载脂蛋白E对实验性自身免疫脑脊髓炎IL．17、IL-6、TNF-α表达的影响

Effect of apolipoprotein E on inflammation in mice with experimental autoimmune encephalomyelitis

目的探讨载脂蛋白E（ApoE）对实验性自身免疫脑脊髓炎（EAE）白介素17（IL-17）、白介素6（IL-6）、肿瘤坏死因子-α（TNF-α）表达的影响。 方法选取10只ApoE基因敲除的C57BL/6J小鼠设为ApoE基因敲除组（ApoE-/-组），同时将40只正常C57BL/6J小鼠按随机数字表法分成野生组（WT组）、ApoE拟肽（COG1120）治疗组、生理盐水组和正常对照组，每组10只。ApoE-/-组、WT组、COG112治疗组和生理盐水组均采用背部皮下多点注射完全抗原（MOG35-55的PBS缓冲液与等量的含卡介苗的完全弗氏佐剂混合而成）方法诱导制作EAE模型，正常对照组用生理盐水代替MOG35-55。COG112治疗组在第2-35天每隔2天背部皮下注射COG112 5 mg/（kg·d），生理盐水组用生理盐水代替COG112。在第0-35天对各组小鼠进行神经功能缺损评分；第35天评分后大脑和脊髓取材，应用HE染色观察炎性细胞浸润情况，应用免疫组化染色检测IL-17、IL-6、TNF-α表达水平。结果ApoE-/-组神经功能缺损峰值评分较WT组明显增高，COG112治疗组神经功能缺损峰值评分较生理盐水组明显降低，差异均有统计学意义（P〈0.05）。ApoE-/-组的炎性细胞浸润程度重于WT组，COG112治疗组的炎性细胞浸润程度较生理盐水组轻。ApoE-/-组、WT组、COG112治疗组和生理盐水组小鼠大脑和脊髓IL-17、IL-6、TNF-α平均吸光度值均明显高于正常对照组，差异均有统计学意义（P〈0.05）；ApoE-/-组小鼠大脑和脊髓IL-17、IL-6、TNF-α平均吸光度值均明显高于WT组，COG112治疗组小鼠大脑和脊髓IL-17、IL-6、TNF-α平均吸光度值均明显低于生理盐水组，差异均有统计学意义（P〈0.05）。结论ApoE缺乏会加重EAE炎性细胞浸润及增加IL-17、IL-6、TNF-α表达；ApoE拟肽的干预可减少炎性细胞浸润及IL-17、IL-6、TNF-α表达，从而在EAE的免疫炎症反应中起保护作用。

Objective To explore the effect of apolipoprotein E （ApoE） on expressions of interleukin （IL）-IT, IL-6 and tumor necrosis factor （TNF）-α in central nervous system of mice with experimental autoimmune encephalomyelitis （EAE）. Methods Ten C57BL/6J ApoE-deficient （ApoE-/-） mice were selected randomly as ApoE-/- group; 40 normal C57BL/rJ mice were randomly divided into wild-type （WT） group, peptide （COG1120）-treated group, vehicle group and normal control group （n=10）, and mice in the ApoE-/- group, WT group, COGll20-treated group and vehicle group were injected with completeantigen （MOG35-55 in an PBS buffer with complete Freund adjuvant containing Bacillus Calmette-Guerin vaccine） to induce EAE models. Mile in normal control group were not induced EAE models. Mice of COG1120-treated group were injected subcutaneously with COG112 （5 mg/kg body weight in saline） every two days from day 2 to day 35 postimmunization. Mice of vehicle group were injected with physiological saline instead of COG112. Neurological severity scale scores from day 0 to day 35 were recorded. On day 35 after immunization, mice were sacrificed; spinal cord and brain tissues were harvested; the inflammatory infiltration was observed through HE staining; the expressions ofIL-17, IL-6 and TNF-α were quantified by immunohistochemistry. Results The peak symptoms at neurological severity scale and degrees of inflammatory cell infiltration in ApoE-/- group were significantly worse than those in WT group （P〈0.05）. The average optical density of IL-17, IL-6 and TNF-α in the ApoE-/- group, WT group, COG1120-treated group and vehicle group was significantly higher than that in the normal control group （P〈0.05）; that in the ApoE-/- group was significantly higher than that in the WT group （P〈0.05）; that in the COG1120-treated group and was significantly higher than that in the vehicle group （P〈0.05）. Conclusions ApoE deficiency in the EAE mile could enhance inflammatory infiltrates into CNS, and increase the levels oflL-17, IL-6 and TNF-ct in CNS. After the ApoE pepfide treatment, the levels ofIL-17, IL-6 and TNF-α in CNS decrease, therefore, ApoE and COG112 in EAE mile may play a protective role.