Unravelling chromosomal instability in mammalian preimplantation embryos using single-cell genomics
Date
2018-10-10
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Abstract
Viljatus on kogu maailmas kiiresti kasvavaks probleemiks ning iga kuues paar seisab silmitsi olukorraga, et soovitud rasedust ei teki. Esmasünnitajate keskmine vanus kasvab samuti, kuid naise vananedes tema viljakus langeb ning seetõttu pöördub tänapäeval aina rohkem paare viljatusravi poole, et leida oma probleemile lahendus. Kehaväline viljastamine (in vitro fertilization, IVF), mille korral toimub munarakkude viljastamine ning embrüote areng katseklaasis, on maailmas kõige sagedamini kasutatav lastetusravi meetod. Paraku, IVF embrüote sagedased kromosomaalsed aberratsioonid on kõige suurem väljakutse IVF ravil, sest see on üks peamisi raseduse varajase katkemise (kuni 75%) ja IVF ebaõnnestumise põhjuseid. Samas, embrüotega seotud teadustöö on piiratud eetiliste põhjuste tõttu, seega sõltuvad inimese embrüote uuringud suurel määral sobivatest loommudelitest. Kasutades uudset üksikraku kogu-genoomi analüüsi metoodikat, näidati antud doktoritöös, et ka veise IVF embrüotes esinevad väga sagedased kromosomaalsed aberratsioonid, mille tulemusena on suurem osa varajasi embrüoid mosaiiksed. Lisaks avastati, et embrüo on võimeline segregeerima ema- ja isapoolse genoomi eraldi rakuliinidesse, mis võib-olla aluseks eriploidsuse (mixoploidy) ja kimääride tekkele, kuid selle nähtuse esinemissagedus on hetkel teadmata. Kromosoomi aberratsioonid eksisteerivad samuti loomulikul teel saadud embrüotes, kuid munarakkude ja embrüote in vitro manipuleerimine suurendab genoomi ebastabiilsuse sagedust veise varajastes embrüotes, vähendades nende elujõulisust. Kui sama trend kehtib ka inimese embrüote puhul, peaks see julgustama teadus- ja meditsiinikogukonda mitte ainult parandama in vitro kultuuri tingimusi, vaid ka tõstma inimeste teadlikkust viljakuse ja IVF ravi osas. Selleks, et tuvastada embrüo aneuploidiat kasutatakse tänapäeval embrüote sõeluuringut, analüüsides kas kolmanda päeva või viienda päeva biopsiat. Viimasel ajal on aga hakatud pöörama suuremat tähelepanu ka blastotsööli vedeliku (blastotsüsti sees olev vedelik) analüüsile, kuid antud töö käigus selgus, et blastotsööli vedeliku analüüsi tulemused on vastuolulised ning seda meetodit ei saa hetkel kasutada embrüote genoomi analüüsis.
Fertility issues have become a plague of modern society, as one in six couples will encounter problems to achieve clinical pregnancy. Nowadays, natural conception is often taken for granted, leading to an increased number of couples that turn to assisted reproduction to help conceive and ultimately give birth to a healthy baby. In vitro fertilization (IVF) and embryo transfer is the most commonly performed procedure in ART. However, the uprising era of single-cell research has highlighted the alarming fact that human IVF embryos have high prevalence of chromosomal instability (CIN), which represents one of the most serious challenges in IVF. At the same time, the use of in vitro fertilization (IVF) treatment provides a unique opportunity to study the fundamentals of preimplantation human development, from fertilization to blastocyst stage of development. At the same time, embryo-related research is ethically one of the most complex areas of reproductive science that greatly relies on the use of appropriate animal models. By using novel single-cell genomics approaches, the current thesis has demonstrated that the nature and frequency of genomic abnormalities in in vitro cleavage-stage embryos is highly conserved between cattle and human. Additionally, heterogoneic cell division was discovered, by which the zygote segregates entire parental genomes into separate blastomere lineages, giving rise to mixoploidy and chimaerism, although the frequency of this phenomenon and the fate of mixoploid embryos remain to be investigated. By using the established bovine model it was demonstrated that chromosomal abnormalities are also present in naturally conceived embryos, but in vitro procedures exacerbate CIN in early embryos, compromising their survival rate. If this is the same for human this should encourage scientific and medical communities not only to refine and improve in vitro culture conditions, but also raise awareness among men and women of reproductive age. To tackle the issues of embryonic aneuploidy, embryo chromosome screening is performed, by analysing either day 3 or day 5 biopsy. Recently, the use of blastocoel fluid (BF) biopsy was proposed, but the data presented here shows that BF-DNA is not suitable for diagnostic purposes, and day 5 biopsy remains the safest option with the most reliable results.
Fertility issues have become a plague of modern society, as one in six couples will encounter problems to achieve clinical pregnancy. Nowadays, natural conception is often taken for granted, leading to an increased number of couples that turn to assisted reproduction to help conceive and ultimately give birth to a healthy baby. In vitro fertilization (IVF) and embryo transfer is the most commonly performed procedure in ART. However, the uprising era of single-cell research has highlighted the alarming fact that human IVF embryos have high prevalence of chromosomal instability (CIN), which represents one of the most serious challenges in IVF. At the same time, the use of in vitro fertilization (IVF) treatment provides a unique opportunity to study the fundamentals of preimplantation human development, from fertilization to blastocyst stage of development. At the same time, embryo-related research is ethically one of the most complex areas of reproductive science that greatly relies on the use of appropriate animal models. By using novel single-cell genomics approaches, the current thesis has demonstrated that the nature and frequency of genomic abnormalities in in vitro cleavage-stage embryos is highly conserved between cattle and human. Additionally, heterogoneic cell division was discovered, by which the zygote segregates entire parental genomes into separate blastomere lineages, giving rise to mixoploidy and chimaerism, although the frequency of this phenomenon and the fate of mixoploid embryos remain to be investigated. By using the established bovine model it was demonstrated that chromosomal abnormalities are also present in naturally conceived embryos, but in vitro procedures exacerbate CIN in early embryos, compromising their survival rate. If this is the same for human this should encourage scientific and medical communities not only to refine and improve in vitro culture conditions, but also raise awareness among men and women of reproductive age. To tackle the issues of embryonic aneuploidy, embryo chromosome screening is performed, by analysing either day 3 or day 5 biopsy. Recently, the use of blastocoel fluid (BF) biopsy was proposed, but the data presented here shows that BF-DNA is not suitable for diagnostic purposes, and day 5 biopsy remains the safest option with the most reliable results.
Description
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Keywords
embryo cells, genomes, chromosome aberrations, in vitro