A combination of growth factors and cytokines alter preimplantation mouse embryo development, foetal development and gene expression profiles

Abstract Within the maternal tract, the preimplantation embryo is exposed to an array of growth factors (GFs) and cytokines, most of which are absent from culture media used in clinical IVF. Whilst the addition of individual GFs and cytokines to embryo culture media can improve preimplantation mouse embryo development, there is a lack of evidence on the combined synergistic effects of GFs and cytokines on embryo development and further foetal growth. Therefore, in this study, the effect of a combined group of GFs and cytokines on mouse preimplantation embryo development and subsequent foetal development and gene expression profiles was investigated. Supplementation of embryo culture media with an optimised combination of GFs and cytokines (0.05 ng/ml vascular endothelial GF, 1 ng/ml platelet-derived GF, 0.13 ng/ml insulin-like GF 1, 0.026 ng/ml insulin-like GF 2 and 1 ng/ml granulocyte colony-stimulating factor) had no effect on embryo morphokinetics but significantly increased trophectoderm cell number (P = 0.0002) and total cell number (P = 0.024). Treatment with this combination of GFs and cytokines also significantly increased blastocyst outgrowth area (P < 0.05) and, following embryo transfer, increased foetal weight (P = 0.027), crown-rump length (P = 0.017) and overall morphological development (P = 0.027). RNA-seq analysis of in vitro derived foetuses identified concurrent alterations to the transcriptional profiles of liver and placental tissues compared with those developed in vivo, with greater changes observed in the GF and cytokine treated group. Together these data highlight the importance of balancing the actions of such factors for the regulation of normal development and emphasise the need for further studies investigating this prior to clinical implementation.

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111. THE ROLE OF L-PROLINE IN PREIMPLANTATION MOUSE EMBRYO DEVELOPMENT IN VITRO

Reproduction Fertility and Development ◽

10.1071/srb10abs111 ◽

2010 ◽

Vol 22 (9) ◽

pp. 29 ◽

Cited By ~ 1

Author(s):

S. Ozsoy ◽

M. B. Morris ◽

M. L. Day

Keyword(s):

Amino Acids ◽

Embryo Development ◽

Preimplantation Embryo ◽

Blastocyst Stage ◽

Low Density ◽

Mouse Embryo Development ◽

Preimplantation Mouse Embryo ◽

Preimplantation Mouse ◽

Development In Vitro

Amino acids are known to play important roles in preimplantation embryo development, including regulation of cell volume and metabolism. Inclusion of l-glutamine, glycine and betaine in embryo culture medium has been shown to improve development in vitro by acting as organic osmolytes, thereby regulating cell volume. The purpose of the present study was to examine the effect of l-proline on preimplantation mouse embryo development in vitro. One-cell stage embryos were cultured in modified HTF, at low density (1 embryo/100 μL) and high density (1 embryo/μL) in the presence and absence of amino acids. Development of the embryos was scored every 24 h until the blastocyst stage. At low density, l-proline significantly increased the proportion of embryos developing to the blastocyst stage. This effect was abolished by culture at high density, suggesting that l-proline was activating a pathway similar to that involved in autocrine signalling by trophic factors in the preimplantation embryo. The improvement in development observed in the presence of l-proline was not due to its action as an organic osmolyte since the osmolality of the medium was 270 mOsm. Furthermore, glycine and betaine, which are known to act as osmolytes in embryos, had no effect on blastocyst development. In embryonic stem cells L-proline is taken up by an amino acid transporter and is involved in the regulation of growth and differentiation (1). The present data suggest that l-proline may have a similar, important role in preimplantation development. (1) JM Washington, J Rathjen, F Felquer, A Lonic, MD Bettess, N Hamra, L Semendric, BSN Tan, J-A Lake, RA Keough, MB Morris and PD Rathjen (2010) Am J Physiol Cell Physiol 298: C982–C992.

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