TWIN Longitudinal Investigation of FEtal discordance

Lifelong health is in part set during intrauterine life. An adverse
intrauterine environment can induce persistent epigenetic changes that are
thought to cause long-term health effects. There is an urgent need for human
studies that can identify the epigenetic alterations that underlie the impact
of intrauterine adversity on disease, in particular cardiovascular disease
(CVD) and neurodevelopmental impairment (NDI). This study will focus on
identical twin pairs who shared a single placenta i.e., monochorionic (MC)
twins. Every year, over 600 MC twins are born in The Netherlands and they are
at high risk of experiencing an adverse intrauterine environment. In one third
of pairs, one fetus has significantly less access to nutrients and resources
during pregnancy than its co-twin, conditions known to be linked to increased
CVD risk and impaired neurodevelopment in adults. Thus, although genetically
identical, great differences in intrauterine exposure exist within twin pairs,
providing an unique natural experiment allowing a robust assessment of the
development of cardiac- and neurodevelopmental risk factors in childhood and
probe the underlying epigenetic mechanisms. Instead of relying on commonly used
blood samples, this study will examine altered epigenetic regulation in
mesenchymal stromal cells (MSCs), an enhanced proxy for other tissues involved
in CVD and NDI. These multipotent cells are known to display metabolic changes
in newborns exposed to an adverse intrauterine environment and can be
differentiated into other cell types. The hypothesis is that twins discordant
for pregnancy complications display a distinct epigenetic signature in MSCs.
This signature contributes to cellular metabolic alterations and is associated
with future cardiovascular and neurodevelopmental outcome in childhood and
beyond. Our results will not only address an unmet clinical need in the
high-risk group of MC twins, but may also advance early-life CVD prevention
strategies and underpin their efficacy in the general population.

The main objective is to pinpoint the biological pathways that set long-term
risk of CVD and NDI when epigenetically disturbed during fetal development by
(1) establishing a longitudinal cohort of MC twins, (2) defining epigenetic
alterations induced by an adverse intrauterine environment by comparing
genome-wide DNA methylation of MC twins in MSCs, and (3) evaluating the role of
these epigenetic alterations in mediating the effects of intrauterine adversity
on cardiovascular and neurodevelopmental risk factors at birth and at the age
of 2, 5 and 8 years.

A longitudinal observational study.

MC twins are at high risk of experiencing adverse prenatal conditions. The
focus in current research, however, has been on the resulting acute
complications during pregnancy. In this study, we will focus on the postnatal
period as a first and essential step to understand and address the long-term
health consequences of complicated MC pregnancies. The LUMC is the national
referral center for the management, treatment and follow-up of complicated MC
twin pregnancies, and therefore has ample experience in performing (clinical)
assessments of MC twins and their parents with minimal burden and risk. The
majority of assessments already is part of standard clinical care for specific
sub-groups of MC twins and without risk. In this study, we will perform these
assessments to the whole population of MC twins born in the LUMC.
Antenatal, a burden for the pregnant mother during routine ultrasound might be
prolonged examination time (~10 minutes) for the additional cardiac
measurements. At birth, placenta, cord and cord blood is easy to collect with
no risk to the mother or new-borns. Postnatally (between day 0-3), cranial and
cardiac ultrasound and, at 2, 5 and 8 years, neurodevelopmental evaluation are
standard of care for all MC twins complicated by Twin-Twin Transfusion Syndrome
(TTTS), Twin Anemia-Polycythemia Sequence (TAPS), prematurity (gestational age
at birth <30 weeks or birth weight <1000 grams), severe growth restriction
(birth weight are assessed as part of standard care at our outpatient clinic. After informed
consent from both parents, a control group of uncomplicated twins (N~35) will
be assessed (from prenatal to birth to follow-up at 2, 5 and 8 years)
specifically for the purpose of this study. These assessments are not
associated with any risk and the burden is minimal (e.g. 120 minutes at 2 and 5
years and 150 minutes at 8 year follow-up). Neurodevelopmental assessment at 2,
5 and 8 years is generally experienced as enjoyable for children. A mouth swab
and a few scalp hairs will be taken postnatally and at follow-up which is
without risk. In addition to routine cranial ultrasound scan in the first
postnatal week, the infants will undergo neuro-imaging, consisting of a cranial
ultrasound and MRI-examination around TEA. Cranial ultrasound is a safe and
non-burdening bedside procedure. MRI is also considered a low-risk and
minimally invasive procedure (CCMO) and we will follow the standard precautions
for neonatal MRI examinations in our hospital. For extremely preterm born
infants (<28 weeks), infants with TAPS, TTTS and those with brain lesions
already detected on routine cranial ultrasound, the MRI scan at TEA is part of
standard clinical care. Infants with a clinical indication for an MRI scan will
receive a light oral sedative (chloral hydrate) according to our current NICU
guideline to achieve optimal diagnostic scan quality. We will scan all other
infants, where MRI is not part of routine clinical care, during natural sleep
using the *feed and wrap* method.