RNAa-RNAi 双靶向调控VEGF-KDR/sFlt-1信号通路治疗PE

Preeclampsia is an extremely common pregnancy complication, which is still eager for effective therapies. Existing researches have confirmed that increased binding of VEGF to sFlt-1 and decreased binding of VEGF to KDR are the major pathological mechanisms of PE. Thus, both VEGF and sFlt-1 are therapeutic targets of PE. RNA activation (RNAa) is a new mechanism of gene regulation. Special small activating RNAs are designed to targeting promoter regions, which could sustained activate the expression of target genes. This study intends to utilize RNAa and RNAi to activate VEGF and silence sFlt-1 separately, thereby sustained activate VEGF-KDR signaling pathway, and suppress VEGF-sFlt-1 signaling pathway, and subsequently promote secretion of nitric oxide and decrease oxidative damage. Furthermore, we also intends to use G4-PAMAM dendrimer, which is a nanocarrier with non-toxic and little effect on the fetus, to co-deliver VEGF-saRNA and sFlt-1-siRNA into preeclamptic rats, and then in vivo analyze the therapeutic effect of VEGF-saRNA-sFlt-1-siRNA-G4-PAMAM dendrimer on PE. Our study may finally demonstrate an effective therapeutic method for gene therapy of PE.

子痫前期(PE)是妊娠期常见疾病，目前尚缺乏有效治疗方法。研究证实VEGF与KDR结合减少而VEGF与sFlt-1结合增加是导致PE的主要原因；因此，VEGF-KDR/sFlt-1通路成为PE治疗的靶点。RNA激活(RNAa)是最近发现的一种新分子现象：小激活RNA(saRNA)可以靶向目的基因启动子区，持续有效上调目的基因的表达。我们拟将RNAa和现有的RNA干扰技术相结合，通过saRNA靶向激活VEGF表达并下调sFlt-1表达，持续有效的活化VEGF-KDR通路，抑制VEGF-sFlt-1通路，从而通过促进NO分泌、减少ROS等逆转PE病理生理改变。同时利用安全且对胎儿影响小的G4-PAMAM纳米颗粒搭载VEGF-saRNA和sFlt-1-siRNA，在PE动物模型上验证该纳米复合物通过双靶向调控VEGF-KDR/sFlt-1信号通路对PE的治疗作用，以期找到PE基因治疗的新方法。

背景：现有的大量研究证实VEGF与其可溶性受体sFlt1的结合增加是子痫前期的主要病理机制。因此，VEGF和sFlt1是治疗子痫前期的关键靶点。.方法：本研究拟利用RNAa和RNAi技术分别激活VEGF和沉默sFlt1的表达。此外，我们利用树枝状聚合物PAMAM纳米颗粒搭载saRNA-VEGF和siRNA-sFlt1，并注射到子痫前期大鼠体内，探讨其对子痫前期的治疗作用。.结果：滋养细胞转染saRNA-VEGF后，VEGF、Nrf2和HO1的表达显著升高，并且显著缓解了TNF-α对滋养细胞的负性作用。滋养细胞转染saRNA-VEGF/siRNA-sFlt1后的迁移能力和成管能力均显著增强。PAMAM具有较高的细胞转运效率，是转运小分子RNA的良好纳米载体。研究结果显示siRNA-sFlt1-PAMAM可以在不影响滋养细胞增殖能力的同时，显著降低滋养细胞分泌的sFLT1水平。分组动物实验研究结果显示：TNF-α干预组的孕鼠血清sFlt1水平、平均动脉压、尿蛋白水平与对照组相比明显升高，而胎鼠和胎盘重量与对照组相比明显降低。接受siRNA-sFlt1-PAMAM/saRNA-VEGF-PAMAM治疗的孕鼠组中这些PE症状均有显著缓解。.结论：这些数据表明siRNA-sFlt1-PAMAM/saRNA-VEGF-PAMAM具有治疗PE的潜在价值，为PE的治疗提供了新策略。