行政院國家科學委員會專題研究計畫成果報告:實驗性自體免疫前葡萄膜炎之基因治療

Abstract

葡萄膜炎一般認為是一種自體免疫疾病，其確切的病因與致病機轉不 明。反覆發作可因各種併發症如角膜病變、白內障、青光眼、網膜水腫或損傷 而造成失明，是失明的重要原因之一。 一般認為葡萄膜炎是一種延發型過敏反應（Delayed type hypersensitivity ) 所造成的，在人類葡萄膜炎的組織病理檢查許多實驗動物模型可以得到證實，而最近的許多證據顯示DTH 是由可分泌γ-INF、IL -2 、IL -12 的T helper:1 ( Th1）所引導，這相對於分泌IL -4 , IL6 、IL -l 0為主的T helper2 ( Th2）。後者可能是導致Type 1 hypersensitivity 如氣喘等的重要原因。Thl 與Th2 相關的細胞問素可互相抗衡。因此在局部環境中耗予較大量的IL -4 可使原來會形成Thl 的反應轉成Th2，而較多量的IL -12 也可使原來會形成Th2反應轉成TH1。 本研究將使用帶有Interleukin 4 plasmid的 脂小體與治病 抗原牛的黑色素相關抗原，先後打入實驗動物路易士鼠（Lewis Rat）的後腳。 我們的假設是當脂小體匯流到腳附近的淋巴結，而被其中的抗原呈現細胞所吞噬之後，便 可以在細胞附近的微綑環境中有較多的IL-4，這將有助於Th2免疫反應。於是在其後吸取並呈現抗原給T 細胞時便能引導該T 細胞成Th2型。因而抑制Thl 的免疫反應與延發型過敏反應，從而抑制或防止實驗性自體反應前葡萄膜炎的發生• 而且在其後數週至數月當中，當有相同的抗原進入體內時，便能以Th2而不是Thl 的方式反應• 於是以Thl 反應為主的實驗性自體免疫葡萄膜炎便能得到預防。而脂小體可以使IL - Plasmid 在組織間滯留時問延長，並增加APC 的攝取，有助於增長疫苗的效期與效價。 本法所用的只是為量的核甘酸，可避免全身大量投與細胞介質而達到免疫調整的目的。而注射黃甘酸與致病抗原於同一地方則可使該免疫調節較具抗原專一性。本法應可稱為”治療性疫苗”。如能應用在葡萄膜炎病人而能在數月或數年之中防止葡萄膜炎的發作或再發，當是葡萄炎患者的一大福音。

Experimental autoimmune anterior uveitis (EAAU) is an inflammatory autoimmune disease of anterior uveal tract, which serves as a iodel for human acute anterior uveitis. It has been shown that EAAU is mediated by CD4+ T cells and delayed type hypersensitivity, We have shown that intraocular injection of soluble melanin associated antigen can prevent the development cf primary onset and recurrence of EAAU. To test the possibility of gene therapy for EAAU, we will inject IL-4 plasmid entrapping liposome and bovine MAA (uveitoger ic antigen in the model of EAAU) into the hind footpad of Lewis rat. Our rational is the injected liposome-entrapped IL- 4 plasmid will be taken to regional lymph node ar d transfect the lymphoid cells and antigen presenting cells over there. This will develcip an IL-4 rich microenvironment that favors T helper 2 (Th2) response when antigen is presented to and activate specific T cell clones. T helper 2 pattern of cytoki e secretion will downregulate T helper 1 (Thi) response and delayed type hypersensitivity and therefore prevent and/or attenuate the severity of the autoimmune inflammatory disease. The liposome entrapped plasmid will stay longer in tissue and al c easier to be taken along with lymph to regional lymph node, and will make this gene therapy last longer and more effective. This gene therapy will only involve the use of tiny amount of nucleic acid, and will avoid large amount of systemic administration of cytokines to modulate immune response. The injection of the mixture of antigen and nucleic acid at the same site can make this immunomodulation more antigen specific. This approach could be called ¡¥therapeutic vaccine¡¦, If it works and can be applied to uveitis patients, it will be of great benefit to them, who usually suffer from repe ited recurrences and various complications that will lead to blindness.