介白素20(Interleukin-20, IL-20)是IL-10家族之一的成員，而IL-10家族包含了有IL-10、IL-19、IL-20、IL-22、IL-24、IL-26、IL-28和IL-29等成員。在IL-10家族中，IL-19, IL-20, IL-24共用了相同的受體次單元；其中IL-19, IL-20, IL-24可以透過IL-20R1/IL-20R2進行下游訊息傳遞，而IL-20, IL-24還可透過IL-22R/IL-20R2進行傳遞。目前已經有許多研究顯示，在乾癬，類風濕性關節炎，以及粥狀動脈硬化等等疾病的致病機轉中，IL-20不僅參與其中也扮演著重要角色。而實驗室之前也證實在in vitro情形下，由IL-20所引起的致病反應可以被IL-20的單株抗體(7E)所中和。由這個實驗結果顯示，IL-20與其單株抗體7E結合的motif對IL-20所調控的細胞反應非常重要，因此我想要找出IL-20與7E的結合部位。首先我構築並表現各種不同的截短IL-20蛋白質，並利用來分析IL-20與7E結合的位置。經由西方墨點法的實驗證實，我已經找到了IL-20與7E所結合的位置，而藉由專一性的突變掉IL-20上的兩個胺基酸序列，可以破壞IL-20與7E之間的結合。利用同樣的策略，我也同時找出了IL-20被C2所辨識之抗原決定位。為了証實7E抗原決定位之重要性，我接下來分析比較了突變型IL-20與野生型IL-20蛋白質的生物功能與引起之訊息傳遞是否不同。由實驗結果顯示，突變型IL-20蛋白質在引起軟骨細胞(chondrocyte)IL-6基因表現及引起類風濕性關節炎動物模式之關節滑囊纖維母細胞(collagen induced arthritis synovial fibroblast，CIA synovial fibroblast)RANKL基因表現的作用皆有降低；在訊息傳遞部份，相較於野生型IL-20，CIA synovial fibroblast處理突變型IL-20蛋白質後所引起之STAT-3磷酸化程度也較低。由以上結果顯示出，IL-20上被7E辨識之位置，對IL-20能否正常執行功能扮演重要角色。

Interleukin-20 (IL-20) belongs to the IL-10 family, which includes IL-10, IL-19, IL-20, IL-22, IL-24, IL-26, IL-28, and IL-29. Within this subfamily, IL-19, IL-20, and IL-24 share their receptor subunits; all three members are capable of signaling through IL-20R1/IL-20R2, while IL-20 and IL-24 can also use IL-22R/IL-20R2. Previous studies showed that IL-20 is involved in several diseases, such as psoriasis, rheumatoid arthritis, and atherosclerosis. The IL-20-mediated responses involved in these diseases can be neutralized by monoclonal antibody (7E) in vitro. These results suggest that the binding motif of IL-20 by 7E must be very important in the IL-20-mediated cellular responses. Therefore, we were aimed to determine the vital motif of IL-20 recognized by 7E. We performed a serial deletion on IL-20 coding region and expressed the recombinant proteins of these deleted mutants. The truncated IL-20 proteins were analyzed for 7E recognition. After performing Western blot of truncated IL-20 proteins with 7E, we have mapped the epitope of IL-20 which was recognized by 7E. Specific mutation of two amino acids renders IL-20 (IL-20-mutant) unrecognizable by 7E. Similarly, we used the same strategies to approach the epitope of IL-20 recognized by C2, another monoclonal antibody against IL-20 generated in our lab. To confirm the importance of the 7E-binding motif, we further analyzed and compared the biological function and intracellular signaling of IL-20-mutant and wild-type IL-20 (IL-20-WT). The effects of IL-20-mutant on induction of IL-6 in chondrocytes and RANKL in synovial fibroblasts of rats with collagen-induced arthritis (CIA) were decreased. In addition, IL-20-mutant showed lower activity in STAT-3 phosphorylation of CIA synovial fibroblasts than IL-20-WT did. Therefore, substitution of 7E-binding motif in IL-20 indeed impaired its biological activities.

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