12/15-脂氧合酶抑制剂黄芩素对软骨细胞凋亡的抑制作用研究  

Inhibition of 12/15 lipoxygenase by baicalein alleviates chondrocyte apoptosis

作　　者：陈恺哲 燕宇飞[1,2] 袁俊[1,2] 郭蕾[2] 何川[1,2] 

机构地区：[1]上海交通大学医学院附属瑞金医院骨科 [2]上海市伤骨科研究所,上海200025

出　　处：《生物骨科材料与临床研究》2017年第3期5-8,I0003,I0004,共6页Orthopaedic Biomechanics Materials and Clinical Study

基　　金：上海市卫生和计划生育委员会科研基金项目(201540275);上海市中西医结合临床重大项目(ZY3-LCPT-2-2003)

摘　　要：目的探讨12/15-脂氧合酶抑制剂黄芩素(Baicalein)对硝普钠(SNP)诱导的软骨细胞凋亡的抑制作用及可能机制。方法取8周雄性SD大鼠膝关节软骨,采用Ⅱ型胶原酶消化法提取软骨细胞并体外培养。设置对照组、SNP凋亡组和Baicalein给药组,以0.5 m M SNP作用24小时诱导软骨细胞凋亡,以5 M,25 M,50 M,100 M不同浓度的Baicalein作用细胞,确定最适浓度,对照组仅加入同体积溶剂(蒸馏水)。CCK8法检测药物对细胞毒性;Annexin V/PI双染法检测细胞凋亡;免疫荧光观测凋亡诱导因子(AIF)在软骨细胞中的定位。结果 Baicalein在各浓度下对软骨细胞无明显毒性,与对照组相比,凋亡组软骨细胞活性明显下降(<0.01),与凋亡组相比,给药组细胞活性浓度依赖性地升高(<0.01);流式检测显示对照组软骨细胞早期凋亡率3.16%,凋亡组27.8%,100 M Baicalein给药组14.1%;免疫荧光检测显示,凋亡组AIF出现核内移位,100 M Baicalein给药组AIF核移位受到抑制。结论 Baicalein通过抑制12/15-脂氧合酶活性,进而抑制AIF从线粒体中的释放及核转位,发挥抗软骨细胞凋亡作用。Objective To investigate the inhibitory effect and possible pathogenesis of Baicalein,12/15-lipoxygenase inhibitor, on apoptosis of chondrocytes induced by sodium nitroprusside （SNP）. MethodsControl group, SNP group and Baicalein group were setting. The 0.5mM SNP was used to induce apoptosis for 24 hours, meanwhile, Baicalein group was treated additionally with a variety of concentration gradient of 5μM,25μM,50μM,100μM to determine the optimum concentration. The control group received only the same volume of solvent. The cytotoxicity of reagents was detected by CCK8. The Annexin V/PI double staining was used to determine the apoptosis rate. The immunofluorescence was used to detect the intracellular localization of apoptosis inducing factor （AIF） in chondrocytes. Results Baicalein has no statistically significant in toxicity to chondrocytes at multiple concentration gradients, the activity of chondrocytes in the SNP group decreased significantlycompared with the control group（P〈0.01）, and Baicalein increased the cytoactive in a concentration-dependent manner compared with the SNP group（P〈0.01）, flow cytometry analysis showed that the early apoptosis rate of chondrocytes was 3.16% in control group, 27.8% in SNP group and 14.1% in 100μM Baicalein group, respectively. Immunofluorescence assay showed that the nuclear translocation of AIF was observed in SNP group compared with the control group, while the nuclear translocation of AIF was inhibited in Baicalein-treated group. Conclusion The 12/15-lipoxygenase inhibitor, Baicalein, has anti-apoptosis effect on chondrocytes through inhibiting the release of AIF from mitochondria and nuclear translocation.

关 键 词：花生四烯酸盐12-脂氧合酶 花生四烯酸盐15-脂氧合酶 脂氧合酶抑制剂 软骨细胞 细胞凋亡 

分 类 号：R684.3[医药卫生—骨科学]