丹参酮ⅡA对兔纤维环细胞能量代谢保护作用.doc

丹参酮ⅡA对兔纤维环细胞能量代谢保护作用 　 作者：陈勇　杨述华　张其川　周建国　黄吉军　熊蠡茗 【摘要】 ：［目的］考察丹参酮ⅡA(TSⅡA)对白细胞介素-1β(IL-1β)诱导的兔纤维环细胞能量代谢障碍的保护作用。［方法］藻酸盐串珠立体培养兔纤维环细胞，将细胞分为7组，在培养过程中加入不同浓度的药物：A组为空白对照不加入药物，B组加入4 μg／ml TSⅡA，C组加入10ng／ml IL-1β，D～G组在给予10 ng／mlIL-1β同时分别加入0．5、1、2和4 μg／ml TSⅡA。于培养3 d后行Na+-K+-ATP酶活性检测、，琥珀酸脱氢酶活性检测、MTT法细胞增殖情况检测以及细胞凋亡的流式细胞仪检测。［结果］G组Na+-K+-ATP酶活性(3.23±0.28U/mgprot)较C组(1.118±0.15U／mgprot)明显增高(Plt;0.01)，与A组接近(3.57±0.15 U／mgprot)(Pgt;0.05)。G组琥珀酸脱氢酶活性(12.48±0.97U／mgprot)较c组(3.03±0.60 U／mgprot)明显增高(Plt;0.01)，与A组接近(14.24±1.56 U／mgprot)(Pgt;0.05)。G组MTT试验吸光度(0.77±0.06)较C组(0.31±0.07)明显增高(P＜0.01)，随着TSⅡA浓度的升高，D～G组吸光度随着TsIIA上升而上升。G组细胞死亡细胞比例和凋亡细胞比例分别为21.08±1.46%和8.99±0.33%，均显著低c组(43.11±2.7，Plt;0.01和11.71±0.32，Plt;0.01)。［结论］TSIIA能够减轻IL-1β对纤维环细胞能量代谢的抑制作用，从而改善纤维环细胞的增殖、死亡及凋亡。 【关键词】 丹参酮ⅡA　椎间盘　白细胞介素-1β　Na+-K+1-ATP酶　琥珀酸脱氢酶 　　1 Materials and methods 　　1.1 Materials and animals: TSⅡA standard preparation was purchased from the National Institute for the control of pharmaceutical and biological products; Fetal bovine serum, DMEM-F12 culture medium and collagenase II from Gibco Co. (USA); MTT kit from Lingfei Company of Science and Technology (Wuhan); pancreatin, DMSO, interleukin-1β (IL-1β), alginate from Sigma Co. (USA); the ATPase test kit and the succinate dehydrogenase (SDH) test kit from Jiancheng Biological Company of Science and Technology (Nanjin); BCA quantitative test for protein kit (Wuhan Boster Biological Technology Co. Ltd.), annexin V-FITC kit (Jingmei Biotech Co. Ltd.), Ordinary Japanese rabbit was provided by animals center of Tongji medical college of Huazhong University of Science and Technology. 　　1.2 Experimental methods 　　1.2.1 Isolation culture of rabbit annulus fibrosus (AF) cells: Back skins was incised from posterior in sterile environment and exposed lumbar in paragraph 1 to 6 after killed rabbit by venous air embolism, and then take out the lumbar which has exposed paragraph. After that stripping the intervertebral disc, pulposus and non-annulus fibrosus tissue were removed. An