臺灣博碩士論文加值系統

介紹：類風濕性關節炎是自體免疫疾病，受到遺傳因子及環境因素的影響。F11R與發炎細胞移動有關，也是芳香烴受體的目標基因，而芳香烴受體受到很多環境因子的作用。F11R可能是連接環境因素與類風濕性關節炎一個橋樑。
目的:我們想了解： (1)F11R在類風濕性關節炎病人表現是否確實增加？(2)F11R是否與類風濕性關節炎某些症狀或徵象相關？(3) F11R與發炎指標關係為何？
材料與方法: 從類風濕性關節炎病人及正常人中，抽取周邊血液單核球中核糖核酸及周邊血液去氧核醣核酸。利用即時定量聚合酶連鎖反應偵測F11R之mRNA表現量。以SNP genotyping assay進行基因多型性鑑定。以酵素連結免疫分析法測定血漿中IFN-γ表現量。
結果: F11R 在類風濕性關節炎患者的周邊血液單核球表現量比普通人多（p =0.018）。患者有唾腺侵犯時，F11R數值較低 (p= 0.037)。但F11R與其他次發性乾燥症症狀相關性不顯著。F11R啟動子-688 A＞C，C carrier有較低的anti-CCP (p=0.002) 及較少的淚腺問題(p=0.009)。校正DR4的干擾後，這現象依然存在。在類風濕關節炎病人，F11R啟動子-688 A＞C 和 -436 A>G兩個位置基因多型性與基因表現量不相關。IFN-γ在類風濕性關節炎病人的表現量和F11R並沒有顯著相關。
討論：F11R在類風濕性關節炎的病人表現量較高，與幫助發炎細胞通過細胞間隙的功能有關。F11R病人唾腺功能不良時數值較低，可能與它在此族群細胞間隙表現量減少有關。F11R啟動子 -688 A變異為C時，對病人有保護作用。IFN-γ在類風濕性關節炎病人與F11R作用，需要進一步實驗才能釐清。

Introduction: Rheumatoid arthritis is an autoimmune disease, which is affected by genetic and environmental factors. F11R is associated with inflammatory cells migration and also the target gene of aryl hydrocarbon receptor. The aryl hydrocarbon receptor is triggered by multiple environmental factors. F11R may link environmental factors to rheumatoid arthritis.
Aim: We investigate whether (1) F11R expression increases in rheumatoid arthritis? （2）F11R is associated with rheumatoid arthritis characteristics ? and（3）F11R is associated with inflammatory markers?
Material and Methods: We extracted RNA from peripheral blood mononuclear cells and DNA from peripheral blood in rheumatoid arthritis patients and control group. F11R mRNA expression was determined by real time PCR. SNP genotyping assay was done for single nucleotide polymorphism. IFN-γ was quantified by enzyme-link immunosorbent assay.
Results: F11R expression in peripheral blood mononuclear cells is more in rheumatoid arthritis patients than control group (p = 0.018). F11R expression is lower in patients who has impaired salivary gland function than control group (p= 0.037). F11R association with other secondary Sjögren’s syndrome characteristics is not significant. In F11R promoter -688 A＞C, C carrier possesses lower anti-CCP（p=0.002）and lacrimal gland problems(p=0.009). Both are significant after correcting DR4 existence. Different SNP genotype in F11R promoter -688 A＞C and -436 A>G doesn’t lead to different gene expression in rheumatoid arthritis patients. IFN-r amount in rheumatoid arthritis is not correlated with F11R.
Discussion: F11R’s role in inflammatory cell migration explains its higher expression in rheumatoid arthritis patients. The lower F11R expression in subgroup with impaired salivary gland function may be related with less intercellular junction expression. C carrier in F11R promoter -688 A＞C locus seems protective. The relation of IFN-γ and F11R in rheumatoid arthritis patients needs more studies to clarify.

