Soluble vascular endothelial growth factor receptor-1 (sFlt-1) is an anti-angiogenic protein which is secreted by numerous cell types and acts as a decoy receptor for the angiogenic protein vascular endothelial growth factor (VEGF). Despite its physiologic importance in maintaining angiogenic balance, excess sFlt-1 levels are associated with the pathogenesis of many diseases, especially those with angiogenic imbalance, endothelial dysfunction, and hypertension. Although sFlt-1 is a soluble protein, it contains a binding site for the extracellular matrix component heparan sulfate. This allows cells to retain and localize sFlt-1 in order to prevent excessive VEGF signaling. During pregnancy, placental syncytiotrophoblasts develop a large extracellular matrix which contains significant amounts of heparan sulfate. Consequently, the placenta becomes a potential storage site for large amounts of sFlt-1 bound to extracellular heparan sulfate. Additionally, it should be noted that sFlt-1 can bind to the anticoagulant unfractionated heparin due to its molecular mimicry to heparan sulfate. However, it remains unknown whether unfractionated heparin can compete with heparan sulfate for binding of localized sFlt-1. In this study, we hypothesized that administration of unfractionated heparin would displace and solubilize placental extracellular matrix(ECM)-bound sFlt-1. If unfractionated heparin can displace this large reservoir of sFlt-1 in the placenta and mobilized it into the maternal circulation, we should be able to observe its effects on maternal angiogenic balance and blood pressure. To test this hypothesis, we utilized in vitro, ex vivo, and in vivo methods. Using the BeWo placental trophoblast cell line, we observed increased sFlt-1 in the media of cells treated with unfractionated heparin compared to controls. The increase in media sFlt-1 was found in conjunction with decreased localized cellular Flt (sFlt-1 and Flt-1) as measured by total cell fluorescence. Similar results were observed using ex vivo placental villous explants treated with unfractionated heparin. Real-time quantitative PCR of the explants showed no change in sFlt-1 or heparanase-1 mRNA expression, eliminating increased production and enzymatic cleavage of heparan sulfate as causes for sFlt-1 media increase. Timed-pregnant rats given a continuous infusion of unfractionated heparin exhibited an increased mean arterial pressure as well as decreased bioavailable VEGF compared to vehicle-treated animals. These data demonstrate that chronic unfractionated heparin treatment is able to displace matrix-bound sFlt-1 into the maternal circulation to such a degree that mean arterial pressure is significantly affected. Here we have shown that the placental ECM is a storage site for large quantities of sFlt-1, and that it should be carefully considered in future studies concerning angiogenic balance in pregnancy.

中文翻译：

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普通肝素从胎盘细胞外基质中置换出sFlt-1。

可溶性血管内皮生长因子受体1（sFlt-1）是一种抗血管生成蛋白，由多种细胞类型分泌，并充当血管生成蛋白血管内皮生长因子（VEGF）的诱饵受体。尽管其在维持血管生成平衡方面具有重要的生理意义，但过量的sFlt-1水平却与许多疾病的发病机理有关，尤其是那些具有血管生成失衡，内皮功能障碍和高血压的疾病。尽管sFlt-1是一种可溶性蛋白，但它包含细胞外基质成分硫酸乙酰肝素的结合位点。这允许细胞保留和定位sFlt-1，以防止过度的VEGF信号传导。在怀孕期间，胎盘合体滋养层细胞会形成大的细胞外基质，其中含有大量的硫酸乙酰肝素。所以，胎盘成为与细胞外硫酸乙酰肝素结合的大量sFlt-1的潜在储存位点。另外，应该注意的是，由于sFlt-1分子模拟硫酸乙酰肝素，因此它可以与抗凝剂普通肝素结合。但是，尚不知道普通肝素能否与硫酸乙酰肝素竞争结合局部sFlt-1。在这项研究中，我们假设未分级肝素的施用会置换并溶解胎盘细胞外基质（ECM）结合的sFlt-1。如果普通肝素可以取代胎盘中大量的sFlt-1储库并动员其进入母体循环，我们应该能够观察到它对母体血管生成平衡和血压的影响。为了验证这一假设，我们利用了体外，离体和体内方法。使用BeWo胎盘滋养层细胞系，与对照组相比，我们观察到用普通肝素处理的细胞培养基中sFlt-1增加。发现培养基sFlt-1的增加与局部细胞Flt（sFlt-1和Flt-1）的减少有关，如通过总细胞荧光所测。使用未分级肝素处理的离体胎盘绒毛外植体观察到相似的结果。外植体的实时定量PCR显示sFlt-1或乙酰肝素酶1 mRNA表达没有变化，消除了由于sFlt-1培养基增加而引起的硫酸乙酰肝素的产量增加和酶促裂解。与赋形剂处理的动物相比，连续注入普通肝素的定时妊娠大鼠表现出平均动脉压升高和生物利用性VEGF降低。这些数据表明，慢性普通肝素治疗能够将结合基质的sFlt-1置换到母体循环中，从而使平均动脉压受到显着影响。在这里，我们已经表明，胎盘ECM是大量sFlt-1的储存位点，在有关妊娠期血管生成平衡的未来研究中应仔细考虑。