Preparation for acellular human amniotic membrane matrix and assessment the immunogenic property
Preparation for acellular human amniotic membrane matrix and assessment the immunogenic property
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DOI：
10.16098/j.issn.0529-1356.2016.04.021
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发表时间：
2016-08-01
期刊：
影响因子：
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通讯作者：
Zhang Hai-yan
中科院分区：
文献类型：
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作者：
Lu Xin;Yuan Jie;Zhang Hai-yan
目的制备脱细胞人羊膜基质（AHAM）并评价其作为细胞移植支架的免疫原性。方法将胎盘剥去，切成片，用胰蛋白酶和EDTA孵育，使其脱细胞。为了生成冷冻保存的AHAM，将新鲜的AHAM片放入含有1:1的甘油和硫酸软骨素的memm - neaa培养皿中，保存在-80℃。使用前，冷冻保存的AHAM片用无菌PBS复水。采用免疫荧光法检测人白细胞抗原在HAM和AHAM中的表达。采用免疫荧光和流式细胞术检测小鼠AHAM移植4周后脾脏CD4（+）和CD8(+) T细胞的比例。结果免疫荧光分析证实新鲜和低温保存的AHAM对人白细胞抗原抗体均为阴性。免疫荧光和流式细胞术分析证实，AHAM移植后小鼠脾脏CD4（+）和CD8(+) T细胞比例没有变化。结论AHAM的免疫原性较低，不会引起小鼠移植后T细胞介导的免疫排斥反应。这些结果支持了AHAM作为治疗和管理疾病的细胞递送平台的潜力。
Objective To prepare of the acellular human amniotic membrane matrix (AHAM) and to assess the immunogenic property of AHAM as a scaffold for cell transplantation. Methods To prepare the AHAM, the HAM was peeled from the placenta, cut into pieces and incubated in trypsin with EDTA for decellularization. For generating cryopreserved AHAM, the fresh AHAM pieces were placed in dishes with a 1:1 mixture of glycerol and chondroitin sulfate in MEM-NEAA and stored at -80 degrees C. Before using, the cryopreserved AHAMs were rehydrated with sterile PBS. The expression of human leucocyte antigen in HAM and AHAM was determined by immunofluorescence. The ratio of CD4(+) and CD8(+) T cells in mouse spleen after AHAM transplantation for four weeks was assessed by immunofluorescence and flow cytometry. Results Immunofluorescence analysis confirmed that the fresh and cryopreserved AHAM were negative for human leucocyte antigen antibody. Immunofluorescence and flow cytometry analysis confirmed that the ratio of CD4(+) and CD8(+) T cells in mouse spleen after AHAM transplantation was not changed. Conclusion The immunogenic property of AHAM is relatively low and does not cause the T cells mediated immunological rejection after transplantation in the mouse model. These results support the potential of AHAM as a cell delivery platform to treat and manage disease.