槟榔多酚对大鼠肺微血管内皮细胞低氧损伤的保护作用

目的：探究槟榔多酚对大鼠肺微血管内皮细胞（PMVEC）低氧损伤的保护作用。 方法：采用氧化应激指标丙二醛和超氧化物歧化酶筛选肺部缺氧损伤细胞模型最适条件；采用细胞计数试剂盒法筛选槟榔多酚最适给药剂量。将PMVEC分为常氧对照组、模型对照组和槟榔多酚组，采用BCA法检测蛋白浓度并测定PMVEC中的氧化应激水平，蛋白质印迹法检测炎症和凋亡相关蛋白的表达，免疫荧光染色法检测闭合蛋白和闭锁小带1等紧密连接蛋白的表达，Transwell小室实验检测跨内皮细胞膜电阻，罗丹明荧光染料检测PMVEC屏障的通透性。 结果：1%氧浓度培养PMVEC细胞 48?h后成功构建肺部缺氧损伤细胞模型 ；20?μg/mL的槟榔多酚可以显著逆转缺氧损伤细胞模型中PMVEC的存活率下降以及氧化应激水平的升高（均 P<0.05）；槟榔多酚对于低氧诱导下PMVEC中炎症相关蛋白核因子κB（NF-κB）和核因子E2相关因子（Nrf）2的上调有显著抑制作用（均P<0.05）；槟榔多酚可以下调低氧诱导下PMVEC中凋亡相关蛋白胱天蛋白酶（caspase）3、Bcl-2相关X蛋白（Bax）的表达（均P<0.05），从而减轻低氧诱导下细胞的凋亡；槟榔多酚显著上调低氧诱导下PMVEC闭合蛋白和闭锁小带1的表达（均P<0.05），提高跨膜电阻值并降低罗丹明透过量，改善肺微血管内皮屏障的通透性。结论：槟榔多酚可以通过减轻PMVEC的氧化应激损伤、降低炎症相关蛋白表达、减少细胞凋亡及提高细胞屏障的通透性，改善PMVEC的低氧损伤。

Protective effects of areca nut polyphenols on hypoxic damage of rat pulmonary microvascular endothelial cells

Objective: To investigate the protective effects of areca nut polyphenols on hypoxic damage of rat pulmonary microvascular endothelial cells (PMVECs). Methods: Malondialdehyde and superoxide dismutase (SOD) were used to determine the optimal modeling of lung hypoxic injury cells. CCK-8 method was used to detect cell viability for determining the effective dose of areca nut polyphenols. Rat PMVECs were divided into control group, hypoxia model group and areca nut polyphenols group. BCA method was used to detect the protein concentration of each group, and the oxidative stress level in PMVECs was measured. Western blotting was used to detect the expression of inflammatory and apoptosis-related proteins. Immunofluorescence staining was used to detect the expression of occludin and zonula occludens (ZO) 1. Transwell chamber was used to detect transendothelial electrical resistance, and rhodamine fluorescent dye was used to detect PMVECs barrier permeability. Results: The hypobaric hypoxia-induced cell injury model was established by culturing PMVECs for 48?h at 1% oxygen concentration. The 20?μg/mL areca nut polyphenols significantly reversed the survival rate and the oxidative stress of PMVECs in hypoxia model group (all P<0.05). Areca nut polyphenols had significant inhibitory effect on the up-regulation of inflammation-related proteins, including nuclear factor-κB (NF-κB) and nuclear factor-E2-related factor (Nrf) 2 in hypoxia model group (allP<0.05). And areca nut polyphenols could reduce hypoxia-induced PMVECs apoptosis by down-regulating the expressions of apoptosis-related proteins, including cysteine aspartic acid specific protease (caspase) 3, Bcl-2 associated X protein (Bax) in PMVECs (allP<0.05). In addition, areca nut polyphenols effectively improves the transendothelial electrical resistance and barrier permeability of PMVECs through elevating the expression of occludin and ZO-1 (allP<0.05).Conclusion: Areca nut polyphenols can inhibit the hypoxic damage of PMVECs by reducing oxidative stress and apoptosis down-regulating the expression of inflammatory proteins and reducing membrane permeability.