臺灣博碩士論文加值系統

川崎氏症為一種原因不明的急性全身性血管發炎症候群，主要好發於亞洲兒童，是後天性心臟病的主要元兇。先前研究指出在日本族群中，發現一些基因座與川崎氏症有相關，其中ITPKC和CASP3已被證實在日本族群及台灣族群皆與川崎氏症有關連。而ITPKC與CASP3為鈣池調控鈣離子通道 (Stroe-operated calcium channels；SOCs) 調控的鈣池調控鈣離子流 (Store-operated calcium entry) 的上下游，在發炎反應中扮演著重要的角色。因此，本研究對調控SOCs開啟的鈣離子濃度感應子STIM1和SOCs的重要組成分子ORAI1的基因多形性，以及將ITPKC與CASP3以2-locus analysis與川崎氏症之相關性進行探討。結果顯示，STIM1 (rs2304891) 與川崎氏症之併發症冠狀動脈損傷的形成 (於隱性遺傳模式中， P = 0.019) 有顯著的相關性，而攜帶G/A/A/A單體型組合者也有較高形成冠狀動脈損傷的危險 (OR：1.51，95% C.I.：0.98-2.34，P = 0.068)。而ORAI1的變異與川崎氏症之易感性、冠狀動脈損傷的形成及免疫球蛋白治療成效在我們的研究中皆未出現相關性。另一方面，在ITPKC與CASP3的2-locus analysis中，發現其基因間的交互作用在冠狀動脈損傷會有顯著的影響 (OR：3.06，95% C.I.：1.12-8.33，P = 0.023)；而對免疫球蛋白治療成效的分析，結果並未出現明顯的差異 (OR：3.41，95% C.I.：0.82-14.2，P = 0.106)。在2011年發表在Circulation: Cardiovascular Genetics的文獻指出，以美國/英國/澳洲等族群對川崎氏症的研究發現TGF-beta signaling pathway扮演了重要的角色。因此我們首度以亞洲族群作為探討，結果顯示在TGF-beta signaling pathway中，當SMAD3 (rs1438386) 攜帶T對偶基因型時，與川崎氏症之易感性有非常顯著的相關 (於對偶遺傳模式中，P = 0.001)。進一步以單體型做統計，TGF-β2基因rs10482751/rs2027567/rs1209576攜帶T/G/G單體型有較高罹患川崎氏症的風險 (OR：1.24，95％ CI：1.01-1.52，P = 0.04)；在SMAD3 基因rs4776338/rs7162912/rs12901071攜帶T/A/A或C/C/G單體型則分別有4倍(P <0.0001)和37倍(P <0.0001)較高的風險罹患川崎氏症。

Kawasaki disease (KD) is an acute systemic vasculitis syndrome of an unknown etiology. KD occurs worldwide especially in Asia. In developed countries, KD is the leading cause of acquired heart disease in children. Previous studies identified several candidate regions by carring out Genome-wide association study for KD susceptibility. Functional SNPs in ITPKC (rs28493229) and CASP3 (rs72689236) had been confirmed the association with KD both in Japan and in Taiwan population indivisually. In non-excitable cells such as T cells and mast cells, the calcium influx through IP3-mediated store-operated channels (SOCs) plays an impotant role in regulating inflammatory reaction. Besides, SOCs are in the middle of the stream between ITPKC and CASP3. STIM1 is a calcium concentration sensor, activation of STIM1 would conjugate with SOCs major component-ORAI1 then trigger calcium influx. In this study, we aim to determine the association between STIM1 gene polymorphisms, ORAI1 gene polymorphisms, ITPKC/CASP3 2-locus analysis and KD. Our results showed that a SNP (rs2304891) in STIM1 may contribute to the risk of coronary artery formation, however there was no evidence indicated ORAI1 genetic variations could affected KD pathology. In addition, we were first supported the similar result in the Japanese population that ITPKC and CASP combinatorial effect to coronary artery formation in Taiwan population. In 2011, an article from the “Circulation: Cardiovascular Genetics” indicated that TGF-β pathways polymorphisms had significant association with KD in the Europe and U.S. population. We picked 12 SNPs that had significant influences in the multiethnic TDT or CAL formation from this study, however, our results indicated that only a SNP (rs1438386) and its haplotypes associated with the susceptibility of KD.

