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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/82801完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 李建南(Chien-Nan Lee) | |
| dc.contributor.author | Yu-Fen Chih | en |
| dc.contributor.author | 池郁棻 | zh_TW |
| dc.date.accessioned | 2022-11-25T07:59:53Z | - |
| dc.date.copyright | 2021-11-09 | |
| dc.date.issued | 2021 | |
| dc.date.submitted | 2021-09-16 | |
| dc.identifier.citation | [1] Z. Rosenwaks et al., 'The pros and cons of preimplantation genetic testing for aneuploidy: clinical and laboratory perspectives,' Fertility and sterility, vol. 110, no. 3, pp. 353-361, 2018. [2] R. Makhijani et al., 'Impact of trophectoderm biopsy on obstetric and perinatal outcomes following frozen–thawed embryo transfer cycles,' Human Reproduction, vol. 36, no. 2, pp. 340-348, 2021. [3] M. Montag, K. van der Ven, B. Rösing, and H. van der Ven, 'Polar body biopsy: a viable alternative to preimplantation genetic diagnosis and screening,' Reproductive biomedicine online., vol. 18, pp. 6-11, 2009, doi: 10.1016/S1472-6483(10)60109-5. [4] L. Huang, B. Bogale, Y. Tang, S. Lu, X. Xie, and C. Racowsky, 'Noninvasive preimplantation genetic testing for aneuploidy in spent medium may be more reliable than trophectoderm biopsy,' Proceedings of the National Academy of Sciences of the United States of America, vol. 116, no. 28, p. 14105, 2019, doi: 10.1073/pnas.1907472116. [5] J. Xu et al., 'Noninvasive chromosome screening of human embryos by genome sequencing of embryo culture medium for in vitro fertilization,' Proceedings of the National Academy of Sciences, vol. 113, no. 42, pp. 11907-11912, 2016. [6] M. Leaver and D. Wells, 'Non-invasive preimplantation genetic testing (niPGT): the next revolution in reproductive genetics?,' Human reproduction update, vol. 26, no. 1, pp. 16-42, 2020. [7] J. Jiao et al., 'Minimally invasive preimplantation genetic testing using blastocyst culture medium,' Human Reproduction, vol. 34, no. 7, pp. 1369-1379, 2019. [8] V. Kuznyetsov et al., 'Evaluation of a novel non-invasive preimplantation genetic screening approach,' PLoS ONE, vol. 13, no. 5, 2018, doi: 10.1371/journal.pone.0197262. [9] D. K. Gardner, M. Lane, J. Stevens, T. Schlenker, and W. B. Schoolcraft, 'Blastocyst score affects implantation and pregnancy outcome: towards a single blastocyst transfer,' Fertility and sterility., vol. 73, no. 6, pp. 1155-1158, 2000, doi: 10.1016/S0015-0282(00)00518-5. [10] C. Robinson and M. J. Jasper, 'Embryo culturing conditions and the development of non-invasive preimplantation genetic testing for aneuploidy detection (NI-PGT-A),' Fertility and sterility., vol. 112, no. 3, pp. e239-e240, 2019, doi: 10.1016/j.fertnstert.2019.07.1370. [11] K. Rule, R. J. Chosed, T. A. Chang, J. D. Wininger, and W. E. Roudebush, 'Relationship between blastocoel cell-free DNA and day-5 blastocyst morphology,' Journal of assisted reproduction and genetics, vol. 35, no. 8, pp. 1497-1501, 2018. [12] B. M. Hanson et al., 'Noninvasive preimplantation genetic testing for aneuploidy exhibits high rates of deoxyribonucleic acid amplification failure and poor correlation with results obtained using trophectoderm biopsy,' Fertility and Sterility, 2021. [13] Q. S. Yeung et al., 'A prospective study of non-invasive preimplantation genetic testing for aneuploidies (NiPGT-A) using next-generation sequencing (NGS) on spent culture media (SCM),' Journal of assisted reproduction and genetics, vol. 36, no. 8, pp. 1609-1621, 2019. [14] E. Darwish and Y. Magdi, 'Artificial shrinkage of