Yang Changjun, DeMars Kelly M, Hawkins Kimberly E, Candelario-Jalil Eduardo
Department of Neuroscience, McKnight Brain Institute, University of Florida, Gainesville, FL 32610, USA.
Department of Neuroscience, McKnight Brain Institute, University of Florida, Gainesville, FL 32610, USA.
Peptides. 2016 Jul;81:29-37. doi: 10.1016/j.peptides.2016.03.009. Epub 2016 Mar 25.
Adropin is a peptide encoded by the energy homeostasis associated gene (Enho) and plays a critical role in the regulation of lipid metabolism, insulin sensitivity, and endothelial function. Little is known of the effects of adropin in the brain and whether this peptide modulates ischemia-induced blood-brain barrier (BBB) injury. Here, we used an in vitro BBB model of rat brain microvascular endothelial cells (RBE4) and hypothesized that adropin would reduce endothelial permeability during ischemic conditions. To mimic ischemic conditions in vitro, RBE4 cell monolayers were subjected to 16h hypoxia/low glucose (HLG). This resulted in a significant increase in paracellular permeability to FITC-labeled dextran (40kDa), a dramatic upregulation of vascular endothelial growth factor (VEGF), and the loss of junction proteins occludin and VE-cadherin. Notably, HLG also significantly decreased Enho expression and adropin levels. Treatment of RBE4 cells with synthetic adropin (1, 10 and 100ng/ml) concentration-dependently reduced endothelial permeability after HLG, but this was not mediated through protection to junction proteins or through reduced levels of VEGF. We found that HLG dramatically increased myosin light chain 2 (MLC2) phosphorylation in RBE4 cells, which was significantly reduced by adropin treatment. We also found that HLG significantly increased Rho-associated kinase (ROCK) activity, a critical upstream effector of MLC2 phosphorylation, and that adropin treatment attenuated that effect. These data indicate that treatment with adropin reduces endothelial cell permeability after HLG insult by inhibition of the ROCK-MLC2 signaling pathway. These promising findings suggest that adropin protects against endothelial barrier dysfunction during ischemic conditions.
内脂素是一种由能量稳态相关基因（Enho）编码的肽，在脂质代谢、胰岛素敏感性和内皮功能的调节中起关键作用。关于内脂素在大脑中的作用以及这种肽是否调节缺血诱导的血脑屏障（BBB）损伤，人们知之甚少。在这里，我们使用大鼠脑微血管内皮细胞（RBE4）的体外血脑屏障模型，并假设内脂素会在缺血条件下降低内皮通透性。为了在体外模拟缺血条件，将RBE4细胞单层置于16小时的缺氧/低糖（HLG）环境中。这导致对异硫氰酸荧光素标记的葡聚糖（40kDa）的细胞旁通透性显著增加，血管内皮生长因子（VEGF）急剧上调，以及紧密连接蛋白闭合蛋白和血管内皮钙黏蛋白的丢失。值得注意的是，HLG还显著降低了Enho表达和内脂素水平。用合成内脂素（1、10和100ng/ml）处理RBE4细胞后，HLG后的内皮通透性呈浓度依赖性降低，但这不是通过对紧密连接蛋白的保护或通过降低VEGF水平介导的。我们发现HLG显著增加了RBE4细胞中肌球蛋白轻链2（MLC2）的磷酸化，而内脂素处理可显著降低这种磷酸化。我们还发现HLG显著增加了Rho相关激酶（ROCK）的活性，ROCK是MLC2磷酸化的关键上游效应器，而内脂素处理减弱了这种效应。这些数据表明，内脂素处理通过抑制ROCK-MLC2信号通路降低了HLG损伤后的内皮细胞通透性。这些有前景的发现表明，内脂素在缺血条件下可防止内皮屏障功能障碍。
