输注胰岛抗原特异性Treg细胞延长同系NOD小鼠移植胰岛的存活时间

Prolonged islet isograft survival in NOD mice treated with islet antigen-specific regulatory T cells

目的探讨输注胰岛抗原特异性调节性T淋巴细胞（Treg细胞）对非肥胖糖尿病（NOD）小鼠同系胰岛移植物存活时间的影响。方法n以未成熟树突状细胞（imDc）联合谷氨酸脱羧酶一65在体外诱导童贞T淋巴细胞分化成胰岛抗原特异性Treg细胞。以已发生糖尿病的NOD小鼠为受者，将分离得到的尚未进展为糖尿病的NOD小鼠的胰岛（500胰岛当量）移植至受者的肾包膜下，对照组不行移植，只观察血糖变化；单纯胰岛移植组只进行胰岛移植，不输注胰岛抗原特异性Treg细胞；实验组于术前1d静脉输注1×10^6个胰岛抗原特异性Treg细胞，然后进行胰岛移植。术后检测受者的血糖，以判断移植胰岛的存活时间，观察胰岛移植物的病理学变化。结果对照组血糖持续高于11．1m01／L；单纯胰岛移植组小鼠的血糖于术后1～2d降至正常，到7～17d时开始陆续升高，并维持在术前水平，移植物存活时间为（12．2±2．6）d；实验组小鼠的血糖于术后1～2d降至正常，至第27天开始有小鼠血糖升高超过11．1mmol／L第43天时，所有小鼠的血糖均超过11．1mmol／L，移植物的存活时间为（35．2±4．3）d，明显长于单纯胰岛移植组（P〈0．01）。单纯胰岛移植组的移植胰岛有明显的淋巴细胞浸润，并伴有胰岛细胞严重破坏，胰岛素染色未见完整的胰岛存在，仅有极少量残存的分泌胰岛素的胰岛细胞；实验组第15天时移植胰岛形态完整，仅有少量淋巴细胞浸润，分泌胰岛素的胰岛大量存在。结论体外诱导产生的胰岛抗原特异性Treg细胞可以延缓自身免疫系统对移植胰岛的破坏，明显延长NOD小鼠移植胰岛的存活时间。

Objective To investigate the survival of islet isograft in NOD mice treated with islet antigen-specific regulatory T cells. Methods GAD-65 antigen pulsed immature dendritic cells （imDC） were used to induce naive T cells into islet antigen-specific regulatory T cells. NOD mice which had progressed to type 1 diabetes （T1DM）, as the recipients, received islet isografts （500 IEQ） under renal capsule from NOD mice without T1DM. In NOD mice in control group without transplantation, the changes in blood glucose （BG） were observed. NOD mice in simple islet transplantation group were given islet isograft without Treg infusion. In experiment group, NOD mice were infused with 1 x 10^6 islet antigen-specific regulatory T cells on the 1st day before transplantation, subsequently underwent islet isotransplantation. The survival of the islet isograft was evaluated by BG levels and the pathological changes were observed. Results BG levels were sustained above 11. I mmol/L in control group. In simple islet transplantation group, BG level was decreased to the normal level in 1- 2 days after transplantation, and began to rebound in 7- 17 days posttransplantation and maintained at the preoperative level. The mean survival of the islet isograft in the NOD mice was （12. 2 ±2. 6） day; In experiment group, 13G level was decreased to the normal level in 1 -2 days after transplantation, rebounded above 11.1 mmol/L in some mice on the 27th day after transplantation, and rebounded above 11.1 mmol/L on the 43th day in all mice. The mean survival of the islet isograft in the NOD mice was （35.2 ± 4. 3） days, which was significantly prolonged compared to simple islettransplantation group （P 〈 0. 01）. In simple islet transplantation group, the islet isograft was infiltrated by many lymph cells and damaged severely, and only few residual islet cells secreted insulin without complete islet existing in insulin staining. The islet isograft in experiment group was intact on the 15th day, with little lymph cell infiltration and a great number of islets secreting insulin. Conclusion Infusion of islet antigen-specific regulatory T cells induced by imDC and islet antigen GAD65 in vitro could delay the destruction of autoimmune system and prolong the islet isograft survival in NOD mice.