Low Peak Estradiol Levels Associated with Increased Miscarriage and Decreased Live Birth in Non-Letrozole IUI cycles: A Multi-center Retrospective Cohort Study

Low Peak Estradiol Levels Associated with Increased Miscarriage and Decreased Live Birth in Non-Letrozole IUI cycles: A Multi-center Retrospective Cohort Study | Authorea try { document.documentElement.classList.add('js'); } catch (e) { } var _gaq = _gaq || []; _gaq.push(['_setAccount', 'G-8VDV14Y67G']); _gaq.push(['_trackPageview']); (function() { var ga = document.createElement('script'); ga.type = 'text/javascript'; ga.async = true; ga.src = ('https:' == document.location.protocol ? 'https://ssl' : 'http://www') + '.google-analytics.com/ga.js'; var s = document.getElementsByTagName('script')[0]; s.parentNode.insertBefore(ga, s); })(); Skip to main content Preprints Collections Wiley Open Research IET Open Research Ecological Society of Japan All Collections About About Authorea FAQs Contact Us Quick Search anywhere Search for preprint articles, keywords, etc. Search Search ADVANCED SEARCH SCROLL This is a preprint and has not been peer reviewed. Data may be preliminary. 10 February 2025 V1 Latest version Share on Low Peak Estradiol Levels Associated with Increased Miscarriage and Decreased Live Birth in Non-Letrozole IUI cycles: A Multi-center Retrospective Cohort Study Authors : Kexin FAN 0009-0006-4942-4869 , Xinru GU , Aniguli SHABIER , Jiaqi XIAO , Kai XU , Yilin CHEN , Yang Fu , Chunyan An , Zhi LIU , and Mingzhu Cao [email protected] Authors Info & Affiliations https://doi.org/10.22541/au.173917810.07488353/v1 357 views 183 downloads Contents Abstract Information & Authors Metrics & Citations View Options References Figures Tables Media Share Abstract Objectives : To evaluate the relationship between peak estradiol levels on the ovulation trigger day and reproductive outcomes in IUI cycles. Design : A retrospective cohort study. Setting : China. Sample : The cohort consisted of 7525 IUI cycles during the period 2019-2023. Methods: We used data from three institutions. Outcomes included live birth, clinical pregnancy, miscarriage, ectopic pregnancy, multiple pregnancy, preterm birth and neonatal growth parameters. Statistical analyses, including multivariate stepwise regression and mediation analysis, were conducted to evaluate estradiol’s impact on outcomes. Main Outcome Measures : The miscarriage rate and live birth rate with low peak estradiol levels. Results: The highest estradiol quartile exhibited higher live birth (13.5%, P < 0.001) and clinical pregnancy rates (17.3%, P = 0.004). Conversely, the lowest estradiol quartile had increased miscarriage risk (OR = 2.15, P = 0.012) and reduced live birth rates (OR = 0.60, P = 0.070). Specifically, non-letrozole cycles increased miscarriage risk (OR = 4.30, P = 0.005) and decreased live birth rates (OR = 0.31, P = 0.010), whereas no significant impact was observed in letrozole cycles. Estradiol significantly mediated the effect of ovulation stimulation type on reproductive outcomes, with a suppression effect observed (PM = 180.5%). Funding : Medical Research Fund of Guangdong Province (A2024003), and Xinjiang Support Rural Science and Technology Program in Guangdong Province (KTPYJ 2023014). Conclusions: Higher estradiol levels on the ovulation trigger day enhance live birth and clinical pregnancy rates. Low estradiol levels, especially in non-letrozole cycles, are associated with higher miscarriage risks and poorer reproductive outcomes. Running Title: Low Peak Estradiol Increase Miscarriage and Decrease Live Birth Article Title: Low Peak Estradiol Levels Associated with Increased Miscarriage and Decreased Live Birth in Non-Letrozole IUI cycles: A Multi-center Retrospective Cohort Study Kexin FAN 1# , Xinru GU 2# , Aniguli SHABIER 3# , Jiaqi XIAO 4# , Kai XU 5# , Yilin CHEN 6 , Yang FU 7 , Chunyan AN 7 , Zhi LIU 8* , Mingzhu CAO 7* 1. Nanfang Hospital, Southern Medical University/The First School of Clinical Medicine, Southern Medical University, Guangzhou, Guangdong Province, China. 2. Reproductive Center, Maoming People’s Hospital, Maoming, Guangdong Province, China. 3. Reproductive Center, People’s Hospital of Kashgar Prefecture, Xinjiang Uygur Autonomous Region, China. 4. Department of Clinical Medicine, The Nanshan College of Guangzhou Medical University, Guangzhou, Guangdong Province, China. 5. Department of Thoracic Surgery, West China Hospital, Sichuan University, Chengdu, Sichuan, China; Lung Cancer Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China 6. School of Computer Science, The University of Sydney, NSW 2006, Australia 7. Department of Obstetrics and Gynecology, Reproductive Medicine Center; Guangdong Provincial Key Laboratory of Major Obstetric Diseases; Guangdong Provincial Clinical Research Center for Obstetrics and Gynecology; Guangdong-Hong Kong-Macao Greater Bay Area Higher Education Joint Laboratory of Maternal-Fetal Medicine; The Third Affiliated Hospital, Guangzhou Medical University, Guangzhou, Guangdong Province, China. 8. Department of Ultrasound, Nanfang Hospital, Southern Medical University # The authors consider that the first five authors should be regarded as joint First Authors *Corresponding authors: 1. Author Name: Dr. Mingzhu Cao; Academic Affiliation: The Third Affiliated Hospital, Guangzhou Medical University, Guangzhou, Guangdong Province, China; Mailing Address: No. 63, Duobao Road, Liwan District, Guangzhou, Guangdong Province, China; Email: [email protected] ; Phone: 020-81292233; ORCID: 0000-0003-1329-4215. 2. Author Name: Dr. Zhi Liu; Academic Affiliation: Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong Province, China; Mailing Address: No. 1838, Guangzhou Avenue North, Baiyun District, Guangzhou, Guangdong Province, China; Email: [email protected] ; Phone: 020-61642100; ORCID: 0000-0002-8464-3235. Abstract Objectives : To evaluate the relationship between peak estradiol levels on the ovulation trigger day and reproductive outcomes in IUI cycles. Design : A retrospective cohort study. Setting : China. Sample : The cohort consisted of 7525 IUI cycles during the period 2019-2023. Methods: We used data from three institutions. Outcomes included live birth, clinical pregnancy, miscarriage, ectopic pregnancy, multiple pregnancy, preterm birth and neonatal growth parameters. Statistical analyses, including multivariate stepwise regression and mediation analysis, were conducted to evaluate estradiol’s impact on outcomes. Main Outcome Measures : The miscarriage rate and live birth rate with low peak estradiol levels. Results: The highest estradiol quartile