四环素对糖皮质激素诱导的骨髓间充质干细胞损伤的保护作用

Protection of tetracycline on damage of rat bone mesenchymal stem cells induced by glucocorticosteroid

目的探讨四环素对糖皮质激素诱导大鼠骨髓间充质干细胞(BMSCs)损伤的保护作用。方法体外建立糖皮质激素诱导大鼠BMSCs损伤模型,分离与体外培养大鼠BMSCs。将大鼠BMSCs分为正常对照组、激素对照组,四环素4、20、100、500μg/ml浓度组。采用吖啶橙/溴化乙锭双荧光染色法观察四环素对激素诱导的BMSCs凋亡的影响;采用MTT法检测各组BMSCs活性,绘制细胞生长曲线;用免疫组织化学方法对细胞凋亡因子bcl-2、bax进行分析;Western blot测定细胞凋亡因子caspase-3表达。结果吖啶橙/溴化乙锭染色显示细胞损伤为凋亡。四环素对激素抑制BMSCs的增殖有明显保护作用,并与四环素的浓度成一定的量效关系;100μg/ml的四环素能明显阻断激素诱导的BMSCs凋亡,其机制可能与提高bcl-2表达、降低bax、caspas-3表达有关。结论四环素具有保护激素诱导的BMSCs损伤的作用,这为早期干预治疗激素性骨坏死提供了依据。

Objective To investigate the effect of tetracycline on rat bone mesenchymal stem cells （BMSCs） injury indued by glucocorticosteroid. Methods The model of rat BMSCs injury induced by glucoconicosteroid in vitro was established, and then isolately cultured in vitro. The BMSCs of rats were divided into six groups ： normal control group, hormonone control group, 4, 20, 100,500μg/ml tetracycline group. The injury of BMSCs induced by the glucocorticosteroid was detected by Acridine orange-ethidium bromide double staining. The cytotoxicity of BMSCs was measured by MTT assay, and then drawing cell growth curve. Staining and analysis of bcl-2, bax epressions were detected by immunohistoehemical. Western blot was used to detect the expression of caspase-3. Results Acridine orange-ethidium bromide double staining showed that the cell damage was apoptosis. Tetracycline had protection effect on cell proliferation of BMSCs inhibited by glueocorticosteroid by dose-dependent mode. 100μg/ml tetracycline could induce the apoptosis of BMSCs by increasing the bcl-2, decreasing bax,caspas-3. Conclusion Tetracycline can protect the injury of BMSCs induced by glucocorticosteroid. This provide some experimental pharmaco-foundation for intervention therapy of early osteonecrosis.