未成熟树突状细胞诱导辅助型T细胞偏移抑制大鼠高危角膜移植排斥反应的研究

Immature dendritic cells induced by T helper cell migration inhibiting high-risk corneal transplantation rejection reaction of rat

目的探讨未成熟树突状细胞(immatuere dendritic cell,im DC)诱导辅助型T细胞偏移在大鼠高危角膜移植排斥反应的的抑制作用及其机制。方法本研究im DC来源于小骨骨髓,受体选择SD大鼠48只,供体选择Wistar大鼠30只,采取碱烧伤的方式建立同种异体高危角膜移植模型。随机将受体SD大鼠分为成熟树突状细胞(matuere dendritic cell,m DC)组、im DC组及对照组,每组均为16只。m DC组和im DC组:术前7 d将体外培养的供体骨髓来源的m DC或im DC经尾静脉注射,且细胞悬浮于0.1 m L的体外培养液中;对照组:术前7 d将0.1 m L体外培养液经尾静脉注射。术后每天对大鼠角膜植皮进行裂隙灯检查,对各组大鼠角膜植皮存活情况进行准确评估;分别于术后3 d、7 d、14 d每组各将入组的4只大鼠处死并取角膜植皮组织进行HE染色,对各组大鼠脾脏CD4+CD25+调节性T细胞表面的Scurfin蛋白表达进行Western blot检测分析。结果 im DC组角膜植片存活时间为(18.67±2.46)d,均较m DC组(6.78±0.56)d、对照组(9.28±1.24)d明显延长,差异均有统计学意义(均为P<0.05);im DC组角膜新生血管评分为(4.54±1.02)分,低于m DC组(8.67±2.11)分、对照组(14.87±3.24)分,差异均有统计学意义(均为P<0.05);im DC组术后3 d、7 d、14 d Scurfin蛋白表达水平分别为2.16±0.74、2.35±0.83、2.28±0.76,均高于m DC组(1.42±0.17、1.56±0.42、1.48±0.12)及对照组(1.39±0.21、1.42±0.26、1.69±0.28),差异均有统计学意义(均为P<0.05)。结论 im DC可抑制角膜移植免疫排斥反应发生,而诱导CD4+CD25+调节性T细胞的产生,进而促进免疫耐受的形成。

Objective To study the inhibitory effect and mechanism of T cell mi- gration induced by immature dendritic cells in high risk corneal allograft rejection in rats. Methods The immature dendritic cells were derived from bone marrow, the re- ceptor choose 48 SD rats, and 30 Wistar rats as donor. The allogeneic high-risk corneal transplantation model by alkali burn was established. The receptor SD rats were ran- domly divided into mDC group （mature dendritic cells, mature dendritic cell, 16 cases）, imDC group （immature dendritic cells, imDC, immature dendritic cell, 16 cases） and control group （ 16 cases）. In imDC and mDC group,the donor bone marrow derived im- DC cultured in vitro was injected via tail vein at preoperative 7 days,the cells were and suspended in 0.1 mL culture liquid. In control group ,0.1 mL culture liquid was injected via tall vein at preoperative 7 days. The rat corneal graft was examined by slit lamp, and the corneal graft survival was accurately assessed, four rats were sacrificed at 3 days, 7 days, 14 days in each group, respectively,the corneal graft tissue were stained with HE, the Scurfm protein expression of CD4＋ CD25 ＋ regulation T cell surface in rats spleen was detected by Western blot. Results The survival time of corneal graft in the imDC group was （ 18.67 ± 2.46） days, compared with the mDC group （ 6.78 ±0.55 ） days, the control group （9.28 ± 1.24） days, was significantly prolonged, the differences were sta- tistically significant （ all P 〈 0.05 ）. The neovascularization score in imDC group was 4. 54 ± 1.02, were lower than those in the mDC group （ 8.67± 2. 11 ） and the control group （ 14.87 ± 3.24 ）, the differences were statistically significant （ all P 〈 0.05 ）. Scurf- in protein expression levels in the imDC group at postoperative 3 days,7 days, 14 days were 2. 16 ±0.74,2.35±0.83 and 2.28 ±0.76 ,which were higher than those in the mDC group （1.42 ±0. 17,1.56 ±0.42,1.48±0.12）,the control group （1.39 ±0.21,1.42± 0. 26,1.69 ± 0.28 ） ,the differences were statistically significant （ all P 〈 0.05 ）. Conclu- sion The immature dendritic ceils can inhibit the immune rejection of corneal trans- plantation,and induce the production of CD4 ＋ CD25 ＋ regulatory T cells, which can promote the formation of immune tolerance.