Sakimoto Tohru, Yamada Ai, Sawa Mitsuru
Department of Ophthalmology, Nihon University School of Medicine, Tokyo, Japan.
Invest Ophthalmol Vis Sci. 2009 Oct;50(10):4618-21. doi: 10.1167/iovs.08-2669. Epub 2009 Apr 30.
An involvement of tumor necrosis factor-alpha-converting enzyme (TACE)-dependent ectodomain shedding in the release of soluble tumor necrosis factor receptor 1 (sTNFR1) from corneal epithelium was evaluated.
In vitro experiments were performed using the human SV40-transformed human corneal epithelial cell (HCEC) line. Ectodomain shedding was stimulated by phorbol myristate acetate (PMA, 3 microM) or peptidoglycan (PGN, 100 microg/mL), with or without TACE inhibition, using TNF-alpha processing inhibitor-1 (TAPI-1, 250 microg/mL) or tissue inhibitor of metalloproteinase-3 (TIMP-3, 2 microg/mL) by addition to the HCEC culture medium. The concentrations of sTNFR1 in culture medium were analyzed by enzyme-linked immunosorbent assay. To induce an inflammatory response in the ocular surface, corneal alkali burn of BALB/c mice was made from a filter paper dipped in 1 N NaOH solution. TNFR1 expression in corneal and conjunctival epithelia was evaluated by immunohistochemistry 28 days after wounding.
In HCEC culture medium, sTNFR1 release was significantly increased by the addition of PMA (t-test, P < 0.01) or PGN (P < 0.01). The increased release of sTNFR1 was significantly inhibited by the addition of TAPI-1 or TIMP-3, indicating the possibility of TACE-dependent ectodomain shedding of TNFR1. In the corneal alkali burn model, TNFR1 was expressed in corneal and conjunctival epithelia.
TACE-dependent ectodomain shedding of sTNFR1 was recognized in corneal epithelium. In the inflamed ocular surface, TNFR1 was expressed in the corneal and conjunctival epithelia after alkali burn treatment. sTNFR1, released from the ocular surface, may play an anti-inflammatory role in the inflammatory condition.
评估肿瘤坏死因子-α转化酶（TACE）依赖性胞外域脱落参与可溶性肿瘤坏死因子受体1（sTNFR1）从角膜上皮释放的情况。
使用人SV40转化的人角膜上皮细胞（HCEC）系进行体外实验。通过添加佛波酯肉豆蔻酸酯乙酸盐（PMA，3 microM）或肽聚糖（PGN，100 microg/mL）刺激胞外域脱落，在有或没有TACE抑制的情况下，通过向HCEC培养基中添加肿瘤坏死因子-α加工抑制剂-1（TAPI-1，250 microg/mL）或金属蛋白酶组织抑制剂-3（TIMP-3，2 microg/mL）。通过酶联免疫吸附测定分析培养基中sTNFR1的浓度。为了在眼表诱导炎症反应，用浸有1 N NaOH溶液的滤纸对BALB/c小鼠进行角膜碱烧伤。在受伤28天后通过免疫组织化学评估角膜和结膜上皮中TNFR1的表达。
在HCEC培养基中，添加PMA（t检验，P < 0.01）或PGN（P < 0.01）可显著增加sTNFR1的释放。添加TAPI-1或TIMP-3可显著抑制sTNFR1释放的增加，表明TNFR1存在TACE依赖性胞外域脱落的可能性。在角膜碱烧伤模型中，TNFR1在角膜和结膜上皮中表达。
在角膜上皮中发现了sTNFR1的TACE依赖性胞外域脱落。在炎症性眼表中，碱烧伤治疗后角膜和结膜上皮中表达TNFR1。从眼表释放的sTNFR1可能在炎症状态中发挥抗炎作用。
