Presence of endometrioma decreased blastocyst formation rate but not impair Assisted Reproductive Technology (ART) outcome

This study aims to assess the impact of endometrioma on patients who undergo ART treatment due to endometriosis. A retrospective study was conducted on women ≤ 40 years of age who underwent ART treatment at an academic medical center between January 2014 and December 2020. Two-hundred-and-eight women had received IVF/ICSI treatment due to endometriosis and there were 89 patients presence of endometrioma. Patients were further divided into primary endometrioma, recurrent endometrioma and those having received cystectomy for endometrioma prior to IVF/ICSI. The control group included 624 infertile women without endometriosis. In the endometrioma subgroup (B) the blastocyst formation rate was significantly lower when compared with the endometriosis (A) and control groups (C). The cumulative live birth rates (CLBRs) (60.5% versus 49.4% versus 56.9%, p = 0.194 in A versus B, p = 0.406 in A versus C, p = 0.878 in B versus C) were comparable. Multiple logistic regression analysis revealed that female age, total FSH dose and blastocyst formation rate were the significant variables in predicting CLBR (OR 0.89, CI 0.80–0.99, p < 0.025, OR 0.68 CI 0.53–0.88, p = 0.003 and OR 30.04, CI 9.93–90.9, p < 0.001, respectively). The CLBRs were comparable at 47.1%, 60% and 57.9% in the primary endometrioma, s/p cystectomy and recurrent endometrioma group. Although the blastocyst formation rate was lower in the endometrioma group, CLBR was not worse than those who were in the endometriosis or control group. Cystectomy for endometrioma did not alter IVF/ICSI outcomes if the ovarian reserve was comparable. Recurrent endometrioma did not worsen IVF/ICSI outcomes than primary endometrioma.


Introduction
Endometriosis is a chronic disease known to be detrimental to fertility [1]. Approximately 25-50% of women experiencing infertility may be affected by endometriosis, with 30-50% of women diagnosed with endometriosis being infertile [2]. A significant number of women with endometriosis eventually seek In Vitro Fertilization (IVF) with or without Intracytoplasmic Sperm Injection (ICSI) for conception. An endometrioma is the formation of a cyst within the ovary with an ectopic endometrial tissue lining [3]. It is found in 17-44% of patients with endometriosis [4]. The exact mechanism through which endometrioma causes infertility remains uncertain. Several proposed mechanisms include distorting tubo-ovarian anatomy [5], reduction in the quality and quantity of developing follicles by anatomical proximity of the ovarian cyst to the nearby follicular pool [6,7], and a reduced ovarian reserve and embryo quality due to local inflammation milieu, toxic content and oxidative damage [8,9].
Although with a higher cancel rate and lower oocyte retrieval, current evidence shows that women with endometrioma have similar IVF/ICSI outcomes in terms of live birth rate when compared to those without endometriosis [10]. However, there have been only a few studies comparing IVF/ICSI outcomes between women with endometrioma and women with endometriosis. Current molecular, histological and morphological evidence suggests that endometrioma is detrimental to the ovaries [6,11]. Endometrioma, being different from endometriosis, likely has a specific impact on IVF/ICSI outcomes. Therefore, the first part of this study was aimed to compare IVF/ICSI outcomes between women with endometrioma and women with endometriosis.
In the second part of our study, we aimed to determine the impact of cystectomy on IVF/ICSI outcomes. Current guidelines are unclear regarding the surgical removal of an endometrioma prior to IVF/ICIS [12]. Surgical treatment of endometrioma has been shown to increase natural fecundity [13]. Additionally, an endometrioma left in situ during IVF/ ICSI may cause technical difficulties during oocyte retrieval, progression of the endometrioma after artificial reproductive technology (ART), and higher risks of infection [14][15][16]. Evidence has shown that surgical treatment of endometrioma could be detrimental to ovarian reserve [17,18], subsequently adversely affecting reproductive outcomes of IVF/ICSI [19]. Therefore, the second part of our study aimed to determine the impact of cystectomy for endometrioma on the IVF/ICSI outcomes.

