Fetus-derived DLK1 is required for maternal metabolic adaptations to pregnancy and is associated with fetal growth restriction

Abstract

Pregnancy is a state of high metabolic demand. Fasting diverts metabolism to fatty acid oxidation, and the fasted response occurs much more rapidly in pregnant women than in non-pregnant women. The product of the imprinted DLK1 gene (delta-like homolog 1) is an endocrine signaling molecule that reaches a high concentration in the maternal circulation during late pregnancy. By using mouse models with deleted Dlk1, we show that the fetus is the source of maternal circulating DLK1. In the absence of fetally derived DLK1, the maternal fasting response is impaired. Furthermore, we found that maternal circulating DLK1 levels predict embryonic mass in mice and can differentiate healthy small-for-gestational-age (SGA) infants from pathologically small infants in a human cohort. Therefore, measurement of DLK1 concentration in maternal blood may be a valuable method for diagnosing human disorders associated with impaired DLK1 expression and to predict poor intrauterine growth and complications of pregnancy.

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Figure 1: The conceptus is the source of elevated maternal plasma DLK1 levels in late gestation.
Figure 2: Fetus, not placenta, is the source of maternal circulating DLK1.
Figure 3: Maternal genotype and conceptus-derived DLK1 alter maternal metabolism.
Figure 4: Low DLK1 levels in human pregnancy are associated with pathological SGA.

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Acknowledgements

M.A.M.C. was supported by a PhD studentship from the Cambridge Centre for Trophoblast Research. Research was supported by grants from the MRC (MR/J001597/1 and MR/L002345/1), the Medical College of Saint Bartholomew's Hospital Trust, a Wellcome Trust Investigator Award, EpigeneSys (FP7 Health-257082), EpiHealth (FP7 Health-278414), a Herchel Smith Fellowship (N.T.) and NIH grant RO1 DK89989. The contents are the authors' sole responsibility and do not necessarily represent official NIH views. We thank G. Burton for invaluable support, and M. Constância and I. Sandovici (University of Cambridge) for the Meox2-cre mice. We are extremely grateful to all of the participants in the Pregnancy Outcome Prediction study. This work was supported by the NIHR Cambridge Comprehensive Biomedical Research Centre (Women's Health theme) and project grants from the MRC (G1100221) and Sands (Stillbirth and Neonatal Death Charity). The study was also supported by GE Healthcare (donation of two Voluson i ultrasound systems for this study) and by the NIHR Cambridge Clinical Research Facility, where all research visits took place.

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M.C., M.A.M.C., A.C.F.-S. and G.C.S.S. conceived and designed the experiments. M.C., M.A.M.C., J.A.C., M.H., I.G., N.T., C.L.D. and D.S.C.-J. performed the experiments. M.C., M.A.M.C., A.C.F.-S., C.L.D., F.G. and G.C.S.S. analyzed the data. M.C. and U.S. performed statistical analysis. S.R.B., T.L.P., A.C.F.-S. and G.C.S.S. contributed reagents. M.C., M.A.M.C., A.C.F.-S. and G.C.S.S. wrote the manuscript. M.C., A.C.F.-S. and G.C.S.S. provided supervision.

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Correspondence to Marika Charalambous.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Altered body composition in Null females can be detected during embryogenesis.

(a) Total bodyweight of virgin females at 12 weeks does not differ between genotypes. Null females have larger abdominal WAT deposits (b) and reduced muscle (combined gastrocnemius/soleus) (c) than WT and Mat females at 12 weeks. WT and Null n = 8, Mat n = 7, compared by One-way ANOVA, with Bonferroni's Multiple comparison post-hoc test with WT vs Null and WT vs Mat, *p<0.05. Body weight (d), crown rump (C-R) length (e) and tissue weights (f) in embryos at E18.5. Null embryos have reduced mass and length, and reduced lean (hindlimb and forelimb) mass than embryos expressing Dlk1, WT n > 21, Null n > 13, Mat n > 10, compared by Kruskall-Wallace test with Dunn's Multiple comparison post-hoc test comparing WT vs Null and WT vs Mat, *p<0.05, ***p<0.001. (g) Null mice are born small but catch up in the preweaning period. Serial measurements of pup weight from birth to weaning, Nulls weigh significantly less at birth but not thereafter, n = 4-11 per genotype, each time point compared as above.

Supplementary Figure 2 Summary of experimental crosses

Experimental crosses used to determine which compartment of the conceptus (embryo, placental fetal endothelium or placental trophoblast) is the source of maternal circulating DLK1 in pregnancy.

Supplementary Figure 3 Levels of pituitary Gh mRNA do not differ between the groups.

Expression levels of Gh (a) measured by real-time quantitative PCR in pituitaries from females in the cohort. Gh levels did not differ between the groups. Data was normalised to β-actin levels, then expressed as WT = 1. Groups were compared by One-way ANOVA, with Bonferroni's Multiple comparison post-hoc test with WT vs Null and WT x WT vs Null x WT and Null x Null, ***p<0.001.

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Cleaton, M., Dent, C., Howard, M. et al. Fetus-derived DLK1 is required for maternal metabolic adaptations to pregnancy and is associated with fetal growth restriction. Nat Genet 48, 1473–1480 (2016). https://doi.org/10.1038/ng.3699

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