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背景:随着磁分选技术的完善,体外分选、扩增足量的对移植抗原具有特异性的细胞已成为可能,但就其在体内应用剂量及免疫耐受的效能问题目前鲜有报道。目的:探索供体抗原特异性CD4+CD25+Treg细胞在体内应用诱导移植免疫耐受的量效关系。方法:以SD大鼠为供体、Wistar大鼠为受体,建立同种异体肾移植动物模型;体外分选、富集Wistar大鼠脾脏CD4+CD25+Treg细胞,并诱导其对SD大鼠供体抗原的特异性表型;根据不同数量(2×105、5×105、1×106、2×106)供体抗原特异性CD4+CD25+Treg细胞在肾移植中单剂量尾静脉注射,并以未注射组为对照。术后15d分析移植肾脏存活状况。术后4,9,15d采血检测各组肌酐水平,同时进行移植肾脏病理检查,按照BanffSchema标准进行诊断,并根据Watanabe的方法进行半定量评分。结果与结论:术后15d内对照组死亡率最高83.3%,2×105组次之66.7%,2×106组为58.3%,5×105组为33.3%,1×106组则全部存活;各实验组术后4,9,15d血肌酐水平均明显低于对照组(P<0.05,P<0.01);术后第9,15天,2×105组、5×105组血肌酐水平均明显高于1×106组、2×106组(P<0.05);术后第4,9,15天移植肾脏病理检查的半定量评分结果显示,各时间段5×105组、2×105组与对照组间差异无显著性意义,各时间段1×106组与2×106组优于对照组(P<0.05)。结果初步证实供体抗原特异性CD4+CD25+Treg细胞受体内应用能够改善大鼠移植肾功能,延长移植肾存活时间,1×106为相对理想的单次应用剂量。
Background: With the improvement of magnetic separation technology, it has become possible to sort out and amplify enough cells specific for the transplanted antigen in vitro, but little is known about its efficacy in the application of dose and immune tolerance in vivo . Objective: To explore the dose-response relationship of donor antigen-specific CD4 + CD25 + Treg cells in vivo to induce immune tolerance of transplantation. Methods: Spleens of Wistar rats were used as donors and Wistar rats as recipients. Allogeneic renal allograft models were established. The CD4 + CD25 + Treg cells from spleens of Wistar rats were enriched in vitro and induced in SD rats Donor antigen specific CD4 + CD25 + Treg cells in different numbers (2 × 105, 5 × 105, 1 × 106, 2 × 106) of donor antigen-specific tail vein injection in renal transplantation, And no injection group as a control. Fifteen days after transplantation, the survival of renal allografts was analyzed. At 4, 9 and 15 days after surgery, blood creatinine levels were measured in each group. At the same time, renal pathological examination was performed. According to BanffSchema criteria, semi-quantitative scoring was performed according to Watanabe’s method. RESULTS AND CONCLUSION: The mortality rate of the control group was 83.3% at 15 days after operation, 66.7% at 2 × 105 group, 58.3% at 2 × 106 group, 33.3% at 5 × 105 group and all at 1 × 106 group. The levels of serum creatinine in experimental group at 4, 9 and 15 days after operation were significantly lower than those in control group (P <0.05, P <0.01). On the 9th and 15th days after operation, serum creatinine levels in 2 × 105 group and 5 × 105 group were significantly higher (P <0.05). The results of semi-quantitative grading of pathological examination on the 4th, 9th and 15th days after transplantation showed that in the 5 × 105 group, 2 × 105 group and There was no significant difference between control groups. Each time period, 1 × 106 group and 2 × 106 group were better than the control group (P <0.05). The results initially confirmed that donor antigen-specific CD4 + CD25 + Treg cell receptor application can improve renal allograft function in rats and prolong the survival time of renal allograft, 1 × 106 is a relatively ideal single dose.