Different signaling pathways involved in the anti-inflammatory effects of unfractionated heparin on lipopolysaccharide-stimulated human endothelial cells
Different signaling pathways involved in the anti-inflammatory effects of unfractionated heparin on lipopolysaccharide-stimulated human endothelial cells
复制标题
普通肝素对脂多糖刺激的人内皮细胞的抗炎作用涉及不同的信号通路
DOI：
10.1186/s12950-020-0238-7
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发表时间：
2020-02-10
影响因子：
5.1
通讯作者：
Ma, Xiaochun
中科院分区：
文献类型：
--
作者：
Li, Xu;Li, Lu;Ma, Xiaochun
在脓毒症中，炎症反应和凝血之间存在复杂的相互作用。肝素被用作公认的抗凝血剂，并具有多种可能影响败血症的生物学特性。本研究旨在探讨普通肝素对内毒素刺激的人肺微血管内皮细胞(HPMECs)抗炎作用的可能信号转导途径。方法将靶向I-κB-α的小干扰RNA导入HPMECs。在加入脂多糖(10 g/ml)之前，先用超滤(0.0 1 U/ml~ 10 U/ml)处理15 。检测全身炎症反应标志物。用酶联免疫吸附试验检测3 h和1 h时IL-6、IL-8的释放。1 h后，用Western印迹法检测核因子-κB(NF-κB)、磷酸化抑制物κB-α(IκB-α)、信号转导和转录激活因子3(STAT3)及ERK1/2、jnk、p38丝裂原活化蛋白激酶的表达。结果在HPMEC中，UFH明显抑制脂多糖刺激的IL-6和IL-8的产生，尤其是在10 U/ml时，抑制脂多糖诱导的IκB-α、ERK1/2、JNK、p38MAPK和STAT3的磷酸化。UFH还能抑制脂多糖刺激的NF-κB的核转位。更重要的是，针对I-κB-α的小干扰RNA可诱导更明显的炎症反应。UFH抑制I-κB-α沉默细胞中细胞因子的产生和不同信号通路的磷酸化。结论UFH通过不同的信号通路发挥抗炎作用。
BackgroundThere is a complex interplay between inflammatory response and coagulation in sepsis. Heparin is used as a recognized anticoagulant and possesses multiple biological properties that possibly affect sepsis. This study aimed to determine the possible signaling pathways involved in the anti-inflammatory effects of unfractionated heparin (UFH) on lipopolysaccharide (LPS)-stimulated human pulmonary microvascular endothelial cells (HPMECs).MethodsHPMECs were transfected with siRNA targeting IκB-α. Cells were treated with UFH (0.01 U/ml~ 10 U/ml) 15 min before adding LPS (10 μg/ml). We detected the markers of systemic inflammatory response. Release of interleukin (IL)-6, IL-8 were evaluated at 3 h by ELISA and at 1 h by qRT-PCR. After 1 h, nuclear factor-κB (NF-κB) as well as phosphorylated inhibitor κB-α (IκB-α), signal transducer and activator of transcription-3 (STAT3) and ERK1/2, JNK, p38 mitogen-activated protein kinase (MAPK) expressions were evaluated by Western blot. DNA binding was conducted to further prove the activation of NF-κB pathway.ResultsIn HPMECs, UFH obviously inhibited LPS-stimulated production of IL-6 and IL-8, especially in 10 U/ml. UFH inhibited LPS-induced phosphorylation of IκB-α, ERK1/2, JNK, p38 MAPK and STAT3. UFH also suppressed LPS-stimulated nuclear translocation of NF-κB. Importantly, transfection with siRNA targeting IκB-α induced more obvious inflammatory response. UFH suppressed cytokines production and phosphorylation of different signaling pathways in IκB-α silencing cells.ConclusionThese results demonstrate that UFH exerts the anti-inflammatory effects on LPS-stimulated HPMECs by different signaling pathways.