Hughes Jason M, Neese Olivia R, Bieber Dylan D, Lewis Kirsten A, Ahmadi Layla M, Parsons Dustin W, Canfield Scott G
Department of Anatomy, Cell Biology and Physiology, Indiana University School of Medicine, Terre Haute, IN, United States.
Department of Biology, Indiana State University, Terre Haute, IN, United States.
Front Cell Neurosci. 2022 May 11;16:835649. doi: 10.3389/fncel.2022.835649. eCollection 2022.
Recently, the safety of repeated and lengthy anesthesia administration has been called into question, a subset of these animal studies demonstrated that anesthetics induced blood-brain barrier (BBB) dysfunction. The BBB is critical in protecting the brain parenchyma from the surrounding micro-vasculature. BBB breakdown and dysfunction has been observed in several neurodegenerative diseases and may contribute to both the initiation and the progression of the disease. In this study we utilize a human induced pluripotent stem cell (iPSC) derived-BBB model, exhibiting near properties, to evaluate the effects of anesthetics on critical barrier properties.
iPSC-derived brain microvascular endothelial cells (BMECs) expressed near barrier tightness assessed by endothelial electrical resistance and para-cellular permeability. Efflux transporter activity was determined by substrate transport in the presence of specific inhibitors. cellular transport was measured utilizing large fluorescently tagged dextran. Tight junction localization in BMECs was evaluated with fluorescent microscopy. The anesthetic, propofol was exposed to BMECs at varying durations and concentrations and BBB properties were monitored post-exposure.
Following propofol exposure, BMECs displayed reduced resistance and increased permeability indicative of a leaky barrier. Reduced barrier tightness and the dysregulation of occludin, a tight junction protein, were partly the result of an elevation in matrix metalloproteinase (MMP) levels. Efflux transporter activity and cellular transport were unaffected by propofol exposure. Propofol induced barrier dysfunction was partially restored following matrix metalloproteinase inhibition.
For the first time, we have demonstrated that propofol alters BBB integrity utilizing a human BBB model that displays key characteristics. A leaky BBB enables otherwise impermeable molecules such as pathogens and toxins the ability to reach vulnerable cell types of the brain parenchyma. A robust human BBB model will allow for the evaluation of several anesthetics at fluctuating clinical scenarios and to elucidate mechanisms with the goal of ultimately improving anesthesia safety.
最近，反复长时间麻醉给药的安全性受到质疑，部分动物研究表明麻醉剂会导致血脑屏障（BBB）功能障碍。血脑屏障对于保护脑实质免受周围微血管系统的影响至关重要。在几种神经退行性疾病中都观察到了血脑屏障的破坏和功能障碍，这可能在疾病的发生和发展中都起到作用。在本研究中，我们利用人诱导多能干细胞（iPSC）衍生的血脑屏障模型（具有近乎天然的特性）来评估麻醉剂对关键屏障特性的影响。
iPSC衍生的脑微血管内皮细胞（BMECs）通过内皮电阻和细胞旁通透性评估显示出近乎天然的屏障紧密性。通过在特定抑制剂存在下的底物转运来测定外排转运体活性。利用大型荧光标记葡聚糖测量细胞转运。用荧光显微镜评估BMECs中紧密连接的定位。将麻醉剂丙泊酚以不同的持续时间和浓度作用于BMECs，并在暴露后监测血脑屏障特性。
丙泊酚暴露后，BMECs显示出电阻降低和通透性增加，表明屏障渗漏。屏障紧密性降低和紧密连接蛋白occludin的失调部分是基质金属蛋白酶（MMP）水平升高的结果。外排转运体活性和细胞转运不受丙泊酚暴露的影响。基质金属蛋白酶抑制后，丙泊酚诱导的屏障功能障碍得到部分恢复。
我们首次证明，利用具有关键天然特征的人血脑屏障模型，丙泊酚会改变血脑屏障的完整性。渗漏的血脑屏障使病原体和毒素等原本无法透过的分子能够接触到脑实质中易受影响的细胞类型。一个强大的人血脑屏障模型将允许在不同的临床情况下评估多种麻醉剂，并阐明其机制，最终目标是提高麻醉安全性。
