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Endometrial pattern, thickness and growth in predicting pregnancy outcome following 3319 IVF cycle
Reproductive BioMedicine Online (2014) 29, 291–298
w w w. s c i e n c e d i r e c t . c o m w w w. r b m o n l i n e . c o m
ARTICLE
Endometrial pattern, thickness and growth in predicting pregnancy outcome following 3319 IVF cycle Jing Zhao, Qiong Zhang, Yonggang Wang, Yanping Li * Reproductive Medicine Center, Xiangya Hospital, Central South University, Changsha, Hunan, China * Corresponding author.
E-mail address: [email protected] (Y Li). Yanping Li, MD, is a professor and doctoral supervisor of Central South University, Changsha, China and the Director of Reproductive Medicine Center, Department of Obstetrics and Gynecology, Xiangya Hospital. She performs more than 1000 IVF cycles per year and the clinical pregnancy rate is almost 50%. She conducts clinical work on infertility, IVF, preimplantation genetic diagnosis, intrauterine insemination and microsurgery, and research on reproductive endocrinology, poor ovarian response, thin endometria and embryonic stem cells.
A retrospective study of 3319 women was conducted to assess predictive ability of endometrial characteristics for outcomes of IVF and embryo transfer. Endometrial thickness, growth and pattern were assessed at two time points (day 3 of gonadotrophin stimulation and day of HCG administration). Endometrial patterns were classified as pattern A: triple-line pattern comprising a central hyperechoic line surrounded by two hypoechoic layers; pattern B: an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and poorly defined central echogenic line; and pattern C: homogenous, hyperechogenic endometrium. The endometrium of pregnant women was thinner on day 3 of stimulation, thicker on the day of HCG administration, and showed greater growth in thickness compared with non-pregnant women. Clinical pregnancy rates differed according to endometrial pattern on the day of HCG administration (55.2%, 50.9% and 37.4% for patterns A, B and C, respectively). A positive linear relationship was found between endometrial thickness on the day of HCG administration and clinical pregnancy rate. Endometrial thickness, change and pattern were independent factors affecting outcome. Receiver operator characteristic curves showed that endometrial pattern, thickness and changes were not good predictors of clinical pregnancy. Discriminant analysis indicated that 58.7% of original grouped cases were correctly classified. Although endometrium with triple-line or increased thickness may favour pregnancy, combined endometrial characteristics do not predict outcomes.
Abstract
© 2014 Published by Elsevier Ltd on behalf of Reproductive Healthcare Ltd. KEYWORDS: endometrial growth, endometrial pattern, endometrial thickness, pregnancy rate, transvaginal ultrasound
http://dx.doi.org/10.1016/j.rbmo.2014.05.011 1472-6483/© 2014 Published by Elsevier Ltd on behalf of Reproductive Healthcare Ltd.
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Introduction Assisted reproductive technology has been commonly used in infertility treatment over the past 3 decades. In addition to embryo quality, the receptivity of the endometrium plays an important role in clinical outcome. Sonography is a noninvasive and simple method that has been used to evaluate endometrial receptivity. Several sonographic parameters have been evaluated. These include endometrial thickness, endometrial pattern, endometrial volume and endometrial and subendometrial blood flow (Basir et al., 2002; Dickey et al., 1992, Fanchin, 2001; Wang et al., 2010; Yuval et al., 1999). Endometrial thickness and pattern have been evaluated as possible predictors of pregnancy in multiple studies, with conflicting results. Some investigators have reported significant correlations between pregnancy rate, endometrial thickness, pattern, or both (Al-Ghamdi et al., 2008; Chen et al., 2010; Noyes et al., 1995; Richter et al., 2007; Rinaldi et al., 1996), whereas others have not shown such a relationship (Basir et al., 2002; De Geyter et al., 2000; Dietterich et al., 2002; Rashidi et al., 2004). Only a few studies (Gonen and Casper, 1990; Gonen et al., 1989; McWilliams and Frattarelli, 2007); however, have evaluated the change in endometrial thickness occurring during IVF stimulation, and these studies reached different conclusions with small samples. This study was designed to assess the relationship between endometrial features (endometrial pattern and thickness and endometrium growth) and clinical outcome based on 3319 IVF stimulation cycles.
Materials and methods Patient recruitment and counselling The study was reviewed and approved by the Institutional Review Board and the Ethics Committee of Xiangya Hospital, Changsha, China (IRB reference number 201212047, dated 27 August, 2011). The study was conducted in accordance with the Declaration of Helsinki, as revised in 1983. We conducted a retrospective cohort study of 3319 consecutive infertile women. Briefly, the women underwent fresh IVF-ET between January 2009 and May 2011 at the Reproductive Medicine Centre of Xiangya hospital, Central South University (Changsha, China). Before undergoing IVF, patients were examined with ultrasonography more than once at the different stages of their menstrual cycles. If any subtle cavity abnormalities were found, those women were recommended to undergo further examination or treatment with hysteroscopy or saline sonohysterogram. Exclusion criteria included presence of known endometrial polyp, uterine anomaly, and insemination method other than IVF, cycles using donor oocytes or cryopreserved embryos. Patients underwent no therapeutic intervention other than routine procedures.
J Zhao et al. response and co-existing medical conditions. Ovarian stimulation was initiated when the serum oestradiol concentration level was ≤50 pg/ml, and the largest follicle diameter was <10 mm without ovarian cysts. Urofollitiopin injection (HMG; LiZhu, China), human menopausal gonadotrophin, or both, were used to achieve ovarian stimulation. The initial dosage of gonadotrophin ranged from 150 to 450 IU, depending on the basal FSH level, antral follicular count and maternal age. When at least two follicles were 18 mm or wider in diameter, 10,000 IU of HCG (HCG; Profasi; Serono, Italy) was administered. Oocyte retrieval was carried out 36 h after the administration of HCG, and was followed by conventional IVF. Up to three high-quality embryos were transferred 72 h after oocyte collection according to the centre’s standard practice. When available, surplus good-quality embryos were cryopreserved. The luteal phase was supported using 80 mg progesterone in oil (Progesterone; Tongyong, Shanghai, China) intramuscular injection daily lasting 75 days. Clinical pregnancy was defined as identification of a gestational sac 4–5 weeks after embryo transfer.
