression of TF.
CONCLUSIONS: Treatment with tafamidis lowers the thrombotic potential in human primary endothelial cells by reducing TF expression and activity. This previously unknown off-target effect may provide a novel mechanistic explanation for the lower number of thromboembolic complications in ATTR cardiomyopathy patients treated with tafamidis.
摘要：
背景：Tafamidis是一种分子伴侣，可稳定甲状腺素运载蛋白（TTR）同四聚体，防止其解离和随后在器官组织中沉积为淀粉样原纤维。Tafamidis降低了TTR淀粉样蛋白（ATTR）心肌病患者的死亡率和因心血管原因住院的发生率。由于ATTR心肌病与血栓栓塞并发症的高风险相关，我们假设tafamidis可能具有直接的辅助抗血栓作用.
方法：原代人主动脉内皮细胞（HAECs）用临床相关浓度的tafamidis和患者血浆进行治疗，在开始用tafamidis治疗之前和之后。通过与肿瘤坏死因子α（TNFα）孵育诱导TF的表达。通过蛋白质印迹测量组织因子(TF)的细胞内表达。通过比色测定法测量TF活性。通过定量聚合酶链反应测量TF的基因表达。
结果：用他法米治疗剂量依赖性地降低了TNFα诱导的TF的表达和活性。在用患者血浆处理的细胞中证实了这种作用。信号转导和转录激活因子3（STAT3）的磷酸化被tafamidis显着抑制。HAECs与tafamidis和STAT3激活剂colivelin的孵育部分挽救了TF的表达。
结论：用tafamidis治疗通过降低TF表达和活性来降低人原代内皮细胞的血栓形成潜能。这种先前未知的脱靶效应可能为使用tafamidis治疗的ATTR心肌病患者的血栓栓塞并发症数量减少提供了新的机制解释。
方法：原代人主动脉内皮细胞（HAECs）用临床相关浓度的tafamidis和患者血浆进行治疗，在开始用tafamidis治疗之前和之后。通过与肿瘤坏死因子α（TNFα）孵育诱导TF的表达。通过蛋白质印迹测量组织因子(TF)的细胞内表达。通过比色测定法测量TF活性。通过定量聚合酶链反应测量TF的基因表达。
结果：用他法米治疗剂量依赖性地降低了TNFα诱导的TF的表达和活性。在用患者血浆处理的细胞中证实了这种作用。信号转导和转录激活因子3（STAT3）的磷酸化被tafamidis显着抑制。HAECs与tafamidis和STAT3激活剂colivelin的孵育部分挽救了TF的表达。
结论：用tafamidis治疗通过降低TF表达和活性来降低人原代内皮细胞的血栓形成潜能。这种先前未知的脱靶效应可能为使用tafamidis治疗的ATTR心肌病患者的血栓栓塞并发症数量减少提供了新的机制解释。
