Project Title:

A novel intervention strategy to prevent IUGR

Discipline:

Foetal Medicine

Chief Investigator:

Dr Kathryn Gatford

Funding Amount:

$75,000

Recipient:

The University of Adelaide

Overview:

Intrauterine growth restriction (IUGR) increases risks of perinatal death >5-fold, and has lifelong adverse effects on health. There is currently no treatment to prevent or cure IUGR. We wish to test a novel approach to stimulate the mother’s own production of hormones that promote placental function and fetal growth. We are requesting support to test active ghrelin treatment in mice with variable litter size and prenatal constraint. Importantly, diet can be used to induce activation of ghrelin, providing a pathway for developing a therapy to prevent IUGR which is acceptable to patients.

Research Outcomes:

Researchers:

Kathryn Gatford, Claire Roberts, Beverly Muhlhausler, Hannah Brown, Jeremy Thompson.

Research Completed:

2018

Research Findings:

Intrauterine growth restriction (IUGR) increases risks of perinatal death >5-fold, and has lifelong adverse effects on health. There is currently no treatment to prevent or cure IUGR. In this project, we tested a new dietary approach to promote fetal growth. Unfortunately, the dietary supplement we trialled did not promote fetal growth in normal pregnancies or stimulate responses in the mother that would promote fetal growth in restricted pregnancies. However, we also developed a new mouse model of intrauterine growth restriction, which we will next use to test other interventions. We also tested new methods to measure placental function, since a poorly-functioning placenta is a major cause of restricted growth of the baby before birth. This project has generated valuable new tools to help us solve the problem of poor growth before birth.

Key Outcomes:

Intrauterine growth restriction (IUGR) increases risks of perinatal death >5-fold, and has lifelong adverse effects on health. There is currently no treatment to prevent or cure IUGR.

The original project aims were to therefore to characterise effects of maternal acyl-ghrelin infusion throughout normal pregnancy and those complicated by Intrauterine Growth Restriction (IUGR) in the mouse on:

(i) placental size, structure and function, specifically glucose and amino acid transport, and

(ii) fetal growth.

Since submitting the CRF application, we had conducted a pilot study and shown that feeding C8-fatty acid (octanoic acid), which is bound to and activates ghrelin, was able to increase circulating GH in non-pregnant mice. This led us to substitute this dietary approach for the originally-proposed surgical implantation of infusion pumps to deliver acyl-ghrelin. This dietary approach is how we hoped to translate any effects of direct acyl-ghrelin administration, since this will be far more acceptable to patients than infusions or injections of acyl-ghrelin. In the first experiment conducted within the CRF project, we therefore fed non-pregnant and pregnant mice with octanoic acid, and measured maternal circulating acyl-ghrelin, growth hormone (which we know promotes fetal growth) and fetal growth. Unfortunately, the dietary supplement we trialled did not promote fetal growth in normal pregnancies or stimulate responses in the mother that would promote fetal growth in restricted pregnancies. We found decreased activation of ghrelin in pregnancy, although its local action may be increased, since we also discovered that ghrelin-producing cells in the stomach increase during pregnancy. The results of this experiment mean that we will next trial oral administration of synthetic compounds that activate the ghrelin-receptor as an approach to promote fetal growth.

In our second series of experiments, we tested new methods to measure placental function, since a poorly-functioning placenta is a major cause of restricted growth of the baby before birth. Currently, placental transport of nutrients from the mother to the growing fetus is measured in experimental animals using radio-labelled tracers. This only allows one or two nutrients to be measured and is limited to facilities where radioactive materials can be handled. We therefore developed and tested a method using fluorescently-labelled glucose. We established that radiolabelled tracers are needed to measure transport of specific nutrients, but our new method provides a useful marker of blood flow through the placenta. This work has now been published (Kaur et al 2019 Placenta in press doi: 10.1016/j.placenta.2019.01.014).

