托法替布对EAE大鼠的疗效观察及机制探讨

Curative effect of tofacitinib on EAE rats and its mechanism

为观察托法替布对EAE大鼠的疗效,探讨相关机制,揭示托法替布对EAE大鼠的防治作用,将50只Wistar大鼠分为正常对照组,EAE对照组,小、中、大剂量托法替布组,10只/组。用豚鼠脊髓匀浆制备抗原诱导EAE模型。自造模之日起EAE对照组及小、中、大剂量托法替布组分别以生理盐水和托法替布1、2、4 mg/(kg·d)灌胃,连续10 d。于发病高峰期行神经功能障碍评分(neurological dysfunction score,NDS)并处死大鼠,留取血清及脑组织。采用ELISA测定血清IL-21及IL-10水平;苏木精-伊红(hematoxylin-eosin,HE)染色观察脑组织病理改变;免疫组化法检测脑白质脱髓鞘及星形胶质细胞活化情况。结果显示,正常对照组均未发病,EAE对照组出现精神萎靡、肢体瘫痪甚至死亡;各剂量托法替布组大鼠NDS、血清IL-21水平、脑组织炎性细胞浸润程度及抗胶质原纤维酸性蛋白(glial fibrillary acidic protein,GFAP)抗体阳性细胞平均光密度(D)均降低(P<0.05),而IL-10水平及髓鞘碱性蛋白(myelin basic protein,MBP)染色阳性表达平均D则升高(P<0.05),且均呈剂量依赖关系。提示托法替布对EAE大鼠具有治疗作用,且呈剂量依赖关系,可能的机制为托法替布通过降低IL-21水平、升高IL-10水平,明显减轻脑组织炎性细胞浸润程度及脑白质脱髓鞘病变,抑制脑组织内星形胶质细胞活化,从而减轻EAE临床表现,降低NDS。

To study the curative effect of tolfatinib on EAE rats and its mechanism,fifty Wistar rats were assigned into normal control group,EAE control group,low-dose,median-dose and high-dose tofacitinib group(10 for each).The spinal cord homogenate of guinea pig was used as antigen to induce EAE model.The control group was fed saline,and low-dose,median-dose and high-dose tofacitinib group were fed tofacitinib 1,2 and 4 mg/(kg·d)for 10 days.The neurological dysfunction score(NDS)of the onset of the peak was recorded,and rats were executed to collect serum and brain tissue.The levels of IL-10 and IL-21 in serum were evaluated by ELISA.The pathologic changes of rat brain tissue were observed by hematoxylin-eosin(HE)staining.The degree of demyelination and active astrocytes in brain tissues were detected via immunohistochemistry.The results showed that the normal control group had no disease.In the EAE control group,mental depression,limb paralysis and even death occurred.After intervention with tofacitinib,the NDS of the onset of the peak were shortened,the level of IL-21 declined,the inflammatory cell infiltration and the average optical density(D)of anti-glial fibrillary acidic protein(GFAP)antibody positive cells decreased(P<0.05).The degree of myelin basic protein(MBP)and active astrocytes were enhanced significantly,while the level of IL-10 increased in each tofacitinib group(P<0.05),and in a dose-dependent manner.So tofacitinib has therapeutic effects in rats with EAE in a dose-dependent manner,and the mechanism could be that tofacitinib inhibits the inflammatory cells infiltration,myelinoclasis and activation of astrocytes by decreasing IL-21 and increasing IL-10.