Ultrasound measurement Endometrial features assessed included endometrial thickness and pattern on day 3 of gonadotrophin stimulation and on the day of HCG administration, as well as the change in endometrial thickness from day 3 to day of HCG administration. All were measured by transvaginal 8 MHz ultrasonography with Doppler ultrasound (Mindray DC-6 Expert; Shenzhen, China). Endometrial thickness was measured in a median longitudinal plane of the uterus as the maximum distance between the endometrial–myometrial interface of the anterior to the posterior wall of the uterus. Endometrial pattern was classified as pattern A (a triple-line pattern consisting of a central hyperechoic line surrounded by two hypoechoic layers), pattern B (an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and a poorly defined central echogenic line), and pattern C (homogenous, hyperechogenic endometrium).
Statistical analysis Continuous data were expressed as mean ± SD values or as median and range, according to the distribution, and were analysed for differences using the Student’s t-test. Categorical data were presented as numbers, and chi-squared test was used for statistical comparison of percentages. Binary logistic regression analysis was also carried out, and the receiver operating characteristic (ROC) curve was applied to determine the predictive value of endometrial parameters. Discriminant analysis was used to evaluate the predictive ability of all of the endometrial parameters combined. The significance level for all analyses was P < 0.05. Statistical analysis was carried out using the Statistical Package for Social Sciences (SPSS Inc, Chicago, IL, USA).
Ovulation induction and IVF–embryo transfer procedures
Results
The choice of stimulation protocol was individualized on the basis of the patient’s age, cause of infertility, ovarian
A total of 3319 IVF cycles (3319 women) were included in the study. The clinical pregnancy rate, defined as identification
Endometrial characteristics and IVF-ET outcome
293 statistically significant differences in embryo quality were found between the groups (data not shown). Endometrial thickness and pattern measured on day 3 of gonadotrophin stimulation and on the day of HCG administration in both pregnant and non-pregnant women are shown in Table 2 and Figure 1. Pregnant women had significantly thinner endometrial linings on day 3 of gonadotrophin stimulation (P = 0.008), significantly thicker endometrial linings on the day of HCG administration (P < 0.001), and a greater
of a gestational sac 4–5 weeks after embryo transfer, was 52.3%. The demographic data and ovarian responses are presented in Table 1. When subdividing the women into those who conceived (n = 1010) and those who did not conceive (n = 923), significant differences were observed between the two groups. These included age, body mass index, basal FSH, basal oestradiol, duration of ovarian stimulation, total gonadotrophin dose, P-value on the day of HCG administration, and total number of embryos produced (P < 0.05). No
Table 1
Parameters for the IVF study group.
Variables Age (years) BMI(kg/m2) Baseline FSH (IU/L) Baseline LH (IU/L) Baseline E2 (pg//mL) Days of stimulation (days) Ampules of gonadotrophin used (IU) Oestradiol on HCG day (pg/mL) P-value on HCG day (ng/mL) Number of oocytes retrieved Number of embryos Number of embryos transferred Cause of infertility; n (%) Tubal factor Endometriosis PCOS Multiple factors
All patients (n = 1933)
Pregnanta (n = 1010)
Non-pregnant (n = 923)
P-value
31.2 ± 4.6 21.73 ± 2.87 6.3 ± 2.9 4.5 ± 2.8 33.2 ± 21.9 11.1 ± 2.3
30.6 ± 4.4 21.6 ± 2.6 6.1 ± 2.9 4.5 ± 3.0 31.9 ± 22.0 11.2 ± 2.3
31.8 ± 4.8 21.9 ± 3.1 6.4 ± 2.9 4.5 ± 2.4 34.6 ± 21.6 11.0 ± 2.3
<0.05 <0.05 <0.05 NS <0.05 <0.05
2115.0 ± 832.5
2040 ± 772.5
2197.5 ± 885.0
<0.05
3489.7 ± 2112.2
3514.2 ± 2026.8
3462.9 ± 2202.7
NS
0.62 ± 0.48
0.60 ± 0.46
0.64 ± 0.53
<0.05
13.0 ± 5.8
13.2 ± 5.7
12.7 ± 6.0
NS
7.3 ± 4.1 2.3 ± 0.2
7.6 ± 3.9 2.3 ± 0.3
7.0 ± 4.3 2.2 ± 0.3
<0.05 NS NS
1767 (91.4) 23 (1.2) 23 (1.2) 120 (6.2)
924 (91.5) 12 (1.2) 12 (1.2) 62 (6.1)
843 (91.3) 11 (1.2) 11 (1.2) 58 (6.3)
BMI, body mass index; NS, not significant; PCOS, polycystic ovary syndrome. a Patients who achieved a clinical pregnancy.
Table 2 Comparison of endometrial thickness at three points in pregnant and nonpregnant women. Variable Endometrial thickness on day 3 of stimulation (mm) Endometrial thickness on day of HCG administration (mm) Change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration (mm) Data shown as mean ± SD. P < 0.05 was statistically significant. a Clinical pregnancy.
Pregnanta (n = 1010)
Non-pregnant (n = 923)
6.0 ± 1.6
6.2 ± 1.7
0.008
11.0 ± 2.2
10.3 ± 2.2
<0.001
5.0 ± 2.5
4.1 ± 2.6
<0.001
P-value
294
J Zhao et al. P < 0.05
60.0
pattern A B C
Clinical pregnancy rate (%)
50.0
40.0
30.0
20.0
10.0
0.0 Day 3
Day HCG
Time
Figure 1 Clinical pregnancy rate by endometrial pattern on day 3 of gonadotrophin stimulation and on the day of HCG administration. Pattern A: a triple-line pattern consisting of a central hyperechoic line surrounded by two hypoechoic layers; pattern B: an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and a poorly defined central echogenic line and pattern C: homogenous, hyperechogenic endometrium.
