Soluble Fas Ligand and Ocular Immune Privilege

描述(申请人提供)：Fas配体(FasL)是一种膜结合的可溶性蛋白质，这两种形式在免疫特权眼内都有表达。在我们以前的研究中，我们用一个眼肿瘤模型证明了不同形式的FasL调节眼睛的天然免疫：(I)膜FasL(MFasL)诱导炎症并终止免疫豁免，而(Ii)可溶性FasL(SFasL)预防炎症并维持免疫豁免。因此，鉴于FasL在眼内是结构性表达的，并且FasL的膜形式是促炎的，我们认为在正常眼中：(I)mFasL的促炎功能被阻断，和/或(Ii)FasL主要以可溶性(抗炎)形式表达。我们的实验数据支持后者。对正常眼、FasL基因敲除眼和慢性炎症眼进行FasL蛋白印迹分析。正常眼有较高的可溶性(27 KDa)/膜(38 KDa)FasL比值(10：1)。在FasL基因敲除小鼠的眼睛中检测到另外三条修饰的sFasL条带(28-31 kDa)，这些条带是完全不存在的。修饰后的sFasL对眼睛是独一无二的，在其他免疫特权部位没有发现，如睾丸。 最后，27kD和31kD sFasL条带在缺乏免疫豁免权和继发于色素弥散综合征的慢性炎症中完全消失。 我们推测，修饰的sFasL在眼睛内表达，并在 保持免疫特权。这一假设将在三个具体目标上得到验证：(I)确定sFasL在眼睛中的表达位置及其修饰方式；(Ii)确定修饰后的sFasL如何刺激天然免疫；以及(Iii)确定修饰后的sFasL是否控制免疫豁免权。我们相信这项研究不仅将促进我们对免疫豁免的理解，而且有助于解释关于FasL在移植和肿瘤中的生理作用的争议。

DESCRIPTION (provided by applicant): Fas ligand (FasL) is produced as a membrane-bound and soluble protein and both forms are expressed within the immune privileged eye. In our previous studies we used an ocular tumor model to demonstrate that the different forms of FasL regulate innate immunity in the eye: (i) membrane FasL (mFasL) induces inflammation and terminates immune privilege, while (ii) soluble FasL (sFasL) prevents inflammation and maintains immune privilege. Therefore, given that FasL is constitutively expressed within the eye, and the membrane form of FasL is pro-inflammatory, we propose that in a normal eye either: (i) the pro-inflammatory function of mFasL is blocked, and/or (ii) FasL is expressed primarily in the soluble (anti-inflammatory) form. Our experimental data support the latter. Western blot analysis of FasL was performed on normal, FasL knockout, and chronically inflamed eyes. A high ratio of soluble (27 kDa) to membrane (38 kDa) FasL (10:1) was detected in normal eyes. Three additional bands (28-31 kDa) of modified sFasL were detected that were completely absent in the eyes of FasL knockout mice. The modified sFasL was unique to the eye and not found in other immune privileged sites, such as the testis. Finally, the 27kD and 31kD sFasL bands were completely absent from eyes that lacked immune privilege and displayed chronic inflammation secondary to pigment dispersion syndrome. We hypothesize that modified sFasL is expressed within the eye and plays a central role in maintaining immune privilege. This hypothesis will be tested in three Specific Aims (i) determine where sFasL is expressed in the eye and how it is modified, (ii) determine how modified sFasL stimulates innate immunity, and (iii) determine if modified sFasL controls immune privilege. We believe this study will advance not only our understanding of immune privilege, but help explain the controversy over the physiological role of FasL in transplants and tumors.