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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 胡忠怡(Chung-Yi Hu) | |
dc.contributor.author | Yen-Ju Chen | en |
dc.contributor.author | 陳衍儒 | zh_TW |
dc.date.accessioned | 2021-06-15T04:02:59Z | - |
dc.date.available | 2015-03-12 | |
dc.date.copyright | 2010-03-12 | |
dc.date.issued | 2010 | |
dc.date.submitted | 2010-02-11 | |
dc.identifier.citation | 1. Borowitz MJ, Chan JKC: B lymphoblastic leukaemia/lymphoma, not otherwise specified, in WHO classification of tumours of haematopoietic and lymphoid tissues 2008, 168-170 p. International Agency for Research on Cancer, Lyon.
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Linderson Y, Eberhard D, Malin S, Johansson A, Busslinger M, Pettersson S: Corecruitment of the Grg4 repressor by PU.1 is critical for Pax5-mediated repression of B-cell-specific genes, EMBO Rep 2004, 5:291-296 22. Eberhard D, Busslinger M: The partial homeodomain of the transcription factor Pax-5 (BSAP) is an interaction motif for the retinoblastoma and TATA-binding proteins, Cancer Res 1999, 59:1716s-1724s; discussion 1724s-1725s 23. Emelyanov AV, Kovac CR, Sepulveda MA, Birshtein BK: The interaction of Pax5 (BSAP) with Daxx can result in transcriptional activation in B cells, J Biol Chem 2002, 277:11156-11164 24. Urbánek P, Wang ZQ, Fetka I, Wagner EF, Busslinger M: Complete block of early B cell differentiation and altered patterning of the posterior midbrain in mice lacking Pax5/BSAP, Cell 1994, 79:901-912 25. Nutt SL, Urbanek P, Rolink A, Busslinger M: Essential functions of Pax5 (BSAP) in pro-B cell development: difference between fetal and adult B lymphopoiesis and reduced V-to-DJ recombination at the IgH locus, Genes Dev 1997, 11:476-491 26. Fuxa M, Busslinger M: Reporter gene insertions reveal a strictly B lymphoid-specific expression pattern of Pax5 in support of its B cell identity function, J Immunol 2007, 178:3031-3037 27. Cobaleda C, Schebesta A, Delogu A, Busslinger M: Pax5: the guardian of B cell identity and function, Nat Immunol 2007, 8:463-470 28. Souabni A, Cobaleda C, Schebesta M, Busslinger M: Pax5 promotes B lymphopoiesis and blocks T cell development by repressing Notch1, Immunity 2002, 17:781-793 29. Nutt SL, Heavey B, Rolink AG, Busslinger M: Commitment to the B-lymphoid lineage depends on the transcription factor Pax5, Nature 1999, 401:556-562 30. Rolink AG, Nutt SL, Melchers F, Busslinger M: Long-term in vivo reconstitution of T-cell development by Pax5-deficient B-cell progenitors, Nature 1999, 401:603-606 31. Kozmik Z, Kurzbauer R, Dorfler P, Busslinger M: Alternative splicing of Pax-8 gene transcripts is developmentally regulated and generates isoforms with different transactivation properties, Mol Cell Biol 1993, 13:6024-6035 32. Zwollo P, Arrieta H, Ede K, Molinder K, Desiderio S, Pollock R: The Pax-5 gene is alternatively spliced during B-cell development, J Biol Chem 1997, 272:10160-10168 33. Santoro A, Bica MG, Dagnino L, Agueli C, Salemi D, Cannella S, Veltroni M, Cetica V, Giarin E, Fabbiano F, Basso G, Arico M: Altered mRNA expression of PAX5 is a common event in acute lymphoblastic leukaemia, Br J Haematol 2009, 146:686-689 34. Sadakane Y, Zaitsu M, Nishi M, Sugita K, Mizutani S, Matsuzaki A, Sueoka E, Hamasaki Y, Ishii E: Expression and production of aberrant PAX5 with deletion of exon 8 in B-lineage acute lymphoblastic leukaemia of children, Br J Haematol 2007, 136:297-300 35. Robichaud GA, Nardini M, Laflamme M, Cuperlovic-Culf M, Ouellette RJ: Human Pax-5 C-terminal isoforms possess distinct transactivation properties and are differentially modulated in normal and malignant B cells, J Biol Chem 2004, 279:49956-49963 36. Arseneau JR, Laflamme M, Lewis SM, Maicas E, Ouellette RJ: Multiple isoforms of PAX5 are expressed in both lymphomas and normal B-cells, Br J Haematol 2009, 147:328-338 37. Zhou CZ, Peng ZH, Zhang F, Qiu GQ, He L: Loss of heterozygosity on long arm of chromosome 22 in sporadic