目錄
表目錄.... ................................................................................................6
圖目錄.....................................................................................................9
中文摘要................................................................................................10
英文摘要................................................................................................12
壹、緒論.......... .....................................................................................14
一、 川崎氏症(Kawasaki disease).......................................................14
二、 鈣池調控鈣離子通道 (Store-operated Ca2+ Channel；SOC)....15
三、 Inositol 1,4,5-trisphosphate 3-kinase C (ITPKC)……………….16
四、 Stromal interaction molecule 1 (STIM1)......................................18
五、 Orai1/CRACM1............................................................................19
六、 Transforming growth factor-beta (TGF-β) signaling pathway....21
貳、材料與方法......................................................................................22
一、病患篩選 (Patient recruitments) .....................................................22
二、DNA萃取 (DNA extraction) ..........................................................23
三、單一核苷酸基因多形性選擇條件...................................................25
四、基因型檢測 (TaqMan Genotyping Assay) .....................................29
伍、統計學分析......................................................................................31
叁、第一節 STIM1及ORAI1基因多形性與川崎氏症之相關性..........33
一、背景....................................................................................................33
二、結果....................................................................................................36
三、結論....................................................................................................42
四、討論....................................................................................................43

第二節 ITPKC和Caspase3 2-locus analysis與川崎氏症之相關性.......45
一、背景....................................................................................................45
二、結果....................................................................................................48
三、結論....................................................................................................50
四、討論....................................................................................................51

第三節TGF-β signaling pathway基因多形性與川崎氏症之相關性..53
一、背景....................................................................................................53
二、結果....................................................................................................55
三、結論....................................................................................................58
四、討論....................................................................................................59
肆、參考文獻............................................................................................61
伍、圖表....................................................................................................67

1.Wang, C.L., et al., Kawasaki disease: infection, immunity and genetics. Pediatr Infect Dis J, 2005. 24(11): p. 998-1004.
2.Liang, C.D., et al., Coronary artery fistula associated with Kawasaki disease. Am Heart J, 2009. 157(3): p. 584-8.
3.Roos, J., et al., STIM1, an essential and conserved component of store-operated Ca2+ channel function. J Cell Biol, 2005. 169(3): p. 435-45.
4.Feske, S., et al., A mutation in Orai1 causes immune deficiency by abrogating CRAC channel function. Nature, 2006. 441(7090): p. 179-85.
5.Onouchi, Y., et al., ITPKC functional polymorphism associated with Kawasaki disease susceptibility and formation of coronary artery aneurysms. Nat Genet, 2008. 40(1): p. 35-42.
6.Liou, J., et al., STIM is a Ca2+ sensor essential for Ca2+-store-depletion-triggered Ca2+ influx. Curr Biol, 2005. 15(13): p. 1235-41.
7.Williams, R.T., et al., Identification and characterization of the STIM (stromal interaction molecule) gene family: coding for a novel class of transmembrane proteins. Biochem J, 2001. 357(Pt 3): p. 673-85.
8.Manji, S.S., et al., STIM1: a novel phosphoprotein located at the cell surface. Biochim Biophys Acta, 2000. 1481(1): p. 147-55.
9.Stathopulos, P.B., et al., Structural and mechanistic insights into STIM1-mediated initiation of store-operated calcium entry. Cell, 2008. 135(1): p. 110-22.
10.Liou, J., et al., Live-cell imaging reveals sequential oligomerization and local plasma membrane targeting of stromal interaction molecule 1 after Ca2+ store depletion. Proc Natl Acad Sci U S A, 2007. 104(22): p. 9301-6.
11.Zhang, S.L., et al., STIM1 is a Ca2+ sensor that activates CRAC channels and migrates from the Ca2+ store to the plasma membrane. Nature, 2005. 437(7060): p. 902-5.
12.Wu, M.M., et al., Ca2+ store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane. J Cell Biol, 2006. 174(6): p. 803-13.
