Association between endometrial thickness and neonatal outcomes in intrauterine insemination cycles: a retrospective analysis of 1,016 live-born singletons

Intrauterine insemination (IUI) has been widely applied for infertility treatment with the reported clinical pregnancy rate varying from 5 to 20% [1‐3]. However, compared with spontaneously conceived children, singletons born after IUI have shown increased risks of adverse perinatal outcomes including preterm birth (PTB), low birthweight (LBW) and small-for-gestational age (SGA) [4‐7]. The major concern is whether these poorer outcomes are associated with the IUI treatment per se or intrinsic parental characteristics related to infertility [6, 8]. Understanding the relative contribution of the multifarious factors would be essential for assessing the safety of assisted reproductive treatment and promoting its continuous advancements in clinical practice.

Adequate endometrial development is of paramount importance not only for early embryo development and implantation, but also for placentation and fetal growth [9‐11]. As the most commonly used indicator for endometrial receptivity, endometrial thickness (EMT) measurement has become part of standard monitoring during infertility treatment due to its simple and non-invasive approach through transvaginal ultrasonography (TVU) [12, 13]. A thin endometrium, either in fresh or frozen-thawed embryo transfer (FET) cycles, is associated with a lower chance to achieve pregnancy [12, 13]. More recently, a growing body of evidence has also suggested the possible relationship between decreased EMT and reduced birthweight of in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) newborns [11, 14‐17].

Contrary to the suboptimal endometrial environment caused by supraphysiologic hormonal milieu during IVF/ICSI cycles [18, 19], ovarian stimulation is milder in IUI treatment with fewer mature follicles and lower estradiol (E₂) production. In addition, considering the potential effects of embryo culture medium, cryopreservation method and other laboratory procedures in IVF/ICSI [6, 8, 20, 21], a study on neonatal outcomes after IUI may be more suitable to investigate the independent influence of parental factors such as EMT. Indeed, results on EMT and pregnancy chances following IUI treatment have been controversial, with two recent meta-analyses concluding no evidence for an association [13, 22]. However, it is still unclear whether EMT has any impact on IUI infant outcomes as in IVF/ICSI cycles.

The aim of the present study was to comprehensively evaluate the association between EMT and neonatal outcomes of live-born singletons in IUI cycles.

A total of 1266 singleton livebirths resulting from IUI were screened from our database and 250 cycles were excluded as detailed in Fig. 1. Of the remaining 1016 cycles, thin endometrium (EMT ≤7.6 mm) was observed in 109 (10.7%) women, while the number of patients with EMT 7.7–13.0 mm and ≥ 13.1 mm was 798 (78.5%) and 109 (10.7%), respectively.

Baseline characteristics according to EMT stratification were shown in Table 1. There was a significant increase from EMT ≤7.6 mm to EMT ≥13.1 mm group in the proportion of nulligravida, length of treatment and peak E₂ level (all P < 0.001). No significant differences were found when parental age and BMI, parity, duration of infertility, infertility diagnosis, rank of IUI attempts, stimulation protocol, postprocessing TMSC, incidence of VTS and pregnancy complications, as well as year of treatment were compared among the three groups. With a range between 4.1 and 20.6 mm, the mean thickness of endometrium was 6.60 ± 0.96, 10.06 ± 1.43 and 14.42 ± 1.27 mm in the EMT ≤7.6, 7.7–13.0 and ≥ 13.1 mm groups, respectively (P < 0.001). The EMT in the LE group was slightly thinner than that in the LE + hMG group (9.68 ± 2.30 vs. 10.21 ± 2.28 mm, P = 0.016) (Supplementary Figure S1).

