Decreased fetal thymus size in pregnancies after assisted reproductive technologies

Abstract

Objective

To compare the size of the fetal thymus, using both fetal thymic-thoracic ratio and fetal thymus transverse diameter values in Assisted reproductive technologies (ART) or naturally conceived pregnancies.

Methods

In this retrospective study, fetal thymic-thoracic ratio and fetal thymus transverse diameter were evaluated in 204 pregnant women. Patients were examined in two groups. The study included 58 Intracytoplasmic sperm injection (ICSI) patients (study group) and 146 healthy pregnant women (control group).

Results

Fetal thymic-thoracic ratio in ART pregnancies were found to be statistically significantly lower than that of the control group (p = .001). Also, the fetal thymus transverse diameter value was found to be statistically significantly lower in ART pregnancies compared to that of the control group (p = .001).

Conclusions

The size of the fetal thymus, manifested with a decrease in both fetal thymic-thoracic ratio and thymus transverse diameter values, decreased in ART pregnancies.

Keywords:

• Assisted reproductive technologies
• fetal thymus
• intracytoplasmic sperm injection
• thymus transverse diameter
• thymic-thoracic ratio

Introduction

Assisted reproductive technologies (ART) are alternative treatment methods used to achieve intrauterine pregnancy in couples who cannot conceive naturally and include the procedures of ovulation induction, intrauterine insemination (IUI), and in vitro fertilization (IVF), and intracytoplasmic sperm injection (ICSI) [1]. It has been suggested that epigenetic changes in ART pregnancies may lead to differences in fetal and placental gene expression compared to spontaneous pregnancies [2]. The hormonal stimulation and embryo culture procedures used in assisted reproductive technologies (ART) can create a stressful environment for gametogenesis and early embryonic development. This environment can potentially lead to epigenetic changes in the genes involved in growth and development, which can have negative effects on fetal development and health [3–6].

Maternal anatomical and physiological adaptations are necessary for healthy embryo implantation and fetal development, and maternal immune response plays a critical role in the coordinated realization of these adaptations. Thymus, T lymphocytes produced in the thymus, and regulatory T cells which are a subset of these T lymphocytes have critical roles in this immune response [7,8]. Fetal thymus size was investigated in healthy naturally conceived [9] and complicated pregnancies [10–14]. However, fetal thymus size assessment has not received enough attention in ART pregnancies, where pregnancy complications are known to be more common.

To the best of our knowledge, there is only one study in the literature examining the fetal thymus size in ART pregnancies. The present study evaluated only the fetal thymic-thoracic ratio in ART pregnancies [15]. However, there is no study evaluating fetal thymus transverse diameter in these pregnancies. For this reason, we aimed to compare the size of the fetal thymus, by using both fetal thymic-thoracic ratio and fetal thymus transverse diameter in ART and naturally conceived pregnancies.

Materials and methods

This retrospective study was approved by Sakarya University Ethics Committee (decision no:40026-378, approval date: 30 June 2021). In this study carried out between 1 November 2018 and 15 June 2021 in Sakarya University Training and Research Hospital, Gynecology and Obstetrics Clinic, Perinatology Department, singleton pregnancies were included with a routine second-trimester screening at 18–23 gestational weeks. Patients who conceived pregnancy with ICSI, an ART procedure (n: 58), were included in the study group, and the patients who conceived a natural pregnancy (n: 146) were included in the control group. The fetal thymus transverse diameter and fetal thymic-thoracic ratio values measured during routine second-trimester screening at 18–23 gestational weeks were obtained from the medical records. Patients with gestational or pregestational diabetes, pregnancy-associated hypertensive diseases, pregnant women with sonographic estimated fetal weight <10 percentile, patients with SLE or other accompanying chronic inflammatory diseases, and those with fetal structural or chromosomal disorders were excluded from the study.

All ultrasound examinations were performed by a single sonographer (Koray Gök) using a Voluson 730 and a Voluson E6 (GE Medical Systems, Milwaukee, WI) ultrasound machine. Thus, the measurements were standardized and the bias was limited. All measurements were carried out in the absence of fetal movements. Fetal thymus transverse diameter measurement was performed according to that previously described by Zalel et al. [16]. The thymus was identified in the three vessels view as a homogeneous structure in the anterior mediastinum. The transverse diameter of the fetus was measured by placing the ultrasound calipers perpendicular to the junction between the sternum and the spine (Figure 1). Fetal thymic–thoracic ratio measurement was performed according to that previously described by Chaoui et al. [14]. The thymus was identified in the three vessels and trachea (3VT) view as a hypoechogenic structure with echogenic dots filling the space between the vessels posteriorly and the anterior chest wall (sternum and ribs) anteriorly. The anteroposterior diameter of the thymus was measured along the midline between the transverse aortic arch border posteriorly and the posterior chest wall anteriorly. Also, the mediastinal axial diameter was measured, along the line traced to measure the thymic diameter, as the distance between the anterior edge of the thoracic vertebral body at the level of the transverse arch posteriorly and the internal edge of the sternum anteriorly. The TT-ratio was then calculated as the ratio of the anteroposterior thymic to the intrathoracic mediastinal diameter (Figure 2).

