Maternal Cytokines CXCL12, VEGFA, and WNT5A Promote Porcine Oocyte Maturation via MAPK Activation and Canonical WNT Inhibition.

Xin Liu,Yuchen Hao,Guowei Bu,Xia Zhang,Zhiting Liu,Jilong Zhou,Xudong Hou,Yi-Liang Miao,Zhekun Li,Hongmei Zhu

doi:10.3389/fcell.2020.00578

Xin Liu, Yuchen Hao + Show 8 more

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https://doi.org/10.3389/fcell.2020.00578

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Abstract

Maternal regulatory factors endow the oocyte with developmental competence in vivo, which might be absent in current in vitro maturation (IVM) systems, thereby compromising oocyte quality. In the present study, by employing RNA sequencing data analysis, we expect to identify potential contributing factors to support porcine oocyte maturation through binding to their receptors on the oolemma. Here, C-X-C motif chemokine ligand 12 (CXCL12), vascular endothelial growth factor A (VEGFA), and Wingless-type MMTV integration site family member 5A (WNT5A), termed CVW, are selected and confirmed to be important maternal cytokines for porcine oocyte maturation. Combined supplementation of CVW promotes the nuclear maturation percentage from 57.2% in controls to 75.9%. More importantly, these maternal cytokines improve the developmental potential of matured oocytes by parthenogenesis, fertilization, and cloning, as their blastocyst formation efficiencies and total cell numbers are increased. CVW supplementation also enlarges perivitelline space and promotes cumulus expansion, which results in a more complete transzonal projection retraction on the zona pellucida, and a reduced incidence of polyspermy in fertilized oocytes. Meanwhile, inhibiting the CVW receptor-mediated signaling pathways severely impairs oocyte meiotic resumption and cumulus expansion during IVM. We further determine that maturation improvement by CVW is achieved through activating the MAPK pathway in advance and inhibiting the canonical WNT pathway at the end of the IVM period. These findings provide a new combination of three cytokines to promote the porcine IVM process, which also holds potential to be used in human assisted reproduction technologies as well as in other species.

Highlights

In vitro maturation (IVM) is an important reproductive technique to obtain viable oocytes for pig embryo and piglet production, which has been widely used to create genetically modified pigs for breed improvement or human disease research
We speculated that at least CXCL12-CXCR4, vascular endothelial growth factor A (VEGFA)/VEGFB-fms-like tyrosine kinase 1 (FLT1), and WNT5AFZD5 interactions may be essential for porcine oocyte and embryo development
We noticed that the CXCR4, FLT1, frizzled class receptor 5 (FZD5) transcripts peaked in morulae, matured oocytes, and zygotes, respectively (Figure 1B)

Summary

Introduction

In vitro maturation (IVM) is an important reproductive technique to obtain viable oocytes for pig embryo and piglet production, which has been widely used to create genetically modified pigs for breed improvement or human disease research. By inserting the mouse UCP1 gene, pigs have been created with reduced fat deposition and improved thermogenic capacity (Zheng et al, 2017) Another pig model with mutant huntingtin expression successfully exhibits features of Huntington’s disease for further clinical study (Yan et al, 2018). Pigs are considered to be the best candidate for human organ generation via xenotransplantation, and their endogenous retrovirus has been genetically inactivated to eliminate safety concerns for future applications (Niu et al, 2017). All these studies require a large number of porcine embryos derived by in vitro fertilization (IVF) or somatic cell nuclear transfer (SCNT) techniques. For reasons not fully clear, oocytes obtained after IVM are less competent than their in vivo-produced counterparts, which is believed to be responsible for the poor developmental phenotype of IVF or SCNT embryos, thereby limiting the practical use of these techniques in agriculture and biomedicine

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