中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2015年
39期
3223-3228
,共6页
郭守刚%张杰%汪春娟%杜怡峰
郭守剛%張傑%汪春娟%杜怡峰
곽수강%장걸%왕춘연%두이봉
实验性自身免疫性脑脊髓炎%LINGO-1%基因沉默%髓鞘
實驗性自身免疫性腦脊髓炎%LINGO-1%基因沉默%髓鞘
실험성자신면역성뇌척수염%LINGO-1%기인침묵%수초
Experimental autoimmune encephalomyelitis%LINGO-1%Gene silencing%Melination
目的 探讨沉默LINGO-1基因表达对自身免疫性脑脊髓炎(EAE)小鼠运动功能的影响.方法 通过MOG35-55诱导雌性C57/BL6小鼠建立EAE小鼠模型.将EAE小鼠(105只)完全随机分为5组:A组(21只):侧脑室注入5μl滴度为5×109 Tu/ml编码LINGO-1shRNA的慢病毒载体(LV/LINGO-1-shRNA);B组(21只):侧脑室注入5μl滴度为5×10s Tu/ml LV/LINGO-1-shRNA;C组(21只):侧脑室注入5μl滴度为5×107 Tu/ml LV/LINGO-1-shRNA;D组(21只):侧脑室注入5μl无义序列的LVCON053;E组(21只):未治疗组.动态观察干预后不同时间点小鼠神经功能评分情况及LINGO-1蛋白表达水平,并行髓鞘快蓝染色了解各组髓鞘脱失情况.结果 侧脑室注射LV/LINGO-1-shRNA组EAE小鼠脑组织内LINGO-1蛋白表达从干预后第7天起可见明显下调,其中以B组和C组下调最明显(1.99±0.13,2.08±0.10,P＜0.05,P＜0.01).从第7天开始,相比较于D组和E组,A组、B组和C组运动功能评分有不同程度地下降[(3.11 ±0.13)分,(2.42 ±0.13)分,(2.96 ±0.10)分比(3.56 ±0.15)分,(3.87 ±0.12)分,P＜0.01,P＜0.01,P＜0.05].30 d时A组和B组髓鞘密度明显高于未治疗组(0.72±0.09,0.83 ±0.11比0.56±0.10,P＜0.05,P＜0.01).结论 侧脑室注射LV/LINGO-1 shRNA是一种沉默EAE小鼠脑内LINGO-1表达的有效方式.LINGO-1下调可改善EAE小鼠运动功能及促髓鞘修复,但效果并不随LV/LINGO-1-shRNA剂量增加而增强.
目的 探討沉默LINGO-1基因錶達對自身免疫性腦脊髓炎(EAE)小鼠運動功能的影響.方法 通過MOG35-55誘導雌性C57/BL6小鼠建立EAE小鼠模型.將EAE小鼠(105隻)完全隨機分為5組:A組(21隻):側腦室註入5μl滴度為5×109 Tu/ml編碼LINGO-1shRNA的慢病毒載體(LV/LINGO-1-shRNA);B組(21隻):側腦室註入5μl滴度為5×10s Tu/ml LV/LINGO-1-shRNA;C組(21隻):側腦室註入5μl滴度為5×107 Tu/ml LV/LINGO-1-shRNA;D組(21隻):側腦室註入5μl無義序列的LVCON053;E組(21隻):未治療組.動態觀察榦預後不同時間點小鼠神經功能評分情況及LINGO-1蛋白錶達水平,併行髓鞘快藍染色瞭解各組髓鞘脫失情況.結果 側腦室註射LV/LINGO-1-shRNA組EAE小鼠腦組織內LINGO-1蛋白錶達從榦預後第7天起可見明顯下調,其中以B組和C組下調最明顯(1.99±0.13,2.08±0.10,P＜0.05,P＜0.01).從第7天開始,相比較于D組和E組,A組、B組和C組運動功能評分有不同程度地下降[(3.11 ±0.13)分,(2.42 ±0.13)分,(2.96 ±0.10)分比(3.56 ±0.15)分,(3.87 ±0.12)分,P＜0.01,P＜0.01,P＜0.05].30 d時A組和B組髓鞘密度明顯高于未治療組(0.72±0.09,0.83 ±0.11比0.56±0.10,P＜0.05,P＜0.01).結論 側腦室註射LV/LINGO-1 shRNA是一種沉默EAE小鼠腦內LINGO-1錶達的有效方式.LINGO-1下調可改善EAE小鼠運動功能及促髓鞘脩複,但效果併不隨LV/LINGO-1-shRNA劑量增加而增彊.
