Expression and Impact of Vaspin on In Vitro Oocyte Maturation through MAP3/1 and PRKAA1 Signalling Pathways.

Patrycja Kurowska,Agnieszka Rak,Iwona Rajska,Ewa Mlyczyńska,Anthony Estienne,Katarzyna Poniedziałek-Kempny,Katarzyna Soból,Joelle Dupont,Alix Barbe

doi:10.3390/ijms21249342

Abstract

Oocyte maturation is a critical stage in embryo production and female reproduction. The aims of this study were to determine: (i) the mRNA and protein expression of vaspin and its receptor 78-kDa glucose-regulated (GRP78) in porcine cumulus–oocyte complexes (COCs) by real-time PCR and Western blot analysis, respectively, and their localisation by immunofluorescence; and (ii) the effects of vaspin on in vitro oocyte maturation (IVM) and the involvement of mitogen ERK1/2 (MAP3/1)- and AMPKα (PRKAA1)-activated kinases in the studied processes. Porcine COCs were matured in vitro for 22 h or 44 h with vaspin at a dose of 1 ng/mL and nuclear maturation assessed by Hoechst 33342 or DAPI staining and the measurement of progesterone (P4) level in the maturation medium. We showed that vaspin and GRP78 protein expression increased in oocytes and cumulus cells after IVM. Moreover, vaspin enhanced significantly porcine oocyte IVM and P4 concentration, as well as MAP3/1 phosphorylation, while decreasing PRKAA1. Using pharmacological inhibitors of MAP3/1 (PD98059) and PRKAA1 (Compound C), we observed that the effect of vaspin was reversed to the control level by all studied parameters. In conclusion, vaspin, by improving in vitro oocyte maturation via MAP3/1 and PRKAA1 kinase pathways, can be a new factor to improve in vitro fertilisation protocols.

Highlights

Oocyte maturation is a critical stage of embryo production in mammals, characterised by periods of meiotic arrest and resumption [1]
In our previous published data, we described the dependence on oestrous phase vaspin expression of porcine ovarian structures, ovarian follicles [19], and corpus luteum (CL) [20]
We showed that (i) vaspin and GRP78 mRNA and protein changed after cumulus–oocyte complexes (COCs) maturation in oocytes and cumulus cells, (ii) vaspin enhanced significantly porcine oocyte maturation and P4 secretion by COCs, (iii) vaspin upregulated MAP3/1 and decreased PRKAA1 phosphorylation, and iv) the stimulatory effect of vaspin on oocyte maturation was mediated by MAP3/1 and PRKAA1 kinase activation

Summary

Introduction

Oocyte maturation is a critical stage of embryo production in mammals, characterised by periods of meiotic arrest and resumption [1]. Balance is needed between factors promoting or inhibiting oocyte maturation for subsequent fertilisation and embryo development [1]. In vitro maturation includes both nuclear and cytoplasmic maturation. Cytoplasmic maturation is characterised by molecular and structural changes, including reorganisation of the cytoskeleton to provide a mature ovum to support fertilisation and early embryonic development [4]. It is important that the in vitro environment must support both nuclear and cytoplasmic maturation; knowledge about factors stimulating oocyte maturation in vitro is a first step to improve in vitro fertilisation protocols. Chappaz et al [9] showed the expression of adiponectin receptors in porcine oocytes and cumulus cells, as well as a decrease in immature oocyte number by adiponectin addition via the p38MAP pathway. Craig et al [7] noted upregulation in leptin receptor protein expression after oocyte maturation and the stimulatory effect of leptin on this process via the MAPK pathway

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