colorectal carcinoma, World J Gastroenterol 2002, 8:668-673 38. Velasco A, Pallares J, Santacana M, Yeramian A, Dolcet X, Eritja N, Puente S, Sorolla A, Llecha N, Matias-Guiu X: Loss of heterozygosity in endometrial carcinoma, Int J Gynecol Pathol 2008, 27:305-317 39. Takeuchi S, Koike M, Seriu T, Bartram CR, Slater J, Park S, Miyoshi I, Koeffler HP: Homozygous deletions at 9p21 in childhood acute lymphoblastic leukemia detected by microsatellite analysis, Leukemia 1997, 11:1636-1640 40. Miller CB, Mullighan CG, Su X, Ma J, Wang M, Zhang J, Williams RT, Downing JR: Pax5 Haploinsufficiency Cooperates with BCR-ABL1 to Induce Acute Lymphoblastic Leukemia, ASH Annual Meeting Abstracts 2008, 112:293 41. Liang DC, Yang CP, Lin DT, Hung IJ, Lin KH, Chen JS, Hsiao CC, Chang TT, Peng CT, Lin MT, Chang TK, Jaing TH, Liu HC, Wang LY, Yeh TC, Jou ST, Lu MY, Cheng CN, Sheen JM, Chiou SS, Wu KH, Hung GY, Chen RL, Chen SH, Cheng SN, Chang YH, Chen BW, Ho WL, Wang JL, Lin ST, Hsieh YL, Wang SC, Chang HH, Yang YL, Huang FL, Chang CY, Chang WH, Lin KS: Long-term results of Taiwan Pediatric Oncology Group studies 1997 and 2002 for childhood acute lymphoblastic leukemia, Leukemia 2009, 24:397-405 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/45060 | - |
dc.description.abstract | PAX5 (Paired box 5)基因屬PAX基因家族的一員,表現於中樞神經及造血系統。PAX5蛋白N端會辨識目標基因之專一性DNA序列,C端會吸引其他轉錄相關蛋白,調控基因表現,並決定B細胞發育(B-cell commitment)。小兒急性淋巴性白血病(childhood acute lymphoid leukemia, childhood ALL)為淋巴系細胞發生癌變增生,使骨髓中的白血球失去正常細胞分化能力,影響正常造血細胞生長及功能的惡性腫瘤。在前人大規模調查研究中發現,小兒與成人B細胞ALL病人中,有近30%的病人在PAX5基因產生單一或雙對偶基因缺失、染色體轉位或點突變,暗示PAX5在B-ALL成病過程中,可能扮演重要角色。本研究希望了解PAX5在B細胞淋巴性白血病血癌細胞中表現的情形,解析PAX5表現變化與PAX5基因失去異合性(loss of heterozygosity, LOH)的關聯性,探討PAX5表現變異在B-ALL病患中是否具有分子特徵或臨床預後價值。
本研究針對採自25名小兒B-ALL患者之白血病細胞及3位正常B細胞對照組進行PAX5 mRNA表現量差異與臨床資料之關聯性分析,並進一步分析部分病人的PAX5蛋白表現量差異,及此差異是否源自PAX5 DNA之變異。結果顯示,雖未達統計上顯著差異,B-ALL白血病細胞內PAX5 mRNA表現普遍偏低(median (25%-75%), B-ALL:0.4 (0-0.9), Normal:0.9 (0.9-1), p=0.1438),半數以上B-ALL病人白血病細胞的PAX5 mRNA表現量低於正常B細胞對照組的一半以下。進一步分析PAX5 mRNA表現量與B-ALL細胞同時表現non-B marker數目之關聯性,雖未達統計上顯著差異,但可看到白血病細胞PAX5表現量較低,與其帶有較多non-B markers有關聯。此外,白血病細胞較正常B細胞擁有較多種長度變異片段。本研究亦分析7位B-ALL病人的白血病細胞PAX5蛋白表現量,發現其中4例PAX5蛋白表現量較低,3例表現量與正常B細胞對照組相似,且PAX5蛋白表現量趨勢與mRNA大致相同。另外利用微衛星標記分析6位B-ALL病人白血病細胞及緩解時期的PAX5 DNA刪除情形,3例為LOH,1例為無改變的異合子;奇怪的是,PAX5 DNA刪除情形與PAX5 mRNA表現量並不一致。綜合以上結果,顯示PAX5 mRNA、蛋白表現量差異,與PAX5 DNA是否發生LOH無直接關聯。結論: 1. 小兒急性B淋巴細胞白血病病人的PAX5基因表現量普遍較低。2. 部份B-ALL病人帶有較多的C端變異片段,包括影響PAX5蛋白質transactivation能力的Δ7/8片段。3. 部分病人的PAX5蛋白質表現量有顯著下降,但與PAX5基因劑量降低無直接相關。因此PAX5基因在DNA轉錄為RNA過程中,除了基因劑量外,或許仍有其他機制參與調控RNA表現。 | zh_TW |
dc.description.abstract | PAX5 (Paired box 5), a member of PAX gene family, is expressed in the central nervous and the hematopoietic systems and is known to play an important role in B cell commitment. Childhood acute lymphoid leukemia (ALL) is a malignant disease resulting from uncontrolled proliferation of lymphoid progenitors. In previous large-scaled study, the PAX5 gene was the most frequent target of somatic mutation, being altered in about 30% in both pediatric and adult patients with B-ALL. However, whether PAX5 mutation plays an important role in B-ALL leukemogenesis remains uncharacterized. In this study, we intend to investigate the molecular or clinical characteristics of PAX5 expression alteration in B-ALL.
The RNA and protein expression, DNA status (loss of heterozygosity, LOH) of PAX5 gene, as well as clinical data were investigated in 25 diagnostic bone marrow samples obtained from childhood B-ALL patients. Peripheral blood mature B cells obtained from 3 healthy subjects were used as control. Decreased PAX5 mRNA expression was noted in 14 out of 25 leukemic samples as compared to normal mature B cells (median (25%-75%), ALL: 0.4 (0-0.9), Normal: 0.9 (0.9-1), p=0.1438). Although it was not statistically significant, it revealed that the leukemic cells with decreased level of PAX5 mRNA expression tend to carry more non-B markers. There were more mRNAs with variant sizes in leukemic cells than in normal