沉默髓系细胞触发受体1基因对神经病理性痛大鼠的影响

Effect of silencing TREM1 on rats with neuropathic pain

目的探讨髓系细胞触发受体1(triggering receptor expressed on myeloid cells 1,TREM1)在大鼠神经病理性痛中的作用及可能机制。方法成年雄性SD大鼠,体重220~300 g,取鞘内置管成功大鼠48只,随机分为四组(n=12):对照组(S组)、神经病理性痛组(CCI组)、TREM1sh RNA组(RNAi组)和阴性慢病毒组(Virus组)。采用慢性坐骨神经压榨性损伤法(CCI)制备神经病理性痛模型。RNAi组于造模前1周鞘内注射p GLVU6/RFP/Puro-sh RNA 30μl(1×109IU/ml);Virus组、CCI组和S组分别鞘内注射等量阴性慢病毒和生理盐水。于造模前1 d和造模后1、3、7、14d测定机械缩足反应阈(MWT)和热缩足潜伏期(TWL)。于造模后14 d痛阈测定结束后,处死大鼠,取L4-5节段脊髓组织,采用Western blot法测定脊髓TREM1、TLR4、My D88、IκBα和p-NF-κB p65蛋白含量,RT-PCR法测定脊髓IL-1β、TNF-α和IL-6 mRNA表达量。结果与S组比较,CCI组和Virus组TREM1蛋白含量明显增加(P<0.05);与CCI组比较,RNAi组TREM1蛋白含量明显降低(P<0.05)。与S组比较,CCI组、RNAi组和Virus组造模后各时点MWT和TWL明显降低(P<0.05);脊髓TLR4、My D88和p-NF-κB p65蛋白含量明显增加,IκBα蛋白含量明显降低(P<0.05);IL-1β、TNF-α和IL-6 mRNA表达量明显升高(P<0.05)。与CCI组比较,RNAi组大鼠造模后各时点MWT和TWL明显升高(P<0.05);脊髓TLR4、My D88和p-NF-κB p65蛋白含量明显降低,IκBα蛋白含量明显增加(P<0.05);IL-1β、TNF-α和IL-6 mRNA表达量明显降低(P<0.05)。结论干扰TREM1基因可以缓解大鼠神经病理性痛,其机制可能与抑制TLR4/My D88/NF-κB通路有关。

Objective To investigate the role of triggering receptor expressed on myeloid cells 1（ TREM1） in rats with neuropathic pain and its possible mechanism. Methods Forty-eight male adult Sprague-Dawley rats,weighing 220-300 g,were successfully placed intrathecal catheters,and then randomly divided into 4 groups（ n = 12） ： sham operation group（ group S）,neuropathic pain group（ group CCI）,TREM1 sh RNA group（ group RNAi） and negative lentivirus group（ group Virus）. The neuropathic pain was induced by chronic sciatic nerve compression injury（ CCI）. In group RNAi,30 μl p GLVU6/RFP/Puro-sh RNA（ 1×10^9 IU/ml） was injected intrathecally 1 week before modeling. Group Virus was injected with 30 μl negative lentivirus,whereas group CCI and group S with equal amount of normal saline. MWT and TWL were measured 1 day before（ baseline） and 1,3,7,14 day after modeling. When behavioral test finished,the expression levels of TREM1,TLR4,My D88,IκBα and p-NF-κB p65 in spinal cord were determined by Western blot. Whereas the mRNA expression levels of IL-1β,TNF-α and IL-6 in spinal cord were measured by RTPCR. Results Compared with group S,the expression levels of TREM1 in groups CCI and Virus significantly increased（ P〈0. 05）. While compared with group CCI,the TREM1 expression of group RNAi in spinal cord significantly decreased（ P〈0. 05）. Compared with group S,MWT and TWL of groups CCI,Virus and RNAi after modeling and the expression of IκBα significantly decreased（ P〈0. 05）,whereas the expression of TLR4,My D88,p-NF-κB p65 increased significantly（ P〈0. 05）,as well as the expression of IL-1β,TNFα and IL-6 mRNA（ P〈0. 05）. Compared with group CCI,the MWT and TWL of group RNAi after modeling and the expression of IκBα remarkably increased（ P〈0. 05）,whereas the expression of TLR4,MyD 88 and p-NF-κB p65 in the spinal cord remarkably decreased（ P〈0. 05）,as well as the expression of IL-1β,TNF-α and IL-6 mR NA（ P〈0. 05）. Conclusion TREM1 knockdown can alleviate neuropathic pain,the underlying mechanism might be the inhibition of TLR4/MyD 88/NF-κB signaling pathway.