目錄
壹、 中文摘要 1
貳、 英文摘要 3
參、 前言 5
一、 F11R 5
二、 類風濕性關節炎：RA (Rheumatoid arthritis) 6
三、 類風濕性關節炎與F11R 6
四、 研究目的 7
肆、 材料與方法 8
一、 從全血中分離單核淋巴球、顆粒性淋巴球與白血球 8
二、 利用real time PCR偵測F11R之表現量 8
1. 研究族群 8
2. 萃取周邊血液單核淋巴球（PBMCs）的total RNA 8
3. 反轉錄（Reverse transcription） 9
4. 即時定量聚合酶連鎖反應 （quantitative real-time PCR） 10
三、 F11R 單一核苷酸多型性 （Single Nucleotide Polymorphism） 11
1.研究族群 11
2. 萃取Buffy coat中的DNA 11
3. 基因型鑑定 12
四、 IFN-γ （Interferon γ）表現量 13
1. 研究族群 13
2. 以ELISA (enzyme-link immunosorbent assay) 測定IFN-γ 13
五、 統計分析 15
伍、 結果 16
一、 F11R gene expression 表現結果 16
1. F11R 在RA患者的PBMC表現量比正常人多 16
2. RA病人有唾腺功能不良時，F11R mRNA數值較低。 16
3. F11R與RA病人其他secondary Sjögren’s syndrome標準相關性不顯著。 16
二、 F11R SNP結果 17
4. promoter -688 A＞C 與promoter -436 A>G 基因多型性在類風濕關節炎病人與正常人間無差別 17
5. promoter -688 A＞C 中C carrier有較低的anti-CCP 17
6. promoter -688 A＞C 中C carrier有較少淚腺功能不良 18
7. promoter -688 A＞C ：校正DR4的干擾後，C並未增加lymphadenopathy發生率。 18
8. promoter -688 A＞C: SNP genotype 與 gene expression 不相關 19
9. promoter -436 A>G：SNP genotype 與 gene expression 不相關 19
三、 IFN-γ 表現量 19
10. 類風濕關節炎病人血中IFN-γ濃度與PBMC F11R mRNA表現量並不相關 19
11.類風濕性關節炎病人和正常人間血中IFN-γ濃度沒有差別 19
12. 類風濕關節炎病人中IFN-γ濃度與CRP值成正相關 20
13. 周邊血中IFN-γ與RA病人合併secondary Sjögren’s syndrome特徵的關係 20
陸、 討論 21
一、 F11R表現量 21
二、 F11R SNP 23
三、 Interferon γ和F11R表現量 24
柒、 參考文獻 27
捌、 表目錄 33
1. RA與對照組之F11R mRNA 表現量的比較 33
2. F11R mRNA表現量與RA病人唾腺功能的關係 33
3. F11R與RA病人其他secondary Sjögren’s syndrome特徵的關係 34
4. RA與對照組之F11R alleles在Genotype frequency的比較 34
5. RA與對照組之F11R alleles在Gene carriage間的比較 35
6. RA與對照組之F11R alleles在Allele frequency間的比較 35
7. promoter -688 A＞C 中C carrier與anti-CCP的關係 36
8. promoter -688 A＞C校正DR4後與anti-CCP的關係 36
9. promoter -688 A＞C 中C carrier與schirmer’s test及lymphadenopathy間的關係 37
10. promoter -688 A＞C校正DR4後與淚腺功能的關係 37
11. promoter -688 A＞C校正DR4的干擾後，與lymphadenopathy關係 38
12. promoter -688 A＞C及promoter -436 A>G: SNP genotype 與 gene expression表現關係 38
13. 類風濕關節炎病人IFN-γ濃度與F11R mRNA表現量的關係 39
14. 類風濕關節炎病人與對照組之IFN-γ 血中濃度的比較 39
15. 類風濕關節炎病人中IFN-γ濃度與CRP值的關係 39
16. 周邊血中IFN-γ與RA病人secondary Sjögren’s syndrome特徵的關係 40

1.Kornecki, E., et al., Activation of human platelets by a stimulatory monoclonal antibody. J Biol Chem, 1990. 265(17): p. 10042-8.
2.Babinska, A., et al., F11-receptor (F11R/JAM) mediates platelet adhesion to endothelial cells: role in inflammatory thrombosis. Thromb Haemost, 2002. 88(5): p. 843-50.
3.Cavusoglu, E., et al., Association of Plasma Levels of F11 Receptor/Junctional Adhesion Molecule-A (F11R/JAM-A) With Human Atherosclerosis. Journal of the American College of Cardiology, 2007. 50(18): p. 1768-1776.
4.Azari, B.M., et al., Silencing of the F11R gene reveals a role for F11R/JAM-A in the migration of inflamed vascular smooth muscle cells and in atherosclerosis. Atherosclerosis, 2010. 212(1): p. 197-205.
5.Salifu, M.O., et al., Relationship between the soluble F11 receptor and markers of inflammation in hemodialysis patients. J Investig Med, 2007. 55(3): p. 115-9.
6.Hooper, N.M., E.H. Karran, and A.J. Turner, Membrane protein secretases. The Biochemical journal, 1997. 321 ( Pt 2): p. 265-79.
7.Martin-Padura, I., et al., Junctional adhesion molecule, a novel member of the immunoglobulin superfamily that distributes at intercellular junctions and modulates monocyte transmigration. J Cell Biol, 1998. 142(1): p. 117-27.
