篩選並分析感染C型肝炎病毒併發蕭格式症候群病人的類風濕性因子

The Isolation and Characterization of Rheumatoid Factors from a Hepatitis C Virus Infected Patient with Sjögren’s Syndrome

李雨青(Lee Reu-Ching)

指導教授 ： 楊沂淵

臺北醫學大學/醫學院/醫學檢驗生物技術學研究所/碩士(2001年)

摘要

我們先前已經從類風濕性關節炎併發ITP的病人中使用噬菌體展現系統篩選出一些類風濕性因子。為了了解IgG的類風濕性因子在其他自體免疫疾病上所扮演的角色，我們從C型肝炎病毒感染併發蕭格式症候群Sjögren’s syndrome (SS) 的病人上做了一個噬菌體展現抗體基因庫。蕭格式症候群的特徵是會侵害外分泌腺的淋巴細胞導致乾眼症 (xerostomia) 和淋巴性唾腺炎 (heratoconjunctivitis sicca)。經過針對人類IgG分子所做的重組基因庫篩選之後，我們利用西方墨點分析法在14個隨機挑選的噬菌體菌株，觀察是否有Fab分子50 kd蛋白的表現。我們找到了其中的13個是含有lambda輕鏈基因，而找到的類風濕性因子都會對人類Fc片段分子呈現特異性的結合，相對於牛血清白蛋白 (bovine serum albumin) 則不會有結合的反應。我們也使用酵素連接免疫吸附分析法 (ELISA) 去分析這些篩選出來的類風濕性因子對於人類Fc片段的特異性和先前從ITP病人上篩選的GG3和GG48兩個類風濕性因子有何不同。從DNA的序列分析上發現了這些類風濕性因子菌株分成3個族群，分別呈現的代表號碼為hcvrf1、hcvrf2和hcvrf11。有趣的是，我們發現hcvrf11的重鏈V基因和hcvrf2是相同的，而輕鏈V基因則是和hcvrf1相同。然而hcvrf11的類風濕性因子結合力卻不會受到這些改變影響。綜合這些實驗結果顯示在蕭格式症候群的病人身上會產生IgG類風濕性因子，並且這些類風濕性因子可能在這些自體免疫的病人病因上扮演重大的角色。

關鍵字

C型肝炎病毒 ； 蕭格式症候群 ； 類風濕性因子

並列摘要

We have previously isolated several IgG rheumatoid factors (RFs) from both patients with rheumatoid arthritis (RA) and idiopathic thrombocytopenia purpura (ITP) using phage display system. To study IgG RFs in patients with other autoimmune disease, we have constructed phage display antibody libraries from a hepatitis C virus infected patient with Sjögren’s syndrome (SS) which is characterized by lymphocytic invasion of exocrine glands resulting in xerostomia and heratoconjunctivitis sicca. After panning against human IgG molecules, the expression of Fab fragments from 14 randomly selected phage clones was identified as a band of 50 kd in a western blotting analysis. Of which, 13 clones contain lambda light chain gene. All representative RFs bound specifically to Fc-coated plates, but not to bovine serum albumin (BSA). ELISA data also revealed that their Fc binding activity is comparable to that of GG3 and GG48 clones previously isolated from ITP patient. Sequence determination suggested that these RFs consisted of 3 groups of clones, represented by hcvrf1, hcvrf2 and hcvrf11. Interestingly, the heavy chain V gene of hcvrf11 is identical to that of hcvrf2 and its light chain V gene is the same as that of hcvrf1. However, the RF activity of hcvrf11 is retained regardless of this promiscuity. Taken together, these results suggested that IgG RFs may be generated in patients with Sjögren’s syndrome and may play an important role in the pathogenesis of this autoimmune disease.

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