13.Luik, R.M., et al., The elementary unit of store-operated Ca2+ entry: local activation of CRAC channels by STIM1 at ER-plasma membrane junctions. J Cell Biol, 2006. 174(6): p. 815-25.
14.Potier, M. and M. Trebak, New developments in the signaling mechanisms of the store-operated calcium entry pathway. Pflugers Arch, 2008. 457(2): p. 405-15.
15.Huang, G.N., et al., STIM1 carboxyl-terminus activates native SOC, I(crac) and TRPC1 channels. Nat Cell Biol, 2006. 8(9): p. 1003-10.
16.Penna, A., et al., The CRAC channel consists of a tetramer formed by Stim-induced dimerization of Orai dimers. Nature, 2008. 456(7218): p. 116-20.
17.Ji, W., et al., Functional stoichiometry of the unitary calcium-release-activated calcium channel. Proc Natl Acad Sci U S A, 2008. 105(36): p. 13668-73.
18.Yeromin, A.V., et al., Molecular identification of the CRAC channel by altered ion selectivity in a mutant of Orai. Nature, 2006. 443(7108): p. 226-9.
19.Vig, M., et al., CRACM1 multimers form the ion-selective pore of the CRAC channel. Curr Biol, 2006. 16(20): p. 2073-9.
20.Prakriya, M., et al., Orai1 is an essential pore subunit of the CRAC channel. Nature, 2006. 443(7108): p. 230-3.
21.Lis, A., et al., CRACM1, CRACM2, and CRACM3 are store-operated Ca2+ channels with distinct functional properties. Curr Biol, 2007. 17(9): p. 794-800.
22.Maruyama, Y., et al., Tetrameric Orai1 is a teardrop-shaped molecule with a long, tapered cytoplasmic domain. J Biol Chem, 2009. 284(20): p. 13676-85.
23.Li, M.O., et al., Transforming growth factor-beta regulation of immune responses. Annu Rev Immunol, 2006. 24: p. 99-146.
24.Massague, J., TGFbeta in Cancer. Cell, 2008. 134(2): p. 215-30.
25.Blobe, G.C., W.P. Schiemann, and H.F. Lodish, Role of transforming growth factor beta in human disease. N Engl J Med, 2000. 342(18): p. 1350-8.
26.Taylor, A.W., Review of the activation of TGF-beta in immunity. J Leukoc Biol, 2009. 85(1): p. 29-33.
27.Christ, M., et al., Immune dysregulation in TGF-beta 1-deficient mice. J Immunol, 1994. 153(5): p. 1936-46.
28.Leask, A. and D.J. Abraham, TGF-beta signaling and the fibrotic response. FASEB J, 2004. 18(7): p. 816-27.
29.Chen, C.L., et al., Cholesterol suppresses cellular TGF-beta responsiveness: implications in atherogenesis. J Cell Sci, 2007. 120(Pt 20): p. 3509-21.
30.Matsubara, T., et al., A sibship with recurrent Kawasaki disease and coronary artery lesion. Acta Paediatr, 1994. 83(9): p. 1002-4.
31.de Inocencio, J. and R. Hirsch, The role of T cells in Kawasaki disease. Crit Rev Immunol, 1995. 15(3-4): p. 349-57.
32.Parekh, A.B. and J.W. Putney, Jr., Store-operated calcium channels. Physiol Rev, 2005. 85(2): p. 757-810.
33.Chang, W.C. and A.B. Parekh, Close functional coupling between Ca2+ release-activated Ca2+ channels, arachidonic acid release, and leukotriene C4 secretion. J Biol Chem, 2004. 279(29): p. 29994-9.
34.Vig, M. and J.P. Kinet, Calcium signaling in immune cells. Nat Immunol, 2009. 10(1): p. 21-7.
35.Feske, S., C. Picard, and A. Fischer, Immunodeficiency due to mutations in ORAI1 and STIM1. Clin Immunol, 2010. 135(2): p. 169-82.
36.Kuo, H.C., et al., ITPKC single nucleotide polymorphism associated with the Kawasaki disease in a Taiwanese population. PLoS ONE, 2011. 6(4): p. e17370.
