Correlation between clinical and laboratory parameters and early pregnancy loss in assisted reproductive technology cycles: A cross-sectional study

Abstract Background The miscarriage rate after pregnancy resulting from assisted reproductive technology (ART) is about 20%, roughly half of which is biochemical. The correlations between the number and quality of oocytes, estradiol level and early pregnancy loss have not been fully clarified. Objective This study aimed to examine the clinical and laboratory parameter effects on early abortion in ART cycles. Materials and Methods In this cross-sectional study, 408 women who were ART candidates and were referred to the Yazd Infertility and Research Center, Yazd, Iran during March 2017 to March 2020 participated. Women who had a fresh embryo transferred and who had a positive beta human chorionic gonadotropin serum test were included in the study. The Anti-Müllerian hormone (AMH) level, embryo quality, oocyte number, progesterone level, estradiol level, and maternal age were extracted from the medical records. Results No significant difference was observed in the age, mean estradiol and progesterone levels on trigger day, number of MII oocytes, and embryo quality between the groups (p = 0.19, 0.42, 0.07, 0.34 and 0.20, respectively). No statistically significant difference was found between the 3 groups of AMH level (p = 0.20). After evaluation using logistic regression, the rate of negative clinical pregnancies was higher in the group with AMH < 1 ng/ml. However, this was not found to be statistically significant. Conclusion We did not find any correlation between early abortion and AMH levels, embryo quality, oocyte number, progesterone level, estradiol level, or maternal age.


Introduction
Assisted reproductive techniques (ART) is an important treatment in many infertile couples to get pregnant. The ART pregnancy rate is about 40%, but the rate of delivery is still 20-30%, and 1 of the main reasons is miscarriage (1). The incidence of early pregnancy loss in patients receiving fresh embryos in ART is 14.9% (2) and is higher than the miscarriage incidence after natural conception (3). In natural conception many abortions occur before clinical pregnancy detection and a number of miscarriages that occur following spontaneous pregnancies may not be diagnosed at all. In contrast, after pregnancy following ART, most women have an accurate diagnosis and follow up of pregnancy (4). Therefore, these women who become pregnant through ART are the ideal population in which to study the risk factors for early abortion.
One of the main reasons for the higher incidence of early abortion among women who become pregnant through ART is that these women are often older prime parous women in comparison with women who get naturally pregnant (4,5). Based on previous research, we know that the clinical pregnancy rate is significantly associated with the age of donors (3) and biological age is more crucial than chronological age for childbearing (6).
Ovarian reserve and quality of oocytes significantly decrease with aging. In addition, chromosomal abnormalities increase with ovarian reserve decline and advanced maternal age (7).
Therefore, low levels of Anti-Müllerian hormone (AMH) may be associated with early abortion, poor pregnancy outcome, as well as increased pregnancy comorbidity such as preeclampsia (8)(9)(10)(11). Moreover, serum AMH level may affect in vitro fertilization outcomes (12), but AMH cannot be used to predict the live birth rate (13).
Apart from maternal age and AMH, factors such as the levels of estradiol and progesterone on trigger day, number of retrieved oocytes, number or type of embryos transferred, and ovarian stimulation protocol may also be associated with spontaneous miscarriage (14)(15)(16)(17)(18).
The overall sample sizes of previous studies have been small, and the mentioned factors interact with each other, so there is still no consensus on the effects of these factors.
The purpose of this study was to evaluate various factors that may affect early spontaneous abortion after ART treatment.

Therapeutic protocol
Ovarian stimulation was done according to the agonist, antagonist, or microdose protocol (19).

Ethical considerations
The study protocol was approved by

Statistical analysis
The SPSS software (Statistical Package for the Social Sciences, version 26, Chicago, IL, USA) was used to perform all the statistical analyses. The baseline characteristics were compared between the groups using the student's t test for continuous data and the Chi-square test for categorical data.
A p-value < 0.05 was considered statistically significant. In addition, logistic regression was used for estimating the effect of each variable on pregnancy outcomes.

Results
In this study, 60 women out of 408 had negative clinical pregnancies and were categorized as    (20); these were not transferred.
Considering AMH > 2 ng/ml as the reference group, the odds ratio of the AMH < 1 ng/ml group was estimated to be 1.59 (95% confidence interval = 0.87-2.91) based on logistic regression, which means the chance of early abortion was higher in the AMH < 1 ng/ml group.

Discussion
This study was conducted with 408 women The results showed no significant difference between the positive clinical pregnancy and early abortion groups in terms of the mean age range (p = 0.19). This is inconsistent with the results of 1 study which indicated that maternal age was a predictor of early abortion (5).
Moreover, the estradiol levels on the trigger day were compared between the positive clinical pregnancy group and the early abortion group.
The mean estradiol level in the positive clinical pregnancy group was 1418.38 ± 920.36 pg/ml, and in the early abortion group was 1317.77 ± 680.14 pg/ml. No statistically significant difference was found between the 2 groups (p = 0.42). These results are similar to a study which showed that serum estradiol levels above the 90 th percentile on the trigger day using HCG were associated with a lower fertilization rate, but this estradiol level did not affect embryo growth, implantation, clinical pregnancy, or the early abortion rate (14).
The number of MII oocytes was another factor investigated for its importance in clinical pregnancy, but there was no significant difference between the 2 groups of positive clinical pregnancy and early abortion (p = 0.34). In a previous study it was found that an increased number of oocytes was associated with a reduced rate of clinical abortion, and patients with poor ovarian response and fewer oocytes were at a higher risk for clinical abortion (15), but early abortion was not related to the number of oocytes.
In the present study, embryo quality was assessed as another factor in early abortion.
The women were divided into 3 groups (A, B and C), based on the embryo quality score. was not a prognostic factor in early abortion and clinical pregnancy. After evaluation using logistic regression, the rate of negative clinical pregnancies was found to be higher in the group with AMH < 1 ng/ml. However, this was not statistically significant. Contrary to our results, 1 study has shown a relationship between low AMH and early abortion (8). But another study on the relationship between AMH level and early abortion found that lower AMH levels were not associated with a higher rate of early abortion (22).
The present study showed that the mean progesterone level on trigger day using HCG was 0.43 ± 0.56 ng/ml in the early abortion group and 0.55 ± 0.45 ng/ml in the positive clinical pregnancy group, but no statistically significant difference was found between the 2 groups (p = 0.07). Our results are similar to other studies which also showed that higher progesterone levels were not associated with spontaneous abortion (16,18).
According to our study, prognostic factors that affect ART outcomes are not necessarily prognostic of the abortion rate. We found no correlation between early abortion and AMH levels, embryo quality, oocyte number, progesterone level, estradiol level, or maternal age.

Conclusion
We did not find any correlation between early abortion and AMH levels, embryo quality, oocyte number, progesterone level, estradiol level, or maternal age.