exhibited higher live birth (13.5%, P < 0.001) and clinical pregnancy rates (17.3%, P = 0.004). Conversely, the lowest estradiol quartile had increased miscarriage risk (OR = 2.15, P = 0.012) and reduced live birth rates (OR = 0.60, P = 0.070). Specifically, non-letrozole cycles increased miscarriage risk (OR = 4.30, P = 0.005) and decreased live birth rates (OR = 0.31, P = 0.010), whereas no significant impact was observed in letrozole cycles. Estradiol significantly mediated the effect of ovulation stimulation type on reproductive outcomes, with a suppression effect observed (PM = 180.5%). Funding : Medical Research Fund of Guangdong Province (A2024003), and Xinjiang Support Rural Science and Technology Program in Guangdong Province (KTPYJ 2023014). Conclusions: Higher estradiol levels on the ovulation trigger day enhance live birth and clinical pregnancy rates. Low estradiol levels, especially in non-letrozole cycles, are associated with higher miscarriage risks and poorer reproductive outcomes. Keywords Estradiol levels, intrauterine insemination, live birth rate, letrozole Introduction Infertility affects 8 to 12% of couples globally, posing a significant public health issue(1). Advances in assisted reproduction have improved access to infertility treatments, with intrauterine insemination (IUI) commonly recommended as a first-line minimally invasive, and cost-effective option(2). IUI involves the direct deposition of sperm into the intrauterine cavity(3). This procedure is effective for mild male factors, ovulatory dysfunction, and unexplained infertility, though its efficacy is reduced in women with tubal factor infertility or advanced endometriosis(4). Clinical pregnancy rates following IUI are similar to natural conception, ranging from 10-15% per cycle(5-8). IUI success largely relies on follicle development and sperm quality(9). During an IUI cycle, ovarian stimulation can be achieved through aromatase inhibitors (letrozole, LE), clomiphene citrate (CC), or gonadotropins (Gn), which stimulate follicle growth(10). Follicle development as detected by sonography and serum steroid hormone levels were closely monitored to ensure the presence of mature follicle(s). Among these hormones, estradiol (E2), produced by granulosa cells, plays a crucial role in follicle development and peaks on the ovulation trigger day(11, 12). Hence, monitoring E2 levels during ovarian stimulation is essential, as rising E2 levels signify follicle growth(13). While adequate E2 levels are essential for optimal reproductive outcomes, supra-physiological levels of E2 might adversely affect reproductive outcomes by compromising endometrial receptivity and reducing embryo quality(14). Additionally, elevated E2 levels are also associated with an increased risk of ovarian hyperstimulation syndrome and hormonal imbalances that can hinder successful conception(15). Existing evidence have mainly focused on the impact of high E2 levels, especially in embryo transfer cycles, on reproductive outcomes. Conversely, the clinical implications of low E2 levels on ovulation trigger day or peak E2 levels in IUI cycles remain unclear. Limited evidence suggests that higher peak E2 levels may improve conception rates in IUI cycles, although the precise mechanism are not fully understood(16). Nonetheless, the impact of low peak E2 levels, particularly with LE, which induce low E2 production, on reproductive outcomes during IUI treatment has not been well illustrated. To address this gap, we performed a retrospective study encompassing clinical data from three institutions. The primary purpose of this study was to evaluate the association between diverse peak E2 levels on ovulation trigger day and reproductive outcomes in IUI treatment. By elucidating the role of peak E2 in IUI success, this study aims to provide insights to refine ovarian stimulation protocols and enhance the efficacy of IUI as a fertility treatment. Data source This is a multi-center retrospective cohort study including 7525 IUI cycles from January 2019 to June 2023, with data from The Third Affiliated Hospital of Guangzhou Medical University (accounting for 6396 cycles), The People’s Hospital of Maoming (1002 cycles), and The People’s Hospital of Kashgar Prefecture (127 cycles). The institutional review board approved the study protocol (approval number 2024-153), and all participants had provided written informed consent. Inclusion and exclusion criteria of participants The inclusion criteria of the study were as follows, 1) IUI cycles performed from January 2019 to June 2023, 2) cycles with serum E2 levels tested on the day of luteinizing hormone (LH) surge or human chorionic hormone (HCG) injection for ovulation trigger, 3) cycles with confirmed follow-up results pertaining to live birth, 4) female participants aged from 20 to 40 years old. Cycles met any of the following criteria were excluded from the analysis, 1) participants with abnormal endometrial cavity (including endometrial polyps, submucosal myoma, intrauterine adhesion, etc., 2) untreated hydrosalpinx, or 3) participants with severe systematic diseases including uncontrolled diabetes, hypertension, or systemic lupus. Follow-up data were collected through hospital records, with confirmation of live birth outcomes via medical discharge summaries or clinic follow-up visits. Outcomes measured The primary outcome assessed in the study was the live birth rate (LBR) per cycle. Live births were defined as delivery of a live newborn at least completing gestational 28 weeks. Twins or triplets delivered by one mother was counted as a single live birth. The secondary outcomes included clinical pregnancy rate, spontaneous miscarriage rate, ectopic pregnancy rate, multiple pregnancy rate and preterm birth rate. Clinical pregnancy was defined as detection of intrauterine gestational sac at gestational 6 to 7 weeks. Early miscarriage was defined as miscarriage occurred before gestational 14 weeks, and late miscarriage was determined as spontaneous miscarriage occurred between 14 +1 and 27 +6 gestational weeks. Ectopic pregnancy was diagnosed upon the detection of gestational sac outside of uterine cavity. Multiple pregnancy refers to those with multiple fetus detected with sonography at gestational 6 to 7 weeks. Preterm birth was defined a delivery of a live newborn between gestational 28 to 36 +6 weeks. The rates of clinical pregnancy, spontaneous miscarriage, ectopic pregnancy and live birth were calculated as the proportions of above-mentioned outcomes out of all IUI cycles in each group. The rates of preterm birth, fetal malformation and