siRNA转染对大鼠髓核细胞TRAIL表达和细胞行为的影响

Effect of siRNA transfection on TRAIL expression and behavior of rat nucleus pulposus cells

[目的]探讨siRNA转染对大鼠髓核细胞TRAIL表达和细胞行为的影响。[方法]采用弯曲鼠尾法建立椎间盘退变模型。处死动物。取退变椎间盘组织分离培养髓核细胞。P1髓核细胞分为4组,分别为空白对照组、TRAIL-siRNA转染组(siTRAIL组)、TNF-α处理组(TNF-α组)和TNF-α处理+TRAIL-siRNA转染组(TNF-α+siTRAIL组)。采用MTT法检测P1髓核细胞增殖,流式细胞仪检测细胞凋亡,Western blot法检测髓核细胞Caspase-3的活性和表达。[结果]相较于空白对照细胞,TRAIL siRNA转染对细胞增殖、凋亡、Caspase-3的表达无明显影响(P>0.05);TNF-α处理引发髓核细胞增殖显著下降(P<0.05),而凋亡率显著升高(P<0.05),Caspase-3的表达显著升高(P<0.05)。TNF-α处理后再用TRAIL-siRNA转染可显著逆转TNF-α对细胞增殖、凋亡和Caspase-3表达的影响(P<0.05)。[结论] TRAIL siRNA转染可沉默TRAIL表达,从而逆转TNF-α诱导大鼠髓核细胞增殖抑制、细胞凋亡和Caspase-3表达。

[Objective] To investigate the effect of siRNA transfection on the expression of tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) and behavior of rat nucleus pulposus cells. [Methods] The model of intervertebral disc degeneration was established by the curved tail method in Sprague-Dawley(SD) rat. After the animals were sacrificed, the nucleus pulposus were harvested to isolate and culture the nucleus pulposus cells. The P1 nucleus pulposus cells were divided into 4 groups and treated correspondingly, namely blank control group(control group), TRAIL-siRNA transfection group(siTRAIL group), TNF-α treatment group(TNF-α group) and TNF-αtreatment + TRAIL-siRNA transfection group(TNF-α+siTRAIL group). MTT method was used to detect P1 nucleus pulposus cells proliferation, flow cytometry was used to detect cell apoptosis, and Western blot method was used to detect the expression of Caspase-3 in the rat nucleus pulposus cells. [Results] Compared with blank control cells, TRAIL siRNA transfection had no significant effect on cell proliferation, apoptosis, and Caspase-3 expression(P>0.05). By contrast, TNF-α treatment decreased cell proliferation(P<0.05), while enhanced cell apoptosis(P<0.05), and increased expression of Caspase-3 significantly(P<0.05). However, TRAIL-siRNA transfection after TNF-αtreatment did significantly reverse the effects of TNF-α on cell proliferation, apoptosis and Caspase-3 expression(P<0.05). [Conclusion]TRAIL siRNA transfection might silence the expression of TRAIL, thereby reversing the proliferation inhibition, apoptosis and Caspase-3 expression of the rat nucleus pulposus cells induced by TNF-α.