Somatic cell nuclear transfer (SCNT) is a method with unique ability to reprogram the epigenome of a fully differentiated cell. However, its efficiency remains extremely low. In this work, we assessed and combined two simple strategies to improve the SCNT efficiency in the bovine. These are the use of less-differentiated donor cells to facilitate nuclear reprogramming and the embryo aggregation (EA) strategy that is thought to compensate for aberrant epigenome reprogramming. We carefully assessed the optimal time of EA by using in vitro-fertilized (IVF) embryos and evaluated whether the use of adipose-derived mesenchymal stem cells (ASCs) as donor for SCNT together with EA improves the blastocyst rates and quality. Based on our results, we determined that the EA improves the preimplantation embryo development per well of IVF and SCNT embryos. We also demonstrated that day 0 (D0) is the optimal aggregation time that leads to a single blastocyst with uniform distribution of the original blastomeres. This was confirmed in bovine IVF embryos and then, the optimal condition was translated to SCNT embryos. Notably, the relative expression of the trophectoderm (TE) marker KRT18 was significantly different between aggregated and nonaggregated ASC-derived embryos. In the bovine, no effect of the donor cell is observed on the developmental rate, or the embryo quality. Therefore, no synergistic effect of the use of both strategies is observed. Our results suggest that EA at D0 is a simple and accessible strategy that improves the blastocyst rate per well in bovine SCNT and IVF embryos and influence the expression of a TE-related marker. The aggregation of two ASC-derived embryos seems to positively affect the embryo quality, which may improve the postimplantation development.
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References
1.
AignerB., RennerS., KesslerB., KlymiukN., KuromeM., WünschA., and WolfE. (2010). Transgenic pigs as models for translational biomedical research. J. Mol. Med. (Berl.), 88, 653–664.
2.
AkagiS., YamaguchiD., MatsukawaK., MizutaniE., HosoeM., AdachiN., KuboM., and TakahashiS. (2011). Developmental ability of somatic cell nuclear transferred embryos aggregated at the 8-cell stage or 16- to 32-cell stage in cattle. J. Reprod. Dev. 57, 500–506.
3.
AvilionA.A., NicolisS.K., PevnyL.H., PerezL., VivianN., and Lovell-BadgeR. (2003). Multipotent cell lineages in early mouse development depend on SOX2 function. Genes Dev. 17, 126–140.
BangJ.I., JinJ.I., GhanemN., ChoiB.H., FakruzzamanM., HaA.N., LeeK.L., UhmS.J., KoD.H., KooB.C., LeeJ.G., and KongI.K. (2015). Quality improvement of transgenic cloned bovine embryos using an aggregation method: Effects on cell number, cell ratio, embryo perimeter, mitochondrial distribution, and gene expression profile. Theriogenology, 84, 509–523.
6.
BasiniG., SimonaB., SantiniS.E., and GrasselliF. (2008). Reactive oxygen species and anti-oxidant defences in swine follicular fluids. Reprod. Fertil. Dev. 20, 269–274.
7.
BoianiM., EckardtS., LeuN.A., SchölerH.R., and McLaughlinK.J. (2003). Pluripotency deficit in clones overcome by clone–clone aggregation: Epigenetic complementation?. EMBO J. 22, 5304–5312.
8.
BrackettB.G., and OliphantG. (1975). Capacitation of rabbit spermatozoa in vitro. Biol. Reprod. 12, 260–274.
9.
BuemoC.P., GambiniA., MoroL.N., HiriartM.I., Fernández-MartínR., CollasP., and SalamoneD.F. (2016). Embryo aggregation in pig improves cloning efficiency and embryo quality. PLoS One 11,e0146390.
10.
BuiL.C., EvsikovA.V., KhanD.R., ArchillaC., PeynotN., HenautA., Le BourhisD., VignonX., RenardJ.P., and DuranthonV. (2009). Retrotransposon expression as a defining event of genome reprograming in fertilized and cloned bovine embryos. Reproduction 138,289.
11.
Chavatte-PalmerP., VelazquezM.A., JammesH., and DuranthonV. (2018). Review: Epigenetics, developmental programming and nutrition in herbivores. Animal, 12, s363–s371.
12.
ColleoniS., DonofrioG., LagutinaI., DuchiR., GalliC., and LazzariG. (2005). Establishment, differentiation, electroporation, viral transduction, and nuclear transfer of bovine and porcine mesenchymal stem cells. Cloning Stem Cells, 7, 154–166.
13.
CooperD.K., SatyanandaV., EkserB., van der WindtD.J., HaraH., EzzelarabM.B., and SchuurmanH.J. (2014). Progress in pig-to-non-human primate transplantation models (1998–2013): A comprehensive review of the literature. Xenotransplantation, 21, 397–419.
14.
