Unravelling the mechanisms of chromosomal instability during early embryogenesis
Kuupäev
2025-02-18
Autorid
Ajakirja pealkiri
Ajakirja ISSN
Köite pealkiri
Kirjastaja
Abstrakt
Kromosomaalne ebastabiilsus (ingl. Chromosomal INstability, CIN) on imetajate, näiteks inimeste ja veiste implantatsioonieelsete embrüote küllaltki sage tunnus. Enamasti eemaldatakse kromosoomianomaaliatega embrüod juba varase arengu käigus iseeneslike abortide kaudu, kuid elussünnini jõudmise korral võib kromosomaalne ebastabiilsus viia ka kaasasündinud väärarendite tekkele. Esimeses projektis uurisime, kas inimese implantatsioonieelsetes embrüodes täheldatud CIN esineb ka hobuste implantatsioonieelsetes embrüodes. Selleks kohandasime haplaritmisis (ingl. haplarithmisis) algoritmi nii, et see ühilduks hobuste analüüsiga ja avastasime, et sagedased kromosomaalsed kõrvalekalded esinevad ka hobuste implantatsioonieelsetes embrüodes. Hobuste lõigustuvates embrüodes avastatud kromosomaalsed vead olid sarnased nii inimeste kui veiste leidudele võrrelduna kromosomaalselt normaalsete blastotsüstidega ehk lootepõiekestega. Antud meetodit saaks edukalt kasutada tõuaretuses - huvipakkuvate geneetiliste variantidega kuid ilma geneetiliste vigade ja patogeensete variantideta hobuste embrüote valimiseks. Lisaks aneuploidiatele tuvastati implantatsioonieelsetes embrüodes ka kogu genoomi aberratsioone. Näiteks triploidiaid (kolmekordne kromosoomide arv), ühe vanema täiendava haploidse kromosoomi-komplekti olemasolul. Samuti avastasime samas embrüos koos eksisteerivaid androgeneetilisi, günogeneetilisi ja polüploidseid blastomeere ehk lõigustusrakke. Teises projektis uurisime kimäärsete ja miksoploidsete embrüote päritolu ja arengupotentsiaali. Avastasime, et vanemate genoomid eralduvad viljastatud munaraku ehk sügoodi esimese jagunemise ajal eraldiseisvateks blastomeerideks, mille tulemuseks on ebanormaalse genoomiga rakkude samaaegne esinemine koos normaalsete diploidsete (kahekordne kromosoomide arv) rakkude või muude ebanormaalsete rakkudega. Selle protsessi kirjeldamiseks lõime termini "heterogoonne jagunemine". Geeniekspressiooni muutvad stressireaktsioonid soodustavad ülegenoomsete muutuste tagajärjel tekkinud arenguhäireid ebanormaalsetes rakkudes ja nende erinev arengusaatus võib seletada androgenootide, günogenootide, triploidsuse, kimäärsuse ja miksoploidsuse teket, mida täheldati hilisema arengu käigus. Seetõttu soovitame haplotüüpidel põhinevat preimplantatsioonilist geneetilist testimist (ingl. Preimplantation Genetic Testing, PGT) ülegenoomsete kõrvalekallete tuvastamiseks, et suurendada munaraku kehavälise viljastamise (IVF) programmide edu. Kolmandas projektis uurisime kolmanda põlvkonna sekveneerimistehnoloogiate kiire arenguga seotud pikkadel lugemitel põhineva kogu genoomi järjestamise (lrWGS) PGT tehnoloogiat. Võrdlusuuring, milles kasutati nn. Genome in a Bottle (GIAB) Aškenazi triot, näitas üksikute rakkude lrWGS-i andmete sobivust geenivariantide eristamiseks ja faasimiseks. lrWGS-põhise PGT meetodi kasutamine inimese embrüotel näitas 100% vastavust DNA mikrokiibi põhise PGT-ga ühe nukleotiidi variatsioonide (SNV), indelite ja aneuploidiate tuvastamiseks, tõestades et lrWGS-põhine PGT on sobiv alternatiiv praegustele meetoditele.