multiple pregnancy were calculated as the proportions of those outcomes out of cycles with clinical pregnancies. Birth weight and height were measured and recorded from newborns’ birth medical record. Statistical analysis Descriptive statistics were demonstrated as mean with standard deviation (SD) for normally distributed continuous variables, and median (25 th and 75 th percentile) for those not following normal distribution. Categorical variables were expressed as case numbers and percentages. Statistical analyses were performed using R 4.2.1 software (R Foundation for Statistical Computing, Vienna, Austria). Data were compared using Student’s t test and the chi-square test, as appropriate. In this cohort, cycles were grouped based on peak E2 levels on ovulation trigger day at the 25 th , 50 th , 75 th , and above the 75 th percentiles. Multivariate stepwise regression analyses were performed to identify the independent effect of various E2 levels on reproductive outcomes, adjusting for potential confounding variables including maternal age, BMI, infertility factors, endometrial thickness on trigger day, number of dominant follicles, and the type of ovulation stimulation used. For variables with significant E2 effects, a stepwise regression approach was employed, while for those without significant E2 effects, standard multivariate regression was applied. Missing data were dealt with multiple imputation. Mediation analysis was conducted to assess the role of E2 in mediating the effects of number of dominant follicles, type of ovulation stimulation, and female age on reproductive outcomes. Additional subgroup analyses were performed on cycles using LE, either alone or in combination with Gn, as well as on cycles without LE for ovulation induction. Cycles in both subgroup analyses were stratified similarly as described above. Reproductive outcomes were then compared across these groups and validated through multivariate logistic regression analysis. Results Over the study period, we identified 7,525 IUI cycles, of which 5,980 IUI cycles meeting the inclusion and exclusion criteria for our analysis. The detailed inclusion and exclusion of cycles for analysis were listed in Figure 1. Notably, the mean peak E2 levels on ovulation trigger day was 339.85 pg/mL. All cycles were categorized into four distinct groups based on their peak E2 levels (supplemental Figure 1): Group I (E2 levels below 25 th percentile, ≤ 196.46 pg/mL), Group Ⅱ (E2 levels between 25 th and 50 th percentile, > 196.46 and ≤ 298.64 pg/mL), Group Ⅲ (E2 levels between 50 th and 75 th percentile, > 298.64 and ≤ 426.43 pg/mL) and Group Ⅳ (E2 levels above 75 th percentile, > 426.43 pg/mL). The slight difference in the number of cases in Group Ⅳ (N = 1492) is due to rounding when categorizing values at the quartile boundaries. Specifically, cases with values exactly on the quartile cut-off were assigned to the preceding quartile. Table 1 shows the overview of baseline and treatment cycle related characteristics across various groups. No differences were detected of female participants’ age, duration of infertility, type of infertility and basal FSH. Group Ⅲ exhibited a higher proportion of AID cycles (23.33%), and AIH was prevalent in Group I, Ⅱ and Ⅳ. A notable disparity was found in the proportion of cycles attributed to male factors, which was markedly higher in Group Ⅲ (26.87%) compared to other groups, whereas ovulation disorder was more frequently encountered in Group I (15.57%). Participants’ body mass index (BMI) varied notably ( P < 0.001), with a slightly higher BMI observed in the lower E2 groups. Although the mean number of AFC, AMH levels as well as basal FSH levels shows statistically significantly differences, the differences hold minimal clinical significance. Almost half of all groups constituted the first IUI attempt, and roughly one-third of all groups encompassed the second cycle. The type of ovarian stimulation also differed significantly ( P < 0.001), with LE being more prevalent in the lower E2 quartiles, whereas Gn is more common in the higher quartiles. The number of dominant follicles, defined as those with an average diameter of ≥14 mm, differed substantially among the groups ( P < 0.001). The presence of only one dominant follicle was most prevalent in Group Ⅱ and Ⅲ. In Group IV, the presence of two or more dominant follicles was more common than in other groups. Serum levels and LH levels on ovulation trigger day showed minimal differences among all groups. Endometrial thickness on ovulation trigger day slightly increased from Group I to Group IV and peaking in Group Ⅲ (9.08 ±1.89 mm). Progressive motile sperms among all groups showed no apparent variation. As illustrated in Table 2, elevated E2 levels (>75%) in Group IV were associated with the highest live birth rate (13.54%, P < 0.001), clinical pregnancy rate (17.29%, P = 0.004), and multiple birth rate (9.30%, P = 0.001). However, there were no significant differences observed in spontaneous miscarriage rates ( P = 0.555), preterm birth rate ( P = 0.826), ectopic pregnancy rates ( P = 0.875), fetal malformation rates ( P = 0.856), newborns’ weight ( P = 0.414), or newborns’ height ( P = 0.932) Multivariate logistic regression analysis (Table 3) was performed to assess the independent impacts of varying peak E2 levels on several reproductive outcomes. Group II, representing E2 levels closest to physiological ranges, was designated as the reference group. Higher E2 levels (>75 th percentile) in Group Ⅳ were associated with a significantly increased likelihood of clinical pregnancy (OR = 1.32, 95% CI: 1.05, 1.68, P = 0.020), and slightly increased live birth rate but no statistical significance (OR =1.24, 95% CI: 0.70, 2.17, P = 0.500). The lowest E2 levels in group I showed minimal impact on clinical pregnancy rate (OR = 0.85, 95% CI: 0.66, 1.08, P = 0.200) and live birth rate (OR = 0.60, 95% CI: 0.34, 1.04, P = 0.070), but were associated with a significantly higher risk of spontaneous miscarriage (OR = 2.15, 95% CI: 1.19, 3.97, P = 0.012). The birth weight and height of the newborns were not influenced by varying levels of E2. The mediation analysis (Table 4) revealed a strong mediating role of E2 in the effect of ovulation stimulation type on reproductive outcomes. The indirect effect (ACME = 0.011) was positive, while the direct effect (ADE = -0.005) was negative, resulting in a total effect of 0.006. Notably, the proportion mediated was 180.5%, indicating a suppression effect, where the indirect pathway through E2 reversed the negative direct effect. Therefore, the further analysis