P7C3-A20对创伤性脑损伤的PC12细胞凋亡和氧化的影响

Effects of P7C3-A20 on Apoptosis and Oxidation of PC12 Cells in Traumatic Brain Injury

旨在研究3,6-二溴-beta-氟-N-(3-甲氧基苯基)-9H-咔唑-9-丙胺(P7C3-A20)对大鼠肾上腺髓质嗜铬瘤分化细胞株(PC12细胞)创伤性脑损伤(TBI)的修复作用。将细胞分为对照组(A组)、模型组(B组)、0.03μmol·L^(-1)药物治疗组(C组)、0.3μmol·L^(-1)药物治疗组(D组)、3μmol·L^(-1)药物治疗组(E组)和药物空白组(F组),TBI细胞模型通过使用10μL枪头划出横竖相间4 mm的直线来制备。使用CCK8试剂盒检测细胞活力,荧光显微镜观察细胞凋亡、活性氧(ROS)情况,荧光定量PCR仪检测半胱氨酸蛋白酶-3(Caspase-3)、B细胞淋巴瘤/白血病-2基因(Bcl-2)、血红素氧合酶1(HO-1)、NAD(P)H:醌氧化还原酶1(NQO 1)和谷氨酸半胱氨酸连接酶催化亚基(GCLC)的mRNA相对表达量。结果显示:TBI造模后细胞活力极显著降低(P<0.01),0.03μmol·L^(-1)浓度的P7C3-A20能显著提高TBI后的细胞活力(P<0.05),0.3μmol·L^(-1)浓度的P7C3-A20能极显著提高TBI后的细胞活力(P<0.01)。TBI造模后细胞早晚期凋亡细胞比例极显著增加(P<0.01),0.03μmol·L^(-1)浓度的P7C3-A20能显著减少TBI后的早期凋亡细胞比例(P<0.05),极显著减少TBI后的晚期凋亡细胞比例(P<0.01),0.3和3μmol·L^(-1)浓度的P7C3-A20能极显著减少TBI后的早晚期凋亡细胞比例(P<0.01)。TBI造模后活性氧细胞比例极显著增加(P<0.01),0.03和3μmol·L^(-1)浓度的P7C3-A20能显著减少TBI后的活性氧细胞比例(P<0.05),0.3μmol·L^(-1)浓度的P7C3-A20能极显著减少TBI后的活性氧细胞比例(P<0.01)。0.3μmol·L^(-1)浓度的P7C3-A20能极显著减少TBI后Caspase-3的mRNA相对表达量(P<0.01),显著提升TBI后Bcl-2、HO-1、NQO 1和GCLC的mRNA相对表达量(P<0.05)。P7C3-A20对TBI后的PC12细胞有抗凋亡、减轻氧化应激的修复作用。

The study was conducted to explore the repair effect of 3,6-Dibromo-beta-fluoro-N-(3-methoxyphenyl)-9H-carbazole-9-propanamine(P7C3-A20)on traumatic brain injury(TBI)of rat adrenal medullary pheochromoma differentiated cell line(PC12 cells).The cells were divided into control group(group A),model group(group B)and 0.03μmol·L^(-1)drug treatment group(Group C),0.3μmol·L^(-1)drug treatment group(Group D),3μmol·L^(-1)drug treatment group(Group E)and drug control group(Group F).TBI cell models were prepared by scribing straight lines 4 mm apart horizontally and vertically using a 10μL gun tip.The cell viability was detected by CCK8 kit,the apoptosis and reactive oxygen(ROS)situation were observed by fluorescence microscope,and the relative mRNA expression of cysteine aspartate-specific protease-3(Caspase-3),B-cell lymphoma/leukemia-2(Bcl-2),Heme Oxygenase-1(HO-1),NAD(P)H:quinone oxidoreductase 1(NQO 1)and glutamate-cysteine ligase catalytic subunit(GCLC)genes were detected by fluorescence quantitative PCR instrument.The results showed that the cell viability was extremely significantly decreased after TBI modeling(P<0.01).P7C3-A20 at concentrations of 0.03μmol·L^(-1)could significantly increase the cell viability after TBI(P<0.05).P7C3-A20 at a concentration of 0.3μmol·L^(-1)could extremely significantly increase the cell viability after TBI(P<0.01).After TBI modeling,the proportion of early and late apoptotic cells increased extremely significantly(P<0.01).P7C3-A20 at a concentration of 0.03μmol·L^(-1)could significantly reduce the proportion of early apoptotic cells after TBI(P<0.05),and extremely significantly reduce the proportion of late apoptotic cells after TBI(P<0.01),P7C3-A20 at concentrations of 0.3 and 3μmol·L^(-1)could extremely significantly reduce the proportion of early and late apoptotic cells after TBI(P<0.01).After TBI modeling,the proportion of reactive oxygen species increased extremely significantly(P<0.01).P7C3-A20 at concentrations of 0.03 and 3μmol·L^(-1)could significantly reduce the proportion of reactive oxygen species after TBI(P<0.05),0.3μmol·L^(-1)concentration of P7C3-A20 can significantly reduce the proportion of reactive oxygen species after TBI(P<0.01).P7C3-A20 at a concentration of 0.3μmol·L^(-1)can extremely significantly reduce the relative mRNA expression of Caspase-3 after TBI(P<0.01),and significantly increase the relative mRNA expression of Bcl-2,HO-1,NQO 1 and GCLC genes expression level after TBI(P<0.05).P7C3-A20 has the repair effect of anti apoptosis and reducing oxidative stress on PC12 cells after traumatic brain injury.