Chatard Morgane, Puech Clémentine, Roche Frederic, Perek Nathalie
Université de Lyon, UJM-Saint-Etienne, SNA-EPIS, EA4607, F-42023, Saint-Etienne, France.
Université de Lyon, UJM-Saint-Etienne, INSERM, SAINBIOSE U1089 Team DVH, F-42023, Saint-Etienne, France.
PLoS One. 2016 Jun 23;11(6):e0158010. doi: 10.1371/journal.pone.0158010. eCollection 2016.
Understanding cellular and molecular mechanisms induced by hypoxic stress is crucial to reduce blood-brain barrier (BBB) disruption in some neurological diseases. Since the brain is a complex organ, it makes the interpretation of in vivo data difficult, so BBB studies are often investigated using in vitro models. However, the investigation of hypoxia in cellular pathways is complex with physical hypoxia because HIF-1α (factor induced by hypoxia) has a short half-life. We had set up an innovative and original method of induction of hypoxic stress by hydralazine that was more reproducible, which allowed us to study its impact on an in vitro BBB model. Our results showed that hydralazine, a mimetic agent of the hypoxia pathway, had the same effect as physical hypoxia, with few cytotoxicity effects on our cells. Hypoxic stress led to an increase of BBB permeability which corresponded to an opening of our BBB model. Study of tight junction proteins revealed that this hypoxic stress decreased ZO-1 but not occludin expression. In contrast, cells established a defence mechanism by increasing expression and activity of their efflux transporters (Pgp and MRP-1). This induction method of hypoxic stress by hydralazine is simple, reproducible, controllable and suitable to understand the cellular and molecular mechanisms involved by hypoxia on the BBB.
了解缺氧应激诱导的细胞和分子机制对于减少某些神经系统疾病中的血脑屏障（BBB）破坏至关重要。由于大脑是一个复杂的器官，这使得体内数据的解释变得困难，因此BBB研究通常使用体外模型进行。然而，由于缺氧诱导因子1α（HIF-1α，一种由缺氧诱导的因子）半衰期较短，细胞途径中缺氧的研究与物理性缺氧一样复杂。我们建立了一种由肼苯哒嗪诱导缺氧应激的创新且独特的方法，该方法更具可重复性，这使我们能够研究其对体外BBB模型的影响。我们的结果表明，作为缺氧途径模拟剂的肼苯哒嗪与物理性缺氧具有相同的效果，对我们的细胞几乎没有细胞毒性作用。缺氧应激导致BBB通透性增加，这与我们的BBB模型开放相对应。紧密连接蛋白的研究表明，这种缺氧应激降低了ZO-1的表达，但未降低闭合蛋白的表达。相反，细胞通过增加其外排转运蛋白（Pgp和MRP-1）的表达和活性建立了一种防御机制。这种由肼苯哒嗪诱导缺氧应激的方法简单、可重复、可控，适用于了解缺氧对BBB所涉及的细胞和分子机制。