based on ovulation stimulation type, especially cycles involved with or without LE, is necessary. LE, an aromatase inhibitor, could reduce the production of E2 and hence stimulate the FSH pulse through hypothalamic-pituitary-ovarian (HPO) axis(17). In IUI cycles with the involvement of LE, lower levels of E2 were a common clinical observation. However, whether low E2 levels, in particular stimulated with LE, on ovulation trigger day could affect the reproductive outcomes were not clearly illustrated. Here, we conducted two subgroup analyses: one focusing on cycles stimulated with LE alone or in combination with Gn, and the other for cycles without LE. Each subgroup was stratified using the same E2-based categorization as applied in the overall analysis: Group Ⅰ (E2 ≤ 196.46 pg/mL), Group Ⅱ (196.46 < E2 ≤ 298.64 pg/mL), Group Ⅲ (298.64 426.43 pg/mL). Among the cycles, 1,712 used LE alone, 453 combined LE and Gn, and 3,815 cycles did not involve with LE. The logistic regression analyses of the above-mentioned subgroups are presented in Table 5. In cycles with LE alone or in combination with Gn, higher E2 levels in Subgroup IV were associated with increased odds of live birth (OR: 3.51, 95% CI: 1.09, 14.05, P = 0.050), but minimal impact on clinical pregnancy, spontaneous miscarriage and multiple pregnancy rates. The lowest E2 levels in Subgroup I showed no obvious impact on reproductive outcomes. In cycles without the involvement of LE, the lowest E2 levels in subgroup I demonstrated reduced odds of live birth (OR: 0.31, 95% CI: 0.13, 0.76, P = 0.010) compared to the reference group, Subgroup II. While the lowest E2 levels in subgroup I did not significantly affect the clinical pregnancy rate, they were significantly correlated with markedly higher odds of spontaneous miscarriage (OR: 4.30, 95% CI: 1.53, 12.20, P = 0.005). The birth weight and height of the newborns were not differed significantly in various subgroups. Discussion This study aimed to investigate the correlation between peak E2 levels on ovulation trigger day (HCG/LH day) and pregnancy rates in IUI cycles. Our findings provided robust evidence that higher peak E2 levels are associated with enhanced clinical pregnancy rates. In contrast, while not statistically significant, the lowest E2 levels observed in Group I were associated with reductions in both clinical pregnancy and live birth rates. This trend could be attributed to an increased risk of miscarriage associated with the lowest E2 levels in Group I. To be specific, in subgroups of cycles without the involvement of LE, lowest E2 levels, as indicated in Subgroup I, were associated with an increased spontaneous miscarriage rate and consequently, a reduced live birth rate, while the impact on clinical pregnancy rates was minimal. In those cycles with LE alone or in combination of Gn, no such association of lowest E2 levels and reproductive outcomes were found. The findings in the present study suggested that in the absence of LE, insufficient peak E2 levels may be linked with poorer reproductive outcomes, specifically, the increased risk of spontaneous miscarriage and decreased likelihood of live birth. These findings underscore the critical role of E2 in optimizing reproductive outcomes and highlight the need for tailored ovarian stimulation protocols. Clinicians should be aware of the potential impact of low peak E2 levels on reproductive outcomes in non-LE IUI cycles. Monitoring E2 levels during ovarian stimulation and adjusting treatment protocols accordingly may help improve the success rate of IUI procedures. In the present study, we highlighted the importance of independent impact of heightened E2 levels on ovulation trigger day in fostering improved reproductive outcomes in IUI cycles, namely, elevated clinical pregnancy rates. In cycles with embryo transfer following in-vitro fertilization (IVF) and intra-cytoplasmic sperm injection (ICSI), the importance of varying E2 levels has been evaluated in several studies. In fresh embryo transfer cycles, a positive correlation between every increase of E2 at 1 ng/ml and live birth rate was identified (OR: 1.19, 95% CI: 1.057, 1.334) (18). This positive effect of E2 levels in ovarian stimulation cycles could extend to the following frozen embryo transfer cycles, which demonstrated elevated cumulative live birth rate (65.1% vs. 81.6% as in the lowest vs. the highest quartile of E2 levels)(19). However, the clinical value of peak E2 levels was not fully illustrated in IUI cycles. New et al. identified that elevated E2 levels on ovarian trigger day in cycles stimulated with LE could improve live birth rates, with LBRs ranging from 9.4% to 11.1% in the lower E2 cohorts and from 12.5% to 13.5% in the higher E2 cohorts, as the E2 level quartile increased from the 25 th (110 pg/mL) through the 75 th percentile (225 pg/mL) (20). Unlike the present study, New’s data involved with timed intercourse and IUI cycles. Additionally, E2 levels in other ovarian stimulation agents, such as CC and Gn, was not determined in the above-mentioned study. Moreover, the impact of varying peak E2 levels on various pregnancy rate in IUI cycles were not elucidated yet. Here, we provided one of the first evidence to address the research gaps. Our study provided one pioneer evidence to suggest that lower peak E2 levels (in particular, < 25 th and 25 th -50 th quartiles) on ovulation trigger day had minimal impact on reproductive outcomes in IUI cycles. As indicated by the mediation analysis, type of ovulation stimulation could be one critical influencer of peak E2 levels. Given that LE could induce E2 levels lower than physiological levels, cycles involved with LE might induce extremely low levels of E2 on ovulation trigger day. To further confirmed the potential impact of low E2 on reproductive outcomes, we re-analyzed cycles using LE, either alone or in combination with Gn, as well as on cycles without LE for ovulation induction. LE, an aromatase inhibitor, is widely used in IUI cycles to induce ovulation. By inhibiting the aromatase enzyme, LE reduces the conversion of androgens to estrogens, resulting in lower concentration of circulating E2(21). Lower E2 concentration induced by LE are found to be sufficient for successful follicular development and ovulation(22). This reduction in E2 levels can help to avoid the excessive estrogenic environment associated with other ovulation induction agents such as CC(23). The application of LE in IUI cycles has been studied extensively, demonstrating its efficacy in inducing mono-follicular ovulation, thereby reducing the risk of multiple