CzernikM., AnzaloneD.A., PalazzeseL., OikawaM., and LoiP. (2019). Somatic cell nuclear transfer: Failures, successes and the challenges ahead. Int. J. Dev. Biol. 63, 123–130.
15.
da SilvaC.G., MartinsC.F., CardosoT.C., Da CunhaE.R., BesslerH.C., MartinsG.H.L., PivatoI., and BáoS.N. (2016). Production of bovine embryos and calves cloned by nuclear transfer using mesenchymal stem cells from amniotic fluid and adipose tissue. Cell. Reprogram. (Formerly “Cloning Stem Cells”), 18, 127–136.
16.
DegrelleS.A., JaffrezicF., CampionE., Lê CaoK.A., Le BourhisD., RichardC., RoddeN., FleurotR., EvertsR.E., LecardonnelJ., HeymanY., VignonX., YangX., TianX.C., LewinH.A., RenardJ.-P., and HueI. (2012). Uncoupled embryonic and extra-embryonic tissues compromise blastocyst development after somatic cell nuclear transfer. PLoS One 7,e38309.
17.
EckardtS., and McLaughlinK.J. (2004). Interpretation of reprogramming to predict the success of somatic cell cloning. Anim. Reprod. Sci. 82, 97–108.
18.
FaastR., HarrisonS.J., BeebeL.F., McilfatrickS.M., AshmanR.J., and NottleM.B. (2006). Use of adult mesenchymal stem cells isolated from bone marrow and blood for somatic cell nuclear transfer in pigs. Cloning Stem Cells, 8, 166–173.
19.
FatehiA.N., RoelenB.A., ColenbranderB., SchoeversE.J., GadellaB.M., BeversM.M., and Van Den HurkR. (2005). Presence of cumulus cells during in vitro fertilization protects the bovine oocyte against oxidative stress and improves first cleavage but does not affect further development. Zygote 13,177.
20.
GambiniA., De StefanoA., BevacquaR.J., KarlanianF., and SalamoneD.F. (2014). The aggregation of four reconstructed zygotes is the limit to improve the developmental competence of cloned equine embryos. PLoS One 9,e110998.
21.
GambiniA., JarazoJ., OliveraR., and SalamoneD.F. (2012). Equine cloning: in vitro and in vivo development of aggregated embryos. Biol. Reprod. 87,15.
22.
GasparR.C., ArnoldD.R., CorrêaC.A., da RochaC.V.Jr., PenteadoJ.C., Del ColladoM., Del ColladoM., VantiniR., GarciaJ.M., and LopesF.L. (2015). Oxygen tension affects histone remodeling of in vitro–produced embryos in a bovine model. Theriogenology, 83, 1408–1415.
23.
GerriC., McCarthyA., Alanis-LobatoG., DemtschenkoA., BruneauA., LoubersacS., FogartyN.M.E., HampshireD., ElderK., SnellP., ChristieL., DavidL., Van de VeldeH., Fouladi-NashtaA.A., and NiakanK.K. (2020). Initiation of a conserved trophectoderm program in human, cow and mouse embryos. Nature, 587, 443–447.
24.
Gonzalez-MunozE., and CibelliJ.B. (2018). Somatic cell reprogramming informed by the oocyte. Stem Cells Dev. 27, 871–887.
25.
GopichandranN., and LeeseH.J. (2006). The effect of paracrine/autocrine interactions on the in vitro culture of bovine preimplantation embryos. Reproduction, 131, 269–277.
26.
GouveiaC., HuyserC., EgliD., and PepperM.S. (2020). Lessons learned from somatic cell nuclear transfer. Int. J. Mol. Sci. 21,2314.
27.
HallV., HinrichsK., LazzariG., BettsD.H., and HyttelP. (2013). Early embryonic development assisted reproductive technologies, and pluripotent stem cell biology in domestic mammals. Vet. J. 197, 128–142.
28.
KnottJ.G., and PaulS. (2014). Transcriptional regulators of the trophoblast lineage in mammals with hemochorial placentation. Reproduction 148,R121.
29.
LiZ., HeX., ChenL., ShiJ., ZhouR., XuW., LiuD., and WuZ. (2013). Bone marrow mesenchymal stem cells are an attractive donor cell type for production of cloned pigs as well as genetically modified cloned pigs by somatic cell nuclear transfer. Cell. Reprogram. (Formerly “Cloning Stem Cells”), 15, 459–470.
30.
LimH., AlvarezY.D., GasnierM., WangY., TetlakP., BissiereS., WangH., BiroM., and PlachtaN. (2020). Keratins are asymmetrically inherited fate determinants in the mammalian embryo. Nature, 585, 404–409.
31.
LiuW, LiuX., WangC., GaoY., GaoR., KouX., ZhaoY., LiJ., WuY., XiuW., WangS., YinJ., LiuW., CaiT., WangH., ZhangY., and GaoS. (2016a). Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing. Cell Discov. 2,16010.
