丹红有效成分配伍对缺氧损伤脑微血管内皮细胞的保护作用  

Protective effect of combined administration of active ingredients of Danhong on cerebral micro-vascular endothelial cell injured by hypoxia

作　　者：周惠芬[1] 何昱[1] 张宇燕[1] 杨洁红 赵涛 付巍 周鹏[2] 万海同[1] 

机构地区：[1]浙江中医药大学心脑血管病研究所,浙江杭州310053 [2]山东步长制药有限公司,山东菏泽274000

出　　处：《中国中药杂志》2014年第22期4399-4404,共6页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金项目(81173647,81373898,81473587);浙江省自然科学基金项目(LR12H27001,LZ14H270001);浙江省中医药(中西医结合)重点学科(2012-XK-A06)

摘　　要：为研究丹红有效成分配伍对缺氧致原代培养的乳鼠脑微血管内皮细胞(r BMECs)损伤的保护作用。原代培养乳鼠脑微血管内皮细胞并对细胞进行缺氧4 h损伤模型的同时,丹红有效成分(原儿茶醛、丹酚酸B、羟基红花黄色素A、丹参素)配伍作用于r BMECs,MTT确定细胞非毒性剂量,采用比色法测定LDH,SOD活性和MDA水平,RT-PCR法检测ICAM-1,MMP-9,P53 mRNA表达,流式细胞术检测r BMECs细胞周期及早期凋亡的变化。丹红有效成分配伍1,2,3,7,8,9组方显著抑制缺氧细胞上清液中LDH的增加;1,2,4,6,7组方显著增强缺氧细胞内SOD活性;1,2,3,8,9组方显著降低MDA水平;1,2,6,7,9组方显著抑制缺氧诱导的r BMECs早期凋亡,缺氧后细胞P53 mRNA表达明显上调的后果是能引起G1期阻滞和促进细胞凋亡,证实P53基因的调节功能体现在G1期校正点的监测;1,2,5,6,7,8,9组方显著下调P53 mRNA表达;1,4,7,8,9组方显著下调ICAM-1 mRNA表达;1,3,6,9组方显著下调MMP-9 mRNA表达。丹红有效成分配伍对缺氧所致原代培养的r BMECs损伤具有明显的保护作用,其机制与增强细胞抗氧化能力,抑制炎症反应和细胞凋亡有关。为研究方剂配伍规律提供思路,指导临床用药。To study the protective effect of combined administration of active ingredients of Danhong on cultured primary mice＇s brain micro-vascular endothelial cells（ r BMECs） injured by hypoxia. Primary mice＇s brain micro-vascular endothelial cells were cultured to establish the 4 h hypoxia model. Meanwhile,active ingredients（ protocatechuic aldehyde,salvianolic acid B,hydroxysafflor yellow A and tanshinol） of Danhong were administered in r BMECs. The non-toxic dosage was determined by MTT. The leakage of lactate dehydrogenase（ LDH）,cell superoxide dismutase（ SOD） activity and MDA level were detected by the colorimetric method. The expressions of ICAM-1,MMP-9,P53 mRNA were detected by RT-PCR method. Changes in r BMECs cell cycle and early apoptosis were detected by flow cytometry. Danhong＇s active ingredients and prescriptions 1,2,3,7,8,9 could be combined to significantly restrain LDH in hypoxic cells supernatant. Prescriptions 1,2,3,7,8,9 could significantly enhance SOD activity in anoxic cells; Prescriptions 1,2,3,8,9 could significantly decrease the MDA level; Prescriptions 1,2,6,7,9 could significantly inhibit the early r BMECs apoptosis induced by hypoxia. After hypoxia,the up-regulated P53 mRNA expression could cause retardation in G1 phase and promote cell apoptosis. This proved that the regulatory function of P53 gene lay in monitoring of calibration points in G1 phase. Prescriptions1,2,5,6,7,8,9 could significantly down-regulate the P53 mRNA expression; Prescriptions 1,4,7,8,9 could significantly downregulate the ICAM-1 mRNA expression; Prescriptions 1,3,6,9 could significantly down-regulate the MMP-9 mRNA expression. The combined administration of Danhong＇s active ingredients showed a significant protective effect on primary cultured r BMECs injury induced by hypoxia. Its mechanism may be related to the enhancement of cellular antioxidant capacity and the inhibition of inflammatory response and cell apoptosis. This study could provide ideas for researching prescription compatibility,and

关 键 词：有效成分配伍 乳鼠脑微血管内皮细胞 细胞凋亡 抗氧化 抗炎症 

分 类 号：R285[医药卫生—中药学] R285.5[医药卫生—中医学]