摘要
目的初步探索补阳还五汤对大鼠髓核细胞(nucleus pulposus cell,NPC)退变的抑制效应与Nod样受体蛋白3(Nod-like receptor protein 3,NLRP3)/胱天蛋白酶-1(caspase-1)信号通路之间的关系。方法离体培养大鼠NPC,得到的细胞根据处理方法分为空白组(0.9%氯化钠溶液预处理)、应力拉伸组(应力拉伸刺激+0.9%氯化钠溶液预处理)和补阳还五汤组(应力拉伸刺激+1.0g/L补阳还五汤制剂预处理),采用细胞体外拉伸应力刺激法构建大鼠NPC退变模型。采用实时荧光定量聚合酶链反应(real time fluorescent quantitative polymerase chain reaction,qRT-PCR)和蛋白质印迹法检测3组细胞聚集蛋白聚糖(aggrecan)、NLRP3、caspase-1的mRNA及蛋白表达情况;采用酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)检测3组细胞上清液白细胞介素(interleukin,IL)-1β、IL-18的表达水平。结果qRT-PCR和蛋白质印迹法检测结果显示,应力拉伸组NLRP3、caspase-1的mRNA与蛋白表达较空白组显著上升,aggrecan的mRNA与蛋白表达较空白组显著下降,差异均有统计学意义(P<0.05);补阳还五汤组NLRP3、caspase-1的mRNA与蛋白表达较应力拉伸组显著下降,aggrecan的mRNA与蛋白表达较应力拉伸组显著上升,差异均有统计学意义(P<0.05)。ELISA检测结果显示,应力拉伸组IL-1β、IL-18的表达水平较空白组显著上升,差异均有统计学意义(P<0.05);补阳还五汤组IL-1β、IL-18的表达水平较应力拉伸组显著下降,差异均有统计学意义(P<0.05)。结论补阳还五汤可通过抑制NLRP3/caspase-1信号通路的激活,减轻细胞炎症反应,延缓细胞外基质降解重构,从而发挥防治NPC退变的作用。
Objective To explore the relationship between the inhibitory effect of Buyang Huanwu Decoction on the degeneration of rat nucleus pulposus cell(NPC)and the signaling pathway of Nod-like receptor protein 3(NLRP3)/caspase-1.Methods Rat NPC were cultured in vitro.The obtained cells were divided into blank group(pre-treatment with 0.9%sodium chloride solution),stress stretching group(stress stretching stimulation+0.9%sodium chloride solution pretreatment)and Buyang Huanwu Decoction group(stress stretching stimulation+1.0g/L Buyang Huanwu Decoction preparation pretreatment)according to the treatment method.The rat NPC degeneration model was established by cell tensile stress stimulation.Real time fluorescent quantitative polymerase chain reaction(qRT-PCR)and Western blotting were used to detect the mRNA and protein expression of aggrecan,NLRP3 and caspase-1 in three groups.The expression levels of interleukin(IL)-1βand IL-18 in cell supernatant of three groups were measured by enzyme-linked immunosorbent assay(ELISA).Results The results of qRT-PCR and Western blot-ting showed that the mRNA and protein expressions of NLRP3 and caspase-1 in stress stretching group were signif-icantly higher than those in blank group,and the mRNA and protein expressions of aggrecan in stress stretching group were significantly lower than those in blank group,the differences were statistically significant(P<0.05).The mRNA and protein expressions of NLRP3 and caspase-1 in Buyang Huanwu Decoction group were significantly lower than those in stress stretching group,and the mRNA and protein expressions of aggrecan in Buyang Huanwu Decoction group were significantly higher than those in stress stretching group,the differences were statistically sig-nificant(P<0.05).The results of ELISA showed that the expression levels of IL-1βand IL-18 in stress stretching group were significantly higher than those in blank group,the differences were statistically significant(P<0.05).The expression levels of IL-1βand IL-18 in Buyang Huanwu Decoction group
作者
叶辛
梅正峰
李陶冶
蒋啸为
张贻春
YE Xin;MEI Zhengfeng;LI Taoye;JIANG Xiaowei;ZHANG Yichun(Department of Orthopaedics,Hangzhou Third People's Hospital,Hangzhou 310009,China)
出处
《中国现代医生》
2022年第21期66-70,共5页
China Modern Doctor
基金
杭州市科技发展计划项目(20191203B110)。