8.Ueki, T., et al., Expression of junctional adhesion molecules on the human lymphatic endothelium. Microvascular Research, 2008. 75(2): p. 269-278.
9.Ostermann, G., et al., JAM-1 is a ligand of the β2 integrin LFA-1 involved in transendothelial migration of leukocytes. Nature Immunology, 2002. 3(2): p. 151-158.
10.Ozaki, H., et al., Cutting edge: combined treatment of TNF-alpha and IFN-gamma causes redistribution of junctional adhesion molecule in human endothelial cells. J Immunol, 1999. 163(2): p. 553-7.
11.Sobocka, M.B., et al., Cloning of the human platelet F11 receptor: a cell adhesion molecule member of the immunoglobulin superfamily involved in platelet aggregation. Blood, 2000. 95(8): p. 2600-9.
12.Scott, D.L., F. Wolfe, and T.W. Huizinga, Rheumatoid arthritis. Lancet, 2010. 376(9746): p. 1094-108.
13.Arnett, F.C., et al., The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis. Arthritis Rheum, 1988. 31(3): p. 315-24.
14.Morita, A., et al., Molecular basis of tobacco smoke-induced premature skin aging. J Investig Dermatol Symp Proc, 2009. 14(1): p. 53-5.
15.van Venrooij, W.J., J.J. van Beers, and G.J. Pruijn, Anti-CCP Antibody, a Marker for the Early Detection of Rheumatoid Arthritis. Ann N Y Acad Sci, 2008. 1143: p. 268-85.
16.Turesson, C., et al., The impact of HLA-DRB1 genes on extra-articular disease manifestations in rheumatoid arthritis. Arthritis Res Ther, 2005. 7(6): p. R1386-93.
17.Sobocki, T., et al., Genomic structure, organization and promoter analysis of the human F11R/F11 receptor/junctional adhesion molecule-1/JAM-A. Gene, 2006. 366(1): p. 128-44.
18.Nourshargh, S., F. Krombach, and E. Dejana, The role of JAM-A and PECAM-1 in modulating leukocyte infiltration in inflamed and ischemic tissues. J Leukoc Biol, 2006. 80(4): p. 714-8.
19.Ley, K., et al., Getting to the site of inflammation: the leukocyte adhesion cascade updated. Nature reviews. Immunology, 2007. 7(9): p. 678-89.
20.Ewert, P., et al., Disruption of tight junction structure in salivary glands from Sjogren''s syndrome patients is linked to proinflammatory cytokine exposure. Arthritis Rheum, 2010. 62(5): p. 1280-9.
21.Gutwein, P., et al., Downregulation of junctional adhesion molecule-A is involved in the progression of clear cell renal cell carcinoma. Biochem Biophys Res Commun, 2009. 380(2): p. 387-91.
22.Ohyama, Y., et al., Cytokine messenger RNA expression in the labial salivary glands of patients with Sjogren''s syndrome. Arthritis Rheum, 1996. 39(8): p. 1376-84.
23.Fox, R.I., et al., Cytokine mRNA expression in salivary gland biopsies of Sjogren''s syndrome. J Immunol, 1994. 152(11): p. 5532-9.
24.Dolhain, R.J., et al., Increased expression of interferon (IFN)-gamma together with IFN-gamma receptor in the rheumatoid synovial membrane compared with synovium of patients with osteoarthritis. Br J Rheumatol, 1996. 35(1): p. 24-32.
25.Machold, K.P., K. Neumann, and J.S. Smolen, Recombinant human interferon gamma in the treatment of rheumatoid arthritis: double blind placebo controlled study. Ann Rheum Dis, 1992. 51(9): p. 1039-43.
26.Veys, E.M., et al., Interferon gamma in rheumatoid arthritis--a double blind study comparing human recombinant interferon gamma with placebo. J Rheumatol, 1988. 15(4): p. 570-4.
27.Cannon, G.W., et al., Prospective two-year followup of recombinant interferon-gamma in rheumatoid arthritis. J Rheumatol, 1990. 17(3): p. 304-10.
28.Vermeire, K., et al., Accelerated collagen-induced arthritis in IFN-gamma receptor-deficient mice. J Immunol, 1997. 158(11): p. 5507-13.
29.Andrews, H.J., et al., Modulation of human chondrocyte metabolism by recombinant human interferon gamma: in-vitro effects on basal and IL-1-stimulated proteinase production, cartilage degradation and DNA synthesis. Biochim Biophys Acta, 1989. 1012(2): p. 128-34.
30.Naik, M.U., et al., Attenuation of junctional adhesion molecule-A is a contributing factor for breast cancer cell invasion. Cancer Res, 2008. 68(7): p. 2194-203.