芍药苷抑制FasL诱导的Fas/FasL信号转导通路的实验研究

目的探讨芍药苷对Fas配体（FasL）诱导大鼠颈椎间盘纤维细胞的Fas/FasL信号转导通路的影响。方法采用1月龄SD大鼠（雌雄不拘）的颈椎间盘纤维环,采用酶消化法,建立椎间盘纤维环细胞体外培养。传至3代纤维环细胞,建立FasL诱导体外培养纤维环细胞凋亡模型。实验中,将细胞随机分为空白组、模型组和药物组,模型组应用适宜浓度的Fas配体（FasL）诱导其凋亡,药物组在FasL诱导的同时给予适宜浓度芍药苷培养液干预。Annexin V/FITC法检测细胞凋亡率,分别鉴定模型复制情况及观察芍药苷保护作用;荧光定量PCR技术检测Fas、Caspase-3、Caspase-8基因表达水平。结果药物组纤维环细胞的凋亡率明显低于模型组（P〈0.01）,药物组的Fas、Caspase-3、Caspase-8凋亡因子基因的表达明显低于模型组（P〈0.01）。结论芍药苷可通过下调纤维环细胞的Fas、Caspase-3、Caspase-8的基因表达,抑制Fas/FasL信号转导通路,从而降低纤维环细胞的凋亡率。

Experimental Research of Paeoniflorin Inhibition on Fas/FasL Signal Transduction Pathway Induced by FasL

Objective To explore the impacts of paeoniflorin on Fas/Fasl signal transduction pathway of the cervical disc fiber cells induced by Fasl in the rats.Methods The cervical intervertebral disc annulus fibrosus was collected from SD rats(either sex)of 1 month old.The enzyme digestion was adopted to establish the in vitro culture of intervertebral disc annulus fibrosus cells.When the annulus fibrosus cells were passed down till the 3rd generation,the apoptosis model of the in vitro annulus fibrosus cells induced by FasL was set up.In the experiment,the cells were randomized into a blank group,a model group and a medicine group.In the model group,FasL of the proper concentration was used to induce apoptosis.In the medicine group,at the same time of Fasl induction,paeoniflorin culture solution of a proper concentration was given for the intervention.Annexin V/FITC was adopted to detect the apoptosis rate,identify the model duplication and observe the protection of paeoniflorin.The fluorescence quantitative polymerase chain reaction(FQ-PCR)technique was used to detect the expression levels of Fas,Caspase-3 and Caspase-8.Results In the medicine group,the apoptosis rate of annulus fibrosus cells was lower apparently than that in the model group(P<0.01).In the medicine group,the expressions of apoptotic factor genes as Fas,Caspase-3 and Caspase-8 were lower obviously than those in the model group(P<0.01).Conclusion The paeoniflorin inhibits the transduction pathway of Fas/FasL signals through the down-regulation of the genetic expressions of Fas,Caspase-3 and Caspase-8 of annulus fibrosus cells so that the apoptosis rate of annulus fibrosus cells can be decreased.