change with endometrial thickness (P < 0.001). The clinical pregnancy rates in women with endometrial patterns A, B, and C on day 3 of gonadotrophin stimulation were similar (49.8% v 51.7% v 53.6%). The clinical pregnancy rate in women with endometrial patterns A (55.2%) and B (50.9%) on the day of HCG administration was significantly greater (P < 0.05) than that of women with endometrial pattern C (37.4%). Endometrial thickness was further evaluated at threshold increments of 1 mm to assess its discriminatory ability for clinical pregnancy. Pregnancy rates ranged from 28.6% among women with an endometrial thickness of 6 mm or less to 67.7% among women with an endometrial thickness of over 6 mm (Table 3). Of the 14 women with endometrial thickness below 6 mm, four achieved clinical pregnancy. One of these pregnancies was in a woman with the lowest endometrial thickness (4.8 mm). Binary logistic regression analysis was used to assess the effect on clinical pregnancy of age, endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial pattern on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, basal FSH, oestradiol, P-values on the day of HCG administration, number of oocytes retrieved and the number of embryos produced. The analysis indicated that age (R = −0.047, P < 0.001), endometrial thickness on day 3 of gonadotrophin stimulation (R = −0.097, P < 0.05), endometrial pattern on the day of HCG administration (R = −0.228, P < 0.05) were negatively correlated with clinical pregnancy, and that increasing endometrial
thickness on the day of HCG administration (R = 0.150, P < 0.001), and the number of embryos (R = 0.046, P < 0.05) were associated with improved clinical pregnancy rates. Endometrial pattern on day 3 of gonadotrophin stimulation, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, basal FSH, oestradiol and P-values on the day of HCG administration and the number of oocytes retrieved were not significantly correlated with clinical pregnancy (Table 4). The ROC curve of endometrial thickness was analysed on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, and the change in endometrial thickness 3 days after gonadotrophin stimulation to the day of HCG administration to assess their predictive value for pregnancy. In the present study, the ROC curve analysis revealed that the area under the curve (AUC) was 0.528. As the increased endometrial thickness on day 3 of gonadotrophin stimulation was negatively associated with clinical pregnancy rate, the area under the curve was 0.428 (1–0.472) (95% confidence interval, 0.503–0.554) for endometrial thickness on day 3 of gonadotrophin stimulation compared with pregnancy, which was 0.596 (95% confidence interval, 0.571 to 0.621) for endometrial thickness on the day of HCG administration versus pregnancy, and was 0.606 (95% confidence interval, 0.580–0.630) for the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration versus pregnancy (Figure 2). Additionally, discriminant analysis was also applied to evaluate the predictive ability of all the endometrial parameters combined, including endometrial thickness on day 3 of gonadotrophin stimulation, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on the day of HCG administration and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration. The results indicated that the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration was excluded from the analysis, because the within-group variance was 6.249, higher than the minimum tolerance. The standardized canonical discriminant function coefficients of endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial pattern on day of HCG administration were −0.399, 0.855, 0.074, −0.421, respectively (not shown). According to the discriminant analysis, 58.7% of the original grouped cases and 58.4% of cross-validated grouped cases were correctly classified (Table 5).
Discussion So far, only four studies have evaluated the change in endometrial thickness occurring during IVF stimulation (Bassil, 2001; Gonen and Casper, 1990; Gonen et al., 1989; McWilliams and Frattarelli, 2007). To our knowledge, the present study has the largest sample size, with 3319 IVF cycles, and assesses the effect of endometrial thickness, pattern and change in endometrial thickness during ovarian stimulation on clinical pregnancy rate.
Endometrial characteristics and IVF-ET outcome
295
Table 3 Cycle outcomes according to endometrial thickness on the day of HCG administration. Endometrial thickness (mm) ≤6 >6 to ≤7 >7 to ≤8 >8 to ≤9 >9 to ≤10 >10 to ≤11 >11 to ≤12 >12 to ≤13 >13 to ≤14 >14 to ≤15 >15 to ≤16 >16 Table 4
Cycles n
Clinical pregnancy rate (n) %
Number of embryos transferred (n)
Implantation rate n (%)
14 33 137 269 360 357 297 207 122 60 43 34
4/13 (28.57) 8/33 (24.24) 45/137 (32.85) 126/269 (46.84) 177/360 (49.17) 190/357 (53.22) 172/297 (57.91) 120/207 (57.97) 81/122 (66.39) 40/60 (66.67) 24/43 (55.81) 23/34 (67.65)
31 69 290 581 759 766 634 433 251 129 91 69
4/31 (12.90) 9/69 (13.04) 60/290 (20.69) 175/581 (30.12) 239/759 (31.49) 255/766 (33.29) 234/634 (36.91) 164/433 (37.88) 110/251 (43.82) 49/129 (37.98) 33/91 (36.26) 31/69 (44.93)
Binary logistic regressiona (model R2 = 0.099, P < 0.001).
Independent variable Maternal age Endometrial thickness on day 3 of stimulation Endometrial pattern on day 3 of stimulation Baseline FSH Endometrial thickness on day of HCG administration Endometrial pattern on day of HCG administration Change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration Oestradiol level on HCG day P-value on HCG day Number of oocytes retrieved Number of embryos
Rb
P-valuec
−0.047 −0.097 0.057 −0.002 0.150 −0.228 0.007
0.000 0.002 NS NS 0.000 0.003 NS
0.000 −0.197 −0.020 0.046
NS NS NS 0.010
NS, not statistically significant. a Clinical pregnancy served as dependent variable; Method = enter. b Partial coefficients. c P-values. R values were considered statistically non-significant unless P < 0.05.
The day of stimulation within each cycle on which the endometrial characteristics were measured varies between studies. Some investigators measured them on the day of HCG administration (Basir et al., 2002; Bassil, 2001; Puerto et al., 2003; Sharara et al., 1999), oocyte retrieval (Bassil, 2001; Gonen and Casper, 1990; Sharara et al., 1999) or embryo transfer (Bassil, 2001; Kovacs et al., 2003; Puerto et al., 2003; Raga et al., 1999), and after pituitary suppression, on the sixth day of gonadotrophin stimulation, and on the day of HCG administration (McWilliams and Frattarelli, 2007), which makes it difficult to compare between studies. The endometrial thickness on day 3 of gonadotrophin stimulation can be seen as the basal status when menstrual bleeding has just stopped, and the endometrial thickness on the day of HCG administration is often used to document adequate endometrial development. So, we used the measurement taken at these two defined points. Recently, Al-Ghamdi et al. (2008) demonstrated a significant increase in pregnancy rate as endometrial thickness
increased in a 2464-cycle cohort study. McWilliams and Frattarelli (2007) confirmed this result, reporting that pregnant women had significantly greater endometrial thickness on day 6 of gonadotrophin stimulation and on the day of HCG administration, and had a greater change in endometrial thickness compared with non-pregnant women. In reviewing our data, pregnant women have a significantly but slightly thinner endometrial thickness on day 3 of gonadotrophin stimulation than that of non-pregnant women (6.03 ± 1.55 mm versus 6.23 ± 1.7 2 mm, P = 0.008), but the difference is not clinically significant, with the results falling within the range of measurement error. A significant difference was observed in endometrial thickness on the day of HCG administration, and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration between pregnant women and non-pregnant women. This finding indicates that adequate endometrial development is favourable for improved pregnancy rate. We postulate that the endometrium of non-pregnant women may be
296
J Zhao et al. ROC Curve 1.0 Source of the curve The endometrial thickness on day 3 of stimulation. The endometrial thickness on day of HCG administration.
