对羟基苯甲醛对体外神经血管单元氧化应激损伤的保护作用研究

Protective effects of p-hydroxybenzaldehyde on oxidative stress-induced injury in neurovascular units in vitro

目的探讨对羟基苯甲醛(p-HBA)对过氧化氢(H_(2)O_(2))诱导的体外神经血管单元(NVU)氧化应激损伤模型的保护作用。方法建立神经元、血管内皮细胞和星形胶质细胞共培养NVU体系,分为空白组、对照组、模型组、阿托伐他汀组(1μmol/L)、p-HBA低、中、高剂量组(1、10、100μmol/L)。阿托伐他汀组、p-HBA低、中、高剂量组采用相应浓度受试药干预NVU体系24 h后,除对照组外,其余各组诱导NVU氧化应激损伤模型。检测血脑屏障渗漏值及神经营养因子等指标。结果干预4 h后,与对照组比较,模型组渗漏值、脑源性神经营养因子(BDNF)、神经营养素3(NT-3)、血管内皮生长因子(VEGF)水平及神经元凋亡率明显升高,胶质纤维酸性蛋白(GFAP)、闭合蛋白(occludin)、突触素蛋白表达明显降低(P<0.05)。与模型组比较,阿托伐他汀组、p-HBA低、中、高剂量组渗漏值、神经元凋亡率明显降低,阿托伐他汀组BDNF、occludin、突触素表达明显升高(P<0.05),p-HBA低剂量组VEGF,p-HBA中剂量BDNF、VEGF、occludin、突触素以及p-HBA高剂量组BDNF、NT-3、VEGF、GFAP、occludin、突触素表达水平明显升高(P<0.05)。p-HBA低剂量组BDNF明显低于阿托伐他汀组[(15.50±10.00)pg/ml vs(24.70±8.07)pg/ml,P<0.05],p-HBA高剂量组VEGF水平明显高于阿托伐他汀组[(38.40±5.05)pg/ml vs(22.50±7.04)pg/ml,P<0.05]。结论p-HBA对体外NVU氧化应激损伤具有保护作用,其机制与减轻紧密连接蛋白的破坏从而降低血脑屏障通透性,升高脑神经营养因子和突触素,减少神经元凋亡,促进神经修复有关。

Objective To investigate the protective effect of p-HPA on hydrogen peroxide(H_(2)O_(2))-induced injury in NVU in vitro.Methods A NVU model was established with co-culture of neurons,vascular endothelial cells and astrocytes.Then the following experiment was performed on the blank group,control group,model group and atorvastatin group(1μmol/L),and low-,medium-and high-dose p-HPA groups(1,10 and 100μmol/L).The NVU model was firtstly treated with corresponding drugs,and then exposed to 500μmol/L H_(2)O_(2) for 4 h except the control group.The leakage of BBB was observed,and related nutritional factors were detected.Results After 4 h of intervention,the leakage value,neuronal apoptotic rate,and levels of BDNF,NT-3 and VEGF were significantly increased,and the expression levels of GFAP,occludin,and synaptoin were obviously reduced in the model group than the control group(P<0.05).When compared with the model group,the atorvastatin group and the low-,medium-and high-dose p-HBA groups had significantly decreased leakage value and neuronal apoptotic rate;the atorvastatin group had elevated levels of BDNF,occluding and synaptosin(P<0.05);the low-dose p-HBA group had increased VEGF level,the medium-dose p-HBA group had elevated levels of BDNF,VEGF,occludin and synaptoin,and the high-dose p-HBA group had higher levels of BDNF,NT-3,VEGF,GFAP,occluding and synaptosin(P<0.05).The BDNF level in the low-dose p-HBA group was significantly lower than that in the atorvastatin group(15.50±10.00 pg/ml vs 24.70±8.07 pg/ml,P<0.05)and the VEGF in the high-dose p-HBA group was significantly higher than that in the atorvastatin group(38.40±5.05 pg/ml vs 22.50±7.04 pg/ml,P<0.05).Conclusion p-HPA shows protective effects on NVU against oxidative stress injury in vitro,and the related mechanisms involve in alleviating the destruction of tight junction protein and then decreasing the BBB permeability,and reducing the apoptosis of neurons by increasing neurotrophic factors and synaptosin to promote the neurorestoration.