General discussion

Pregnancy complications derived from impaired placentation, including gestational hypertension and IUGR are the main causes of perinatal morbidity and mortality and a risk for different diseases in the newborn (314). Even though there are some established genetic and environmental factors that contribute to these adverse outcomes, they can appear in women who were healthy before pregnancy, without a history of complications themselves or in their family (315). The risk factors are usually studied in the context of the maternal-foetal-placental unit, without considering paternal factors (19). However, maternal risk factors alone may not explain the cause of the complication in many cases. In this thesis, we have proposed some paternal factors as contributors to the development of pregnancy complications. As important adverse pregnancy outcomes can trace their origin to the placenta, paternal factors affecting placentation may influence the risk of adverse pregnancy outcome. The results of the Reus Tarragona Birth Cohort reported in this thesis show that paternal MTHFR C677T genotype and elevated tHcy levels are associated with impaired placentation and lower birthweight. In addition, we observed that in some cases maternal factors were not associated as a risk factor for those adverse outcomes. Nevertheless, this applied to elevated tHcy during early pregnancy and the MTHFR 677TT genotype and it is possible that their underlying associations with adverse outcomes were not detected due to the protective effect of folic acid supplementation during the first trimester of pregnancy in 94.2% of the mothers, and from preconception in about a 36% of them. Previous studies have also focused on the important role of paternal genetic and nutritional status on the success of placenta and fetal development. In a study with male rats exposed to a high-fat diet and females on a control diet, Sheau-Fang et al (2010) have shown that the offspring had early onset of impaired insulin

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secretion and glucose tolerance, as well as altered gene expression in pancreatic islets (316). Bielawski et al (2002) in a study performed also in rats, observed that paternal chronic alcohol intake decreased messenger RNA expression of cytosine methyltransferase in sperm, which could result in hypomethylated DNA which is then passed on to the offspring (21). In the same line, Lundi et al (2017) have observed that mice exposed to both a folate deficient diet and a high dose of folic acid supplementation have male descendants (F1) with lower sperm counts which leads to increased postnatal death in the offspring (F2) (189). Therefore, it is also important to consider that paternal lifestyle, apart from genetics, can play a role in pregnancy complications and fetal programming. We consider that it will be important to include paternal factors in the focus of interest to learn more about their implication in complications developed during pregnancy that we cannot explain only with maternal factors. In a context without mandatory folic acid fortification like Spain, where 61% of the men adult have folic acid deficiency, it would be interesting to establish a protocol of folic acid supplementation in fertile man planning to have a baby to avoid pregnancy complications if scientific evidences support it.

Strengths and Limitations

The principal strength of our study is the time on data collection. The prospective design from early pregnancy allows as to have clinical, obstetric and lifestyle data from the first trimester of pregnancy and a blood sample before 12 GW with a confirmed viable foetus. It’s very difficult to obtain first trimester data because it depends on the timely initiation of prenatal care by the mother. Most studies have data from the mid or final trimesters of pregnancy. We also have the participation of the fathers, which is difficult to

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obtain because they are not visited as part of prenatal care and they have to agree to come voluntarily and especially for the interview and blood sample. Many are uninterested for work reasons or don’t want to come to the hospital to give a blood sample. In the fathers, the absence of folic acid food fortification in Spain and the very low use of folic acid supplementation use, allowed us to study the effect of the MTHFR C677T polymorphism in 1C metabolism and pregnancy complications without the masking effect of folic acid.

One of the limitations of this thesis is that observational studies do not allow to assume causality effect in the associations studied. However, we adjusted for potential confounding factors. Over 60% of the fathers participated, but it is possible that these were more motivated for research and possibly healthier. However, this did prevent us from testing our hypothesis as shown by the novel associations we observed. Another limitation is our participant recruitment. We could not access participants having the prenatal check-up outside the hospital, before 12 GW. Participants visiting the hospital for the prenatal care were mostly people with more risk of developing pregnancy complications and some hospital / university staff.

Future perspectives

Further investigation will be useful to replicate the results regarding paternal involvement in pregnancy adverse outcomes. Our study reports ground breaking results about paternal MTHFR C677T genotype and 1C metabolism influence on pregnancy complications. More studies focused on this topic would inform future recommendations or protocols. If the father is associated with pregnancy complications that we cannot explain by maternal risk factors and thus not covered by primary and secondary prevention measures, the

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associated complications likely emerge when it is too late. We could focus on recommendations for the father at preconception. Recommendations on diet and healthy lifestyle or folic acid supplement use in the father might contribute to reducing the incidence of pregnancy complications. In addition, further investigation will be useful to know the mechanisms by which paternal factors are affecting those adverse outcomes. Epigenetic factors have been proposed as a possible mechanism (206). Micronutrients involved in one carbon metabolism can contribute to DNA methylation and affect foetal programming (228).