37.Lin, M.T., et al., Clinical Implication of the C Allele of the ITPKC Gene SNP rs28493229 in Kawasaki Disease: Association With Disease Susceptibility and BCG Scar Reactivation. Pediatr Infect Dis J, 2011. 30(2): p. 148-52.
38.Onouchi, Y., et al., Common variants in CASP3 confer susceptibility to Kawasaki disease. Hum Mol Genet, 2010. 19(14): p. 2898-906.
39.Kuo, H.C., et al., CASP3 gene single-nucleotide polymorphism (rs72689236) and Kawasaki disease in Taiwanese children. J Hum Genet, 2011. 56(2): p. 161-5.
40.Lewis, R.S., The molecular choreography of a store-operated calcium channel. Nature, 2007. 446(7133): p. 284-7.
41.Shimizu, C., et al., Transforming growth factor-beta signaling pathway in patients with Kawasaki disease. Circ Cardiovasc Genet, 2011. 4(1): p. 16-25.
42.Chi, H., et al., ITPKC gene SNP rs28493229 and Kawasaki disease in Taiwanese children. Hum Mol Genet, 2010. 19(6): p. 1147-51.
43.Onouchi, Y., Identification of susceptibility genes for Kawasaki disease. Nihon Rinsho Meneki Gakkai Kaishi, 2010. 33(2): p. 73-80.
44.Hata, A. and Y. Onouchi, Susceptibility genes for Kawasaki disease: toward implementation of personalized medicine. J Hum Genet, 2009. 54(2): p. 67-73.
45.Parekh, A.B., A. Fleig, and R. Penner, The store-operated calcium current I(CRAC): nonlinear activation by InsP3 and dissociation from calcium release. Cell, 1997. 89(6): p. 973-80.
46.Abdullaev, I.F., et al., Stim1 and Orai1 mediate CRAC currents and store-operated calcium entry important for endothelial cell proliferation. Circ Res, 2008. 103(11): p. 1289-99.
47.Nishizuka, Y., The molecular heterogeneity of protein kinase C and its implications for cellular regulation. Nature, 1988. 334(6184): p. 661-5.
48.Emoto, Y., et al., Proteolytic activation of protein kinase C delta by an ICE-like protease in apoptotic cells. EMBO J, 1995. 14(24): p. 6148-56.
49.Majumder, P.K., et al., Mitochondrial translocation of protein kinase C delta in phorbol ester-induced cytochrome c release and apoptosis. J Biol Chem, 2000. 275(29): p. 21793-6.
50.Albert, A.P. and W.A. Large, Activation of store-operated channels by noradrenaline via protein kinase C in rabbit portal vein myocytes. J Physiol, 2002. 544(Pt 1): p. 113-25.
51.Ghayur, T., et al., Proteolytic activation of protein kinase C delta by an ICE/CED 3-like protease induces characteristics of apoptosis. J Exp Med, 1996. 184(6): p. 2399-404.
52.Yi, Q.J., C.R. Li, and X.Q. Yang, Effect of intravenous immunoglobulin on inhibiting peripheral blood lymphocyte apoptosis in acute Kawasaki disease. Acta Paediatr, 2001. 90(6): p. 623-7.
53.Burns, J.C., et al., Intravenous gamma-globulin treatment and retreatment in Kawasaki disease. US/Canadian Kawasaki Syndrome Study Group. Pediatr Infect Dis J, 1998. 17(12): p. 1144-8.
54.Wallace, C.A., et al., Initial intravenous gammaglobulin treatment failure in Kawasaki disease. Pediatrics, 2000. 105(6): p. E78.
55.Durongpisitkul, K., et al., Immunoglobulin failure and retreatment in Kawasaki disease. Pediatr Cardiol, 2003. 24(2): p. 145-8.
56.Uehara, R., et al., Analysis of potential risk factors associated with nonresponse to initial intravenous immunoglobulin treatment among Kawasaki disease patients in Japan. Pediatr Infect Dis J, 2008. 27(2): p. 155-60.