Materials and methods
We retrospectively analyzed patients (n = 2153) who had undergone IVF/ICSI between January 2014 and December 2018, at the Center for Reproductive Medicine, Division of Reproductive Endocrinology and Infertility, Department of Obstetrics, Gynecology and Women's Health, Taichung Veterans General Hospital, Taichung, Taiwan. We included those cycles with the patient's major reason for IVF/ICSI being endometriosis. Endometriosis was confirmed by laparoscopic surgery, the presence of endometrioma or when a patient with dysmenorrhea and pelvic examination revealed palpable tender induration or nodules located in cul-de-sac. Endometrioma was confirmed by visualizing two or more separate examinations done at intervals were at least one month apart, an ovarian cyst with regular margins and ground-glass echogenicity on transvaginal ultrasonography (TVUS) [20]. The endometrioma was measured along three dimensions, with the average diameter calculated during baseline ultrasonography on D2/ D3 in the month of controlled ovarian stimulation (COS). We excluded those cycles with patients aged > 40 years or < 20 years, had simulation duration < 5 days, with severe male factor (including Testicular Sperm Extraction), uterine factor (including adenomyosis as defined by TVUS), preimplantation genetic testing for aneuploidies (PGT-A), donor eggs or immunological factors. Patients whose embryos were not completely transferred back, or who had received embryo transfer from different oocyte pick-up cycles were also excluded.
The control group included women who underwent IVF/ICSI for non-endometriosis-related infertility during the same study period. Endometriosis was ruled out in a pre-ART work-up assessment, including a clinical examination, questioning (regarding the extent of pelvic pain and a prior history of surgery) and TVUS. The control group was matched in age and Anti-mullerian Hormone (AMH) levels in 1 to 3 subject ratio. Blind matching to the results was performed.
In the first analysis, we compared reproductive outcomes amongst patients with endometrioma present during IVF/ICSI treatment, those with only endometriosis, and the control group. In the second analysis, we compared outcomes between patients in the "Recurrent endometrioma group" who were defined as patients with endometrioma during IVF/ICSI and had a prior history of cystectomy for endometrioma at least once. The "Primary endometrioma group" was defined as those who did not have a prior cystectomy and was with endometrioma during IVF/ICSI. Finally, the "status post (s/p) cystectomy group" was defined as patients who had received cystectomy for endometrioma prior to IVF/ICSI treatment and had shown no presence of endometrioma during IVF/ ICSI. This study was approved by the Institutional Review Board of Taichung Veterans General Hospital (IRB No. CE21306B).
Patients underwent COS, oocyte retrieval and embryo transfer according to procedures as previously described [21]. Which protocol (either agonist or antagonist protocol) to use for the patient was at the discretion of the care physician. Stimulation was monitored by transvaginal ultrasound. Ovulation triggering was induced by injecting the recombinant hCG (Ovidrel, Merck Serono) and/or GnRH agonist. Ultrasound-guided transvaginal oocyte retrieval was carried out approximately 36 h post-triggering. Oocytes were either inseminated or underwent ICSI approximately 4 h after collection. We directly perform fresh ET or immediate FET if the patient's CA-125 level is < 40 Units/ml and no other contraindication. If the CA-125 level is higher than 40 Units/ml we would give GnRH agonist (Leuplin Depot 1 M 3.75 mg S.C. Injection) for 2-6 cycles depending on the CA-125 level before ET. For the fresh group, cleavagestage embryos or blastocysts were transferred. The surplus embryos were cryopreserved through vitrification. The endometrial preparations during the following frozen embryo transfer (FET) cycles were programmed by either hormone replacement cycles or modified natural cycles, depending on the individual condition of each patient. Details of the endometrium priming and luteal phase support have been reported earlier [21].

