B7H4在间充质干细胞C3H10T1／2移植治疗小鼠实验性变态反应性脑脊髓炎中的作用研究

The preliminary study for the role of B7-H4 for the treatment of experimental autoimmune encephalo- myelitis by mesenchymal stem cell C3H10 T1/2 transplantation

目的探讨B7．H4分子在C3H10T1／2细胞（C3H10）移植治疗小鼠实验性变态反应性脑脊髓炎（EAE）中的作用。方法①用慢病毒将B7-H4靶向shRNA转染至C3H10细胞（C3H10-shRNA）并观察其对小鼠脾脏细胞活化增殖的调节作用；②利用MOG35-55和完全弗氏佐剂制备小鼠EAE模型，将50只小鼠分为正常对照组、EAE模型组、C3H10细胞组、C3H10-NC细胞组、C3H10-shRNA细胞组，通过HE染色、Fast-blue染色观察病理学改变，用免疫组化法检测B7-H4表达，结合神经功能评分观察不同干预方式对EAE损伤组织结构和功能的影响。结果①下调表达B7-H4的C3H10细胞可使其对小鼠脾细胞的免疫抑制作用减弱；②C3H10移植于EAE小鼠可减少、延缓发病，而移植C3H10-shRNA细胞的EAE小鼠发病时间、神经缺损评分和炎性细胞浸润、脱髓鞘改变及B7-H4的阳性细胞数均介于EAE组与其他对照组之间。结论C3H10细胞移植治疗EAE的安全、有效，B7-H4分子可能通过影响C3H10的免疫调节作用等生物学行为而影响移植疗效。

Objective To investigate the role of the negative co-stimulatory molecule B7-H4 in mesen- chymal stem cell line C3H10 T1/2 （C3H10） in the treatment of experimental allergic encephalitis（EAE）. Methods （1）The lentiviral vectors with mouse B7-H4 target shRNA were transfected into mouse mesenehymal stem cell （C3H10-shRNA） ; the flow cytometry was used to detect the proliferation of splenocyte that was cocultured with C3H10 cell. （2）EAE mice models were immunized with peptide MOG35-55 in complete Freund＇ s adjuvant （CFA） ; C57BL/6 miee（n = 50） were divided into Sham-operated group, EAE model group, C3H10 cell group,C3H10-NC cell group, and C3H10-shRNA cell group. Through HE and luxol fast blue staining to observe the pathologic changes of mice. Immunohistochemistry was used to label B7-H4. Combining with the neural function defect score , the effect of transplantation of mice with different cells was evaluated. Results （1） By knocking down B7-H4 on C3H10, the inhibitory effect of C3H10 on splenocyte proliferation was partly revised ;（2）Transplantation of C3H10 into EAE mice can decrease and delay the attack. The onset time of the attack, the neural function defect score, the inflammatory cell infiltration, the demyelination and the number of B7-H4 positive cells of the C3H10-shRNA cell group was between EAE model group and other control groups. Conclusions The treatment of EAE by C3H10 transplantation is safe and effective. The co-inhibitor molecule B7-H4 expressed on C3H10 cell involved in the effect of the treatment of EAE by C3H10 transplantation.