Research in my laboratory aims at: 1) understanding the cellular and molecular mechanisms underlying mammalian oocyte meiosis, activation, and aging; 2) discovering novel genes, factors, and signaling pathways that are functionally required during mammalian preimplantation embryo development; 3) creating animal models of human genetic diseases using knock-out, knock-in, and transgenesis strategies.
Mammalian oocyte meiosis, activation, and aging
Mammalian oocytes enter meiosis during fetal development, and arrest at diplotene stage of meiotic prophase I - also called germinal vesicle (GV) stage, until puberty. During this prolonged arrest, oocytes grow, differentiate and stockpile maternal components. Following puberty, oocytes resume meiosis under the action of gonadotropin hormones. For most mammals, after going through germinal vesicle breakdown (GVBD), metaphase I (MI), as well as anaphase/telophase I, oocyte extrudes the first polar body and reaches metaphase II (MII). The matured MII oocyte is then ovulated and is ready for fertilization. In some cases, if fertilization or parthenogenetic activation cannot take place right away, oocyte will arrest at MII stage but postovulatory aging then occurs. However, in some species (rat, hamster, some mouse strains, and some patients), spontaneous exit from MII arrest - also named spontaneous activation, can occur quickly in vitro and/or in vivo. A tremendous investment and complicated network (cell-cell interactions, hormones, second messenger molecules, cell cycle regulators, organelle and cytoskeletal systems, transcription and translation factors, ion channels, etc.) has been generated by the body to ensure a high quality oocyte, however, this machinery is also being challenged by environmental exposures, diet, lifestyle, and age. Thus, only understanding the full developmental network, specifically under current challenging situations, can guide our management of human reproduction, including assisted reproductive technology (ART).
Mammalian preimplantation embryo development
Preimplantation development refers to the period from fertilization to implantation, during which the fertilized oocyte progresses through a number of cleavage divisions and major transcriptional and morphogenetic events that lead to the first cell-fate decision (inner cell mass [ICM] and trophectoderm [TE]) and development into a blastocyst capable of implantation. During this brief but dynamic time window, several events occur, which include: 1) maternal-to-zygotic transition (MZT), namely, degradation of maternal mRNAs and replacement with zygotic transcripts; 2) embryo compaction and polarization, which initiate during the 8-cell stage in mouse embryos; 3) blastomere allocation and ICM/TE separation, which is the very first cell fate determination of our life. Previous microarray and current RNA-seq both confirmed more than 10,000 genes expressed during this period; however, for most of these genes, we do not know their function. More importantly, this time window is right clinically relevant: after in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI), human preimplantation embryos need to be cultured in a dish and incubator for several days, then transferred into mother’s uterus. Thus, understanding the role of each expressed gene and what happens during the culture is an essential next step for elucidating developmental networks at play and improving human assisted reproduction.
Animal model generation
There are multiple strategies to create genetically modified animal models. The earliest one is microinjection of foreign DNA into pronucleus to make transgenic animals. This method is straightforward (direct injection into 1-cell embryo); however, integration locus and copy numbers are random. When gene targeting technology is available, we can accurately modify specific locus of interest; but, due to the very low rate of homologous recombination, we have to rely on an excess of somatic cells or stem cells cultured in vitro, then, positive cells are selected to perform somatic cell nuclear transfer (SCNT) to get cloned animals, or through stem cell blastocoel injection to make chimera. In other words, these methods are effective but indirect and not straightforward, thus time consuming. When engineered nucleases (ZFN, TALEN, CRISPR/Cas9) are utilized, things become easier. These nucleases can accurately find precise locus of interest with high efficiency, so we can do direct microinjection into 1-cell embryos for directed gene editing. Compared with ZFN and TALEN, CRISPR/Cas9 system is easier to design, more flexible and cost effective. Currently, we are taking advantage of CRISPR/Cas9 system to perform gene editing. In addition to knock-out, knock-in and transgenic models, my laboratory also works on generation of patient-derived xenograft (PDX) cancer models.
Berberine alleviates LPS-induced apoptosis, oxidation, and skewed lineages during mouse preimplantation development. Biology of Reproduction, 106, 699–709. doi:10.1093/biolre/ioac002. (2022).
Surface functionalization of poly(dimethylsiloxane) substrates facilitates culture of pre-implantation mouse embryos by blocking non-selective adsorption. J R Soc Interface, 19(189), 20210929. presented at the 2022 04. doi:10.1098/rsif.2021.0929. (2022).
Early embryonic lethality of mice lacking POLD2. Molecular Reproduction and Development, 90, 98–108. doi:10.1002/mrd.23663. (2022).
Oocyte Spontaneous Activation: An Overlooked Cellular Event That Impairs Female Fertility in Mammals. Front Cell Dev Biol, 9, 648057. presented at the 2021. doi:10.3389/fcell.2021.648057. (2021).