pregnancies compared to other agents(17). While studies on frozen embryo transfer cycles have shown that LE-induced low E2 levels do not compromise endometrial receptivity or implantation rates, there remains a lack of comprehensive research exploring these effects in IUI cycles(24). Clinical outcomes, such as pregnancy rates and live birth rates, are comparable to or even better than those achieved with higher E2 levels induced by other agents following embryo transfer(25, 26). Furthermore, the influence of low E2 levels induced by LE on reproductive outcomes following IUI outcomes has been a subject of interest but not clearly illustrated. This highlights the novelty of our study, which aims to address this gap. In cycles with LE alone or in combination with Gn, low E2 levels (≤ 196.46 pg/ml) showed minimal impact on reproductive outcomes. However, in New and colleagues’ research, extremely low E2 (≤110 pg/ml, ≤ 157 pg/ml) might be related with reduced chance of clinical pregnancy rate and live birth rates in cycles with LE alone(20). This discrepancy might be attribute to the differences in stratification of E2 concentration. Besides, unlike the specific focus on LE in New’s study, the present study examined a broad range of ovulation agents, reflecting a more practical approach aligned with real-world clinical practice. As demonstrated in subgroup analysis, low E2 levels, particularly in cycles without LE, may be associated with reduced reproductive success, as evidenced by higher miscarriage risk and consequently, lower live birth rates. Therefore, the use of LE in IUI cycles effectively induces ovulation at lower E2 levels without adversely impacting reproductive outcomes. The reduced E2 levels in the LE group may due to the medication’s mechanism and do not increase miscarriage risk. In contrast, in the non-LE cycles, naturally low physiological E2 levels may be associated with a higher risk of miscarriage. For those with live birth, varying peak E2 levels showed no significant impact on their newborns’ birth weight and height. E2, a potent form of estrogen, plays a crucial role in various reproductive processes, including follicular development, endometrial receptivity, and the quality of oocytes, all of which are essential for successful conception and pregnancy. During the follicular phase of the menstrual cycle, E2 is primarily produced by the growing follicles in the ovaries. It promotes the proliferation and differentiation of granulosa cells, which are essential for the maturation of the follicle(27, 28). Additionally, E2 prepares the endometrium for implantation by stimulating the proliferation of the endometrial lining. It modulates the expression of various genes and growth factors that enhance the receptivity of the endometrium to the implanting embryo(29). Furthermore, E2 is also involved in the regulation of oocyte quality. Adequate E2 levels are necessary for the proper chromosomal alignment and spindle formation during oocyte meiosis, which are crucial for genetic stability(30). Given its roles in follicular development, endometrial receptivity, and oocyte quality, E2 might significantly modulate pregnancy rates. Fluctuation in E2 levels can lead to suboptimal conditions for ovulation, implantation, and embryo development. However, higher E2 levels are not always the better. For instance, low E2 levels in natural cycles might indicate poor follicular development, and lead to poor endometrial receptivity, while excessively high levels can cause premature luteinization and endometrial desynchrony(31). Joo et al. found that although pregnancy rates increased with elevated E2 levels up to 3000-4000 pg/mL but decreased when E2 levels exceeding 4000 pg/mL(32). Specifically, in the group with E2 levels above 4000 pg/mL, the pregnancy rate dropped to 29.9% from a peak of 50.0% observed in the 3000-4000 pg/mL group(32). Both scenarios can decrease the likelihood of conception and successful pregnancy. However, in IUI cycles, mild ovarian stimulation can hardly result in extremely supra-physiological levels of E2. As observed in the current study, the median value of E2 in group IV was only 559.81 pg/mL, which is still below the optimal level 3000-4000 pg/mL as observed in embryo transfer cycles. Therefore, it is no wonder higher levels of E2 on ovulation trigger day might be indicative of improved reproductive outcomes. Lower E2, on the other hand, might reflect underlying inadequate ovarian stimulation, insufficient follicular development and endometrial receptivity, leading to reduced pregnancy chance and higher miscarriage risk, further compromising the chances of a successful live birth. The findings of this study hold substantial practical implications in IUI treatment. Elevated production of E2 from multi-follicles could be one of the targets of IUI treatment(33). Tailored treatment protocols with proper ovulation stimulation agents including LE could be provided appropriately to achieve optimal levels of E2. More importantly, the present findings highlight the critical role of maintaining adequate E2 levels on ovulation trigger day. Notably, our mediation analysis suggests that E2 plays a compensatory role in mitigating potential negative direct effects of certain stimulation protocols on reproductive outcomes, emphasizing the complex hormonal regulation involved. Therefore, meticulous monitoring of follicle development and E2 levels is necessary to balance the benefits and the risks of high E2 levels. Clinicians should consider this when designing stimulation protocols and counseling patients about the potential risks and benefits. A key strength of this study lies in its multi-center design, involving data from three different hospitals, which enhances the generalizability and reliability of the findings. The study analyzed a substantial number of IUI cycles, thereby increasing the statistical power and validity of the results. Additionally, mediation analysis, multivariate logistic regression and subgroup analysis allowed for a more nuanced understanding of how E2 levels influenced reproductive outcomes in IUI cycles. These analyses helped identify independent predictors of successful pregnancy outcomes and ensured the robustness of findings. Potential sources of bias included selection bias due to the retrospective nature of the study and information bias related to outcome reporting. To minimize these biases, we included only cycles with complete follow-up data and confirmed outcomes using multiple sources, such as hospital records, medical discharge summaries, and clinic follow-up visits. Despite