哺乳動物藉由自身的免疫系統來抵抗外來病原菌的入侵。而這些免疫系統，我們可大略分為innate immunity和 adaptive immunity兩大類，其中，innate immunity最近被發現是利用Toll-like receptor (TLR)來分辨自體和外來的物質。TLR是一種固定去辨認病原菌上特定物質的受體，並不會像T細胞或B細胞的受體會藉由體細胞基因重組去改變其對辨認物的特異性。TLR9是TLR的一員，它辨認的物質是一種很少出現在脊椎動物基因體的CpG DNA，TLR9除了在對抗病源菌上是很重要的，它也被發現在自體免疫疾病的病程中扮演重要角色；在紅斑性狼瘡的病程上，活化TLR9可以促使plasmacytoid dendritic cell分泌α干擾素(interferon-α)並且讓B細胞的成熟分化而使其產生對身體有害的自體抗體。另一方面，在先前的研究中發現，T helper 1 (Th1) immunity在Pseudomonas aeruginosa所引起的角膜炎病程中也是扮演重要角色，而TLR9的活化會引起強烈的Th1 immunity。所以，我們假設TLR9對於紅斑性狼瘡和P. aeruginosa角膜炎的發生的確是佔很重要的地位，並且研究如果減少TLR9基因表現是否對於這兩種疾病是有療效的。我們藉由Internal ribosome entry site後接green fluorescent protein (IRES-GFP)的方法找出抑制TLR9基因表現最有效的small-hairpin RNA (shRNA)，並且將這段shRNA接進vesicular stomatitis virus GP (VSV-G) pseudotyped 的lentivirus產出Lt-shTLR9這隻病毒。從我們的結果得知，Lt-shTLR9可以在活體外有效地抑制TLR9基因的表現，但是，不幸地我們在以汞加速小鼠產生紅斑性狼瘡的模型中，沒有發現Lt-shTLR9對紅斑性狼瘡有任何的療效。而在小鼠P. aeruginosa角膜炎模型方面，我們已經完成它的架構，最近會進行利用Lt-shTLR9治療小鼠P. aeruginosa角膜炎模型的實驗。總而言之，希望我們的研究對於TLR9在自體免疫疾病的地位中，能提供一點小小的線索，並且對於紅斑性狼瘡和P. aeruginosa角膜炎能有新方向的療法。

In mammals host defense against invading microbial pathogens is elicited by the immune response, which can be broadly categorized into innate and adaptive immunity. The innate immunity discriminates between self and non-self via the family of Toll-like receptors (TLRs), which have been recently identified. TLRs are germ line-encoded non-rearranging receptors and recognize conserved pathogen-associate molecules found in microorganisms but rare or absent in vertebrates. TLR9, one of TLR family, is essential for recognition of CpG DNA, which is usually absent in the vertebrate genome. In addition to pathogen defense, TLR9 is presumably involved in pathogenesis of autoimmune disorders. Activation of TLR9 contributes to systemic lupus erythematosus (SLE) progression by T cell-independent B cell proliferation, pathogenic autoantibody production, and plasmacytoid dendritic cell production of interferon-α. On the other hand, previous studies have provided evidence that T-helper 1 (Th1) immunity is important in the pathogenesis of Pseudomonas aeruginosa keratitis and activation of TLR9 induces a strong Th1 inflammatory response. From these observations, we hypothesized that TLR9 may play a critical role in the pathogenesis of SLE and P. aeruginosa keratitis and investigated whether reduction of TLR9 gene expression may be beneficial in these two diseases. We screened efficient short-hairpin RNA (shTLR9) to knock-down murine TLR9 gene expression by the IRES-GFP system and constructed VSV-G pseudotyped lentiviral vector (Lt-shTLR9) expressing shTLR9 for the delivery of small interfering RNA (siRNA) into mammalian cells. Our results demonstrated that Lt-shTLR9 efficiently knocked down TLR9 gene expression in vitro, but it did not exert therapeutic efficacy mercury-exposed NZBW F1 mice. The murine P. aeruginosa keratitis model was established and the therapeutic efficacy of Lt-shTLR9 on this model will be evaluated in the future. Taken together, our study might provide a clue for TLR9-mediated disease and alternative therapeutic approaches for the treatment of SLE and P. aeruginosa keratitis.

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