Figure 1. Fetal thymus transverse diameter measurement method.

Figure 2. Fetal thymic-thoracic ratio measurement method. (A) Thymus diameter (anteroposterior thymus diameter) and (B) thorax diameter (intrathoracic mediastinal diameter), Fetal thymic-thoracic ratio: (A)/(B).

The statistical evaluations were carried out utilizing the SPSS 24.0 software (SPSS Inc. and Lead Tech. Inc. Chicago. the USA). The Kolmogorov–Smirnov test was adopted to examine the normality of the distribution of the data. Nonparametric Mann–Whitney U test was used to compare the groups because the distribution of the data was not normal. Nonparametric data were presented as the median and interquartile range (IQR). p < .05 was regarded as significant in all analyses.

Results

In the present study, 58 patients with ART pregnancy and 146 patients who naturally conceived were evaluated. The demographic characteristics of the patient group with ART pregnancy and the control group are presented in Table 1. There were no statistically significant differences between ART pregnancies and the control group in terms of age, body mass index (BMI), and the gestational week at which fetal thymic-thoracic ratio and fetal thymus transverse diameter was measured.

Table 1. Demographic characteristics of the study and control groups.

	Study group (n:58)	Control group (n:146)	p Value
Maternal age (years)	32 (23–43)	30 (19–43)	.088^a
BMI (kg/m^2)	26.55 (23.8–28.9)	26.1 (23–28.4)	.139^a
Gestational age at time of study (weeks)	20.92 (19.42–22.7)	21.14 (19.42–22.84)	.096^a

BMI: body mass index.

^aMann–Whitney U test.

The comparison of fetal thymus transverse diameter and fetal thymic-thoracic ratio values in the ART pregnancy group and the control group are presented in Table 2. The fetal thymic-thoracic ratio in ART pregnancies was found to be statistically significantly lower than that of the control group (p = .001). Also, the fetal thymus transverse diameter value was found to be statistically significantly lower in ART pregnancies compared to that of the control group (p = .001).

Table 2. Comparison of fetal thymus transverse diameter and fetal thymic-thoracic ratio in the study and control groups.

	Study group (n:58)	Control group (n:146)	p value
Transverse diameter (mm)	16.8 (13.8–19)	20.8 (18.1–23.1)	.001^a
Thymic-thoracic ratio	0.306 (0.258–0.361)	0.432 (0.392–0.46)	.001^a

^aMann–Whitney U test.

Discussion

The present study aimed to compare the fetal thymus size in ART pregnancies and naturally conceived pregnancies. It was found that both fetal thymic-thoracic ratio and fetal thymus transverse diameter were statistically significantly lower in ART pregnancies compared to those conceived naturally.

Numerous methods have been reported for the ultrasonographic measurement of fetal thymus size [14,17,18], however, it remains unclear which method is the best for the detection of maternal and fetal diseases. In a study evaluating fetal thymus transverse diameter, thymus circumference, and thymic-thoracic ratio in uncomplicated pregnancies, it has been reported that transverse diameter is a convenient and reproducible measurement and, therefore, it is considered the best method in clinical practice [9]. In other previous studies, it has been reported that the fetal thymic-thoracic ratio was not affected by the body mass index and fetal sex [10,19], and this ratio remained constant throughout the pregnancy [14]. Therefore, in the present study, in the fetal echocardiographic examination as a component of routine screening at 18–23 gestational weeks, both fetal thymic-thoracic ratio and fetal thymus transverse diameter were utilized to evaluate the fetal thymus size.