Outcomes and statistical analyses
We followed up these patients through June 2021. Our primary outcome was cumulative live birth rate (CLBR), determined in those who had achieved through fresh and/or vitrified embryos obtained from the same oocyte retrieval cycle. Clinical pregnancy was defined as the presence of at least one embryo having cardiac activity. Live birth was defined as the delivery of a live infant after at least 24 weeks of gestation. Live birth rates (LBR) were calculated individually in the fresh ET cycle and in the first FET cycle if no fresh ET had been performed. Blastocyst formation rates only included those patients who were intended to receive blastocyst transfer.
Data were presented as mean ± standard deviation (SD), or as a percentage. Group comparisons were performed using Mann-Whitney and Pearson's Chi-square tests in SPSS (Version 18). Logistic regression analysis was conducted on clinical covariates assessed for their association with CLBR. Multivariable logistic regression (MLR) analysis was performed on variables that were significant at univariable analysis (P < 0.05). Differences were considered significant at p < 0.05.

Results
We analyzed a total of 208 IVF/ICSI cycles which had indications of endometriosis. These cycles consisted of 119 with endometriosis and 89 with endometrioma. The flow chart of these cycles is shown in Fig. 1. In 89 cycles with endometrioma, 19 had received cystectomy for endometrioma prior to IVF/ICSI and were included in the "recurrent endometrioma group". Five of them received cystectomy twice before and the remainders received cystectomy once. The remaining 64 cycles were included in the "primary endometrioma group". In 119 cycles with no endometrioma during IVF/ICSI, 40 had priorly received cystectomy for endometrioma, and were included in the "s/p cystectomy group". After matching, we had 624 cycles without endometriosis during IVF/ICSI as the control group.
First analyses of the baseline characteristics and outcomes are presented in Table 1. Comparable characteristics in the endometriosis (A), endometrioma (B) and control (C) group Patients in the control group had a significantly higher Body Mass Index (BMI) and received more ICSI In the endometrioma group, the mean size of endometrioma was 3.9 cm (range 1 to 9 cm).
The gonadotropin dose given was significantly higher in the endometrioma group compared with the other groups (FSH dosage 3047 ± 1037.9 IU versus 3619 IU ± 1223 versus 3130 ± 1065.5 IU, p = 0.001 in A versus B, p = 0.4 in A versus C, p = 0.001 in B versus C; LH dosage 941 ± 573.9 IU versus 1224 ± 721.5 IU versus 963 ± 601.2 IU, p = 0.009 in A versus B, p = 0.823 in A versus C, p = 0.003 in B versus C). The number of follicles ≥ 14 mm on the day of triggering, number of oocytes retrieved, and number of mature oocytes were lower in the endometrioma group than in the control group. The blastocyst formation rate was significantly lower in the endometrioma group when compared with the other two groups (57.7% versus 49.4% versus 56%, p = 0.005 in A versus B, p = 0.376 in A versus C, p = 0.008 in B versus C).
Clinical pregnancy rate (CPR) in fresh ET, LBR in fresh ET, and CLBR were comparable in the three groups. Total 1-4 times FETs were included in the CLBR according to the patient's clinical condition who had achieved live birth or ran out of all the vitrified embryos obtained from the same oocyte retrieval cycle. LBR in the first FET in the freeze-all cycle was significantly lower in the endometrioma group when compared with the control group (53.8% versus 36.8% versus 59.2%, p = 0.134 in A versus B, p = 0.536 in A versus C, p = 0.011 in B versus C) ( Fig. 2; Table 2).
In the second analysis, the baseline characteristics and clinical outcomes of primary endometrioma (D), s/p cystectomy (E) and recurrent endometrioma (F) are shown in Table 3. The female age was lower in the s/p cystectomy group than in the endometrioma group ( CPR in fresh ET, LBR in fresh ET, LBR in first FTE in the freeze-all cycle and CLBR were all similar in the primary endometrioma, s/p cystectomy and recurrent endometrioma groups (Fig. 3).