As presented in Table 2, comparisons among the three groups did not reveal any significant difference in gestational age, birthweight and birthweight Z-score (P = 0.503, 0.732 and 0.914, respectively). These findings were not altered in multivariable analyses after controlling for a variety of confounding factors (Table 3). Compared with the EMT 7.7–13.0 mm group, the incidences of PTB, LBW and SGA were 5.5% (adjusted odds ratio [aOR] 0.81, 95% confidence interval [CI] 0.33–2.01), 6.4% (aOR 1.44, 95% CI 0.58–3.58) and 7.3% (aOR 1.21, 95% CI 0.53–2.76) in the EMT ≤7.6 mm group, respectively. Similarly, EMT ≥13.1 mm was not significantly associated with risks of PTB (aOR 0.63, 95% CI 0.24–1.65), LBW (aOR 0.57, 95% CI 0.17–1.95) and SGA (aOR 0.73, 95% CI 0.28–1.92). With regard to other adverse neonatal outcomes including macrosomia, LGA and major congenital malformations, no significant differences were found in both EMT ≤7.6 and ≥ 13.1 mm groups before and after adjustment.

Multiple regression analyses of risk factors for PTB, LBW and SGA were detailed in Supplementary Table S1. Pregnancy complications were found to be the common influencing factor, while higher risks of PTB and LBW were observed in women with VTS. Duration of infertility was demonstrated to be associated with an increased risk of SGA (aOR 1.17, 95% CI 1.03–1.32).

In this retrospective analysis of 1016 live-born singletons, we provided the first evidence that EMT was not associated with adverse perinatal outcomes in IUI cycles. Neither a thin or thickened endometrial lining (EMT ≤7.6 or ≥ 13.1 mm) at trigger day would increase the infant risks of PTB, LBW, SGA and major congenital anomalies.

Despite significant progress in ultrasonographic, immunological and molecular diagnostic techniques of endometrial receptivity, the EMT still remains to be the mainly used marker as part of standard cycle monitoring in assisted reproductive treatment [12, 13]. Over the past decades, much efforts have been made to explore its prognostic capacity in pregnancy chances. While it has been well-established that the probability of clinical pregnancy was lower in IVF/ICSI patients with thin EMT defined by different cut-off values from 6 to 10 mm [12], only few studies have investigated this issue in the setting of IUI treatment with conflicting results [13, 22]. The pooled data revealed no significant difference in mean EMT between pregnant and non-pregnant IUI women, and no difference in clinical pregnancy was observed after association analysis using EMT cut-offs ranging from 3 to 12 mm [13, 22]. Although the discrepancy remains unclarified, it does highlight the necessity to separately assess the influence of EMT on reproductive outcomes between IUI and IVF/ICSI treatment.

The relationship between EMT and neonatal morbidity in IVF/ICSI cycles has been evaluated in several prior studies [11, 14‐17]. In 2006, Chung et al. [14] first reported that patients with an EMT ≤10 mm had a two-fold higher risk of experiencing PTB, LBW or intrauterine fatal demise beyond first trimester than those with an EMT > 12 mm. This study, however, was flawed by a high proportion of multiple gestations (37%) out of 435 viable pregnancies. By retrospectively analyzing 764 singleton deliveries, a later study by Moffat et al. [15] showed that EMT was strongly predictive for neonatal birthweight in pregnancies with obstetric complications but not in uneventful pregnancies. Consistently, Ribeiro and colleagues [16] found that an EMT less than 7 mm was significantly associated with a decrease in neonatal birthweight Z-score. However, they did not detect an increase in clinically relevant outcomes such as PTB or LBW, and attributed it to the limited size of the live-born singleton sample subset. This was subsequently validated in another study by Oron et al. [11] showing that women with an EMT < 7.5 mm had a higher incidence of placenta-related pregnancy complications including SGA. In addition to the aforementioned four studies on fresh embryo transfer cycles, a recent analysis of 6181 singletons resulting from FET also demonstrated that a thin endometrium (EMT < 8 mm) was independently related to a lower birthweight and Z-score, while the modest difference failed to translate into a significantly higher risk of SGA [17].