0.8
Sensitivity
The change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration.
0.6
Reference line.
0.4
0.2
0.0 0.0
0.2
0.4
0.6
0.8
1.0
1 - Specificity Diagonal segments are produced by ties.
Figure 2 Receiver operator characteristic curve of endometrial thickness parameters (endometrial thickness on the day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration). The area under the curve was 0.428 for endometrial thickness on the day 3 of gonadotrophin stimulation, 0.596 for endometrial thickness on the day of HCG administration, 0.606 for the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration. Diagonal segments are produced by ties.
Table 5
Classification of results with discriminant analysis.a,b Predicted group membership
Clinical outcome
Original
Count %
Cross-validatedc
Count %
Non-pregnancy Pregnancy Non-pregnancy Pregnancy Non-pregnancy Pregnancy Non-pregnancy Pregnancy
Non-pregnancy
pregnancy
560 435 60.7 43.1 557 438 60.3 43.4
363 575 39.3 56.9 366 572 39.7 56.6
Total
923 1010 100.0 100.0 923 1010 100.0 100.0
a
58.7% of original grouped cases correctly classified. 58.4% of cross-validated grouped cases correctly classified. c Cross-validation is performed only for those cases in the analysis. In cross validation, each case is classified by the functions derived from all cases other than that case. b
associated with pathological abnormalities, such as squamous cell metaplasia or lacking a normal proliferative response to the rising oestradiol level. In a binary logistic regression model, the endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration were independent predictive factors for pregnancy. A few studies (Al-Ghamdi et al., 2008; Richter et al., 2007; Weissman et al., 1999), including the present study, have shown a steady and gradual increase in pregnancy rate as endometrial thickness
increases, and do not show a detrimental effect of thicker endometrium on pregnancy rate; however, Weissman et al. 1999 reported a high miscarriage rate with increased endometrial thickness (>14 mm). Most studies (Noyes et al., 2001; Rashidi et al., 2004) have reported a significant difference in pregnancy rate above and below a threshold thickness, but did not show a continuous relationship as reported in this study. In the present study, no threshold values for endometrial thickness were identified. To assess the ability of endometrial thickness to predict pregnancy, ROC curves were used. The area under the curve, calculated as (sensitivity/[1-specificity]), is a measure that
Endometrial characteristics and IVF-ET outcome shows how ‘good’ (AUC close to 1) or ‘bad’ (AUC close to 0.5) a test is. In the present study, the results from ROC curve analysis (Figure 2) suggest that the data analysed in this study cannot establish a true endometrial thickness ‘cut-off’ value, and these endometrial parameters have no predictive value for pregnancy; however, adequate endometrial development is required for pregnancy to occur. In contrast to our results, several investigators, such as Gonen and Casper (1990) and Bassil (2001), showed that endometrial responsiveness and thickness during the IVF stimulation seem to be better prognostic predictors of success. Additionally, some studies (Kovacs et al., 2003; Noyes et al., 1995; Wang et al., 2010) reached the same conclusion as the current report, in which the authors believe that endometrial thickness or morphology has no relationship with IVF–embryo transfer outcome. In addition to endometrial thickness, many published studies have assessed the correlation between endometrial pattern and the outcome of assisted reproductive techniques, with conflicting conclusions (Gonen et al., 1989; Khalifa et al., 1992; Mercé et al., 2008; Noyes et al., 2001; Oliveira et al., 1997; Sharara et al., 1999; Chen et al., 2010; Zaidi et al., 1995). In the present study, we found that women with endometrial pattern A or B on day of HCG administration had a significantly higher clinical pregnancy rate than that of women with pattern C, whereas the clinical pregnancy rates were similar in women with different endometrial patterns on day 3 of stimulation. These data indicate that the basal endometrial pattern on day 3 of gonadotrophin stimulation has no positive or negative correlation with clinical pregnancy rate, and the endometrial pattern on the day of HCG administration is correlated with clinical pregnancy rates. Our data contrast with the results of other authors (Bassil, 2001; Khalifa et al., 1992), who showed similar pregnancy rates between endometrial patterns A and B. The present data, however, were in accordance with the results described by Sher et al. (1991) and Bohrer et al. (1996). The former found a significant difference in pregnancy rates (33% versus 7%) in favour of the triple-line endometrium associated with an endometrial thickness of over 9 mm. The latter concluded that a homogeneous endometrium (pregnancy rate, 2.9%) that fails to convert to the trilaminar pattern (pregnancy rate, 23%) is less receptive to implantation. The contradictory conclusion among these authors may partly be due to the differences in laboratory and stimulation methods, heterogeneity of the various studies, and differences in statistical evaluation. A triple-line pattern at ultrasound examination reflects endometrial proliferation. The presence of such a pattern on the day of HCG injection has been found to be associated with a higher pregnancy rate than is the absence of this pattern. The absence of a triple-line pattern may be a sign of premature secretory changes in the endometrium, and that the time of maximal endometrial receptivity has passed (Bourgain and Devroey, 2003). In addition to independent evaluation of endometrial characteristics on clinical outcome, we also assessed the combined predictive effect of the endometrial characteristics, including endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, endometrial pattern on the day 3 of
297 gonadotrophin stimulation, and endometrial pattern on the day of HCG administration, on the clinical outcome with discriminant analysis. The result indicated that combined endometrial thickness, changes and endometrial pattern could not predict the outcome of IVF–embryo transfer correctly. What we found in the present study was not in accordance with an earlier study (Chen et al., 2010). One earlier study showed that the pathological changes in the endometrium may be a cause of infertility (Ruız-Velasco et al., 1997). A thin endometrium or a homogenous, hyperechogenic endometrium on the day of HCG administration may be associated with pathological abnormalities, which may interfere with normal embryo implantation, and ultrasound examination may not be adequate to identify pathology abnormalities in the endometrium. An important limitation of this study is that it is retrospective in nature. We believe, however, that the results are of interest because similar studies have been published with conflicting results. A well-designed and powered randomized clinical trial will be needed to determine which results, if any, are substantiated.
