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目的探讨纳米氧化铝颗粒对小鼠血脑屏障通透性的影响。方法 3月龄ICR小鼠用滴鼻法染毒,为了比较纳米铝作用的颗粒毒性和化学毒性,将小鼠分为对照组、微米铝组、纳米碳组、纳米铝组和铝离子组;为了比较纳米铝作用的剂量-效应关系,将小鼠分为对照组、低剂量组、中剂量组和高剂量组。每只小鼠每次染毒10μl,3次/d,实验时间为30 d。染毒结束后,用硝酸镧染色法检测血脑屏障通透性;用免疫荧光法检测紧密连接蛋白ZO-1、Occludin蛋白表达;用western blot法检测ZO-1,Claudin-5蛋白表达。结果硝酸镧染色结果表明,对照组的硝酸镧颗粒聚集在毛细血管中央,未见对血管内皮的穿透;微米铝组和纳米碳组有部分高电子密度的硝酸镧颗粒位于毛细血管内壁中央,显示血脑屏障的通透性增高;纳米铝和铝离子组有大量硝酸镧颗粒位于血管内壁中央,形成致密的黑色条带,显示血脑屏障受损和通透性增高。免疫荧光法检测紧密连接蛋白ZO-1、Occludin蛋白表达和western blot法检测ZO-1,Claudin-5蛋白表达结果表明,紧密连接蛋白ZO-1、Occludin和Claudin-5蛋白表达在对照组均为高表达,微米铝和纳米碳组表达下调(P<0.0001),纳米铝和铝离子组的表达量最低(P<0.0001)。且这种紧密连接蛋白表达的降低与纳米铝的染毒剂量间有剂量-反应关系(P<0.0001)。结论纳米氧化铝可以降低血脑屏障通透性,这种作用表现为纳米的颗粒作用和铝化学毒性作用的相加。其发生机制与紧密连接蛋白ZO-1、Occludin和Claudin-5蛋白表达降低有关。
Objective To investigate the effect of nano-alumina particles on permeability of blood-brain barrier in mice. Methods The 3 - month - old ICR mice were intranasally inoculated. In order to compare the particle toxicity and chemical toxicity of aluminum nanoparticles, the mice were divided into control group, micron aluminum group, carbon nano group, nano aluminum group and aluminum ion group. To compare the dose-effect relationship of nanoaluminum, mice were divided into control group, low dose group, middle dose group and high dose group. Each mouse exposure to 10μl, 3 times / d, the experimental time was 30 d. After the exposure, the permeability of blood-brain barrier was detected by lanthanum nitrate staining; the expression of tight junction protein ZO-1 and Occludin was detected by immunofluorescence; and the expression of ZO-1 and Claudin-5 protein was detected by western blot. Results The results of lanthanum nitrate staining showed that the lanthanum nitrate particles in the control group gathered in the center of the capillaries and no penetration into the vascular endothelium was observed. Part of the high-electron-density lanthanum nitrate particles in the microparticulate aluminum group and the nanocarbon group were located at the center of the capillaries, Showing increased permeability of the blood-brain barrier; a large number of lanthanum nitrate particles in the nanoaluminum and aluminum ion group were located in the center of the inner wall of the blood vessel to form a dense black band, indicating that the blood-brain barrier is impaired and the permeability is increased. The expression of ZO-1 and Occludin proteins and the expression of Claudin-5 protein in ZO-1 and western blot were detected by immunofluorescence. The results showed that the expressions of tight junction proteins ZO-1, Occludin and Claudin-5 in control group were Highly expressed, the expression of micronized aluminum and carbon nano group was down-regulated (P <0.0001), and the lowest expression of nano-aluminum and aluminum ion group (P <0.0001). Moreover, the decrease of the expression of Claudin was dose-response relationship with the dose of nano aluminum (P <0.0001). Conclusion nano-alumina can reduce the permeability of the blood-brain barrier. This effect is manifested by the addition of nano-particles and aluminum chemical toxicity. The mechanism is related to the decrease of tight junction protein ZO-1, Occludin and Claudin-5 protein expression.