南蛇藤总萜抑制中性粒细胞胞外诱捕网形成的作用与机制研究

The effect and mechanism of the total terpenoids of Celastrus orbiculatus Thunb. on inhibiting the formation of neutrophil extracellular traps

为探索南蛇藤总萜(the total terpenoids of Celastrus orbiculatus Thunb.,TTC)对中性粒细胞胞外诱捕网(neutrophil extracellular traps,NETs)形成及其作用机制,通过分离与制备中性粒细胞,建立佛波酯(phorbol 12-myristate 13-acetate,PMA)诱导的NETs形成细胞模型。利用瑞氏-姬姆萨染色、扫描电镜与免疫荧光技术研究南蛇藤总萜对NETs形成及其生物学标志的影响,并利用Annexin V/PI试验研究南蛇藤总萜对中性粒细胞凋亡的影响。结果表明:南蛇藤总萜能剂量依赖性地抑制PMA诱导的NETs形成;南蛇藤总萜(80μg·mL^(-1))可阻断中性粒细胞弹性蛋白酶(neutrophil elastase,NE)的释放,并破坏NETs相关DNA-NE复合物的形成;南蛇藤总萜(80μg·mL^(-1))可显著抑制PMA诱导的瓜氨酸化组蛋白H3(citrullinated histone H3,CitH3)的表达水平;南蛇藤总萜可剂量依赖性地促进中性粒细胞凋亡。综上,南蛇藤总萜对中性粒细胞NETs的形成具有抑制作用,其机制可能与其直接诱导中性粒细胞凋亡有关。

To investigate the mechanisms and the effect of the total terpenoids of Celastrus orbiculatus Thunb.(TTC) on NETs, the cell model of NETs formation induced by phorbol 12-myristate 13-acetate(PMA) was established by isolating and preparing neutrophils. The effects of TTC on the formation of NETs and their biological markers were studied by Wright-Giemsa Staining, scanning electron microscope and immunofluorescence. The Annexin V/PI experiment was used to study TTC on neutrophil apoptosis. Results showed that TTC could dose-dependently inhibit the formation of NETs induced by PMA;TTC(80 μg·mL^(-1)) could block the release of neutrophil elastase(NE) and disrupt the formation of Nets associated DNA-NE complexes. TTC(80 μg·mL^(-1)) significantly inhibited PMA-induced CitH3 expression;TTC can promote neutrophil apoptosis in a dose-dependent manner. This study suggests that TTC can inhibit the formation of neutrophil NETs, and its mechanism may be related to its direct induction of neutrophil apoptosis.