Conclusion The endometrium, with adequate growth, triple-line on HCG day, or both, seems to be favourable for pregnancy. The endometrial thickness and the change, however, are not prognostically useful in predicting the occurrence of pregnancy, and the combined endometrial characteristics cannot predict the clinical outcome correctly.
Acknowledgement The authors thank everyone in the department of Reproductive Medicine in the Centre South University Xiangya Hospital for their scientific advice and encouragement.
References Al-Ghamdi, A., Coskun, S., Al-Hassan, S., AL-Rejjal, R., Awartani, K., 2008. The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET) outcome. Reprod. Biol. Endocrinol. 6, 37. Basir, G.S., O, W.S., So, W.W., Ng, E.H., Ho, P.C., 2002. Evaluation of cycle-to-cycle variation of endometrial responsiveness using transvaginal sonography in women undergoing assisted reproduction. Ultrasound Obstet. Gynecol. 19, 4844– 4889. Bassil, S., 2001. Changes in endometrial thickness, width, length and pattern in predicting pregnancy outcome during ovarian stimulation in in vitro fertilization. Ultrasound Obstet. Gynecol. 18, 258– 263. Bohrer, M.K., Hock, D.L., Rhoads, G.G., Kemmann, E., 1996. Sonographic assessment of endometrial pattern and thickness in patients treated with human menopausal gonadotropins. Fertil. Steril. 66, 2442–2447. Bourgain, C., Devroey, P., 2003. The endometrium in stimulated cycles for IVF. Hum. Reprod. Update 9, 515–522. Chen, S.-L., Wu, F.-R., Luo, C., Chen, X., Shi, X.-Y., Zheng, H.-Y., Ni, Y.-P., 2010. Combined analysis of endometrial thickness and
298 pattern in predicting outcome of in vitro fertilization and embryo transfer: a retrospective cohort study. Reprod. Biol. Endocrinol. 8, 30. De Geyter, C., Schmitter, M., De Geyter, M., Nieschlag, E., Holzgreve, W., Schneider, H.P., 2000. Prospective evaluation of the ultrasound appearance of the endometrium in a cohort of 1,186 infertile women. Fertil. Steril. 73, 106–113. Dickey, R.P., Olar, T.T., Curole, D.N., Taylor, S.N., Rye, P.H., 1992. Endometrial pattern and thickness associated with pregnancy outcome after assisted reproduction technologies. Hum. Reprod. 7, 418–421. Dietterich, C., Check, J.H., Choe, J.K., Nazari, A., Lurie, D., 2002. Increased endometrial thickness on the day of human chorionic gonadotropin injection does not adversely affect pregnancy or implantation rates following in vitro fertilization-embryo transfer. Fertil. Steril. 77, 7817–7886. Fanchin, R., 2001. Assessing uterine receptivity in 2001. Ann. N. Y. Acad. Sci. 943, 185–202. Gonen, Y., Casper, R.F., 1990. Prediction of implantation by the sonographic appearance of the endometrium during controlled ovarian stimulation for in vitro fertilization (IVF). J. In Vitro Fert. Embryo Transf. 7, 146–152. Gonen, Y., Casper, R.F., Jacobson, W., Blankier, J., 1989. Endometrial thickness and growth during ovarian stimulation: a possible predictor of implantation in in vitro fertilization. Fertil. Steril. 52, 446–450. Khalifa, E., Brzyski, R.G., Oehninger, S., Acosta, A.A., Muasher, S.J., 1992. Sonographic appearance of the endometrium: the predictive value for the outcome of in-vitro fertilization in stimulated cycles. Hum. Reprod. 7, 6776–6780. Kovacs, P., Matyas, S., Boda, K., Kaali, S.G., 2003. The effect of endometrial thickness on IVF/ICSI outcome. Hum. Reprod. 18, 2337– 2341. McWilliams, G.D., Frattarelli, J.L., 2007. Changes in measured endometrial thickness predict in vitro fertilization success. Fertil. Steril. 88, 74–81. Mercé, L.T., Barco, M.J., Bau, S., Troyano, J., 2008. Are endometrial parameters by three-dimensional ultrasound and power Doppler angiography related to in vitro fertilization/embryo transfer outcome? Fertil. Steril. 89, 1111–1117. Noyes, N., Liu, H.C., Sultan, K., Schattman, G., Rosenwaks, Z., 1995. Endometrial thickness appears to be a significant factor in embryo implantation in in-vitro fertilization. Hum. Reprod. 10, 919– 922. Noyes, N., Hampton, B.S., Berkeley, A., Licciardi, F., Grifo, J., Krey, L., 2001. Factors useful in predicting the success of oocyte donation: a 3-year retrospective analysis. Fertil. Steril. 76, 92–97. Oliveira, J.B., Baruffi, R.L., Mauri, A.L., Petersen, C.G., Borges, M.C., Franco, J.G., 1997. Endometrial ultrasonography as a predictor of pregnancy in an in-vitro fertilization programme after ovarian stimulation and gonadotrophin-releasing hormone and gonadotrophins. Hum. Reprod. 12, 25152–25518. Puerto, B., Creus, M., Carmona, F., Civico, S., Vanrell, J.A., Balasch, J., 2003. Ultrasonography as a predictor of embryo
J Zhao et al. implantation after in vitro fertilization: a controlled study. Fertil. Steril. 79, 1015–1022. Raga, F., Bonilla-Musoles, F., Casan, E.M., Klein, O., Bonilla, F., 1999. Assessment of endometrial volume by three-dimensional ultrasound prior to embryo transfer: clues to endometrial receptivity. Hum. Reprod. 14, 28512–28854. Rashidi, B.H., Sadeghi, M., Jafarabadi, M., Nejad, E.S.T., 2004. Relationships between pregnancy rates following in vitro fertilization or intracytoplasmic sperm injection and endometrial thickness and pattern. Eur. J. Obstet. Gynecol. Reprod. Biol. 120, 179– 184. Richter, K.S., Bugge, K.R., Bromer, J.G., Levy, M.J., 2007. Relationship between endometrial thickness and embryo implantation, based on 1,294 cycles of in vitro fertilization with transfer of two blastocyst-stage embryos. Fertil. Steril. 87, 535–539. Rinaldi, L., Lisi, F., Floccari, A., Lisi, R., Pepe, G., Fishel, S., 1996. Endometrial thickness as a predictor of pregnancy after in-vitro fertilization but not after intracytoplasmic sperm injection. Hum. Reprod. 11, 15381–15541. Ruız-Velasco, V., Alfani, G.G., Sanchez, L.P., Marcela, V.A., 1997. Endometrial pathology and infertility. Fertil. Steril. 67, 6876– 6892. Sharara, F.I., Lim, J., McClamrock, H.D., 1999. Endometrial pattern on the day of oocyte retrieval is more predictive of implantation success than the pattern or thickness on the day of hCG administration. J. Assist. Reprod. Genet. 16, 523– 528. Sher, G., Herbert, C., Maassarani, G., Jacobs, M.H., 1991. Assessment of the late proliferative phase endometrium by ultrasonography in patients undergoing in-vitro fertilization and embryo transfer (IVF/ET). Hum. Reprod. 6, 2322–2337. Wang, L., Qiao, J., Li, R., Zhen, X., Liu, Z., 2010. Role of endometrial blood flow assessment with color Doppler energy in predicting pregnancy outcome of IVF-ET cycles. Reprod. Biol. Endocrinol. 8, 122. Weissman, A., Gotlieb, L., Casper, R.F., 1999. The detrimental effect of increased endometrial thickness on implantation and pregnancy rates and outcome in an in vitro fertilization program. Fertil. Steril. 71, 147–149. Yuval, Y., Lipitiz, S., Dor, J., Achiron, R., 1999. The relationships between endometrial thickness, and blood flow and pregnancy rates in in-vitro fertilization. Hum. Reprod. 14, 1067–1071. Zaidi, J., Campbell, S., Pittrof, R., Tan, S.L., 1995. Endometrial thickness, morphology, vascular penetration and velocimetry in predicting implantation in an in vitro fertilization program. Ultrasound Obstet. Gynecol. 6, 1911–1998.