32.
LiuX., LiuH., WangM., LiR., ZengJ., MoD., CongP., LiuX., Chen,Y., and HeZ. (2019). Disruption of the ZBED6 binding site in intron 3 of IGF2 by CRISPR/Cas9 leads to enhanced muscle development in Liang Guang Small Spotted pigs. Transgenic Res. 28, 141–150.
33.
LiuX., WangY., GaoY., SuJ., ZhangJ., XingX., ZhouC., YaoK., AnQ., and ZhangY. (2018a). H3K9 demethylase KDM4E is an epigenetic regulator for bovine embryonic development and a defective factor for nuclear reprogramming. Development 145,dev158261.
34.
LiuZ., CaiY., WangY., NieY., ZhangC., XuY., ZhangX., LuY., WangZ., PooM., and SunQ. (2018b). Cloning of macaque monkeys by somatic cell nuclear transfer. Cell, 172, 881–887.
35.
LocatelliP., OleaF.D., HnatiukA., SepúlvedaD., Pérez SáezJ.M., ArgüelloR., and CrottoginiA. (2013). Efficient plasmid-mediated gene transfection of ovine bone marrow mesenchymal stromal cells. Cytotherapy, 15, 163–170.
36.
LongC.R., WesthusinM.E., and GoldingM.C. (2014). Reshaping the transcriptional frontier: Epigenetics and somatic cell nuclear transfer. Mol. Reprod. Dev. 81, 183–193.
37.
MadejaZ.E., SosnowskiJ., HryniewiczK., WarzychE., PawlakP., RozwadowskaN., PlusaB., and LechniakD. (2013). Changes in sub-cellular localisation of trophoblast and inner cell mass specific transcription factors during bovine preimplantation development. BMC Dev. Biol. 13, 1–17.
38.
MatobaS., InoueK., KohdaT., SugimotoM., MizutaniE., OgonukiN., NakamuraT., AbeK., NakanoT., IshinoF., and OguraA. (2011). RNAi-mediated knockdown of Xist can rescue the impaired postimplantation development of cloned mouse embryos. Proc. Natl. Acad. Sci. U. S. A. 108, 20621–20626.
39.
MatobaS., LiuY., LuF., IwabuchiK.A., ShenL., InoueA., and ZhangY. (2014). Embryonic development following somatic cell nuclear transfer impeded by persisting histone methylation. Cell, 159, 884–895.
40.
MatobaS., and ZhangY. (2018). Somatic cell nuclear transfer reprogramming: Mechanisms and applications. Cell Stem Cell, 23, 471–485.
41.
Misica-TurnerP.M., ObackF.C., EichenlaubM., WellsD.N., and ObackB. (2007). Aggregating embryonic but not somatic nuclear transfer embryos increases cloning efficiency in cattle. Biol. Reprod. 76, 268–278.
42.
MoroL.N., HiriartM.I., BuemoC., JarazoJ., SesteloA., VeraguasD., Rodriguez-AlvarezL., and SalamoneD.F. (2015). Cheetah interspecific SCNT followed by embryo aggregation improves in vitro development but not pluripotent gene expression. Reproduction, 150, 1–10.
43.
NicholsJ., ZevnikB., AnastassiadisK., NiwaH., Klewe-NebeniusD., ChambersI., SchölerH., and SmithA. (1998). Formation of pluripotent stem cells in the mammalian embryo depends on the POU transcription factor Oct4. Cell, 95, 379–391.
44.
NiemannH., KuesW.A., Lucas-HahnA., and CarnwathJ.W. (2011). Somatic cloning and epigenetic reprogramming in mammals. In Principles of Regenerative Medicine. A. Atala, R. Lanza, J.A. Thomson, and R. Nerem, eds. (Elsevier) pp. 129–158. ISBN: 978-0123814227.
45.
OguraA., InoueK., and WakayamaT. (2013). Recent advancements in cloning by somatic cell nuclear transfer. Philos. Trans. R. Soc. Lond. B Biol. Sci. 368,20110329.
46.
OhH.J., ParkJ.E., KimM.J., HongS.G., RaJ.C., JoJ.Y., KangS.K., JangG., and LeeB.C. (2011). Recloned dogs derived from adipose stem cells of a transgenic cloned beagle. Theriogenology, 75, 1221–1231.
47.
OliveraR., MoroL.N., JordanR., LuzzaniC., MiriukaS., RadrizzaniM., DonadeuF.X., and VicheraG. (2016). In vitro and in vivo development of horse cloned embryos generated with iPSCs, mesenchymal stromal cells and fetal or adult fibroblasts as nuclear donors. PLoS One 11,e0164049.
48.
