Ravid Orly, Elhaik Goldman Shirin, Macheto David, Bresler Yael, De Oliveira Raquel Ines, Liraz-Zaltsman Sigal, Gosselet Fabien, Dehouck Lucie, Beeri Michal Schnaider, Cooper Itzik
The Joseph Sagol Neuroscience Center, Sheba Medical Center, Tel Hashomer, Ramat Gan, Israel.
Interdisciplinary Center Herzliya, Herzliya, Israel.
Front Cell Neurosci. 2018 Oct 16;12:359. doi: 10.3389/fncel.2018.00359. eCollection 2018.
Organophosphorus compounds (OPs) are highly toxic chemicals widely used as pesticides (e.g., paraoxon (PX)- the active metabolite of the insecticide parathion) and as chemical warfare nerve agents. Blood-brain barrier (BBB) leakage has been shown in rodents exposed to PX, which is an organophosphate oxon. In this study, we investigated the cellular mechanisms involved in BBB reaction after acute exposure to PX in an established BBB system made of stem-cell derived, human brain-like endothelial cells (BLECs) together with brain pericytes that closely mimic the BBB. Our results show that PX directly affects the BBB both at toxic and non-toxic concentrations by attenuating tight junctional (TJ) protein expression and that only above a certain threshold the paracellular barrier integrity is compromised. Below this threshold, BLECs exhibit a morphological coping mechanism in which they enlarge their cell area thus preventing the formation of meaningful intercellular gaps and maintaining barrier integrity. Importantly, we demonstrate that reversal of the apoptotic cell death induced by PX, by a pan-caspase-inhibitor ZVAD-FMK (ZVAD) can reduce PX-induced cell death and elevate cell area but do not prevent the induced BBB permeability, implying that TJ complex functionality is hindered. This is corroborated by formation of ROS at all toxic concentrations of PX and which are even higher with ZVAD. We suggest that while lower levels of ROS can induce compensating mechanisms, higher PX-induced oxidative stress levels interfere with barrier integrity.
有机磷化合物（OPs）是剧毒化学品，广泛用作杀虫剂（如对氧磷（PX）——杀虫剂对硫磷的活性代谢物）和化学战神经毒剂。在暴露于PX（一种有机磷酸酯氧肟酸）的啮齿动物中已显示出血脑屏障（BBB）渗漏。在本研究中，我们在一个由干细胞衍生的人脑样内皮细胞（BLECs）和脑周细胞组成的成熟BBB系统中，研究了急性暴露于PX后BBB反应所涉及的细胞机制，该系统紧密模拟了BBB。我们的结果表明，PX通过减弱紧密连接（TJ）蛋白表达，在有毒和无毒浓度下均直接影响BBB，并且只有超过一定阈值，细胞旁屏障的完整性才会受到损害。低于此阈值时，BLECs表现出一种形态应对机制，即它们扩大细胞面积，从而防止形成有意义的细胞间间隙并维持屏障完整性。重要的是，我们证明，通过泛半胱天冬酶抑制剂ZVAD - FMK（ZVAD）逆转PX诱导的凋亡性细胞死亡，可以减少PX诱导的细胞死亡并增加细胞面积，但不能防止诱导的BBB通透性增加，这意味着TJ复合体的功能受到了阻碍。PX所有有毒浓度下均会形成活性氧（ROS），ZVAD存在时ROS水平甚至更高，这证实了上述观点。我们认为，虽然较低水平的ROS可以诱导补偿机制，但较高的PX诱导的氧化应激水平会干扰屏障完整性。