Declaration: The authors report no financial or commercial conflicts of interest.
Received 22 February 2014; refereed 11 May 2014; accepted 21 May 2014.
w w w. s c i e n c e d i r e c t . c o m w w w. r b m o n l i n e . c o m
ARTICLE
Endometrial pattern, thickness and growth in predicting pregnancy outcome following 3319 IVF cycle Jing Zhao, Qiong Zhang, Yonggang Wang, Yanping Li * Reproductive Medicine Center, Xiangya Hospital, Central South University, Changsha, Hunan, China * Corresponding author.
E-mail address: [email protected] (Y Li). Yanping Li, MD, is a professor and doctoral supervisor of Central South University, Changsha, China and the Director of Reproductive Medicine Center, Department of Obstetrics and Gynecology, Xiangya Hospital. She performs more than 1000 IVF cycles per year and the clinical pregnancy rate is almost 50%. She conducts clinical work on infertility, IVF, preimplantation genetic diagnosis, intrauterine insemination and microsurgery, and research on reproductive endocrinology, poor ovarian response, thin endometria and embryonic stem cells.
A retrospective study of 3319 women was conducted to assess predictive ability of endometrial characteristics for outcomes of IVF and embryo transfer. Endometrial thickness, growth and pattern were assessed at two time points (day 3 of gonadotrophin stimulation and day of HCG administration). Endometrial patterns were classified as pattern A: triple-line pattern comprising a central hyperechoic line surrounded by two hypoechoic layers; pattern B: an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and poorly defined central echogenic line; and pattern C: homogenous, hyperechogenic endometrium. The endometrium of pregnant women was thinner on day 3 of stimulation, thicker on the day of HCG administration, and showed greater growth in thickness compared with non-pregnant women. Clinical pregnancy rates differed according to endometrial pattern on the day of HCG administration (55.2%, 50.9% and 37.4% for patterns A, B and C, respectively). A positive linear relationship was found between endometrial thickness on the day of HCG administration and clinical pregnancy rate. Endometrial thickness, change and pattern were independent factors affecting outcome. Receiver operator characteristic curves showed that endometrial pattern, thickness and changes were not good predictors of clinical pregnancy. Discriminant analysis indicated that 58.7% of original grouped cases were correctly classified. Although endometrium with triple-line or increased thickness may favour pregnancy, combined endometrial characteristics do not predict outcomes.
Abstract
© 2014 Published by Elsevier Ltd on behalf of Reproductive Healthcare Ltd. KEYWORDS: endometrial growth, endometrial pattern, endometrial thickness, pregnancy rate, transvaginal ultrasound
http://dx.doi.org/10.1016/j.rbmo.2014.05.011 1472-6483/© 2014 Published by Elsevier Ltd on behalf of Reproductive Healthcare Ltd.
292
Introduction Assisted reproductive technology has been commonly used in infertility treatment over the past 3 decades. In addition to embryo quality, the receptivity of the endometrium plays an important role in clinical outcome. Sonography is a noninvasive and simple method that has been used to evaluate endometrial receptivity. Several sonographic parameters have been evaluated. These include endometrial thickness, endometrial pattern, endometrial volume and endometrial and subendometrial blood flow (Basir et al., 2002; Dickey et al., 1992, Fanchin, 2001; Wang et al., 2010; Yuval et al., 1999). Endometrial thickness and pattern have been evaluated as possible predictors of pregnancy in multiple studies, with conflicting results. Some investigators have reported significant correlations between pregnancy rate, endometrial thickness, pattern, or both (Al-Ghamdi et al., 2008; Chen et al., 2010; Noyes et al., 1995; Richter et al., 2007; Rinaldi et al., 1996), whereas others have not shown such a relationship (Basir et al., 2002; De Geyter et al., 2000; Dietterich et al., 2002; Rashidi et al., 2004). Only a few studies (Gonen and Casper, 1990; Gonen et al., 1989; McWilliams and Frattarelli, 2007); however, have evaluated the change in endometrial thickness occurring during IVF stimulation, and these studies reached different conclusions with small samples. This study was designed to assess the relationship between endometrial features (endometrial pattern and thickness and endometrium growth) and clinical outcome based on 3319 IVF stimulation cycles.
