Li Haochuan, Niederkorn Jerry Y, Neelam Sudha, Mayhew Elizabeth, Word R Ann, McCulley James P, Alizadeh Hassan
Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Invest Ophthalmol Vis Sci. 2005 Mar;46(3):900-7. doi: 10.1167/iovs.04-0495.
Amniotic membrane has been applied to the ocular surface to restore corneal function. The beneficial effect of amniotic membrane transplantation may be due to the immunosuppressive effects of amniotic epithelial cells. The purpose of this study was to determine whether amniotic epithelial cells (AECs) secrete anti-inflammatory and antiproliferative factors that affect the chemotaxis of neutrophils and macrophages and suppress both T- and B-cell proliferation in vitro.
Human amniotic cells were isolated from human amniotic membrane and cultured in vitro. The supernatants from AEC cultures were collected after 48 hours of incubation. Neutrophil and macrophage chemotactic activity was tested in the presence of AEC supernatant, using 24-well migration assay chambers. Lymphocyte proliferation was tested by H(3)-thymidine incorporation. Apoptosis was examined by caspase-3 and annexin V assays, and expression of cytokines was assessed by RT-PCR.
AEC supernatant significantly inhibited the chemotactic activity of neutrophils and macrophages toward macrophage inflammatory protein (MIP)-2 (P < 0.05). The supernatant significantly reduced the proliferation of both T and B cells after mitogenic stimulation (P < 0.05). Caspase-3 assays revealed that the supernatant induced apoptosis of T and B cells, but not of corneal epithelial cells and liver cells. In contrast to lymphocytes, macrophages and neutrophils were resistant to apoptosis induced by AEC supernatant. The AECs expressed message for TNFalpha, Fas ligand (FasL), TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), TGFbeta, and macrophage migration-inhibitory factor (MIF). However, AEC induction of apoptosis was inhibited (50%) by anti-FasL antibody but not by anti-TRAIL or anti-TNFalpha antibodies. Moreover, AEC supernatant inhibited macrophage migration in vitro.
AECs secrete soluble factors that inhibit cells in both the innate and adaptive immune systems.
羊膜已应用于眼表以恢复角膜功能。羊膜移植的有益效果可能归因于羊膜上皮细胞的免疫抑制作用。本研究的目的是确定羊膜上皮细胞（AECs）是否分泌抗炎和抗增殖因子，这些因子会影响中性粒细胞和巨噬细胞的趋化性，并在体外抑制T细胞和B细胞的增殖。
从人羊膜中分离出人羊膜细胞并进行体外培养。孵育48小时后收集AEC培养物的上清液。使用24孔迁移分析室在AEC上清液存在的情况下测试中性粒细胞和巨噬细胞的趋化活性。通过H(3)-胸苷掺入法测试淋巴细胞增殖。通过半胱天冬酶-3和膜联蛋白V测定法检测细胞凋亡，并通过逆转录-聚合酶链反应评估细胞因子的表达。
AEC上清液显著抑制中性粒细胞和巨噬细胞对巨噬细胞炎性蛋白（MIP）-2的趋化活性（P < 0.05）。该上清液在有丝分裂原刺激后显著降低T细胞和B细胞的增殖（P < 0.05）。半胱天冬酶-3测定显示，该上清液诱导T细胞和B细胞凋亡，但不诱导角膜上皮细胞和肝细胞凋亡。与淋巴细胞不同，巨噬细胞和中性粒细胞对AEC上清液诱导的凋亡具有抗性。AECs表达肿瘤坏死因子α（TNFalpha）、Fas配体（FasL）、肿瘤坏死因子相关凋亡诱导配体（TRAIL）、转化生长因子β（TGFbeta）和巨噬细胞迁移抑制因子（MIF）的信息。然而，抗FasL抗体可抑制（50%）AEC诱导的凋亡，但抗TRAIL或抗TNFalpha抗体则不能。此外，AEC上清液在体外抑制巨噬细胞迁移。
AECs分泌抑制先天性和适应性免疫系统中细胞的可溶性因子。
