雷公藤内酯醇对实验性自身免疫性脑脊髓炎大鼠血-脑脊液屏障的影响

The Effect of Triptolide on the Blood-Cerebrospinal Fluid Barrier in Wistar Rats with Experimental Autoimmune Encephalomyelitis

摘要:目的探讨雷公藤内酯醇(Tri)对实验性自身免疫性脑脊髓炎(EAE)大鼠血-脑脊液屏障(BCB)的免疫调节作用。方法制备Wistar大鼠EAE模型,应用Tri治疗EAE,评估EAE组、Tri治疗组的临床症状,在不同时间点测定脑脊液(CSF)和外周血白蛋白(Alb)水平并计算二者比值(QA),通过QA值观察BCB变化。于免疫后(p.i.)6、8、10、12、14、16 d处死动物,用免疫组化技术检测脑和脊髓细胞间黏附分子-1(ICAM-1)和干扰素-γ(IFN-γ)表达。结果免疫后14 d为EAE组发病高峰期,Tri治疗组临床症状评分(0.14±0.08)较EAE组(2.19±1.68)低(P<0.01)。免疫后8 d EAE组QA达最大值(0.30±0.05),较佐剂组(0.13±0.05)明显增高(P<0.01)。免疫后6-14 d,EAE组与佐剂组、Tri治疗组比较QA值差异均有统计学意义(F=28.34,P<0.01)。免疫后14 d Tri治疗组QA达最大值(0.19±0.02),QA波峰时间较EAE组延迟且峰值明显较低。免疫后12-16 d,EAE组与佐剂组、Tri治疗组比较ICAM-1阳性表达高(F=17.1l,F=29.87,均P<0.01);免疫后10-16 d,Tri治疗组较EAE组IFN-γ阳性表达低(F=447.92,P<0.01)。结论 EAE中BCB损伤早于临床症状出现。Tri能有效抑制EAE进展,可能与其下调ICAM-1、IFN-γ表达,恢复BCB正常功能有关。

Objective To study the immunoregulatory effect of triptolide（Tri） on the blood-cerebrospinal fluid harrier（BCB） of experimental autoimmune encephalomyelitis（EAE）. Methods EAE was induced in Wistar Rats and treated with Tri. Clinical symptoms were evaluated among EAE and Tri-treated groups. Cerehrospinal fluid （CSF） and peripheral blood at different time points in rats were collected for determining alhumin（Alb）and evaluating BCB permeability. The rats were executed, brains and spinal cords were extracted on the days from 6 to 16 p.i. in order to detect ICAM-1 and IFN -γ by means of immunohistochemistry staining. Results On day 14 p.i. the symptoms reached the peak in EAE group. Meanwhile the mean score for clinical symptoms in Tri group was lower than that in EAE group（ 0.14±0. 08 vs 2.19±1.68, P〈0. 01）. On day 8 p. i. , CSF to serum Alb quotient（QA） reached the peak in EAE group, which was higher than that in Freund adjuvant group （0.30± 0.05 vs0.13±0.05, P〈0.01）. In Tri group QA reached the maximum（0.19±0.02） on the day of 14 p.i. , which was later than that in EAE group. Our result suggested that on days 6-14 p. i the difference of QA was statistically significant between EAE group and Tri group（F=28.34,P=0.01）. On days 12-16 p. i. , the mean photodensity of ICAM-1 in Tri group was lower than that in EAE group（F=17.11, P〈0.01）. Variance analysis indicated on the day 10-16 p. i. the expression of IFN-γ in EAE group was significantly increased compared with Tri group （F = 447.92, P〈 0.01）. Conclusions The injury of BCB is prior to appearance ot clinical symptoms in early EAE stage. Tri could not only down regulated ICAM 1 and IFN-γ but also repaired the normal run.ion of BCB and ameliorate EAE.