**5. Discussion and conclusion**

*Psychopathology - An International and Interdisciplinary Perspective*

clinical pictures with atypical associations.

First, to document in utero exposure to synthetic estrogens, Kebir and Krebs [10] were able to analyze from our data a small number of family records that occurred in HHORAGES testimonies and studied a cohort of 472 exposed subjects. They account for 46.7% of mood disorders, 22.9% of psychotic disorders, 6.6% of anxiety disorders, 11% of eating disorders, and 12.7% of others, which confirms their previous observations published on 2009 and 2010 at the seventh and eighth Congress of the Encephalon in Paris [26, 27] on 43 exposed children highlighting

Second, genetic and epigenetic analyses of HHORAGES siblings have shown in patients suffering from psychotic disorders and exposed in utero to DES and/ or EE that this prenatal exposure is associated with epigenetic processes. Starting from the fact that psychiatric diseases develop from a brain dysfunction during neurodevelopment, and knowing that DES and EE are synthetic hormones (estrogens), endocrine disruptors, and confirming from the HHORAGES data numerous cases of heavy psychiatric disorders in children exposed, the Krebs'team in association with HHORAGES designed 10 years ago, in 2007, a research project Partnership Citizen Institution for Research and Innovation (PICRI), funded by the Ile de France Region, net by the French National REsearch Agency (ANR), that developed the hypothesis that the DES administered during pregnancies could be an environmental risk factor for the development of psychiatric disorders in impregnated children: the epigenome of the foetus could have been modified by in utero exposure to synthetic estrogens. The PICRI project was titled "Influence of hormonal treatments on brain development during pregnancy: study of phenotypic, behavioral and biological changes in informative families." The families of HHORAGES were called to perform peripheral blood sampling after thorough questioning. Many families volunteered to participate in the research: 31 families were selected, satisfying the rigorous inclusion criteria desired. Many more families had come forward during this study, but they could not be included because the psychotic patient refused to come to St. Anne's Hospital for blood sampling. In the selected families, total siblings were composed of first-born unexposed children, exposed children, and post-exposed children, with first-born unexposed serving as intrafamilial control. For the exploration of their epigenome, 485,000 cytosines by genome were studied and analyzed, representing an immense work. To complete this study, a cohort of young adolescents with relational, emotional, and social difficulties was followed for 6 months, some of whom had developed schizophrenic-type psychosis in these 6 months, although not exposed to DES. A comparison of their methylome, analyzed before and after the onset of the disease, was performed. In this study [28], authors reported a global methylation of the

After the analysis of the whole methylome of the selected HHORAGES cohort,

the team of Krebs-Kebir highlighted differential specific methylated regions (DMR): in the zinc finger protein 57 gene ZFP57 and in the ADAM TS9gene and in young psychotic patients exposed in utero to DES/EE [29, 30]. In this work, the authors observed that in exposed individuals, ZFP57 gene methylation may be associated with their psychosis. The ZFP57 gene (located on chromosome 6) is expressed very early in development. It is a transcription regulator, directly related to the phenomenon of methylation and neurodevelopment [31]. The ADAM TS9 gene is implicated in the control of organ shape, especially in the development and function of the uterus and reproductive organs [32] which are often abnormal after in utero DES exposure as well as in the control of the CNS development [33] and in

**14**

psychotic patient genome.

several kinds of cancers [34].

Very few studies have investigated the impact of prenatal exposure to DES and EE on psychiatric outcome. Animal studies on rats or mice allowed us to hypothesize that estrogenic hormones induce neurodevelopmental disturbances in exposed human subjects and may potentially mediate an increased risk of behavioral and psychiatric disorders. Our data therefore strongly suggest that DES/EE exposure during pregnancy is associated with high incidence of behavioral and/or psychiatric disorders. They illustrate a higher risk of schizophrenia, as this disease was 17 times more prevalent than in the general population, with sons being more affected than DES daughters. Regarding the existence of eating disorders (bulimia, anorexia), it should be noted that girls are much more affected than boys, and we often observed the association of eating disorders with bipolarity (manic-depressive disorders), anxiety, and depression. With regard to suicides, our work clearly demonstrates a drastically increased risk of suicide attempts (65.4% versus 0 in the unexposed controls and 0.25% in the general population) and suicides (3.4% versus 0 in the unexposed controls and 0.02% in the general population). It could be noted that, as in the general population, DES sons commit more suicides than DES daughters and the inverse for suicide attempts. Moreover, our data reveal that 50% of the sons who committed suicide suffered from schizophrenia. Psychiatric studies in general have shown that the percentage of suicides is generally higher in individuals with psychiatric disorders than in the general population. But to our knowledge, there is no information or specific studies concerning this association in the context of DES exposure. Sixteen subjects in Group 3 (post-DES children) (**Figure 2** and **Table 1**) had diagnosed psychiatric disorders. An explanation for this finding might be that DES, being a very lipophilic synthetic estrogen, remains in the mothers' fat after estrogenic impregnation in a previous pregnancy and is then released through the placental barrier during the next gestation.