Discussion
Our results show that in patients with endometriosis being the major indication for IVF/ICSI, the CLBRs were similar to the controls (non-endometriosis). Although the presence of endometrioma had comparable CLRBs, the prescribed gonadotropin dose was significantly higher, with the blastocyst formation rate being significantly lower when compared with those patients with only endometriosis. In the second analysis, CLBRs were comparable across the primary endometrioma, s/p cystectomy and recurrent endometrioma groups. The blastocyst formation rate was significantly higher in the s/p cystectomy group when compared with the other two groups.
A number of studies have investigated the effects of endometriosis on ART outcomes. In general, women with endometriosis have similar ART outcomes to those of controls in terms of LBR [22][23][24]. Our results are compatible with these. Regarding the relationship between the presence of endometrioma and ART outcomes, our findings are less consistent. A recent systematic review found that women with endometrioma have fewer oocytes   and MII oocytes retrieved when compared to those without endometriosis, but no such differences in CPR, IR or LBR [25]. Our results are similar to theirs, showing that endometrioma was associated with higher gonadotropin consumption, a lower number of oocytes retrieved, and a lower blastocyst formation rate when compared with controls. The final outcome, CLBR, was the same as the control group. When comparing patients with endometrioma and the control group, we found that the LBR of the first FET is significantly lower in the endometrioma group than in the control group. We do not have good explanations. Since the CLBR were not different, it may be due to embryo selection. More studies are needed to clarify the result. When comparing patients with endometrioma and endometriosis, our results revealed the similarities of LBR and CLBR in patients with either endometrioma or endometriosis. These findings are consistent with previous studies which have shown a similarity in delivery rates and pregnancy rates in patients with unoperated endometrioma and with only endometriosis [26,27]. These studies were, however, conducted in the 2000s, and CLBR was not analyzed at the time.
In comparing primary endometrioma and cystectomy prior to IVF/ICSI, we found that although the patients in the s/p cystectomy group were younger in age, their ovarian reserves were similar, likely due to the surgical excision of endometrioma having lowered the ovarian reserve [17]. In our center, it is our policy is to choose younger patients with moderate ovarian reserves as candidates for receiving cystectomy for endometrioma due to fertility reasons. We encourage these patients to undergo IVF/ICSI if they have not gotten pregnant after a one-year trial post-surgery Despite concerns surrounding a reduced ovarian reserve, surgical removal of endometrioma may improve chances of spontaneous pregnancy through the restoration of the ovarian functional anatomy. This approach has been considered to be a primary treatment method for infertility patients in the case of endometrioma [12]. Additionally, such an operation also improves pain symptoms [28]. Regarding the impact of excised endometrioma on ART outcomes, we found the total FSH dose was significantly lower in s/p cystectomy group. It may be due to the patients in this group were younger. Other ART outcomes show similarities between the primary endometrioma and s/p cystectomy groups in terms of the number of oocytes retrieved, number of mature oocytes, LBR and CLBR. Our findings are consistent with most reports published in the available literature, including a recent meta-analysis [29]. Other studies have reported different results showing that endometrioma surgery diminished ovarian reserves, leading to lower pregnancy rates after IVF [30,31]. However, these studies were conducted on limited numbers of patients, with differences in surgical procedures and varying surgeon expertise. Our results regarding CLBR per oocyte retrieval further confirm that with comparable ovarian reserves, cystectomy prior to operation produced similar IVF/ICSI outcomes when compared with endometrioma in situ.