Materials and methods Patient recruitment and counselling The study was reviewed and approved by the Institutional Review Board and the Ethics Committee of Xiangya Hospital, Changsha, China (IRB reference number 201212047, dated 27 August, 2011). The study was conducted in accordance with the Declaration of Helsinki, as revised in 1983. We conducted a retrospective cohort study of 3319 consecutive infertile women. Briefly, the women underwent fresh IVF-ET between January 2009 and May 2011 at the Reproductive Medicine Centre of Xiangya hospital, Central South University (Changsha, China). Before undergoing IVF, patients were examined with ultrasonography more than once at the different stages of their menstrual cycles. If any subtle cavity abnormalities were found, those women were recommended to undergo further examination or treatment with hysteroscopy or saline sonohysterogram. Exclusion criteria included presence of known endometrial polyp, uterine anomaly, and insemination method other than IVF, cycles using donor oocytes or cryopreserved embryos. Patients underwent no therapeutic intervention other than routine procedures.
J Zhao et al. response and co-existing medical conditions. Ovarian stimulation was initiated when the serum oestradiol concentration level was ≤50 pg/ml, and the largest follicle diameter was <10 mm without ovarian cysts. Urofollitiopin injection (HMG; LiZhu, China), human menopausal gonadotrophin, or both, were used to achieve ovarian stimulation. The initial dosage of gonadotrophin ranged from 150 to 450 IU, depending on the basal FSH level, antral follicular count and maternal age. When at least two follicles were 18 mm or wider in diameter, 10,000 IU of HCG (HCG; Profasi; Serono, Italy) was administered. Oocyte retrieval was carried out 36 h after the administration of HCG, and was followed by conventional IVF. Up to three high-quality embryos were transferred 72 h after oocyte collection according to the centre’s standard practice. When available, surplus good-quality embryos were cryopreserved. The luteal phase was supported using 80 mg progesterone in oil (Progesterone; Tongyong, Shanghai, China) intramuscular injection daily lasting 75 days. Clinical pregnancy was defined as identification of a gestational sac 4–5 weeks after embryo transfer.
Ultrasound measurement Endometrial features assessed included endometrial thickness and pattern on day 3 of gonadotrophin stimulation and on the day of HCG administration, as well as the change in endometrial thickness from day 3 to day of HCG administration. All were measured by transvaginal 8 MHz ultrasonography with Doppler ultrasound (Mindray DC-6 Expert; Shenzhen, China). Endometrial thickness was measured in a median longitudinal plane of the uterus as the maximum distance between the endometrial–myometrial interface of the anterior to the posterior wall of the uterus. Endometrial pattern was classified as pattern A (a triple-line pattern consisting of a central hyperechoic line surrounded by two hypoechoic layers), pattern B (an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and a poorly defined central echogenic line), and pattern C (homogenous, hyperechogenic endometrium).
Statistical analysis Continuous data were expressed as mean ± SD values or as median and range, according to the distribution, and were analysed for differences using the Student’s t-test. Categorical data were presented as numbers, and chi-squared test was used for statistical comparison of percentages. Binary logistic regression analysis was also carried out, and the receiver operating characteristic (ROC) curve was applied to determine the predictive value of endometrial parameters. Discriminant analysis was used to evaluate the predictive ability of all of the endometrial parameters combined. The significance level for all analyses was P < 0.05. Statistical analysis was carried out using the Statistical Package for Social Sciences (SPSS Inc, Chicago, IL, USA).
Ovulation induction and IVF–embryo transfer procedures
Results
The choice of stimulation protocol was individualized on the basis of the patient’s age, cause of infertility, ovarian
A total of 3319 IVF cycles (3319 women) were included in the study. The clinical pregnancy rate, defined as identification
Endometrial characteristics and IVF-ET outcome
293 statistically significant differences in embryo quality were found between the groups (data not shown). Endometrial thickness and pattern measured on day 3 of gonadotrophin stimulation and on the day of HCG administration in both pregnant and non-pregnant women are shown in Table 2 and Figure 1. Pregnant women had significantly thinner endometrial linings on day 3 of gonadotrophin stimulation (P = 0.008), significantly thicker endometrial linings on the day of HCG administration (P < 0.001), and a greater
of a gestational sac 4–5 weeks after embryo transfer, was 52.3%. The demographic data and ovarian responses are presented in Table 1. When subdividing the women into those who conceived (n = 1010) and those who did not conceive (n = 923), significant differences were observed between the two groups. These included age, body mass index, basal FSH, basal oestradiol, duration of ovarian stimulation, total gonadotrophin dose, P-value on the day of HCG administration, and total number of embryos produced (P < 0.05). No
Table 1
Parameters for the IVF study group.
Variables Age (years) BMI(kg/m2) Baseline FSH (IU/L) Baseline LH (IU/L) Baseline E2 (pg//mL) Days of stimulation (days) Ampules of gonadotrophin used (IU) Oestradiol on HCG day (pg/mL) P-value on HCG day (ng/mL) Number of oocytes retrieved Number of embryos Number of embryos transferred Cause of infertility; n (%) Tubal factor Endometriosis PCOS Multiple factors
All patients (n = 1933)
Pregnanta (n = 1010)
Non-pregnant (n = 923)
P-value
31.2 ± 4.6 21.73 ± 2.87 6.3 ± 2.9 4.5 ± 2.8 33.2 ± 21.9 11.1 ± 2.3
30.6 ± 4.4 21.6 ± 2.6 6.1 ± 2.9 4.5 ± 3.0 31.9 ± 22.0 11.2 ± 2.3
31.8 ± 4.8 21.9 ± 3.1 6.4 ± 2.9 4.5 ± 2.4 34.6 ± 21.6 11.0 ± 2.3
<0.05 <0.05 <0.05 NS <0.05 <0.05
2115.0 ± 832.5
2040 ± 772.5
2197.5 ± 885.0
<0.05
3489.7 ± 2112.2
3514.2 ± 2026.8
3462.9 ± 2202.7
NS
0.62 ± 0.48
0.60 ± 0.46
0.64 ± 0.53
<0.05
13.0 ± 5.8
13.2 ± 5.7
12.7 ± 6.0
NS
7.3 ± 4.1 2.3 ± 0.2
7.6 ± 3.9 2.3 ± 0.3
7.0 ± 4.3 2.2 ± 0.3
<0.05 NS NS
1767 (91.4) 23 (1.2) 23 (1.2) 120 (6.2)
924 (91.5) 12 (1.2) 12 (1.2) 62 (6.1)
843 (91.3) 11 (1.2) 11 (1.2) 58 (6.3)
BMI, body mass index; NS, not significant; PCOS, polycystic ovary syndrome. a Patients who achieved a clinical pregnancy.