No work had been reported on the impact of in utero exposure to synthetic progestin hormones administered alone on the occurrence of psychiatric disorders in exposed children before our first presentation in the European Congress of Gynecology in 2017 [24]. For the first time, we described psychiatric disorders that can affect children exposed in utero to progestins. Previously, and during many years, synthetic progestogens were not considered as dangerous during pregnancy or during replacement or contraceptive treatment. Moreover, they have been suggested to exert neuroprotective effects in several animal models of neurological disease [35]. Negative mood symptoms have been reported by Andreen et al. [36] in some women as a result of progesterone during the luteal phase of menstrual cycles. This is believed to be mediated via the action of allopregnanolone on the GABA-A system. A reduction of allopregnanolone circulating levels that correlates to depressive symptoms has been recently reported [37], and conversely, healthy women reported increased anxiety and mood disorders after long-acting subdermal implant of progestogens. In a group of 236 schizophrenic patients at onset, an elevated concentration of progesterone has been found, and authors suggested that steroid hormones may influence brain function, underlying schizophrenia, and major depressive disorders [38]. Moreover, Buoli et al. (2016) found high DHAS levels in patients with a history of psychotic symptoms, suggesting a role of steroids in the etiology of psychosis and mood disorders [39]. Progestins are known to induce GABA receptor activity/neural activation before birth; it is likely that a GABAergic system could contribute to schizophrenia, anxiety, depression, panic disorders, epilepsy, autism, and others [40]. Although some progestins have been banned from the market, others are not: our data demonstrated that caution should

be taken with regard to the use of these progestins during pregnancy and even outside these periods (contraception or hormone replacement therapy).

The brain is a very vulnerable organ because its development covers a very broad period extending from the early prenatal stage (third week of pregnancy) to end around the age of 20. During its development, there are times when its vulnerability is even greater than others; these periods are called "shooting windows," during which the environment can impact the normal process of development. Abdolmaleky et al. [41] as early as 2005 had developed the hypothesis that geneenvironment modulations could be performed via DNA methylations. Krebs' team put forward the hypothesis that DES-induced changes in epigenetic background and alteration of DNA functioning (methylations) could be significant factors to demonstrate a possible origin of psychotic disorders and a link with in utero DES exposure of the children suffering from these illnesses [29, 30].

Numerous studies have shown that, for example, in the rat, early maternal separation or the fact of causing significant stress to the mother changes the methylation signals of certain genes of the rat directly related to the regulation of anxiety. It has also been discovered that the proper environment for changing the methylation signals may be chemical. This is the case of DES recognized by the scientific community as an endocrine disruptor and banned for pregnant women. This change in the level of methylation caused in utero by DES has been demonstrated for urogenital malformations of girls and boys as well as for cancers. On 2015, Harlid et al. published in a pioneer work the first study for evaluation of possible effects of in utero DES exposure on genome-wide DNA methylation in humans [42]. They studied whole blood DNA methylation in 100 40–59-year-old women reporting in utero exposure, compared to 100 unexposed women. They did not find any differential methylation, but the DMR approach was not used in their recent work (2015). On the other hand, in 2017 Rivollier et al. [30] described specific differential methylated regions (DMR) on two genes implicated in neurodevelopment (ZFP57 and ADAMTS9). Surprisingly, they cautiously claim that these DMR are "supposedly" associated with prenatal exposure to DES in young psychotic patients in utero exposed to DES/ EE. Nevertheless, these modifications of methylation are really specific because they do not exist in the methylome of young psychotic patients not exposed to DES [28] in which global methylation of the genome was observed. Moreover authors have compared exposed subjects to their unexposed siblings which do not present these specific methylations although they shared environmental and genetic factors.