The final set of experiments supported by this project allowed us to develop a new mouse model of intrauterine growth restriction, which we will next use to test these and other interventions. By transferring variable numbers of embryos (6-12 per uterine horn) into pseudo-pregnant mice, we generated mouse pregnancies with between 3 and 18 fetuses near the end of pregnancy. This extends the normal litter size range of 3-10 pups), and generates marked IUGR in large litters. Fetuses from the largest litters were ~25% lighter as well as thinner and with evidence of brain-sparing, compared to fetuses from smaller litters. Intriguingly, we have just completed genotyping and shown that the effect of increasing litter size on fetal growth is sex-specific, with the pattern of female growth unaffected by maternal constraint. Males are heavier than females when unconstrained (in small litters), and their placentas are always heavier than those of females. This suggests that the male fetus extracts maximal available nutrients from the mother, whereas the female maintains placental reserve capacity. This strategy is likely to place the male fetus at risk in the event of a “second hit”, consistent with predominantly male loss in utero when the mother is acutely compromised such as during asthma exacerbations. The development and optimisation of this model now allows us to combine genetic models in mice and testing of intervention strategies to prevent IUGR due to chronic restriction.

Overall, this project has generated valuable new tools to help us solve the problem of poor growth before birth. We have eliminated one treatment approach, and developed a mouse model to test others. The experiments funded by this grant have also been valuable for research training, and formed the basis of the PhD being undertaken by Harleen Kaur (supervised by Dr Kathy Gatford, A/Prof Beverly Muhlhausler and Prof Claire Roberts). We were also able to host third year students during their research placement who conducted some measures on placentas from these projects.

Research Papers:

Published in press – available on line:

H Kaur, RL Wilson, AS Care, BS Muhlhausler, CT Roberts, KL Gatford 2019 Validation studies of a fluorescent method to measure placental glucose transport in mice. Placenta, article in press 24 January 2019, doi: 10.1016/j.placenta.2019.01.014

(Two other manuscripts are in preparation; one describing the effects of octanoic acid and pregnancy no the ghrelin-GH axis, and another describing sex-specific IUGR in embryo-transfer mouse pregnancies)

Conference Presentations:

  • Harleen Kaur, Alison Care, Sandra Piltz, Paul Thomas, Beverly Muhlhausler, Claire Roberts, Kathryn Gatford. Sexual dimorphism in fetal growth responses in a mouse model of IUGR. Fetal and Neonatal workshop of Australia and New Zealand, March 2019.
  • Harleen Kaur, Rebecca Wilson, Alison S Care, Beverly S Muhlhausler, Claire T Roberts, Kathryn L Gatford. Validation studies of a fluorescent method to measure placental glucose transport in mice. Society for Reproductive Biology, Annual General Meeting, August 2018. Poster presentation H Kaur.
  • Kaur H, Care AS, Wilson RL, Piltz S, Thomas P, Muhlhausler B, Roberts CT, Gatford KL. A murine model of IUGR induced by embryo transfer. Society for Reproductive Biology, Annual General Meeting, August 2018. Poster presentation KL Gatford.
  • Rebecca Wilson, Harleen Kaur, Beverly S Muhlhausler, Claire T Roberts, Kathryn L Gatford. Developing a fluorescent method to measure placental glucose transport in mice. Fetal and Neonatal workshop of Australia and New Zealand, March 2018. Oral presentation KL Gatford.
  • Harleen Kaur, Hui Li, Pamela Su-Lin Sim, Rebecca L Wilson, Lili Huang, Chen Chen, Johannes D Velhuis, Amanda Page, Beverly S Muhlhausler, Claire T Roberts, Kathryn L Gatford. A potential role for ghrelin as a driver of increased growth hormone during murine pregnancy. Fetal and Neonatal workshop of Australia and New Zealand, March 2018. Oral presentation H Kaur.
  • Wilson RL, Kaur H, Piltz S, Thomas P, Muhlhausler B, Roberts CT, Gatford KL. Pilot study: a murine model of IUGR induced by embryo transfer. Perinatal Society of Australia and New Zealand, March 2018. Poster presentation KL Gatford.

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