目的观察自身免疫性脑脊髓炎(EAE)小鼠发病后不同时间大脑组织中LINGO-1表达水平的变化。方法将C57/BL6小鼠完全随机化分为EAE模型组(15只)、佐剂组(15只)和对照组(15只),在发病第1、7、14、21、30天取大脑半球组织进行实时定量RT—PCR和Western印迹检测,观察LINGO-1的表达差异。同时应用Western印迹检测不同时间点RhoA、RhoA磷酸化形式(P—RhoA)的变化。结果EAE模型组小鼠发病后第1、7、14天LINGO-1mRNA表达水平是对照组的(4.63±0.25)、(2.72±0.12)和(1.98±0.16)倍(P〈0.01,P〈0.01,P〈0.05),至第30天时接近于对照组(P〉0.05);模型组LINGO—1蛋白表达在发病后第1、7、14、21、30天分别为对照组的(2.11±0.15)、(3.15±0.09)、(2.45±0.12)、(1.89±0.17)和(1.21±0.05)倍(P〈0.05,P〈0.01,P〈0.05,P〈0.05,P〈0.05,P〈0.05);模型组小鼠p-RhoA蛋白表达也呈上调趋势,在造模成功1、7d上调最明显(P〈0.01)。而RhoA蛋白的表达无明显变化。结论LINGO-1在EAE小鼠脑组织中表达上调并呈-定规律性改变,可能通过RhoA激活抑制髓鞘修复。临床上针对多发性硬化的LINGO-1阻断治疗应在发病后尽早进行。
Objective To observe the changes of LINGO-1 expression with time after onset in EAE mouse. Methods C57/BL6 mice were completely randomly divided into EAE model group (n = 15 ), adjuvant group( n = 15) and control group( n = 15). LINGO-1 expression of brain tissue was detected on day 1,7,14,21 and 30 after onset by RT-PCR and Western blot. RhoA and p-RhoA expression of brain tissue was analysed by Western blot. Results The LINGO-lmRNA levels in EAE model group were markedly higher than control group on day 1,7and 14 after onset (4. 63 ±0.25,2.72 ±0. 12,1.98 ±0. 16, P 〈0. 01, P 〈 0. 01, P 〈 0. 05 ). On day 30, Lingo-1 mRNA was close to control group. Expression levels of Lingo-1 protein on day 1,7,14,21,30 were higher than control group (2. 11 + 0. 15,3.15 ± 0. 09,2. 45 ± 0. 12,1.89 ± 0. 17,1.21 ± 0. 05,P 〈 0. 05, P 〈 0. 01, P 〈 0.05, P 〈 0. 05,P 〈 0. 05. The levels of p-RhoA protein increased in EAE and the peak appeared on day 1 and day 7 (P 〈 0. 01 ). And there was no difference on RhoA expression among different groups. Conclusions LINGO-1 expression of brain tissue of EAE mouse upregulates and changes with time after onset, which may inhibit myelination by RhoA activation. In clinic, the antagonist of LINGO-1 for MS should be applied as soon as possible.