blastocoel using a laser pulse prior to vitrification improves clinical outcome,' Journal of assisted reproduction and genetics, vol. 33, no. 4, pp. 467-471, 2016. [15] V. Kuznyetsov et al., 'Minimally invasive cell-free human embryo aneuploidy testing (miPGT-A) utilizing combined spent embryo culture medium and blastocoel fluid–towards development of a clinical assay,' Scientific reports, vol. 10, no. 1, pp. 1-12, 2020. [16] C. Rubio et al., 'Embryonic cell-free DNA versus trophectoderm biopsy for aneuploidy testing: concordance rate and clinical implications,' Fertility and sterility, vol. 112, no. 3, pp. 510-519, 2019. [17] P. Vanderzwalmen et al., 'Births after vitrification at morula and blastocyst stages: effect of artificial reduction of the blastocoelic cavity before vitrification,' Human reproduction., vol. 17, no. 3, pp. 744-751, 2002, doi: 10.1093/humrep/17.3.744. [18] 'Noninvasive PGT-A ( NIPGT-A ) Promises and Pitfalls,' 2020. [19] N. Gleicher, J. Metzger, G. Croft, V. A. Kushnir, D. F. Albertini, and D. H. Barad, 'A single trophectoderm biopsy at blastocyst stage is mathematically unable to determine embryo ploidy accurately enough for clinical use,' Reproductive Biology and Endocrinology, vol. 15, no. 1, pp. 1-8, 2017. [20] C. Rubio, C. Racowsky, D. H. Barad, R. T. Scott, and C. Simon, 'Noninvasive preimplantation genetic testing for aneuploidy in spent culture medium as a substitute for trophectoderm biopsy,' Fertility and Sterility, vol. 115, no. 4, pp. 841-849, 2021. [21] L. Huang, S. Lu, C. Racowsky, and X. S. Xie, 'Reply to Gleicher and Barad: noninvasive preimplantation genetic testing may provide the solution to the problem of embryo mosaicism,' Proceedings of the National Academy of Sciences, p. 201912042, 2019. [22] R. C. McCoy, 'Mosaicism in preimplantation human embryos: when chromosomal abnormalities are the norm,' Trends in genetics, vol. 33, no. 7, pp. 448-463, 2017. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/82801 | - |
| dc.description.abstract | "對於習慣性流產、3次以上試管嬰兒療程失敗、高齡婦女、家族有染色體異常病史的夫妻接受試管嬰兒治療時，需要施行胚胎著床前染色體篩檢(preimplantation genetic testing for aneuploidy,PGT-A)，目的是在植入前先了解胚胎染色體是否正常。常規的PGT-A必須採樣囊胚滋養層細胞(TE)作檢驗，這種侵入性的篩檢需要純熟技巧，若胚胎發育不良，則無法進行細胞採樣，採樣過程也可能會對胚胎造成損害，而細胞採樣可能會出現偽陰性和偽陽性，故目前對於PGT-A的應用仍有很多疑慮。 近幾年，有學者在培養人類囊胚培養液中發現細胞游離DNA(cfDNA) ，嘗試用剩餘的囊胚培養液(SCM)做胚胎染色體分析，來比較侵入性與非侵入性的PGT-A中各種參數的一致性。目前的瓶頸為無法確認胚胎的cfDNA是否完整，cfDNA與TE採樣和整個胚胎的各種參數一致性如何，也無法確認cfDNA來自於胚胎本身或胚胎廢棄細胞所排出，或是剔除卵丘細胞過程所遺留下來的DNA(母源汙染)等。2016年開始，有學者以單一SCM分析胚胎的cfDNA，研究結果所得到倍性的一致性偏低，近幾年有些學者將剩餘的培養液SCM與囊胚腔液(BF)作為共同培養液樣本分析胚胎的cfDNA，目前世界上所得到倍性的一致性結果差異性大。 本研究的捐贈冷凍囊胚組樣本為剩餘培養液(SCM)結合囊胚腔液(BF)、內細胞團(ICM)採樣與滋養層(TE)採樣樣本，新鮮培養囊胚組樣本為剩餘培養液(SCM)結合囊胚腔液(BF)與滋養層(TE)採樣樣本，利用NGS進行序列分析，來了解這兩個組別胚胎染色體經序列分析後的一致性，並探討niPGT-A是否將來可以應用在臨床上。在捐贈囊胚組，共收集了55個冷凍囊胚，其中2顆WGA沒有訊號，3顆有雜訊故排除分析，最後分析50個，包括12個整倍體、28個非整倍體及10個鑲嵌型胚胎，這些囊胚先前已經過TE採樣之PGT-A檢測；捐贈的囊胚解凍後，個別以20-μLLifeGlobal培養微滴覆蓋培養油，並於timelapse培養箱獨立培養(37 °C ,5% O2, 6% CO2 )，經培養24小時，收集囊胚腔液(6-μL)，接著以雷射輔助孵化設備(Zilos Tk laser ,Hamilton Thorne Biosciences)將囊胚打洞，並於一小時後收集囊胚腔液與培養液(6-μL)。另外再進行囊胚內細胞團(ICM)之採樣。在新鮮囊胚組，分析35個囊胚，其中10顆WGA沒有訊號，每顆胚胎個別以20-μL LifeGlobal培養微滴覆蓋培養油後於timelapse培養箱獨立培養，培養後第四天進行置換新鮮培養微滴，培養到第五天(24小時)評估囊胚等級，收集完全膨脹囊胚之培養液(6-μL)，接著以雷射輔助孵化設備將囊胚打洞，進行雷射囊胚塌陷後，使囊胚液流出與剩餘的培養液混合，一小時後收集此混合的培養液(6-μL)。另外再進行囊胚滋養層(TE)之採樣。對照組為未接觸囊胚的培養微滴。 所有樣本放置於PCR tube內，冷凍於-20℃直到分析。WGA PGS kit為PerkinElmer PG-Seq Rapide niPGT kit，Sequencing and Data Analysis為Illumina MiSeq v3 Reagent,PG-Find分析軟體。 由結果可看出與單獨收集SCM相比，混合培養液的WGA放大率比較高，捐贈組訊號率明顯高於新鮮組具顯著差異(96% VS 71%，P=0.0002)。捐贈組囊胚ICM vs SB倍性一致性為88%(44/50)，ICM vs TE倍性一致性82%(41/50)，沒有顯著差異(P=0.277)。 