黄绿青霉素对心肌细胞线粒体呼吸链合成相关转录调节基因mRNA表达、膜电位及活性氧的影响  

Impact of citreoviridin on mRNA expression of mitochondrial respiratory chain synthesis related transcriptional regulation gene, membrane potential and reactive oxygen species in rat cardiomyocytes

作　　者：魏灿[1] 孙树秋[1] 

机构地区：[1]哈尔滨医科大学中国疾病预防控制中心地方病控制中心克山病防治研究所,150081

出　　处：《中华地方病学杂志》2013年第4期384-388,共5页Chinese Journal of Endemiology

摘　　要：目的观察黄绿青霉素（CIT）对sD大鼠原代心肌细胞线粒体呼吸链合成相关转录调节基因mRNA表达、膜电位及活性氧的影响。方法用不同浓度CIT[0、1、2、3、4、5、6、7、8、9、10μmol／L]作用SD大鼠原代心肌细胞24h，采用噻唑蓝（MTr）法测定细胞存活率，并根据检测结果，用SPSS13．0软件计算CIT的半数抑制浓度（IC。）。进一步选择心肌细胞存活率为99％、95％、90％时的CIT水平（0．715、1．234、1．650txmol／L）作为低、中、高剂量组，同时以未加CIT作为对照组，分别作用于心肌细胞24h，采用反转录聚合酶链反应（RT—PCR）检测心肌细胞中过氧化物酶体增殖物受体叮共激活因子-1α（PGC-1α）、核呼吸因子1（Nrfl）、核呼吸因子2（Nrf2）mRNA表达；分别以阳离子荧光羰花青染色剂（JC-1）、2’，7＇-二氯荧光素（DCFH2一DA）作为荧光探针，用全能酶标仪检测心肌细胞线粒体膜电位（MMP）和细胞内活性氧（ROS）的变化。结果与0txmoL／LCIT组[（89．4±17．6）％]比较，2～10μmol／LCIT组原代心肌细胞存活率[（80．2±20．2）％、（74．4±18．7）％、（63．2±8．9）％、（51．5±18．8）％、（39．0±15．7）％、（22．6±10．5）％、（19．9±4．9）％、（20．7±4．8）％、（18．5±3-3）％]明显下降（P均〈0．05）。CIT作用于心肌细胞24h的IC。为4．6μmol／L。高、中、低剂量组PGC—letmRNA表达（0．431±0．041、0．619±0．031、0．653±0．037）较对照组（0．776±0．081）明显降低（P均〈0．05）；高剂量组NIflmRNA表达（0．358±0．051）明显低于对照组（0．580±0．098，P〈0．05）；高、中剂量组Nrf2mRNA表达（0．352±0．041、0．472±0．011）明显低于对照组（0．667±0．091，P均〈0．05）。与对照组[（100．00±0．00）％、（100．00±0．00）％]比较，高、中、低剂量组MMP水平[（55．34-3．3）％、（69．8±4．Objective To investigate the impact of citreoviridin（CIT） on mRNA expression ofmitochondrial respiratory chain synthesis related transcriptional regulation gene, mitochondrial membrane（MMP） potential and reactive oxygen species （ROS） in eardiomyocytes of rat. Methods Viability of rat primary cardiomyocytes treated with different concentrations of CIT （0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 Ixmol/L） for 24 h was determined by 3- （4,5-Dimethyhhiazol-2-yl）-2,5-diphenyhetrazolium bromide（MTF） method. Based on the MTI＇ curve, median inhibitory concentration （IC±） was calculated using SPSS 13.0. High-, medium- and low-dose groups of CIT（ 1.650, 1.234, 0.715 μmol/L） were defined corresponding to 99%, 95% and 90% of cardiomyocyte viability, respectively. CIT was not added in as the control group. After 24 hours, the mRNA expression levels of peroxisome-proliferatoractivated receptor /coactivator（PGC-1α）, nuclear respiratory factor l（Nrfl） and nuclear respiratory factor 2（Nrf2） in cardiomyocytes were detected by reverse transcriptase polymerase chain reaction（RT-PCR）. Changes of MMP and intracellular ROS were determined by a fluorescence microplate reader using 5,5＇ ,6,6＇-tetrachloro-1,1＇ ,3,3＇- tetraethylbenzimidazolcarbocyanine iodide （JC-1） and 2,7-dichloroflnorescein diacetate （DCFH2-DA） as fluorescent probes. Results Compared with 0 Ixmol/L CIT group [ （89.4 ± 17.6）%], viabihties of rat primary cardiomyocytes treated with 2 - 10 I±mol/L CIT groups[（80.2 ＋ 20.2）%, （74.4 ± 18.7）%, （63.2 ± 8.9）%, （51.5 ± 18.8）%, （39.0 ± 15.7）%, （22.6±10.5）%, （19.9 ± 4.9）%, （20.7 ± 4.8）%, （18.5 ± 3.3）%] decreased significantly（aU P 〈 0.05）. The ICs0 value of eardiomyocytes after 24 h treatment with CIT was 4.6 p,mol/L. The PGC-lct mRNA expressions of high-, medium- and low-dose groups（0.431 ± 0.041, 0.619 ± 0.031, 0.653 ± 0.037） were significantly lower compared to that of the control group（0.776 ± 0.081, all

关 键 词：黄绿青霉素 肌细胞 心脏 线粒体 膜电位 活性氧 

分 类 号：R542.3[医药卫生—心血管疾病]