Similar to the contradictory findings on pregnancy outcomes, the present study reported no evidence for an association between EMT and IUI infant outcomes as in IVF/ICSI cycles. However, the underlying reasons are still unclear as to why a thin endometrial lining is more clinically relevant in IVF/ICSI than in IUI treatment. During either natural or stimulated cycles, continuous E₂ production from growing follicles induces endometrial proliferation, thus leading to increased EMT as measured by TVU. Compared with a thin endometrium developing under mild ovarian stimulation conditions as in IUI, a thin endometrium developing under maximal stimulation conditions in IVF/ICSI cycles may imply genuine diminished potential for implantation and placentation. Moreover, in the presence of thin endometrium, the embryos are implanted much closer to the spiral arteries of the endometrial basal layer, whose higher vascularity and oxygen concentrations could be detrimental to embryos due to the production of reactive oxygen species [28, 29]. Since embryos are developed in vivo in IUI cycles, we speculate that they may be less susceptible to high oxygen tensions in comparison with in vitro cultured or further vitrified/thawed embryos in IVF laboratories. Nevertheless, these explanations are merely theoretical and further investigations are warranted to provide stronger and more direct evidence.

VTS, defined as the spontaneous reduction of multiple pregnancies to singleton birth [30], was identified as an independent predictor of adverse neonatal outcomes after multivariable regression analysis. This phenomenon has been estimated to occur in 50% of pregnancies with initially three or more gestational sacs, 36% of twin pregnancies and 10–30% of IVF/ICSI pregnancies [31, 32]. In the present study, we reported for the first time that the prevalence of VTS was about 2.1% in IUI treatment, much lower than that in IVF/ICSI cycles. Consistent with previous findings on IVF/ICSI infants [30, 33, 34], VTS is associated with higher risks of PTB and LBW in IUI newborns as well. However, the adjusted odds of SGA and birth defects did not reach statistical significance possibly due to the limitation of relatively small sample size, as reflected by the wide confidence intervals. Therefore, more studies with larger datasets are needed to further assess the incidence, risk factors as well as neonatal outcomes of VTS in IUI cycles.

A major weakness of the study relies on its retrospective design. Some baseline characteristics were not balanced between the stratified groups. Notably, the fact that more patients with thin EMT had been pregnant before but the frequency of nulliparity was similar among the three groups suggested a higher rate of prior pregnancy loss, which was associated with increased chances of adverse neonatal outcomes such as PTB and LBW [35]. While the present study included gravidity and parity number for adjustment, we were unable to retrieve data on the actual causes of pregnancy loss history (e.g., miscarriage, abortion or ectopic pregnancy). In addition, possible unknown or unavailable confounders may not be accounted for in the regression model. For example, over 50% of singletons were delivered via cesarean section, but the indications and timing in relation to neonatal mortality remained unclear [36, 37]. Also, information on endometrial morphology, which has been reported to influence pregnancy outcomes [13], was incomplete in our database and thus not analyzed in this study. Other risk factors for adverse neonatal outcomes, including previous delivery history, medication use, metabolic status, nutrition intake and lifestyle habit [38], were not adequately controlled due to the lack of detailed records. Therefore, future studies with a comprehensive evaluation of women before and during pregnancy are needed for further investigation. Secondly, previous studies have observed significant differences in EMT among various ovarian stimulation protocols and in neonatal outcomes of different sperm sources [7, 22, 23, 39]. To minimize the likelihood of confounding, only IUI cycles using LE and partner semen were included for analysis. Thus, our conclusion should not be extrapolated to IUI singletons born after treatment with donor semen or with clomiphene citrate, gonadotropin only and natural cycles. The single-center nature should also caution generalization of the study finding to other areas in China and other parts of the world. Finally, while the measurement of EMT was performed by highly experienced doctors at our center, some inter-observer inconsistency may still be present. Moreover, most of the neonatal data were collected from parental questionnaires instead of direct access to medical records. In this regard, the detection of some minor problems could be comprised and the actual incidence of congenital malformations may be underestimated.

In summary, our data suggested no association between EMT and adverse neonatal outcomes in IUI cycles. This novel finding would provide reassuring information for patients undergoing IUI treatment with thin endometrial linings regarding their neonatal health. However, further large prospective cohort studies with longer follow-up duration are needed to confirm the conclusion.