Biophysical optimization of preimplantation embryo culture: what mechanics can offer ART. Mol Hum Reprod, 27(1). presented at the 2021 01 22. doi:10.1093/molehr/gaaa087. (2021).
Nanotherapeutics using all-natural materials. Effective treatment of wound biofilm infections using crosslinked nanoemulsions. Mater Horiz, 8(6), 1776-1782. presented at the 2021 06 01. doi:10.1039/d0mh01826k. (2021).
ZC3H4-a novel CCCH-type zinc finger protein-is essential for early embryogenesis in mice†.. Biol Reprod, 104(2), 325-335. presented at the 2021 02 11. doi:10.1093/biolre/ioaa215. (2021).
SOHLHs are essential for fertility regardless of gender or population. Fertil Steril, 114(2), 283-284. presented at the 2020 08. doi:10.1016/j.fertnstert.2020.06.010. (2020).
Loss of POLR1D results in embryonic lethality prior to blastocyst formation in mice. Molecular Reproduction and Development, 87, 1152–1158. doi:10.1002/mrd.23427. (2020).
Loss of RBBP4 results in defective inner cell mass, severe apoptosis, hyperacetylated histones and preimplantation lethality in mice†.. Biol Reprod, 103(1), 13-23. presented at the 2020 06 23. doi:10.1093/biolre/ioaa046. (2020).
MCRS1 is essential for epiblast development during early mouse embryogenesis. Reproduction, 159(1), 1-13. presented at the 2020 01. doi:10.1530/REP-19-0334. (2020).
Med20 is essential for early embryogenesis and regulates Nanog expression. Reproduction. presented at the dec. Retrieved from https://doi.org/10.1530/rep-18-0508. (2018).
MC1568 Enhances Histone Acetylation During Oocyte Meiosis and Improves Development of Somatic Cell Nuclear Transfer Embryos in Pig. Cell Reprogram, 20(1), 55-65. presented at the 2018 02. doi:10.1089/cell.2017.0023. (2018).
Tauroursodeoxycholic acid (TUDCA) alleviates endoplasmic reticulum stress of nuclear donor cells under serum starvation. PLoS One, 13(5), e0196785. presented at the 2018. doi:10.1371/journal.pone.0196785. (2018).
Effects of embryo-derived exosomes on the development of bovine cloned embryos. ( )PLoS ONE, 12, e0174535. presented at the mar. Retrieved from https://doi.org/10.1371/journal.pone.0174535. (2017).
Towards Functional Annotation of the Preimplantation Transcriptome: An RNAi Screen in Mammalian Embryos. Scientific Reports, 6. presented at the nov. Retrieved from https://doi.org/10.1038/srep37396. (2016).
Rethinking progesterone regulation of female reproductive cyclicity. Proceedings of the National Academy of Sciences, 113, 4212–4217. presented at the mar. Retrieved from https://doi.org/10.1073/pnas.1601825113. (2016).
HIF-KDM3A-MMP12 regulatory circuit ensures trophoblast plasticity and placental adaptations to hypoxia. Proceedings of the National Academy of Sciences, 113, E7212–E7221. presented at the nov. Retrieved from https://doi.org/10.1073/pnas.1612626113. (2016).
Cellular Prion Protein Mediates Pancreatic Cancer Cell Survival and Invasion through Association with and Enhanced Signaling of Notch1. The American Journal of Pathology, 186, 2945–2956. presented at the nov. Retrieved from https://doi.org/10.1016/j.ajpath.2016.07.010. (2016).
Optimized Protocols for In Vitro Maturation of Rat Oocytes Dramatically Improve Their Developmental Competence to a Level Similar to That of Ovulated Oocytes. Cellular Reprogramming. presented at the dec. Retrieved from https://doi.org/10.1089/cell.2015.0055. (2015).
Role of Na+/Ca2+ Exchanger (NCX) in Modulating Postovulatory Aging of Mouse and Rat Oocytes. ( )PLoS ONE, 9, e93446. presented at the apr. Retrieved from https://doi.org/10.1371/journal.pone.0093446. (2014).
Control of Spontaneous Activation of Rat Oocytes by Regulating Plasma Membrane Na+/Ca2+ Exchanger Activities. Biology of Reproduction, 88, 160–160. presented at the may/2013. Retrieved from https://doi.org/10.1095/biolreprod.113.108266. (2013).
Mechanisms by which a Lack of Germinal Vesicle (GV) Material Causes Oocyte Meiotic Defects: A Study Using Oocytes Manipulated to Replace GV with Primary Spermatocyte Nuclei. Biology of Reproduction, 89. presented at the oct/2013. Retrieved from https://doi.org/10.1095/biolreprod.113.111500. (2013).
Roles of MAPK and Spindle Assembly Checkpoint in Spontaneous Activation and MIII Arrest of Rat Oocytes. ( )PLoS ONE, 7, e32044. presented at the feb. Retrieved from https://doi.org/10.1371/journal.pone.0032044. (2012).