the rigorous analysis, some variables, for instance, detailed data of newborns’ outcomes and gestational complications, were not included. Future research should investigate the impact of E2 fluctuations throughout the entire IUI cycle, rather than solely focusing on the ovulation trigger day. Besides, in cycles with low E2 levels, whether the supplement of E2 agents before ovulation trigger day might benefit the reproductive outcomes should be explored in future well-designed studies. This could provide deeper insights into optimizing IUI protocols. Additionally, the role of other hormones in conjunction with E2, such as progesterone, LH, and FSH, should be explored to develop more comprehensive and effective treatment strategies in reproductive medicine. Conclusion This study revealed the correlation between E2 levels on ovulation trigger day and pregnancy rates in IUI cycles. Higher E2 levels are independently related with improved clinical pregnancy rates, emphasizing the need for precise E2 monitoring to enhance IUI efficacy. Notably, our study highlights the key mediating role of E2 in the effect of ovulation stimulation type on reproductive outcomes, with the suppression effect indicating that the overall impact is primarily driven by E2-mediated mechanisms. Extremely low levels of E2 induced by LE did not undermine the reproductive outcomes in IUI cycles. However, low E2 levels in cycles without LE, may be associated with reduced reproductive success, as evidenced by higher miscarriage risk and consequently, lower live birth rates. Routine E2 monitoring in clinical practice allows for more precise treatment protocols, potentially increasing IUI success rates. This research offers valuable insights into IUI cycle management, promoting targeted E2 level monitoring and personalized treatment plans to improve pregnancy outcomes. The robust statistical analysis and comprehensive evaluation of patient demographics and treatment protocols further solidify the study’s contributions to optimizing reproductive medicine practices. Acknowledgements Ethics approval and consent to participate The study protocol was approved by the Ethical Committee of The Third Affiliated Hospital of Guangzhou Medical University (approval number 2024-153), and all participants provided written informed consent. Consent for publication All participants had provided written informed consent for the publication of the study results. Attestation Statement We confirm that data regarding any of the subjects in the study has not been previously published unless specified. Furthermore, we agree to provide the data to the editors of the journal for review or query upon request. Availability of data and materials The individual participant data that underlie the results reported in this article, after deidentification (including text, tables, figures, and appendices), will be available beginning 9 months and ending 36 months following article publication. The study protocol will also be available. Data will be shared with investigators whose proposed use of the data has been approved by an independent review committee identified for this purpose. Proposals may be submitted up to 36 months following article publication to our university’s data sharing system, and a data access agreement will be required to gain access to the data. For individual participant data meta-analysis, access will be granted upon submission and approval of a scientifically sound proposal. After 36 months, the data will be available only for meta-analyses through our university’s data access system. Disclosure of Interests None. Funding Medical Research Fund of Guangdong Province (A2024003), and Xinjiang Support Rural Science and Technology (Special Correspondent) Program in Guangdong Province (KTPYJ 2023014). Authors’ contributions K.F. wrote the original draft, reviewed the manuscript, and performed data analysis. X.G., A.S., J.X., K.X., Y.C., C.A., Y.F., Z.L., and M.C. reviewed the manuscript and prepared figures. Z.L. and M.C. also contributed to the conceptualization and data analysis. All authors reviewed the manuscript. Acknowledgements Not applicable. References 1. Agarwal A, Baskaran S, Parekh N, Cho CL, Henkel R, Vij S, et al. Male infertility. Lancet. 2021;397(10271):319-33.2. Allahbadia GN. Intrauterine Insemination: Fundamentals Revisited. Journal of obstetrics and gynaecology of India. 2017;67(6):385-92.3. Allen NC, Herbert CM, 3rd, Maxson WS, Rogers BJ, Diamond MP, Wentz AC. Intrauterine insemination: a critical review. Fertil Steril. 1985;44(5):569-80.4. Starosta A, Gordon CE, Hornstein MD. Predictive factors for intrauterine insemination outcomes: a review. Fertil Res Pract. 2020;6(1):23.5. Starosta A, Gordon CE, Hornstein MD. Predictive factors for intrauterine insemination outcomes: a review. Fertility research and practice. 2020;6(1):23-.6. Boomsma CM, Cohlen BJ, Farquhar C, Boomsma CM. Semen preparation techniques for intrauterine insemination. Cochrane database of systematic reviews. 2019;2019(11):CD004507-CD.7. Potapragada NR, Babayev E, Strom D, Beestrum M, Schauer JM, Jungheim ES. Intrauterine Insemination After Human Chorionic Gonadotropin Trigger or Luteinizing Hormone Surge: A Meta-analysis. Obstetrics and gynecology (New York 1953). 2023;142(1):61-70.8. Kohn TP, Kohn JR, Ramasamy R. Effect of Sperm Morphology on Pregnancy Success via Intrauterine Insemination: A Systematic Review and Meta-Analysis. The Journal of urology. 2018;199(3):812-22.9. Esmailzadeh S, Faramarzi M. Endometrial thickness and pregnancy outcome after intrauterine insemination. Fertil Steril. 2007;88(2):432-7.10. Ayeleke RO, Asseler JD, Cohlen BJ, Veltman‐Verhulst SM, Veltman‐Verhulst SM. Intra‐uterine insemination for unexplained subfertility. Cochrane database of systematic reviews. 2020;2020(3):CD001838-CD.11. Dewailly D, Robin G, Peigne M, Decanter C, Pigny P, Catteau-Jonard S. Interactions between androgens, FSH, anti-Müllerian hormone and estradiol during folliculogenesis in the human normal and polycystic ovary. Human reproduction update. 2016;22(6):709-24.12. Mihm M, Gangooly S, Muttukrishna S. The normal menstrual cycle in women. Animal reproduction science. 2011;124(3-4):229-36.13. Merviel PMDPD, Heraud MHMD, Grenier NMD, Lourdel EMD, Sanguinet PMD, Copin HMDPD. Predictive factors for pregnancy after intrauterine insemination (IUI): An analysis of 1038 cycles and a review of the literature. Fertility and sterility. 2010;93(1):79-88.14. Royster GDMD, Krishnamoorthy KMD, Csokmay JMMD, Yauger BJMD, Chason RJMD, DeCherney AHMD, et al. Are intracytoplasmic sperm injection and high serum estradiol compounding risk factors for adverse obstetric outcomes in assisted reproductive technology? Fertility and sterility. 