mature B cells. PAX5 protein level was also investigated in the leukemic cells of 7 B-ALL patients. Four showed decreased PAX5 expression and the other 3 were of comparable PAX5 level as compared to normal mature B cells. We also investigated LOH of PAX5 locus in 6 paired-samples (leukemic bone marrow cells and remission peripheral blood cells as normal reference) from B-ALL patients via short tandem repeat analysis. Three showed LOH and one remained heterozygous. The DNA status was not correlated with mRNA expression level. In summary, the trend of decreased PAX5 protein expression was parallel to that of PAX5 mRNA expression level, but is not correlated with DNA status. The results indicate that: 1. PAX5 mRNA expression is generally decreased in B-ALL patients; 2. There were more PAX5 C-terminal variant forms present in B-ALL leukemic samples, including transcript without exons 7 and 8, which may influence trans-activating ability of PAX5; 3. Significantly decreased protein level in 14 out of 25 B-ALL patients reveals the association in B-ALL and decreased PAX5 expression; 4. Besides the gene dosage, some other factors could regulate PAX5 transcription through mechanisms awaited further studies. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T04:02:59Z (GMT). No. of bitstreams: 1 ntu-99-R96424017-1.pdf: 1507562 bytes, checksum: e8fab67c71087c1e131589f2201034f6 (MD5) Previous issue date: 2010 | en |
dc.description.tableofcontents | 口試委員會審定書....................................................................................... i
誌謝........................................................................................................... ii 中文摘要..................................................................................................... iii 英文摘要...................................................................................................... v 第一章 緒論...............................................................................................1 1.1 小兒急性淋巴性白血病(childhood acute lymphoid leukemia, ALL)...................1 1.1.1 ALL流行病學資料...........................................................................................1 1.1.2急性淋巴性白血病(acute lymphoid leukemia)簡介.........................................1 1.1.3 ALL臨床症狀與疾病診斷...............................................................................2 1.2基因異常與小兒急性淋巴性白血病之關聯..........................................................4 1.3 PAX5........................................................................................................................5 1.3.1 PAX5簡介.........................................................................................................5 A. PAX5 (paired box 5)............................................................................................5 B. PAX5基因結構...................................................................................................5 C. PAX5 基因在B細胞發育過程中所扮演角色..................................................6 1.3.2 PAX5在B細胞中的splicing pattern................................................................7 1.4研究目的與實驗設計..............................................................................................8 第二章 材料與方法..................................................................................9 2.1實驗材料.................................................................................................................9 2.2成熟B細胞之純化.................................................................................................9 2.3反轉錄酶酵素聚合酶連鎖反應(RT-PCR) ..........................................................10 2.3.1抽取RNA並合成cDNA.................................................................................10 2.3.2聚合酶連鎖反應之引子及反應條件..............................................................10 2.3.3 PAX5 mRNA pattern分析...............................................................................11 2.3.4 PAX5 mRNA表現量分析..............................................................................11 