목적 탐토침묵LINGO-1기인표체대자신면역성뇌척수염(EAE)소서운동공능적영향.방법 통과MOG35-55유도자성C57/BL6소서건립EAE소서모형.장EAE소서(105지)완전수궤분위5조:A조(21지):측뇌실주입5μl적도위5×109 Tu/ml편마LINGO-1shRNA적만병독재체(LV/LINGO-1-shRNA);B조(21지):측뇌실주입5μl적도위5×10s Tu/ml LV/LINGO-1-shRNA;C조(21지):측뇌실주입5μl적도위5×107 Tu/ml LV/LINGO-1-shRNA;D조(21지):측뇌실주입5μl무의서렬적LVCON053;E조(21지):미치료조.동태관찰간예후불동시간점소서신경공능평분정황급LINGO-1단백표체수평,병행수초쾌람염색료해각조수초탈실정황.결과 측뇌실주사LV/LINGO-1-shRNA조EAE소서뇌조직내LINGO-1단백표체종간예후제7천기가견명현하조,기중이B조화C조하조최명현(1.99±0.13,2.08±0.10,P＜0.05,P＜0.01).종제7천개시,상비교우D조화E조,A조、B조화C조운동공능평분유불동정도지하강[(3.11 ±0.13)분,(2.42 ±0.13)분,(2.96 ±0.10)분비(3.56 ±0.15)분,(3.87 ±0.12)분,P＜0.01,P＜0.01,P＜0.05].30 d시A조화B조수초밀도명현고우미치료조(0.72±0.09,0.83 ±0.11비0.56±0.10,P＜0.05,P＜0.01).결론 측뇌실주사LV/LINGO-1 shRNA시일충침묵EAE소서뇌내LINGO-1표체적유효방식.LINGO-1하조가개선EAE소서운동공능급촉수초수복,단효과병불수LV/LINGO-1-shRNA제량증가이증강.
Objective To explore the effect of LINGO-1 silencing on movement function of experimental autoimmune encephalomyelitis (EAE) mice.Methods EAE was established by induction of MOG35-55 in female C57/BL6 mice.Then female EAE mice (n =105) were completely randomly divided into 5 groups:group A (n =21) :5 μl 5 × 109 Tu/ml lentiviral vectors encoding LINGO-1shRNA (LV/ LINGO-1-shRNA) by intracerebroventricular (ICV) injection, group B (n =21) :5 μl 5 × 108Tu/ml LV/LINGO-1-shRNA by ICV injection, group C (n =21) : 5 μl 5 × 107 Tu/ml LV/LINGO-1-shRNA by ICV injection,group D (n =21):5 μl LVCON053 by ICV injection and group E (n =21):untreated.The movement function was scored and the expression of LINGO-1 protein was detected by Western blot on day 1,3,7,14,21,30 after ICV among different groups.Luxol fast blue staining was performed to know about conditions of myelin sheath on day 30.Results The expression of LINGO-1 in EAE mouse was obviously downregulated ever since day 7 after LV/LINGO-1-shRNA implantation.Group B and C achieved the most reduction of LINGO-1 expression (1.99 ± 0.13,2.08 ± 0.10, P ＜ 0.05, P ＜ 0.01).Simultaneously, the movement functional score of group A, B and C was lowered at different levels from day 7 (3.11 ± 0.13, 2.42 ±0.13,2.96 ±0.10 vs 3.56 ±0.15,3.87 ±0.12,P ＜ 0.01,P ＜0.01, P ＜0.05) ,with the most marked decrease in group B.The densities of myelin sheaths in group A and B were higher than untreated group on day 30 (0.72 ± 0.09,0.83 ± 0.11 vs 0.56 ± 0.10, P ＜ 0.05, P ＜ 0.01).Conclusion LV/ LINGO-1shRNA by ICV injection is an effective method to silence LINGO-1 expression.LINGO-1 silencing could ameliorate motor function and promote formation of myelin sheaths.But the effects do not enhance with the increase of LV/LINGO-1-shRNA dose.