OliveraR., MoroL.N., JordanR., PallarolsN., GuglielminettiA., LuzzaniC., MiriukaS.G., and VicheraG. (2018). Bone marrow mesenchymal stem cells as nuclear donors improve viability and health of cloned horses. Stem Cells Cloning 11,13.
49.
OnoY., and KonoT. (2006). Irreversible barrier to the reprogramming of donor cells in cloning with mouse embryos and embryonic stem cells. Biol. Reprod. 75, 210–216.
50.
PicouA. (2009). The isolation and characterization of bovine adult derived adipose stem cells for the use in nuclear transfer. LSU Master's Theses 3541. https://digitalcommons.lsu.edu/gradschool_theses/3541. Last viewed on September 17,2021.
51.
ReikW., DeanW., and WalterJ. (2001). Epigenetic reprogramming in mammalian development. Science, 293, 1089–1093.
52.
RibeiroE.S., GergerR.P., OhlweilerL.U., OrtigariI.Jr., MezzaliraJ.C., ForellF., BertoliniL.R., RodriguesJ.L., AmbrósioC.E., MiglinoM.A., MezzaliraA., and BertoliniM. (2009). Developmental potential of bovine hand-made clone embryos reconstructed by aggregation or fusion with distinct cytoplasmic volumes. Cloning Stem Cells, 11, 377–386.
53.
Rodríguez-AlvarezL., CoxJ., TovarH., EinspanierR., and CastroF.O. (2010). Changes in the expression of pluripotency-associated genes during preimplantation and peri-implantation stages in bovine cloned and in vitro produced embryos. Zygote, 18, 269–279.
54.
RossP.J., WangK., KocabasA., and CibelliJ.B. (2010). Housekeeping gene transcript abundance in bovine fertilized and cloned embryos. Cell. Reprogram. 12, 709–717.
55.
SalamoneD., BarañaoL., SantosC., BussmannL., ArtusoJ., WerningC., PryncA., CarbonettoC., DabsysS., MunarC., SalaberryR., BerraG., BerraI., FernándezN., PapouchadoM., FotiM., JudewiczN., MujicaI., MuñozL., AlvarezS.F., GonzálezE., ZimmermannJ., CriscuoloM., and MeloC. (2006). High level expression of bioactive recombinant human growth hormone in the milk of a cloned transgenic cow. J. Biotechnol. 124, 469–472.
56.
SozenB., JorgensenV., WeatherbeeB.A.T., ChenS., ZhuM., and Zernicka-GoetzM. (2021). Reconstructing aspects of human embryogenesis with pluripotent stem cells. Nat. Commun. 12, 5550. DOI: 10.1038/s41467-021-25853-4. PMID: 34548496.
57.
SuX., WangS., SuG., ZhengZ., ZhangJ., MaY., LiuZ., ZhouH., ZhangY., and ZhangL. (2018b). Production of microhomologous-mediated site-specific integrated LacS gene cow using TALENs. Theriogenology, 119, 282–288.
58.
VajtaG. (2007). Somatic cell nuclear transfer in its first and second decades: Successes, setbacks, paradoxes and perspectives. Reprod. Biomed. Online, 15, 582–590.
59.
VajtaG., PeuraT.T., HolmP., PaldiA., GreveT., TrounsonA.O., and CallesenH. (2000). New method for culture of zona-included or zona-free embryos: The Well of the Well (WOW) system. Mol. Reprod. Dev. 55, 256–264.
60.
WellsK.D., and PowellA.M. (2000). Blastomeres from somatic cell nuclear transfer embryos are not allocated randomly in chimeric blastocysts. Cloning, 2, 9–22.
61.
WhiteM.D., and PlachtaN. (2020). Specification of the first mammalian cell lineages in vivo and in vitro. Cold Spring Harb. Perspect. Biol. 12,a035634.
62.
WilmutI., SchniekeA.E., McWhirJ., KindA.J., and CampbellK.H. (1997). Viable offspring derived from fetal and adult mammalian cells. Nature, 385, 810–813.
63.
WrenzyckiC., Lucas-HahnA., HerrmannD., LemmeE., KorsaweK., and NiemannH. (2002). In vitro production and nuclear transfer affect dosage compensation of the X-linked gene transcripts G6PD, PGK, and Xist in preimplantation bovine embryos. Biol. Reprod. 66, 127–134.
64.
YamanakaK.I., SakataniM., KubotaK., BalboulaA.Z., SawaiK., and TakahashiM. (2011). Effects of downregulating DNA methyltransferase 1 transcript by RNA interference on DNA methylation status of the satellite I region and in vitro development of bovine somatic cell nuclear transfer embryos. J. Reprod. Dev. 57, 393–402.
65.
ZhangS., ChenX., WangF., AnX., TangB., ZhangX., SunL., and LiZ. (2016). Aberrant DNA methylation reprogramming in bovine SCNT preimplantation embryos. Sci. Rep. 6, 1–11.
