摘要
目的:旨在探讨TNFAIP3(TNFα诱导蛋白3)对大鼠腰椎间盘突出症的影响,并探讨其潜在的作用机制。方法:通过自体髓核移植法构建大鼠腰椎间盘突出症模型。8只健康大鼠作为假手术组,除未放置髓核外其余操作同造模大鼠一致。造模5d后,将模型大鼠随机分为4组:模型组、阳性对照组(布洛芬,20mg/kg)、AAV空病毒组、TNFAIP3过表达组(150μL AAV-R-TNFAIP3,1×10^(12)vg/mL),每组8只。阳性对照组以20mg/kg腹腔注射布洛芬注射液。TNFAIP3过表达组腹腔注射AAV-R-TNFAIP3病毒载体,AAV空病毒组腹腔注射等量AAV空病毒载体,每周一次,其余时间与假手术组、模型组每天腹腔注射等量生理盐水。连续治疗4周后,Von Frey纤毛测试大鼠的机械缩足阈值评估治疗镇痛效果,HE染色观察大鼠腰椎组织的病理变化,ELISA检测血清中促炎因子肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)和白细胞介素-6(interleukin-6,IL-6)的水平,qRT-PCR和Western Blot检测腰椎组织中TNFAIP3、Sirt1(沉默调节蛋白1)、FOXO3、Wnt3a、β-catenin(β-连环蛋白)的mRNA和蛋白表达情况。结果:相比于假手术组,模型组和AAV空病毒组大鼠机械性痛觉阈值明显降低(P<0.05),腰椎组织炎性细胞及病理损伤明显增加(P<0.05),血清促炎因子TNF-α和IL-1β、IL-6水平明显升高(P<0.05),腰椎组织中TNFAIP3、Sirt1和FOXO3 mRNA和蛋白表达水平明显下调(P<0.05),Wnt和β-catenin明显上调(P<0.05)。相比与模型组,阳性对照组和TNFAIP3过表达组大鼠机械性痛觉阈值明显升高(P<0.05),腰椎组织炎性细胞及病理损伤明显减少(P<0.05),血清促炎因子TNF-α和IL-1β、IL-6水平明显降低(P<0.05);其中TNFAIP3过表达组大鼠腰椎组织中TNFAIP3、Sirt1和FOXO3 mRNA和蛋白表达水平明显上调(P<0.05),Wnt3a和β-catenin明显下调(P<0.05)。结论:TNFAIP3通过调节Sirt1/FOXO3/Wnt/β-catenin轴改善大鼠腰椎间盘突出症的症状及病理变化。
Objective:To investigate the effects of TNFAIP3(TNFα-induced protein 3)on lumbar disc herniation(LDH)in rats and explore its potential underlying mechanisms.Methods:A rat model of LDH was established through autologous nucleus pulposus transplantation.Eight healthy rats were used as the sham operation group,undergoing the same procedures as the modeling rats except for the placement of the nucleus pulposus.Five days after modeling,the model rats were randomly divided into four groups:model group,positive control group(ibuprofen,20mg/kg),AAV empty virus group,and TNFAIP3 overexpression group(150μL AAV-R-TNFAIP3,1×10^(12) vg/mL),with eight rats in each group.The positive control group received an intraperitoneal injection of ibuprofen at 20mg/kg.The TNFAIP3 overexpression group received an intraperitoneal injection of AAV-R-TNFAIP3 viral vector,while the AAV empty virus group received an equal volume of AAV empty viral vector,both administered once a week.The sham operation and model groups received daily intraperitoneal injections of an equal volume of saline throughout the study period.After four weeks of continuous treatment,the mechanical paw withdrawal threshold(MPWT)was assessed using the Von Frey filament test to evaluate the analgesic effect.Pathological changes in lumbar tissues were observed by hematoxylin and eosin(HE)staining.Serum levels of proinflammatory cytokines,including tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),and interleukin-6(IL-6),were measured by enzyme-linked immunosorbent assay(ELISA).The mRNA and protein expression of TNFAIP3,Sirt1,FOXO3,Wnt3a,and β-catenin in lumbar tissues were detected by qRT-PCR and Western Blot,respectively.Results:Compared with the sham operation group,the model and AAV empty virus groups showed significantly decreased MPWT(P<0.05),increased inflammatory cell infiltration and pathological damage in lumbar tissues(P<0.05),elevated serum levels of TNF-α,IL-1β,and IL-6(P<0.05),and downregulated mRNA and protein expression of TNFAIP3,Sirt1,and FOXO3(P<0.05)with upregulated Wnt3a and β-catenin(P<0.05)in lumbar tissues.Compared with the model group,the positive control and TNFAIP3 overexpression groups exhibited significantly increased MPWT(P<0.05),reduced inflammatory cell infiltration and pathological damage in lumbar tissues(P<0.05),and decreased serum levels of TNF-α,IL-1β,and IL-6(P<0.05).Notably,the TNFAIP3 overexpression group showed significantly upregulated mRNA and protein expression of TNFAIP3,Sirt1,and FOXO3(P<0.05)and downregulated Wnt3a and β-catenin(P<0.05)in lumbar tissues.Conclusion:TNFAIP3 improves the symptoms and pathological changes of LDH in rats by modulating the Sirt1/FOXO3/Wnt/β-catenin axis.
作者
王晶
刘大丰
WANG Jing;LIU Dafeng(The Third Hospital of Heilongjiang,Heilongjiang Bei'an 164092,China)
出处
《河北医学》
2025年第7期1109-1115,共7页
Hebei Medicine
基金
黑龙江省教育厅科学技术研究项目,(编号:13213971)。