ICM vs SB 的FPR為4.8%(1/21) ， ICM vs TE的FPR為42.9%(9/21)，有差異顯著(P=0.0015)。ICM vs SB 的PPV為96%(24/25)， ICM vs TE的PPV為76.3%(29/38)，有顯著差異(P=0.0437)。ICM vs SB 的Specificity為95.2%(20/21)，ICM vs TE的Specificity為57.1%(12/21)，具顯差異著(P=0.0017)。新鮮組25個囊胚的TE vs SB倍性一致性為44%(11/25)； 性別一致性為84% (21/25)：有三個疑似母源DNA汙染，一個是雜訊過多，無法代表真實結果。新鮮組有九個胚胎經iPGT-A篩檢為整倍體的胚胎植入，其中七個胚胎著床，一個已生下正常胎兒，六個持續懷孕，周數大部分皆於第二孕期，另外兩個沒有著床。 目前niPGT-A的疑慮為培養液有多種可能的汙染源，主要可能有母源(卵丘細胞)汙染、或其他外來汙染源等，因此在受精之前必須仔細地去除卵丘細胞，更換培養液時也要嚴謹洗滌胚胎，更要避免外來汙染源。雖然niPGT-A的診斷看起來有參考價值，目前仍無法取代TE，輔助參考可以應用於：1.拯救iPGT-A誤診的胚胎，2.不合適做iPGT-A的囊胚。但目前需要更多證據來證實這項診斷技術在臨床的可行性。" | zh_TW |
| dc.description.provenance | Made available in DSpace on 2022-11-25T07:59:53Z (GMT). No. of bitstreams: 1 U0001-1009202115502500.pdf: 4312066 bytes, checksum: 283f140417d1618035561c7f2e6c7c94 (MD5) Previous issue date: 2021 | en |
| dc.description.tableofcontents | 口試論文審定書..........................i 致謝..................................ii 臨床試驗/研究許可書....................iii 中文摘要..............................iv-v 英文摘要..............................vi-vii 目錄..................................viii 前言..................................1 研究方法與材料.........................6 結果..................................10 討論..................................12 結論..................................21 補充..................................22 參考文獻..............................24 圖目錄 圖一...................................26 圖二...................................26 圖三...................................27 表目錄 表一...................................28 表二...................................29 表三A...................................32 表三B...................................33 表四................................... 34 表五................................... 36 表六. ..................................37 附錄 臨床試驗同意書............................39 NGS告知說明同意書.........................45 | |
| dc.language.iso | zh-TW | |
| dc.subject | 囊胚腔液(BF) | zh_TW |
| dc.subject | 胚胎著床前染色體篩檢(PGT-A) | zh_TW |
| dc.subject | 剩餘的囊胚培養液(SCM) | zh_TW |
| dc.subject | 細胞游離DNA(cfDNA) | zh_TW |
| dc.subject | 侵入性(iPGT-A) | zh_TW |
| dc.subject | 非侵入性(niPGT-A) | zh_TW |
| dc.subject | spent culture medium(SCM) | en |
| dc.subject | Noninvasive preimplantation genetic testing for aneuploidy(niPGT-A) | en |
| dc.subject | invasive preimplantation genetic testing for aneuploidy(iPGT-A) | en |
| dc.subject | preimplantation genetic testing for aneuploidy(PGT-A) | en |
| dc.subject | blastocoel fluid(BF) | en |
| dc.subject | cell-free DNA(cfDNA) | en |
| dc.title | 比較胚胎培養液細胞游離DNA、滋養層採樣以及內細胞團採樣之染色體篩檢成效 | zh_TW |
| dc.title | "Pre-implantation chromosome screening: comparing of cell free DNA from combined blastocoel fluid and spent culture medium, trophoblast biopsy and inner cell mass biopsy" | en |
| dc.date.schoolyear | 109-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 林芯伃(Hsin-Tsai Liu),陳思原(Chih-Yang Tseng) | |
| dc.subject.keyword | 細胞游離DNA(cfDNA),剩餘的囊胚培養液(SCM),囊胚腔液(BF),胚胎著床前染色體篩檢(PGT-A),侵入性(iPGT-A),非侵入性(niPGT-A), | zh_TW |
| dc.subject.keyword | cell-free DNA(cfDNA),spent culture medium(SCM),blastocoel fluid(BF),preimplantation genetic testing for aneuploidy(PGT-A),invasive preimplantation genetic testing for aneuploidy(iPGT-A),Noninvasive preimplantation genetic testing for aneuploidy(niPGT-A), | en |
| dc.relation.page | 46 | |
| dc.identifier.doi | 10.6342/NTU202103111 | |
| dc.rights.note | 未授權 | |
| dc.date.accepted | 2021-09-17 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 分子醫學研究所 | zh_TW |
| dc.date.embargo-lift | 2022-12-31 | - |
| 顯示於系所單位： | 分子醫學研究所 | |
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