57.Sano, T., et al., Prediction of non-responsiveness to standard high-dose gamma-globulin therapy in patients with acute Kawasaki disease before starting initial treatment. Eur J Pediatr, 2007. 166(2): p. 131-7.
58.Egami, K., et al., Prediction of resistance to intravenous immunoglobulin treatment in patients with Kawasaki disease. J Pediatr, 2006. 149(2): p. 237-40.
59.Kobayashi, T., et al., Prediction of intravenous immunoglobulin unresponsiveness in patients with Kawasaki disease. Circulation, 2006. 113(22): p. 2606-12.
60.Burns, J.C. and M.P. Glode, Kawasaki syndrome. Lancet, 2004. 364(9433): p. 533-44.
61.Tremoulet, A.H., et al., Increased incidence and severity of Kawasaki disease among Filipino-Americans in San Diego county. Pediatr Infect Dis J, 2011. 30(10): p. 909-11.
62.Kashef, S., M. Safari, and R. Amin, Initial intravenous gamma-globulin treatment failure in Iranian children with Kawasaki disease. Kaohsiung J Med Sci, 2005. 21(9): p. 401-4.
63.Onouchi, Y., et al., ITPKC and CASP3 polymorphisms and risks for IVIG unresponsiveness and coronary artery lesion formation in Kawasaki disease. Pharmacogenomics J, 2011.
64.Ruiz-Ortega, M., et al., TGF-beta signaling in vascular fibrosis. Cardiovasc Res, 2007. 74(2): p. 196-206.
65.Clark-Greuel, J.N., et al., Transforming growth factor-beta1 mechanisms in aortic valve calcification: increased alkaline phosphatase and related events. Ann Thorac Surg, 2007. 83(3): p. 946-53.
66.Loeys, B.L., et al., A syndrome of altered cardiovascular, craniofacial, neurocognitive and skeletal development caused by mutations in TGFBR1 or TGFBR2. Nat Genet, 2005. 37(3): p. 275-81.
67.Neptune, E.R., et al., Dysregulation of TGF-beta activation contributes to pathogenesis in Marfan syndrome. Nat Genet, 2003. 33(3): p. 407-11.
68.Steinberg, M.H., Genetic etiologies for phenotypic diversity in sickle cell anemia. ScientificWorldJournal, 2009. 9: p. 46-67.
69.Sebastiani, P., et al., Genetic dissection and prognostic modeling of overt stroke in sickle cell anemia. Nat Genet, 2005. 37(4): p. 435-40.
70.Amano, S., F. Hazama, and Y. Hamashima, Pathology of Kawasaki disease: I. Pathology and morphogenesis of the vascular changes. Jpn Circ J, 1979. 43(7): p. 633-43.
71.Rowley, A.H., et al., Searching for the cause of Kawasaki disease--cytoplasmic inclusion bodies provide new insight. Nat Rev Microbiol, 2008. 6(5): p. 394-401.
72.Onouchi, H., et al., Nascent peptide-mediated translation elongation arrest of Arabidopsis thaliana CGS1 mRNA occurs autonomously. Plant Cell Physiol, 2008. 49(4): p. 549-56.
73.Brown, T.J., et al., CD8 T lymphocytes and macrophages infiltrate coronary artery aneurysms in acute Kawasaki disease. J Infect Dis, 2001. 184(7): p. 940-3.
74.Franco, A., et al., Memory T-cells and characterization of peripheral T-cell clones in acute Kawasaki disease. Autoimmunity, 2010. 43(4): p. 317-24.
75.Lan, R.Y., et al., Regulatory T cells: development, function and role in autoimmunity. Autoimmun Rev, 2005. 4(6): p. 351-63.
76.Kim, J.J., et al., A genome-wide association analysis reveals 1p31 and 2p13.3 as susceptibility loci for Kawasaki disease. Hum Genet, 2011. 129(5): p. 487-95.
77.Tsai, F.J., et al., Identification of novel susceptibility Loci for kawasaki disease in a Han chinese population by a genome-wide association study. PLoS ONE, 2011. 6(2): p. e16853.