Lee Jeong Ho, Lee Jinu, Seo Gi Ho, Kim Chul Hoon, Ahn Young Soo
Department of Pharmacology, Yonsei University College of Medicine, Seoul, South Korea.
J Mol Neurosci. 2007;32(2):145-54. doi: 10.1007/s12031-007-0026-3.
Heparin is a classic anticoagulant that is commonly used in the treatment of acute ischemic stroke (AIS). Its use remains controversial, however, due to the risk of cerebral hemorrhagic transformation. In addition to anticoagulant effects, diverse effects on transcription factors can be caused by heparin. Among the transcription factors potentially affected is nuclear factor kappa B (NF-kappaB), a protein that is reportedly related to the survival of cerebral endothelial cells. We investigated the effect of heparin on NF-kappaB activation and cell death following oxygen-glucose deprivation (OGD), an experimental model of AIS. We subjected bEnd.3 cells from a murine cerebral microvascular endothelial cell line to OGD. We examined the effect of heparin on OGD-induced NF-kappaB activation and its mechanism of action, using electrophoretic mobility shift assays, reporter gene analysis, real-time RT-PCR, Western blot analysis, and confocal microscopy. We also measured the effect of heparin on OGD-induced cell death using an MTT assay. Heparin inhibited both tumor necrosis factor alpha- and OGD-induced NF-kappaB activation. Heparin was taken up by endocytosis and then entered the nucleus. Heparin did not affect the nuclear translocation of NF-kappaB, but instead inhibited the DNA binding of NF-kappaB in the nucleus. Cells were more susceptible to OGD-induced cell death after heparin treatment. Besides producing an anticoagulation effect, heparin also inhibits NF-kappaB activation, resulting in increased susceptibility to OGD-induced cell death. This effect may be responsible for hemorrhagic transformation in patients following heparin treatment for AIS.
肝素是一种经典的抗凝剂，常用于治疗急性缺血性卒中（AIS）。然而，由于存在脑出血转化的风险，其使用仍存在争议。除了抗凝作用外，肝素还可对转录因子产生多种影响。潜在受影响的转录因子之一是核因子κB（NF-κB），据报道该蛋白与脑内皮细胞的存活有关。我们研究了肝素对氧糖剥夺（OGD）（一种AIS实验模型）后NF-κB激活和细胞死亡的影响。我们将来自小鼠脑微血管内皮细胞系的bEnd.3细胞进行OGD处理。我们使用电泳迁移率变动分析、报告基因分析、实时逆转录-聚合酶链反应、蛋白质印迹分析和共聚焦显微镜检查，研究了肝素对OGD诱导的NF-κB激活及其作用机制的影响。我们还使用MTT法测量了肝素对OGD诱导的细胞死亡的影响。肝素抑制肿瘤坏死因子α和OGD诱导的NF-κB激活。肝素通过内吞作用被摄取，然后进入细胞核。肝素不影响NF-κB的核转位，但抑制NF-κB在细胞核中的DNA结合。肝素处理后，细胞对OGD诱导的细胞死亡更敏感。除了产生抗凝作用外，肝素还抑制NF-κB激活，导致对OGD诱导的细胞死亡的易感性增加。这种作用可能是AIS患者肝素治疗后出血转化的原因。
