Soluble Fas Ligand and Ocular Immune Privilege

描述（由申请人提供）：Fas配体（FasL）作为膜结合的可溶性蛋白质产生，两种形式都在免疫特权眼内表达。在我们之前的研究中，我们使用眼肿瘤模型来证明不同形式的FasL调节眼睛的先天免疫：（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 在眼内组成型表达，并且 FasL 的膜形式具有促炎性，我们建议在正常眼中：（i）mFasL 的促炎功能被阻断，和/或（ii）FasL 主要以可溶（抗炎）形式表达。我们的实验数据支持后者。对正常、FasL 敲除和慢性发炎的眼睛进行 FasL 的蛋白质印迹分析。在正常眼睛中检测到可溶性 (27 kDa) 与膜 (38 kDa) FasL (10:1) 的高比例。检测到修饰的 sFasL 的另外三个条带 (28-31 kDa)，这些条带在 FasL 敲除小鼠的眼睛中完全不存在。修饰后的 sFasL 是眼睛所独有的，在其他免疫特权部位（例如睾丸）中没有发现。 最后，缺乏免疫特权并表现出继发于色素分散综合征的慢性炎症的眼睛中完全不存在27kD和31kD sFasL带。 我们假设修饰的 sFasL 在眼睛内表达并在 维持免疫特权。该假设将在三个具体目标中进行测试：(i) 确定 sFasL 在眼睛中的表达位置及其修饰方式，(ii) 确定修饰的 sFasL 如何刺激先天免疫，以及 (iii) 确定修饰的 sFasL 是否控制免疫豁免。我们相信这项研究不仅将增进我们对免疫特权的理解，而且有助于解释 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.