摘要
目的探讨高糖条件下培养的牛视网膜微血管内皮细胞、周细胞线粒体活性氧产生量的变化及导致此种变化的原因和所致影响。方法采用选择性培养方法培养牛视网膜微血管内皮细胞、周细胞,通过共聚焦显微镜检测不同葡萄糖浓度下内皮细胞、周细胞线粒体活性氧产生量的变化,同时采用流式细胞仪检测线粒体膜电位、细胞死亡率的变化,RTPCR检测培养细胞锰超氧化物歧化酶及解藕联蛋白(UCP)1、2、3表达情况和不同葡萄糖浓度下的变化。结果(1)随着培养基中葡萄糖浓度的增加,内皮细胞、周细胞线粒体活性氧的产生呈增多趋势。(2)随着培养基中葡萄糖浓度的增加,内皮细胞线粒体膜电位逐渐升高,细胞死亡率逐渐增多,而周细胞则无此变化趋势。(3)培养的视网膜微血管内皮细胞、周细胞中,通过RTPCR检测到UCP1、2mRNA,未检测到UCP3。UCP1、2及锰超氧化物歧化酶表达量随培养基中葡萄糖浓度的变化而变化。结论高糖可以诱导培养的视网膜微血管内皮细胞、周细胞线粒体活性氧产生增多;内皮细胞活性氧产生的增多与线粒体膜电位增高间存在反馈调节;高糖条件下UCP1、2及锰超氧化物歧化酶表达量代偿性增多调节活性氧的产生,但是当糖浓度达到一定程度时,代偿机制消失;内皮细胞、周细胞在高糖条件下表现出的不同特征,说明其在糖尿病性视网膜病变的发生中起着不同的作用。
Objective To study changes of reactive oxygen species （ROS） in mitochondria of bovine retinal endothelial cells and pericytes cultured in high-glucose and its relevance with the pathogenesis of diabetic retinopathy. Methods Bovine retinal endothelial cells and pericytes were cultured with selective culture media, ROS changes in mitochondria of retinal endothelial cells and pericytes in response to different concentrations of glucose were detected with scanning laser confocal microscope. The mitochondria membrane potential （MMP） and cell death ratio （ CDR ） were measured with flow cytometry. Expression of MnSOD and uncoupling protein （UCP） were detected with reverse-transcriptase PCR （RT-PCR）, Results In highglucose, increased ROS was seen in mitochondria of retinal endothelial cells and pericytes. However MMP and CDR only increased in endothelial cells, no change was found in pericytes. UCP1 ,2 mRNA expressed in cultured cells whereas UCP3 was negativ, The expression of UCP1 ,2,MnSOD mRNA changed with the variation of glucose concentrations, Conclusions Increasing of ROS production in mitochondria of retinal endothelial cells and pericytes can be induced by high glucose. There is feedback accommodation mechanism between ROS increasing and MMP elevation in endothelial cells, The different features of endothelial cells and pericytes in high glucose demonstrate their roles in pathogenesis of diabetic retinopathy are different.
出处
《中华眼科杂志》
CAS
CSCD
北大核心
2006年第2期131-138,共8页
Chinese Journal of Ophthalmology
关键词
活性氧
线粒体
细胞
培养的
视网膜血管
内皮细胞
逆转录聚合酶链反应
Reactive oxygen species
Mitochondria
Cell, cultured
Retinal vessels
Endothelial cells
Reverse transcriptase polymerase chain rection
