Quantification of allospecific and nonspecific corneal endothelial cell damage after corneal transplantation
Quantification of allospecific and nonspecific corneal endothelial cell damage after corneal transplantation
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作者：
Chauhan, S. K.;Jurkunas, U.;Dana, R.
目的探讨宿主免疫(同种异体免疫)和手术操作(非同种异体免疫)对角膜移植中角膜内皮细胞(CECs)的影响。方法分别于移植后1、3、8周从同种异体受者、同种异体受体和同种异体排斥反应患者(BALB/c)取引流淋巴结和C57BL/6角膜。我们使用体外角膜杯中试验分析CEC的凋亡，并使用免疫组织化学染色(ZO-1)显示细胞间的连接。用Confoscan软件进行自动细胞分析，通过量化细胞大小(多形性)和形态(多形性)的变异系数来测量CEC的密度和CEC的形态变化。结果杯中角膜实验显示同种异体受体T细胞和更大程度的排斥T细胞(而不是同基因T细胞)诱导了CEC的凋亡。与同种异体角膜移植相比，同种异体角膜移植后细胞密度显著降低(P<；0.001)，排斥反应组细胞密度比同种异体受体组进一步降低。移植后1周，与同基因移植物相比，同种异体移植物的细胞大小变异系数有更大的增加(P=0.001)。结论同种异体角膜移植中CEC损伤的主要原因是宿主的同种异体免疫，而非手术操作，在临床可见排斥反应发生前即可检测到CEC的形态变化。
Purpose To investigate the effect of host immunity (allospecific) and surgical manipulation (non-allospecific) on corneal endothelial cells (CECs) in corneal transplantation.Methods Draining lymph nodes and grafted C57BL/6 corneas were harvested from syngeneic recipients, allograft acceptors, and allograft rejectors (BALB/c) 1, 3, and 8 weeks after transplantation. We analyzed CEC apoptosis using an ex vivo cornea-in-the-cup assay, and visualized cell-to-cell junctions using immunohistochemical staining (ZO-1). Automatic cell analysis using Confoscan software was used to measure CEC density as well as changes in CEC morphology by quantifying the coefficient of variation in cell size (polymegethism) and shape (pleomorphism).Results The cornea-in-the-cup assay showed that allogeneic acceptor T cells and to an even greater extent rejector T cells (but not syngeneic T cells) induced CEC apoptosis. CEC density after corneal transplantation was significantly reduced in allogeneic acceptors compared with syngeneic grafts (P < 0.001), and CEC density was even further reduced in the allo-rejector group compared with the allo-acceptor group. Allogeneic grafts showed a greater increase in the coefficient of variation in cell size (polymegethism) when compared with syngeneic grafts 1 week after transplantation (P = P < 0.001). However, pleomorphism was not significantly different between syngeneic and allo-acceptor grafts, indicating that polymegethism (but not pleomorphism or cell density) is a sensitive indicator of the effect of alloimmunity on CECs.Conclusions Our data demonstrate that host alloimmunity rather than surgical manipulation alone is the major cause of CEC damage in corneal transplantation, and such morphologic changes of CECs can be detected before the clinically visible onset of allograft rejection.