Fetal thymus size was investigated in various maternal and fetal diseases, and researchers have suggested reasons for the changes in thymus size. It has been reported that the small size of the fetal thymus in women with preterm prelabour rupture of membranes (PPROM) may be an ultrasonographically reliable indicator of fetal involvement in the systemic inflammatory response [20,21]. Studies on diabetic pregnant women have suggested that hypoxic and metabolic stress resulting from impaired glucose metabolism may result in a decrease in fetal thymus size [22,23]. Mohamed et al. in their study on preeclamptic pregnant women found that the fetal thymus size decreased. The researchers associated this decrease with the induction of apoptosis in the fetal thymus by binding of cortisol, produced as a result of the activation of the hypothalamic-hypophyseal-adrenal axis due to stress, to its receptor in the cortical thymus [24]. Another study showed the impairment of fetal thymus growth before a clinical disease in preeclampsia, and the researchers have argued that a reduction in fetal thymus size may be a valuable indicator in predicting preeclamptic pregnant women associated with abnormal placentation [12].

Fetal thymus size in ART pregnancies has been previously evaluated only in one study in the literature [15]. In a retrospective study by Nau et al. [15] 774 naturally conceived pregnancies were compared with 162 ART pregnancies. Of the ART pregnancies in this study, 109 were ICSI pregnancies and 53 were IVF pregnancies. In our retrospective study, however, the number of pregnant women was lower and there were 146 naturally conceived pregnancies, while 58 ART pregnancies all consisted of ICSI pregnancies. In the study conducted by Nau et al. only the fetal thymic-thoracic ratio was measured in pregnant women, and it was reported that fetal thymus size was smaller in ART pregnancies compared to naturally conceived pregnancies, similar to ours [15]. Unlike us, in the study of Nau et al. [15] when the fetal thymic-thoracic ratio was compared in IVF and ICSI subgroups, no difference was found between the groups. In our study, unlike the study of Nau et al., we evaluated both fetal thymic-thoracic ratio and fetal thymus transverse diameter together to make a more objective estimation of the fetal thymus. And we found both fetal thymus size parameters to be low in ICSI pregnancies.

How can the small fetal thymus size be explained in pregnancies obtained with ART? Immune tolerance at the maternal-fetal interface and the immune regulatory mechanisms that provide this tolerance are of important for a successful pregnancy since it prevents the growing fetus from being rejected by the mother’s immune system. If these mechanisms fail, immune responses are activated, leading to negative pregnancy outcomes [25]. Regulatory T (Treg) cells have an important role in the proper functioning of all these mechanisms. Regulatory T (Treg) cells have functions including regulating immunity, suppressing inflammation, facilitating maternal vascular adaptations for trophoblast invasion, and placental development [8]. Current evidence has suggested that an insufficient number and function of regulatory T (Treg) cells is associated with idiopathic infertility and recurrent miscarriages, as well as later pregnancy complications from placental insufficiency, including preeclampsia and fetal growth restriction [25]. It has been reported in previous studies that mechanisms including systemic inflammatory response, stress and abnormal placentation result in small fetal thymus size in complicated pregnancies. ART pregnancies are also a complicated form of pregnancy. It has been known that ART includes procedures that are not characteristic of in vivo reproduction, and the oxidative, thermal and mechanical stress brought about by these procedures affects the biological processes of placental growth, development, and function [26]. In line with this argument, Zhang et al. compared placenta samples in pregnancies obtained with ART and in normally conceived pregnancies, and have reported that, in ART pregnancies, different genes are expressed that are closely related to critical placental functions including immune response, metabolism, transmembrane transport, cell differentiation, and oxidative stress [27]. Also, Sacha et al. compared placental pathology in naturally conceived pregnancies to that in ART pregnancies and stated that especially anatomical and vascular placental pathology was more common in ART pregnancies [28]. The same researchers have reported that placental pathology was more common in the ICSI group, however, they did not provide a clear explanation for this result [28]. In light of all these results, it can be argued that there is a problem in the immune response due to the deficiency or functional deficiency of regulatory T cells in ART pregnancies, and this may affect the placentation and eventually manifest itself with a restriction in fetal development, including the thymus.

The retrospective design and perinatal outcomes that were not investigated can be considered as limitations of this study. However, the present study has advantages including having similar demographic characteristics of the groups and the fact that measurements were made by a single expert. Also, measuring the transverse dimension of the fetal thymus apart from the anteroposterior dimension provided more accurate data on the size of the fetal thymus.

In conclusion, the present study determined that the size of the fetal thymus, manifested by a decrease in both fetal thymic-thoracic ratio and thymus transverse diameter, decreased in ART pregnancies. Further studies are needed to assess the relationship between the measurement of decreased fetal thymus size and negative pregnancy outcomes in ART pregnancies.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

The author(s) reported there is no funding associated with the work featured in this article.