2.4微衛星標記分析...................................................................................................11 2.4.1抽取DNA........................................................................................................11 2.4.2 微衛星標記分析之聚合酶連鎖反應............................................................12 2.4.3微衛星標記分析之ABI PRISM 3100毛細管電泳........................................12 2.4.4 LOH定義........................................................................................................13 2.4.5確認PAX5基因第六與第七轉錄子區域聚合酶鏈鎖反應...........................13 2.5西方點墨法 (Western blotting) ...........................................................................13 2.5.1萃取蛋白質......................................................................................................14 2.5.2硫酸十二酯鈉聚丙烯醯胺凝膠電泳(SDS-PAGE)........................................14 2.5.2.1配製SDS-polyacrylamide gel....................................................................14 2.5.2.2檢體前處理................................................................................................14 2.5.2.3電泳分析....................................................................................................14 2.5.3蛋白質轉漬 (Protein transfer) .......................................................................15 2.5.4免疫染色 (Immunoblot) ................................................................................15 2.5.5退染..................................................................................................................15 2.5.6以ImageJ定量蛋白表現量.............................................................................16 2.6統計分析方法及軟體...........................................................................................16 第三章 實驗結果.....................................................................................17 3.1小兒急性淋巴性白血病病人臨床資料...............................................................17 3.2 PAX5 mRNA半定量及定序分析........................................................................18 3.2.1半數ALL病人PAX5 mRNA表現量低於健康對照組..................................18 3.2.2 PAX5 mRNA表現量與non-B markers數目之關聯性分析..........................18 3.2.3 PAX5 exon 1-3與exon 6-10變種片段與風險因子之關聯性分析...............19 3.2.4 PAX5之定序分析...........................................................................................19 3.3 PAX5微衛星標記分析........................................................................................21 3.4 PAX5蛋白質分析................................................................................................22 3.5 PAX5基因微衛星標記及mRNA、蛋白質表現量之關聯性分析......................23 第四章 討論.............................................................................................24 參考文獻.....................................................................................................29 圖.................................................................................................................33 表.................................................................................................................44 附錄.............................................................................................................47 | |
dc.language.iso | zh-TW | |
dc.title | 小兒急性B淋巴細胞白血病中轉錄因子PAX5變異之調查 | zh_TW |
dc.title | Investigating the Variation of Transcription Factor PAX5 in Childhood B-Acute Lymphoblastic Leukemia | en |
dc.type | Thesis | |
dc.date.schoolyear | 98-1 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 林東燦(Dong-Tsamn Lin),林亮音(Liang-In Lin),楊雅倩(Ya-Chien Yang) | |
dc.subject.keyword | 小兒急性B淋巴細胞白血病,PAX5,PAX5 mRNA表現量,PAX5蛋白表現量,失去異合性, | zh_TW |
dc.subject.keyword | acute B-lymphoid leukemia (B-ALL),PAX5 (paired box 5),PAX5 mRNA expression,PAX5 protein expression,loss of heterozygosity (LOH), | en |
dc.relation.page | 82 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2010-02-11 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 醫學檢驗暨生物技術學研究所 | zh_TW |
顯示於系所單位: | 醫學檢驗暨生物技術學系 |
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