The citizen work carried out between the French HHORAGES Patient Association and two major medical research laboratories has provided convincing scientific results: (1) on the detection and confirmation of the existence of psychiatric disorders (accompanied or not of somatic disorders) in children exposed in utero to synthetic hormones and (2) on the mechanisms of action of these synthetic hormones administered to pregnant mothers on the brain of their offspring. The effects of these endocrine disruptors in humans through what is becoming a public health scandal, denied for a long time by doctors, especially psychiatrists, scientists, and specialized journalists, are thus better known. The fact that these synthetic products do not degrade in the human body in the same way as the natural hormones [13] and act on the functioning of genes implicated in neurodevelopment during the fetal life, following an epigenetic mechanism, is a real time bomb. Indeed, this mechanism induces a transgenerational effect already partially demonstrated in the HHORAGES cohort at the third-generation level for hypospadias, a specific genital malformation. So far, only a few third-generation children suffering psychiatric illness are documented in the HHORAGES testimonies. This is understandable because third-generation exposed children are still too young (excepted in some cases) to present psychiatric disorders as schizophrenia which is not the case for hypospadias that are detectable

**17**

*Evidence for Link Between Mental Disorders and in Utero Exposure to Synthetic Hormones…*

usually appear in postadolescence, 18–20 years, and sometimes later.

from birth in male children and grandchildren [43]. In contrast, psychiatric disorders

Mr. René Alexandre, Dr. Henri Pézerat, Pfr. Jean Caston, Mr. Michel Datry, and Dr. Charles Zelwer worked tirelessly to demonstrate the harmful role of the endocrine disruptors and in particular of synthetic hormones; before passing away: we shall never forget their efficient help. This work could not continue without the daily ongoing support of the HHORAGES board especially Mrs. Geneviève A., Mauricette P., Yette B., Annie R., Aimée I., Sylvette C., Denise H., Marie-Noelle P., and Pierre D. We acknowledge Mr. André Cicolella, president of the French Environment-Health network (RES), for his constant support. We are very grateful to Pfr. Michael Dolan, professor at the University of Massachusetts, Amherst (USA), for his help and critical reading during the preparation of the manuscript.

Association HHORAGES-France is registered on the Epidemiological Portal of French Health Databases INSERM (French National Institute for Medical Research) and AVIESAN (National Alliance for Life Sciences and Health) (epidemiologie-

The authors declare they have no competitive financial interests; the finances of HHORAGES-France association come exclusively from the donations of families

*Marie-Odile Soyer-Gobillard:* National Center for Scientific Research (CNRS),

*Charles Sultan:* Unité d'endocrinologie et gynécologie pédiatrique, Hôpital Arnaud-de-Villeneuve, CHU de Montpellier, 34295 Montpellier Cedex 5, France.

*Laura Gaspari:* Unité d'endocrinologie et gynécologie pédiatrique, Hôpital Arnaud-de-Villeneuve, CHU de Montpellier, 34295 Montpellier Cedex 5, France.

Sorbonne University, P. and M. Curie University, Paris 6, and HHORAGES Association (Halt to Artificial Hormones for Pregnancies), The Priory of Baillon, F-95270 Asnieres sur Oise, France. Email: elido66@orange.fr. Doctor of Science (Cell Biology), Honorary Emeritus Research Director at CNRS, laureate of several scientific awards, and author of more than 150 publications in peer-reviewed scientific journals. Former Director of the Department of Cellular and Molecular Biology (UMR 796 "Models in Cell and Evolutive Biology") at the Arago Laboratory (Paris 6 University) and expert at the National Agency for Research (ANR). National president of HHORAGES, France (Stop to Artificial Hormones for Pregnancy) and president of its Scientific Council. Member of the Clinical Projects Reviewers

*DOI: http://dx.doi.org/10.5772/intechopen.80969*

**Acknowledgements**

**Notes**

france.aviesan.fr).

**Author Biography**

and of sympathetic individuals.

Committee, INSERM, Paris.

Email: c-sultan@chu-montpellier.fr

Email: dr.lauragaspari@gmail.com

**Disclaimer**

*Evidence for Link Between Mental Disorders and in Utero Exposure to Synthetic Hormones… DOI: http://dx.doi.org/10.5772/intechopen.80969*

from birth in male children and grandchildren [43]. In contrast, psychiatric disorders usually appear in postadolescence, 18–20 years, and sometimes later.