In comparing primary and recurrent endometriomas, our results show weaker ovarian responses to COS in the recurrent endometrioma group. This finding is inconsistent with a previous study that reported that patients with recurrent endometriomas have the same ovarian response [32]. In contrast, the outcomes of IVF/ICSI, including LBR and CLBR in our study, were compatible between these two groups and was consistent with that previous study [32]. Although there is no significant difference, the pregnancy rates after fresh ET in the recurrent endometrioma group are higher. It may be due to the number of patients in this group being small (n = 19, and only 9 patients of them received fresh ET). More cases are needed to confirm the result. There was a study histologically confirmed surgery for recurrent endometrioma is associated with a greater loss of ovarian tissue, which is more harmful to ovarian reserves when compared with endometriomas that were operated on for the first time [33]. Park et al. also reported that second-line surgery for recurrent endometrioma has a deleterious effect on IVF outcomes, including ovarian response and CPR, when compared with in situ recurrent endometrioma [34]. It has also been reported that spontaneous cumulative pregnancy rates drop to almost half after second-line surgery for endometrioma when compared with those obtained after primary surgery [35]. Therefore, a conservative management plan for the treatment of recurrent endometrioma is more suitable for women who seek fertility. Our study has shown the presence of endometrioma is associated with fewer blastocyst formations when compared with the endometriosis or the control group. Such effects seemed to disappear after performing cystectomy on endometrioma. The poorer ovarian response and lower blastocyst formation rates were unlikely consequences of the deleterious effects that surgery had on the endometriomas and were more likely due to endometrioma per se. There are growing studies that have shown that ovarian quality may be deteriorated by endometrioma [36]. One recent study which used scRNA-seq compared oocytes from patients with endometrioma and healthy oocytes. The results revealed a differential transcriptomic profile indicating a lower oocyte quality in endometrioma [37]. These mechanisms may explain how the blastocyst formation rate we found was lower in the endometrioma group.
In our analysis, the dosage of FSH was relatively high in a patient with a normal AMH level compared with other studies. Since this is real-world data. It may be due to ethnic variations. There were studies that suggested Asian women requiring higher gonadotropin doses to achieve an adequate number of oocytes for IVF [38]. Based on our experience, we use relatively high doses to treat our patients.
Our study possesses certain strengths. First, our primary outcome is CLBR per oocyte retrieval which has been seldom mentioned in previous studies and is a better indicator of quality and success in IVF/ICSI, as cryopreservation is an integral part of IVF [39]. Second, all our patients had undergone ovarian stimulation and IVF performed at a single medical center. In this regard, all measurements (AMH and TVUS) were performed consistently. Third, we compared the presence of endometrioma with endometriosis. This issue is not commonly addressed but still has practical implications. Our results show that for the first time, the presence of endometrioma negatively influenced ovarian responsiveness and blastocyst formation rates. The main limitation of this work are its retrospective design nature and the number size of the recurrent endometrioma group being small (n = 19). In addition, the endometriosis group was heterogeneous, including patients with peritoneal endometriosis or DIE, with some of them having no pathological diagnosis. Nevertheless, it is currently well accepted that TVUS is a highly accurate and reproducible method for the non-invasive diagnosis of DIE and ovarian lesions [40]. Another limitation is that we did not stratify patients according to their endometriosis stages, and only stratified those with endometrioma or endometriosis. As an increasing number of patients are receiving conservative treatment prior to IVF/ICSI, the presence or absence of endometrioma is a more useful characteristic in daily practice.

Conclusion
We have shown that IVF/ICSI outcomes are comparable in those with endometrioma and controls. Although the blastocyst formation rate was lower and gonadotropin consumption was higher in those with endometrioma, IVF/ICSI outcomes in patients with endometrioma were no worse than those with endometriosis. Cystectomy for endometrioma did not alter IVF/ICSI outcomes if the ovarian reserve was comparable. Recurrent endometrioma did not contribute to worse impacts on the IVF/ICSI outcomes when compared with primary endometrioma. Availability of data and materials All data generated and analyzed during this study are included in this published article and its supplementary information files.
Data availability Data of the study are available and can be provided upon request.

Declarations
Competing interests The authors have no relevant financial or nonfinancial interests to disclose.
Ethics approval and consent to participate Institutional Review Board, TCVGH, No. CE21306B. Date of approval: August 30, 2021.

Informed consent statement Not applicable
Consent to publish Authors are responsible for the correctness of the statements provided in the manuscript.