IL-10基因修饰的未成熟树突状细胞在大鼠角膜移植排斥反应中的作用

Role and its mechanism of immature dendritic cells with IL-10 gene modified in rats after keratoplasty

目的:通过建立大鼠角膜穿透性移植模型,探讨IL-10基因修饰的未成熟树突状细胞在大鼠角膜移植排斥反应中的作用及其作用机制。方法:建立大鼠同种异体穿透性角膜移植模型,将受体SD大鼠随机分为:阳性对照组、GFP-DC组、8-DC组及IL-10-GFP-DC组,分别于角膜移植术前3d尾静脉注射等量的PBS、供体Wistar大鼠骨髓源8-DC(培养8d的DC)、转染48h的GFP-DC及IL-10-GFP-DC。术后每天在裂隙灯下观察角膜植片情况,记录排斥反应指数及角膜植片存活时间,在移植术后第14d行各组角膜组织的病理学检查及免疫组织化学检查。结果:IL-10-GFP-DC组角膜植片存活时间较GFP-DC组、8-DC组比较显著延长(P<0.01)。术后第14d时IL-10-GFP-DC组角膜植片的混浊、水肿、新生血管及排斥指数均显著降低(P<0.01)。病理组织学检查结果显示各实验组角膜植片的炎症反应较阳性对照组轻,植片中央未见明显新生血管。免疫组织化学结果显示:IL-10-GFP-DC组的CD4^+、CD8+、CD25^+、IL-2^+、NK^+及NF-κB+阳性细胞数量较阳性对照组、GFP-DC组、8-DC组减少,差异均具有显著统计学意义(P<0.01)。结论:经过供体来源未成熟树突状细胞预处理的受体,角膜植片的存活时间显著延长,成功诱导角膜移植免疫耐受。CD4^+、CD8^+、CD25^+、IL-2^+、NK^+及NF-κB^+阳性细胞参与了同种异体角膜移植排斥反应的调控,IL-10-GFPDC可降低CD4^+、CD8^+、CD25^+、IL-2^+、NK^+及NF-κB^+阳性细胞的浸润,抑制角膜移植排斥反应的发生。

AIM： Through the establishment of penetrating keratoplasty m odel of rats, to detect the role and its m echanism of im m ature dendritic cells with IL-10 gene m odified.METHODS： Allogeneic penetrating corneal transplantation in rat m odel was perform ed. SD rats were random ly divided into positive control group,GFPDC group,8-DC and IL-10-GFP-DC group. At 3d before keratoplasty, the rats were given tail intravenous injection with the sam e am ount of PBS,bone m arrow8-DC（ DC had cultured for 8d） from donor Wistar rats,GFP-DC after 48 h transfection and IL-10-GFP-DC. Rats were observed under slit-lam p for corneal graft cases every day,and recorded rejection index and corneal graftsurvival tim e. At 14 d after keratoplasty,pathologic and im m unohistochem ical exam inations were perform ed.RESULTS： Compared with GFP-DC group and 8-DC group,corneal graft survival tim e of IL-10-GFP-DC group was significantly longer（ P〈0. 01）; at 14 d after keratoplasty,corneal opacity,edem a,neovascularization and rejection index of IL-10-GFP-DC group were significantly lower（ P〈0. 01）. Pathological exam ination showed that in the three experim ental groups corneal inflam m ation was lighter than the positive control group without significant central graft neovascularization.Im m unohistochem istry showed： com pared to the positive control group,GFP-DC group and 8-DC group,CD4~＋,CD8~＋,CD25~＋,IL-2＋,NK＋and NF-κB＋positive cells in IL-10-GFP-DC group were lower（ P〈0. 01）.CONCLUSION： After donor-derived immature dendritic cells pretreated, corneal graft survival was significantly prolonged, successfully induced corneal transplantation tolerance. CD4~＋,CD8~＋,CD25~＋,IL-2＋,NK＋and NF-κB＋positive cells are involved in corneal allograft rejection regulation,IL-10-GFP-DC m ay reduce CD4~＋,CD8~＋,CD25~＋,IL-2＋,NK＋and NF-κB＋positive cell infiltration,inhibit corneal transplant rejection.