2016;106(2):363-70.e3.15. Fang L, Yan Y, Wang S, Guo Y, Li Y, Jia Q, et al. High ovarian GDF-8 levels contribute to elevated estradiol production in ovarian hyperstimulation syndrome by stimulating aromatase expression. International journal of biological sciences. 2021;17(9):2338-47.16. Liu Y, Ye XY, Chan C. The association between endometrial thickness and pregnancy outcome in gonadotropin-stimulated intrauterine insemination cycles. Reproductive biology and endocrinology. 2019;17(1):14-.17. Yang A-M, Cui N, Sun Y-F, Hao G-M. Letrozole for Female Infertility. Frontiers in endocrinology (Lausanne). 2021;12:676133-.18. Xu X, Yang A, Han Y, Wang W, Hao G, Cui N. The Association Between Serum Estradiol Levels on hCG Trigger Day and Live Birth Rates in Non-PCOS Patients: A Retrospective Cohort Study. Frontiers in endocrinology (Lausanne). 2022;13:839773-.19. Huang J, Lu Y, He Y, Wang Y, Zhu Q, Qi J, et al. The effect of peak serum estradiol level during ovarian stimulation on cumulative live birth and obstetric outcomes in freeze-all cycles. Frontiers in endocrinology (Lausanne). 2023;14:1130211-.20. New EP, Kodama S, Devine K, Jahandideh S, Imudia AN, Plosker SM. Live birth associated with peak serum estradiol levels in letrozole intrauterine insemination cycles. Fertility and sterility. 2023;119(5):785-91.21. Oktay K, Hourvitz A, Sahin G, Oktem O, Safro B, Cil A, et al. Letrozole Reduces Estrogen and Gonadotropin Exposure in Women with Breast Cancer Undergoing Ovarian Stimulation before Chemotherapy. The journal of clinical endocrinology and metabolism. 2006;91(10):3885-90.22. Kasuga-Yamashita F, Baba T, Nagao S, Fujibe Y, Morishita M, Kuno Y, et al. Letrozole increases preantral follicle growth and decreases estradiol production without impairing follicle survival. Journal of Ovarian Research. 2022;15(1).23. Amer SA, Smith J, Mahran A, Fox P, Fakis A. Double-blind randomized controlled trial of letrozole versus clomiphene citrate in subfertile women with polycystic ovarian syndrome. Human reproduction (Oxford). 2017;32(8):1631-8.24. Ezoe K, Fukuda J, Takeshima K, Shinohara K, Kato K. Letrozole-induced endometrial preparation improved the pregnancy outcomes after frozen blastocyst transfer compared to the natural cycle: a retrospective cohort study. BMC pregnancy and childbirth. 2022;22(1):1-824.25. Zhang S, Gao F, Fu M, Shen H, Wang Y, Han H. Effects of letrozole co-treatment on the cumulative live-birth rate among normal responders in gonadotropin-releasing hormone antagonist cycles. Frontiers in Medicine. 2022;9.26. Tatsumi T, Jwa SC, Kuwahara A, Irahara M, Kubota T, Saito H. Pregnancy and neonatal outcomes following letrozole use in frozen–thawed single embryo transfer cycles. Human Reproduction. 2017;32(6):1244-8.27. Malathi A, Balakrishnan S, B. S L. Correlation between estradiol levels on day of HCG trigger and the number of mature follicles, number of oocytes retrieved, and the number of mature oocytes after oocyte aspiration in ICSI cycles. Middle East Fertility Society Journal. 2021;26(1):1-10.28. Wolff Mv. Natural Cycle and Minimal Stimulation IVF From Physiology to Clinical Practice. 1st 2022. ed. Cham: Springer International Publishing; 2022.29. Waclawik A, Jabbour HN, Blitek A, Ziecik AJ. Estradiol-17β, Prostaglandin E2 (PGE2), and the PGE2 Receptor Are Involved in PGE2 Positive Feedback Loop in the Porcine Endometrium. Endocrinology (Philadelphia). 2009;150(8):3823-32.30. Zhang Y-R, Yin Y, Guo S-M, Wang Y-F, Zhao G-N, Ji D-M, et al. The landscape of transcriptional profiles in human oocytes with different chromatin configurations. Journal of ovarian research. 2024;17(1):99-.31. Bai X, Zheng L, Li D, Xu Y. Research progress of endometrial receptivity in patients with polycystic ovary syndrome: a systematic review. Reproductive biology and endocrinology. 2021;19(1):122-.32. Joo BS, Park SH, An BM, Kim KS, Moon SE, Moon HS. Serum estradiol levels during controlled ovarian hyperstimulation influence the pregnancy outcome of in vitro fertilization in a concentration-dependent manner. Fertil Steril. 2010;93(2):442-6.33. Ling L, Xia D, Jin Y, Hong R, Wang J, Liang Y. Effect of follicle size on pregnancy outcomes in patients undergoing first letrozole-intrauterine insemination. European journal of medical research. 2024;29(1):184-. Figure legends Figure 1 The flow chart of inclusion and exclusion of cycles Table 1 Comparisons of baseline and cycle related characteristics Characteristics Group I Group II Group III Group IV F/X 2 P-value （≤ 25%, N=1496） (25 to 50%, N=1496) (50 to 75%, N=1496) (> 75%, N=1492) E2 levels on ovulation trigger day ≤ 196.46 > 196.46, ≤ 298.64 > 298.64, ≤ 426.43 > 426.43 Female age (years old) 30.85 ± 3.76 30.77 ± 3.64 31.07 ± 3.73 30.84 ± 3.70 1.82 0.245 Duration of infertility (years) 3.45 ± 2.23 3.39 ± 2.17 3.41 ± 2.18 3.29 ± 2.05 1.42 0.234 Type of infertility Primary infertility 462 (30.88%) 432 (28.88%) 448 (29.95%) 474 (31.77%) 3.27 0.352 Secondary infertility 1034 (69.12%) 1064 (71.12%) 1048 (70.05%) 1018 (68.23%) Type of sperm 30.97 < 0.001 AIH 1262 (84.36%) 1221 (81.62%) 1147 (76.67%) 1226 (82.17%) AID 234 (15.64%) 275 (18.38%) 349 (23.33%) 266 (17.83%) Factors of infertility 36.73 0.001 Male factors 291 (19.45%) 333 (22.26%) 402 (26.87%) 321 (21.51%) Ovulation disorder 233 (15.57%) 184 (12.30%) 189 (12.63%) 208 (13.94%) Endometriosis 65 (4.34%) 65 (4.34%) 62 (4.14%） 61 (4.09%) Sexual dysfunction 22 (1.47%) 22 (1.47%) 27 (1.80%) 22 (1.47%) Unexplained 851 (56.89%) 861 (57.55%) 794 (53.07%) 860 (57.64%) Mixed 34 (2.27%) 31 (2.07%) 22 (1.47%) 20 (1.34%) BMI (kg/m 2 ) 22.3 ± 3.27 22.0 ± 3.27 21.6 ± 3.05 21.3 ± 3.15 22.09 < 0.001 AFC 21.4 ± 9.35 20.8 ± 8.23 20.0 ± 8.20 20.7 ± 8.80 4.50 0.001 AMH (ng/mL) 4.90 ± 3.55 4.74 ± 3.40 4.58 ± 3.35 5.04 ± 3.83 3.48 0.008 Basal FSH (IU/L) 5.91 ± 1.92 5.98 ± 1.93 6.05 ± 1.96 6.03 ± 1.96 1.16 0.327 Cycle number 31.53 <0.001 1 727 (48.60%) 839 (56.08%) 807 (53.94%) 743 (49.80%) 2 579 (38.70%) 514 (34.36%) 488 (32.62%) 562 (37.67%) ≥ 3 190 (12.70%) 143 (9.56%) 201 (13.44%) 187 (12.53%) Type of ovulation stimulation 2263.16 <0.001 NC 231(15.44%) 503(33.62%) 543(36.30%) 210(14.08%) CC 19 (1.27%) 82 (5.48%) 158 (10.56%) 416 (27.88%) LE 964 (64.44%) 456 (30.48%) 197 (13.17%) 95 (6.37%) Gn 88 (5.88%) 273 (18.25%) 425 (28.41%) 546 (36.60%) CC + Gn 14(0.94%) 44(2.94%) 61(4.08%) 106(7.10%) LE + Gn 151 (10.10%) 120 (8.02%) 97 (6.48%) 85 (5.70%) Number of dominant follicles 509.89 <0.001 0 103 (6.89%) 61 (4.08%) 45 (3.01%) 46 (3.08%) 1 1122 (75.00%) 1212 (81.02%) 1263 (84.43%) 861 (57.71%) 2 247 (16.51%) 200 (13.37%) 166 (11.10%) 433 (29.02%) ≥3 24 (1.60%) 23 (1.54%) 22 (1.47%) 152 (10.19%) Serum P levels on trigger Day 18.9 ± 20.8 19.4 ± 15.3 20.2 ± 16.5 19.3 ± 14.7 1.19 0.313 Serum LH levels on trigger Day 6.52 ± 6.84 7.10 ± 8.59 6.12 ± 5.82 6.48 ± 6.73 3.45 0.008 Endometrial thickness on trigger Day (mm) 8.24 ± 1.92 8.91 ± 1.88 9.08 ± 1.89 8.87 ± 2.17 42.44 <0.001 Progressive motile sperms (× 106) 23.0 ±18.0 23.1 ±18.1 23.1 ±17.7 24.0 ± 18.3 0.77 0.543 Abbreviations: E2, estradiol; BMI, body mass index; AFC, antral follicle counting; FSH, follicular stimulating hormone. Table 2 Comparisons of pregnancy outcomes Outcome variable Group I Group II Group III Group IV F/X 2 P-value （75%, N=1492) Clinical pregnancy 194 (12.97%) 202 (13.50%) 232 (15.51%) 258 (17.29%) 22.52 0.004 Biochemical pregnancy loss 11 (0.74%) 24 (1.60%) 14 (0.94%) 24 (1.61%) 7.617 0.055 Spontaneous miscarriage 40 (2.67%) 27 (1.80%) 33 (2.21%) 38 (2.55%) 3.02 0.555 Ectopic pregnancy 7 (0.47%) 6 (0.40%) 7 (0.47%) 10 (0.67%) 1.22 0.875 Live birth 129 (8.62%) 158 (10.56%) 174 (11.63%) 202 (13.54%) 19.18 <0.001 Preterm birth 8 (4.12%) 13 (6.44%) 15 (6.47%) 18 (6.98%) 4.33 0.826 Fetal malformation 2 (1.03%) 1 (0.50%) 2 (0.86%) 4 (1.55%) 1.33 0.856 Multiple pregnancy 4 (2.06%) 5 (2.48%) 9 (3.88%) 24 (9.30%) 17.65 0.001 Newborns’ weight 3073 ± 598 3133 ± 555 3022 ±583 3038 ± 636 0.955 0.414 Newborns’ height 49.5 ± 2.41 49.5 ± 1.73 49.5 ± 2.23 49.4 ± 2.54 0.147 0.932 Abbreviations: Group I refers to E2 < 25%; Group II refers to E2 25 to 50%; Group III refers to E2 50 to 75%; Group IV refers to E2 Table 3 Independent impacts of E2 groups on reproductive outcomes by multivariate logistic regression Live birth Group I 0.60 0.34, 1.04 0.070 Group II Ref Ref Group III 0.96 0.57, 1.61 0.900 Group IV 1.24 0.70, 2.17 0.500 Clinical pregnancy Group I 0.85 0.66, 1.08 0.200 Group II Ref Ref Group III 1.17 0.94, 1.46 0.200 Group IV 1.32 1.05, 1.68 0.020 Spontaneous miscarriage Group I 2.15 1.19, 3.97 0.012 Group II Ref Ref Group III 1.36 0.72, 2.60 0.300 Group IV 1.71 0.93, 3.22 0.087 Multiple pregnancy Group I 0.57 0.12, 2.37 0.500 Group II Ref Ref Group III 2.26 0.76, 7.51 0.200 Group IV 2.16 0.84, 6.67 0.140 Newborns’ weight Group I <0.01 0.00, inf 0.151 Group II Ref Ref Group III <0.01 0.00, inf 0.317 Group IV <0.01 0.00, inf 0.789 Newborns’ height Group I 0.94 0.47, 1.86 0.852 Group II Ref Ref Group III 1.05 0.59, 1.87 0.880 Group IV 0.87 0.47, 1.61 0.659 Abbreviations: OR, Odds Ratio; 95% CI, Confidence Interval; Group I refers to E2 75%; inf, values greater than 1e +5 . Table 4. Mediation Analysis of Number of Dominant Follicles, Type of Ovulation Stimulation, and Female Age on Reproductive Outcomes via E2 Number of Dominant Follicles 0.003 0.018 0.021 0.128 Type of Ovulation Stimulation 0.011 -0.005 0.006 1.805 Female Age -0.0002 -0.010 -0.010 0.016 Abbreviations: ACME, Average Causal Mediation Effect; ADE, Average Direct Effect; PM: Proportion Mediated. Table 5. Subgroup analysis of critical reproductive outcomes in cycles with or without LE using multivariate logistic regression OR 95% CI p-value OR 95% CI p-value Live birth Subgroup Ⅰ 0.89 0.42, 1.84 0.751 0.31 0.13, 0.76 0.010 Subgroup Ⅱ Ref Ref Ref Ref Subgroup Ⅲ 0.98 0.42, 2.37 0.971 0.85 0.43, 1.65 0.645 Subgroup Ⅳ 3.51 1.09, 14.05 0.050 0.83 0.43, 1.54 0.554 Clinical pregnancy Subgroup Ⅰ 0.85 0.62, 1.18 0.323 0.83 0.55, 1.24 0.382 Subgroup Ⅱ Ref Ref Ref Ref Subgroup Ⅲ 1.35 0.90, 2.02 0.143 1.08 0.83, 1.42 0.563 Subgroup Ⅳ 1.45 0.91, 2.28 0.113 1.22 0.92, 1.63 0.176 Spontaneous miscarriage Subgroup Ⅰ 1.25 0.53, 3.06 0.623 4.30 1.53, 12.20 0.005 Subgroup Ⅱ Ref Ref Ref Ref Subgroup Ⅲ 1.57 0.59, 4.22 0.367 1.15 0.50, 2.72 0.700 Subgroup Ⅳ 0.43 0.09, 1.64 0.248 1.69 0.81, 3.79 0.200 Multiple pregnancy Subgroup Ⅰ 0.28 0.02, 3.52 0.339 0 0.00, 1.03*10^21 0.993 Subgroup Ⅱ Ref Ref Ref Ref Subgroup Ⅲ 0.30 0.03, 2.82 0.300 10.09 1.23, 262.20 0.072 Subgroup Ⅳ 2.13 0.12, 39.37 0.595 8.23 1.12, 206.94 0.092 Newborn weight Subgroup I <0.01 0.00, inf 0.230 <0.01 0.00, inf 0.207 Subgroup II Ref Ref Ref Ref Subgroup III <0.01 0.00, inf 0.568 <0.01 0.00, inf 0.284 Subgroup IV <0.01 0.00, inf 0.642 2.50 0.00, inf 0.992 Newborn height Subgroup I 0.93 0.38, 2.26 0.871 0.55 0.15, 2.00 0.368 Subgroup II Ref Ref Ref Ref Subgroup III 0.91 0.29, 2.86 0.867 0.96 0.47, 1.94 0.900 Subgroup IV 0.74 0.22, 2.48 0.624 0.85 0.40, 1.81 0.678 Abbreviations: OR, Odds Ratio; 95% CI, Confidence Interval; Subgroup Ⅰ refers to E2 ≤ 25%; Subgroup Ⅱ refers to E2 25 to 50%; Subgroup Ⅲ refers to E2 50 to 75%; Subgroup Ⅳ refers to E2 >75%; inf, values greater than 1e +5 . Supplemental Figure 1 Boxplot of peak E2 Levels in overall cycles Information & Authors Information Version history V1 Version 1 10 February 2025 Copyright This work is licensed under a Non Exclusive No Reuse License. Keywords fertility and assisted reproduction infertility: assisted conception reproductive science: sex steroids Authors Affiliations Kexin FAN 0009-0006-4942-4869 Southern Medical University Nanfang Hospital View all articles by this author Xinru GU Maoming People's Hospital View all articles by this author Aniguli SHABIER People's Hospital of Xinjiang Uygur Autonomous Region View all articles by this author Jiaqi XIAO Guangzhou Medical University View all articles by this author Kai XU West China Hospital of Sichuan University View all articles by this author Yilin CHEN The University of Sydney School of Computer Science View all articles by this author Yang Fu The Third Affiliated Hospital of Guangzhou Medical University View all articles by this author Chunyan An The Third Affiliated Hospital of Guangzhou Medical University View all articles by this author Zhi LIU Southern Medical University Nanfang Hospital View all articles by this author Mingzhu Cao [email protected] The Third Affiliated Hospital of Guangzhou Medical University View all articles by this author Metrics & Citations Metrics Article Usage 357 views 183 downloads .FvxKWukQNSOunydq8rnd { width: 100px; } Citations Download citation Kexin FAN, Xinru GU, Aniguli SHABIER, et al. Low Peak Estradiol Levels Associated with Increased Miscarriage and Decreased Live Birth in Non-Letrozole IUI cycles: A Multi-center Retrospective Cohort Study. Authorea . 10 February 2025. DOI: https://doi.org/10.22541/au.173917810.07488353/v1 If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click Download. For more information or tips please see 'Downloading to a citation manager' in the Help menu . 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