摘要
目的 研究铝对大鼠原代脉络丛上皮细胞的损伤作用.方法 在无菌条件下,取出生后14 d SD大鼠的脉络丛组织,经眼科剪剪碎,胰酶消化后,将悬浮细胞分别接种于含1％表皮生长因子（EGF）、20％胎牛血清的DMEM培养基中.5d后,玻片上的细胞用特异性转甲状腺素蛋白抗体（anti-TTR）做免疫组化鉴定其纯度.以不同剂量[对照组、低剂量组（100 μmol/L）、中剂量组（400 μmol/L）、高剂量组（1 600 μmol/L）]乳酸铝处理生长状态良好的细胞,48 h后分别测定各组细胞存活率、细胞凋亡率、活性氧（ROS）水平及超氧化物歧化酶（SOD）活力,评估铝对原代脉络丛上皮细胞的损伤作用.结果 经纯度鉴定95％以上培养细胞为脉络丛上皮细胞.与对照组（100％）比较,中（86.74％±4.03％）、高剂量组（81.90％±9.17％）细胞存活率明显下降,差异有统计学意义（P＜0.01）;与低剂量组（92.92％±8.81％）比较,高剂量组细胞存活率明显下降,差异有统计学意义（P＜0.01）.与对照组（1.29％±0.03％）、低剂量（1.68％±0.27％）比较,中（7.26％±0.99％）、高剂量组（22.25％±1.55％）凋亡率均明显升高,差异有统计学意义（P＜0.01）;与中剂量组相比,高剂量组凋亡率明显升高,差异有统计学意义（P＜0.01）.与对照组（17.24％±0.09％）和低剂量组（18.31％±0.21％）比较,中（22.23％±0.41％）、高剂量组（25.10％±1.13％）ROS荧光强度增加,差异有统计学意义（P＜0.05）;与中剂量组相比,高剂量组ROS荧光强度增加,差异有统计学意义（P＜0.05）.与对照组[（37.35±1.05） U/g Hb]比较,低[（28.65±0.74） U/g Hb]、中[（22.75±1.94） U/g Hb]、高剂量组[（13.29±0.64） U/g Hb]SOD活力均下降,差异有统计学意义（P＜0.05）;与低剂量组比较,中、高剂量组SOD活力下降,差异有统计学意义（P＜0.05）;与中剂量组相比,高剂量组SOD活力下降,差异有统计学意义（P＜0.05）.其他各组间差异无统计学意义（P＞0.05）.结论 乳酸铝可能造成大鼠原代脉络丛上皮细胞损伤,使其存活率下降,凋亡率升高,并引起细胞的氧化应激损伤.
Objective To investigate the impairment in primary cultured rat choroid plexus epithelial cells （CPECs） induced by aluminum.Methods The choroid plexus isolated from Sprague-Dawley rats 14 days old was cut into pieces and digested by trypsin in the sterile area.The obtained single cells were cultured in DMEM with 1% epidermal growth factor and 20% fetal calf serum.Five days later,immunohistochemistry with anti-transthyretin antibody was used to identify the purity of cultured cells.The well-grown cells were treated with aluminum lactate at different concentrations （0,100,400,and 1 600 μmol/L for control,low-dose,medium-dose,and high-dose groups）.Forty-eight hours later,the cell viability,apoptotic rate,level of reactive oxygen species （ROS）,and activity of superoxide dismutase （SOD） were measured in each group to evaluate the impairment in primary cultured rat CPECs by aluminum.Results More than 95% of the cultured cells were identified as CPECs.The medium-and high-dose groups had significantly lower cell viability than the control group （86.74%±4.03% vs 100%,P〈0.01;81.90%±9.17% vs 100%,P〈0.01）.The high-dose group had significantly lower cell viability than the low-dose group （81.90%±9.17% vs 92.92%±8.81%,P〈0.01）.The medium-and high-dose groups had significantly higher apoptotic rates than the control group （7.26%±0.99% vs 1.29%±0.03%,P〈0.01;22.25%±1.55% vs 1.29%±0.03%,P〈0.01） and the low-dose group （7.26%±0.99% vs 1.68%±0.27%,P〈0.01;22.25%±1.55% vs 1.68%±0.27%,P〈0.01）.The high-dose group had a significantly higher apoptotic rate than the medium-dose group（22.25%±1.55% vs 7.26%±0.99%,P〈0.01）.The medium-and high-dose groups had significantly higher fluorescence intensity of ROS than the control group （22.23%±0.41% vs 17.24%±0.09%,P〈0.05;25.10%±1.13% vs 17.24%±0.09%,P〈0.05） and the low-dose group （22.23%± 0.41% vs 18.31%±0.21%,P〈0.05;25.10%±1.13% vs 18.31%±0.21%,P〈0.05）.The high-dose group had significantly higher fluorescence intensity of ROS than the medium-dose group （25.10%±1.13% vs 22.23%± 0.41%,P〈0.05）.The low-,medium-and high-dose groups had significantly lower SOD activity than the control group[（28.65±0.74） U/g Hb vs （37.35±1.05） U/g Hb,P〈0.05;（22.75±1.94） U/g Hb vs （37.35±1.05） U/g Hb,P〈0.05;（13.29 ±0.64） U/g Hb vs （37.35 ± 1.05） U/g Hb,P〈0.05].The medium-and high-dose groups had significantly lower SOD activity than the low-dose group [（22.75±1.94） U/g Hb vs （28.65±0.74） U/g Hb,P〈 0.05;（13.29±0.64） U/g Hb vs （28.65±0.74） U/g Hb,P〈0.05],while the high-dose group had had significantly lower SOD activity than the medium-dose group[（13.29±0.64） U/g Hb vs （22.75±1.94） U/g Hb,P〈0.05].There were no significant differences in cell viability,apoptotic rate,level of ROS,or activity of SOD between any other two groups （P〉0.05）.Conclusion Aluminum lactate may induce impairment in primary cultured rat CPECs.It reduces the cell viability,elevates the apoptotic rate,and causes oxidative stress.
出处
《中华劳动卫生职业病杂志》
CAS
CSCD
2016年第4期286-290,共5页
Chinese Journal of Industrial Hygiene and Occupational Diseases
基金
2013年度高等学校博士学科点专项科研基金（20131417110003）
关键词
大鼠
脉络丛
上皮细胞
活性氧
超氧化物歧化酶
Rats
Choroid plexus
Epithelial cells
Reactive oxygen species
Superoxide dismutase