Antud doktoritöös käsitletud neli projekti tutvustasid uuenduslikke lähenemisviise embrüote uurimiseks ja valikuks, aidates oluliselt kaasa meie arusaamisele kromosomaalsete kõrvalekallete päritolu ja mõjude kohta imetajate implantatsioonieelsete embrüote arengus
Chromosomal instability (CIN) is a hallmark of preimplantation embryos for mammals such as human and bovine. These chromosomal abnormalities are known to be selected against during development and cause embryo loss, spontaneous abortion, or lead to congenital abnormalities if compatible with live birth. In the first project, we explored whether CIN observed in human preimplantation embryos also occurs in equine preimplantation embryos. We adapted the haplarithmisis algorithm to be compatible for equine analysis and uncovered frequent chromosomal abnormalities in equine preimplantation embryos, with higher chromosomal errors in arrested cleavage stage embryos compared to blastocysts, similar to findings in human and bovine. The method can be used to select equine embryos devoid of genetic errors and pathogenic variants, while carrying variants of interest. In addition to aneuploidies, whole-genome (WG) aberrations are also detected in preimplantation embryos. For example, triploidy involves the presence of an additional haploid set of chromosomes from one parent. Notably, we recently observed androgenetic, gynogenetic, and polyploid blastomeres coexisting within the same embryo. In the second project, we explored the origin and developmental potential of these chimeric and mixoploid embryos. We confirmed that parental genomes segregate into distinct blastomeres during the first zygotic division, resulting in the co-occurrence of WG abnormal cells with normal diploid cells or other WG abnormal cells, a process we previously discovered and coined “heterogoneic division”. Stress responses in gene expression contribute to developmental impairment in WG abnormal cells, and their differing developmental fates can explain the formation of androgenotes, gynogenotes, triploidy, chimerism and mixoploidy observed later during development. We therefore recommend haplotype-based preimplantation genetic testing (PGT) for WG abnormalities to enhance baby-take home rates during IVF programs. With the rapid development of long-read sequencing technology, we explored the performance of long-read whole-genome sequencing (lrWGS)-based comprehensive PGT in the third project. A benchmark study using the Genome in a Bottle (GIAB) Ashkenazi trio demonstrated the high performance of lrWGS data from single cells for variant calling and phasing. Testing lrWGS-based PGT on human embryos showed 100% consistency with array-based comprehensive PGT for single nucleotide variations (SNVs), indels, and aneuploidies, highlighting lrWGS-based PGT as a promising alternative to current methods. The three projects within this PhD program introduced innovative approaches for embryo study and selection, significantly advancing our understanding of the origins and developmental impacts of chromosomal abnormalities in preimplantation mammalian embryos.
Chromosomal instability (CIN) is a hallmark of preimplantation embryos for mammals such as human and bovine. These chromosomal abnormalities are known to be selected against during development and cause embryo loss, spontaneous abortion, or lead to congenital abnormalities if compatible with live birth. In the first project, we explored whether CIN observed in human preimplantation embryos also occurs in equine preimplantation embryos. We adapted the haplarithmisis algorithm to be compatible for equine analysis and uncovered frequent chromosomal abnormalities in equine preimplantation embryos, with higher chromosomal errors in arrested cleavage stage embryos compared to blastocysts, similar to findings in human and bovine. The method can be used to select equine embryos devoid of genetic errors and pathogenic variants, while carrying variants of interest. In addition to aneuploidies, whole-genome (WG) aberrations are also detected in preimplantation embryos. For example, triploidy involves the presence of an additional haploid set of chromosomes from one parent. Notably, we recently observed androgenetic, gynogenetic, and polyploid blastomeres coexisting within the same embryo. In the second project, we explored the origin and developmental potential of these chimeric and mixoploid embryos. We confirmed that parental genomes segregate into distinct blastomeres during the first zygotic division, resulting in the co-occurrence of WG abnormal cells with normal diploid cells or other WG abnormal cells, a process we previously discovered and coined “heterogoneic division”. Stress responses in gene expression contribute to developmental impairment in WG abnormal cells, and their differing developmental fates can explain the formation of androgenotes, gynogenotes, triploidy, chimerism and mixoploidy observed later during development. We therefore recommend haplotype-based preimplantation genetic testing (PGT) for WG abnormalities to enhance baby-take home rates during IVF programs. With the rapid development of long-read sequencing technology, we explored the performance of long-read whole-genome sequencing (lrWGS)-based comprehensive PGT in the third project. A benchmark study using the Genome in a Bottle (GIAB) Ashkenazi trio demonstrated the high performance of lrWGS data from single cells for variant calling and phasing. Testing lrWGS-based PGT on human embryos showed 100% consistency with array-based comprehensive PGT for single nucleotide variations (SNVs), indels, and aneuploidies, highlighting lrWGS-based PGT as a promising alternative to current methods. The three projects within this PhD program introduced innovative approaches for embryo study and selection, significantly advancing our understanding of the origins and developmental impacts of chromosomal abnormalities in preimplantation mammalian embryos.
Kirjeldus
Doktoritöö kaitstakse Leuveni Katoliiklikus Ülikoolis Belgias
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doktoritööd