半成熟树突状细胞在胚胎干细胞源性肝细胞移植中诱导耐受的研究

Semimature dendritic cells inducing tolerance in allotransplantation of hepatocytes differentiatedfrom mouse embryonic stem cells

目的在小鼠胚胎干细胞（ESC）源性肝细胞移植中，应用半成熟树突状细胞（smDC）诱导免疫耐受。方法绿色荧光蛋白（GFP）标记的129小鼠ESC按前期方法向肝细胞诱导分化，129小鼠smDC亦按前期方法由骨髓单核细胞诱导获得。smDC注入BALB／C小鼠3d后，再将ESC分化的肝细胞移植人BALB／C小鼠肝被膜下。检测肝转氨酶、外周血白细胞介素-2（IL）-2和IL-10水平变化，观察移植细胞生长及淋巴细胞浸润情况。结果移植后7d，smDC组肝功能指标丙氨酸转氨酶（ALT）为212．58U／L，较对照组512．27U／L明显改善（P〈0．05），7d和14d外周血IL-10水平较对照组明显升高，而IL-2水平较对照组明显降低（P〈0．05）。移植细胞存活率在14、21dsmDC组较对照组明显增高，CD4＋T细胞浸润较对照组明显减轻。结论在ESC源性肝细胞异基因移植中，smDC预输注可诱导部分免疫耐受，有望作为一种良好的移植耐受原应用于ESC分化细胞移植研究中。

Objective To induce tolerance in transplantation of hepatocytes differentiated from mouse embryonic stem ceils （ESCs） by using semimature dendritic cells （smD.Cs）. Methods Green fluo- rescent protein （GFP） -stained D3-ESCs in the 129 mice were differentiated into hepatocytes by methods we previously reported. Myeloid smDCs were derived from the 129 mice in vitro also as we previously reported. Then, BALB/C mice （recipients） were firstly injected with smDCs （1.0×10^5 cells per mouse）. Three days later, hepatocytes differentiated from D3-ESCs in the 129 mice as cell suspension were poured into BALB/C mice liver capsule （2. 0× 10^6 cells per mouse）. Thereafter, histopathologieal examinations of the hepatocytes graft were performed. The serum ALT, total bilirubin （TB） and the survival rate of hepatocytes graft were also determined. At the same time, the levels of IL-2 and IL-10 in the recipients were measured by enzyme linked immunosorbent assay （ELISA）. Results As compared with control group, the survival time of the allograft of D3-ESC-hepatocytes was prolonged when BALB/C mice received the 129 mice smDCs. CD4 ＋ lymphoeytes infiltration was significantly reduced in smDC group. The blood levels of IL-10 in smDC group were significantly higher than those in control group （ P 〈 0. 05 ） , but those of IL-2 in smDC group were lower than those in control group （ P 〈 0. 05 ）. Moreover, pathological examination demonstra- ted that transplanted ceils and recipient cells were well arranged and express albumin in smDC group. Con- clusion Donor smDCs could induce partly tolerance in allotransplantation of ESC-hepatocytes. Pre-treat- ment of recipients with donor smDCs has potential for ESC-derived cells allotransplantion.