Monitoring of immune rejection after abdominal aortic patch suture in cynomolgus monkeys
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摘要:
目的 建立腹主动脉补丁缝合术的异种移植模型以及术后免疫排斥监测平台。 方法 将野生型巴马猪的颈动脉，修剪成约2.5 cm×1.0 cm大小的梭形补丁，缝合于食蟹猴腹主动脉，未给予免疫抑制剂。观察受体猴一般情况。术后1年采用病理学检查观察移植物动脉补丁的形态学变化。分别在术前和术后7、14、28、49 d采集受体猴血液样本，利用猪的红细胞、外周血单核细胞（PBMC）检测受体猴血清中针对猪抗原的IgM和IgG抗体水平。采用血常规五分类法和流式细胞术检测受体猴血液中淋巴细胞计数。 结果 3只移植猴存活状态良好。术后1年动脉补丁所在的血管壁外侧组织呈现深红色，苏木素-伊红（HE）染色显示有大量的红细胞和血小板沉积，其中有淋巴细胞的浸润。以猪的红细胞和PBMC为靶细胞，血清中的抗猪IgM和IgG抗体水平在术后28 d达到峰值，术后49 d时水平有所下降。淋巴细胞、T细胞亚群也是在术后28 d达到峰值，49 d开始回落。 结论 动脉补丁缝合术是一种便捷可靠的异种移植模型。没有免疫抑制剂的条件下，受体维持了正常的生理状态。移植物可有效激活受体的免疫系统，诱导抗猪抗体的产生，引发细胞免疫排斥反应，因此该模型可用于监测异种移植过程中的免疫排斥反应。 Abstract:Objective To establish a platform to monitor the immune rejection after abdominal aortic patch suture in a xenotransplantation model. Methods The carotid was excised from wild-type Guangxi Bama pigs, cut into 2.5 cm × 1.0 cm pieces in shuttle shape and subsequently sutured to the abdominal aorta of cynomolgus monkeys. No immunosuppressive agent was administered. General conditions of the recipient monkeys were observed. The morphological changes of the graft artery were assessed by pathological examination at postoperative 1 year. Before and 7, 14, 28 and 49 d after surgery, the blood samples were collected from the recipient monkeys. The serum levels of IgM and IgG antibodies were quantitatively measured by the red blood cell and peripheral blood mononuclear cell (PBMC) from Guangxi Bama pigs. The quantity of lymphocytes in the recipient monkeys was detected by routine blood test and flow cytometry. Results All 3 monkeys undergoing transplantation survived well. At postoperative 1 year, the lateral tissues of the vascular wall at the artery graft were seen in dark red color. Hematoxylin-eosin (HE) staining revealed a large quantity of red blood cell and platelet deposition, accompanied with lymphocyte infiltration. Using porcine red blood cell and PBMC as target cells, the serum levels of anti-pig IgM and IgG antibodies peaked at postoperative 28 d, and slightly declined at postoperative 49 d. The quantity of lymphocytes and T cell subset also peaked at postoperative 28 d and began to decrease at postoperative 49 d. Conclusions Artery patch suture is a simple and reliable xenotransplantation model. The recipients can maintain normal physiological state without the use of immunosuppressive agents. The grafts can effectively activate the immune system of the recipients, induce the production of anti-pig antibodies and provoke cellular immune rejection. Therefore, this model can be utilized to monitor the immune rejection throughout the xenotransplantation process. -
图 1 动脉补丁缝合术示意图及术后图片
A图为动脉补丁缝合术手术方法示意图；B图中左图为手术结束后缝合效果，右图为术后1年的补丁缝合处
Figure 1. Schematic diagram and postoperative images of the artery patch suture
图 2 受体猴动脉切片的病理学图片（HE）
A图为补丁缝合处；B图为非补丁缝合处
Figure 2. Pathological images of the arterial segment of the recipient monkey
图 3 手术后受体猴血清中针对猪IgM和IgG抗体的水平变化
以猪（野生型和GTKO型）的红细胞和PBMC检测个时间点的异种抗体水平。血清中结合到红细胞上的IgM（A图）和IgG（B图）水平；血清中结合到PBMC上的IgM（C图）和IgG（D图）水平；以野生型猪的细胞为靶细胞，与术后0 d比较，aP＜0.05，bP＜0.05；同一时间点的血清样本，与GTKO型猪的细胞比较，cP＜0.05，dP＜0.05
Figure 3. Changes of IgM and IgG antibodies levels in the serum of recipient monkeys after surgery
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