Table 2 Comparison of endometrial thickness at three points in pregnant and nonpregnant women. Variable Endometrial thickness on day 3 of stimulation (mm) Endometrial thickness on day of HCG administration (mm) Change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration (mm) Data shown as mean ± SD. P < 0.05 was statistically significant. a Clinical pregnancy.
Pregnanta (n = 1010)
Non-pregnant (n = 923)
6.0 ± 1.6
6.2 ± 1.7
0.008
11.0 ± 2.2
10.3 ± 2.2
<0.001
5.0 ± 2.5
4.1 ± 2.6
<0.001
P-value
294
J Zhao et al. P < 0.05
60.0
pattern A B C
Clinical pregnancy rate (%)
50.0
40.0
30.0
20.0
10.0
0.0 Day 3
Day HCG
Time
Figure 1 Clinical pregnancy rate by endometrial pattern on day 3 of gonadotrophin stimulation and on the day of HCG administration. Pattern A: a triple-line pattern consisting of a central hyperechoic line surrounded by two hypoechoic layers; pattern B: an intermediate isoechogenic pattern with the same reflectivity as the surrounding myometrium and a poorly defined central echogenic line and pattern C: homogenous, hyperechogenic endometrium.
change with endometrial thickness (P < 0.001). The clinical pregnancy rates in women with endometrial patterns A, B, and C on day 3 of gonadotrophin stimulation were similar (49.8% v 51.7% v 53.6%). The clinical pregnancy rate in women with endometrial patterns A (55.2%) and B (50.9%) on the day of HCG administration was significantly greater (P < 0.05) than that of women with endometrial pattern C (37.4%). Endometrial thickness was further evaluated at threshold increments of 1 mm to assess its discriminatory ability for clinical pregnancy. Pregnancy rates ranged from 28.6% among women with an endometrial thickness of 6 mm or less to 67.7% among women with an endometrial thickness of over 6 mm (Table 3). Of the 14 women with endometrial thickness below 6 mm, four achieved clinical pregnancy. One of these pregnancies was in a woman with the lowest endometrial thickness (4.8 mm). Binary logistic regression analysis was used to assess the effect on clinical pregnancy of age, endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial pattern on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, basal FSH, oestradiol, P-values on the day of HCG administration, number of oocytes retrieved and the number of embryos produced. The analysis indicated that age (R = −0.047, P < 0.001), endometrial thickness on day 3 of gonadotrophin stimulation (R = −0.097, P < 0.05), endometrial pattern on the day of HCG administration (R = −0.228, P < 0.05) were negatively correlated with clinical pregnancy, and that increasing endometrial
thickness on the day of HCG administration (R = 0.150, P < 0.001), and the number of embryos (R = 0.046, P < 0.05) were associated with improved clinical pregnancy rates. Endometrial pattern on day 3 of gonadotrophin stimulation, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, basal FSH, oestradiol and P-values on the day of HCG administration and the number of oocytes retrieved were not significantly correlated with clinical pregnancy (Table 4). The ROC curve of endometrial thickness was analysed on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, and the change in endometrial thickness 3 days after gonadotrophin stimulation to the day of HCG administration to assess their predictive value for pregnancy. In the present study, the ROC curve analysis revealed that the area under the curve (AUC) was 0.528. As the increased endometrial thickness on day 3 of gonadotrophin stimulation was negatively associated with clinical pregnancy rate, the area under the curve was 0.428 (1–0.472) (95% confidence interval, 0.503–0.554) for endometrial thickness on day 3 of gonadotrophin stimulation compared with pregnancy, which was 0.596 (95% confidence interval, 0.571 to 0.621) for endometrial thickness on the day of HCG administration versus pregnancy, and was 0.606 (95% confidence interval, 0.580–0.630) for the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration versus pregnancy (Figure 2). Additionally, discriminant analysis was also applied to evaluate the predictive ability of all the endometrial parameters combined, including endometrial thickness on day 3 of gonadotrophin stimulation, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on the day of HCG administration and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration. The results indicated that the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration was excluded from the analysis, because the within-group variance was 6.249, higher than the minimum tolerance. The standardized canonical discriminant function coefficients of endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, endometrial pattern on day 3 of gonadotrophin stimulation, endometrial pattern on day of HCG administration were −0.399, 0.855, 0.074, −0.421, respectively (not shown). According to the discriminant analysis, 58.7% of the original grouped cases and 58.4% of cross-validated grouped cases were correctly classified (Table 5).
Discussion So far, only four studies have evaluated the change in endometrial thickness occurring during IVF stimulation (Bassil, 2001; Gonen and Casper, 1990; Gonen et al., 1989; McWilliams and Frattarelli, 2007). To our knowledge, the present study has the largest sample size, with 3319 IVF cycles, and assesses the effect of endometrial thickness, pattern and change in endometrial thickness during ovarian stimulation on clinical pregnancy rate.
Endometrial characteristics and IVF-ET outcome
295
Table 3 Cycle outcomes according to endometrial thickness on the day of HCG administration. Endometrial thickness (mm) ≤6 >6 to ≤7 >7 to ≤8 >8 to ≤9 >9 to ≤10 >10 to ≤11 >11 to ≤12 >12 to ≤13 >13 to ≤14 >14 to ≤15 >15 to ≤16 >16 Table 4
Cycles n
Clinical pregnancy rate (n) %
Number of embryos transferred (n)
Implantation rate n (%)
14 33 137 269 360 357 297 207 122 60 43 34
4/13 (28.57) 8/33 (24.24) 45/137 (32.85) 126/269 (46.84) 177/360 (49.17) 190/357 (53.22) 172/297 (57.91) 120/207 (57.97) 81/122 (66.39) 40/60 (66.67) 24/43 (55.81) 23/34 (67.65)
31 69 290 581 759 766 634 433 251 129 91 69
4/31 (12.90) 9/69 (13.04) 60/290 (20.69) 175/581 (30.12) 239/759 (31.49) 255/766 (33.29) 234/634 (36.91) 164/433 (37.88) 110/251 (43.82) 49/129 (37.98) 33/91 (36.26) 31/69 (44.93)
Binary logistic regressiona (model R2 = 0.099, P < 0.001).
Independent variable Maternal age Endometrial thickness on day 3 of stimulation Endometrial pattern on day 3 of stimulation Baseline FSH Endometrial thickness on day of HCG administration Endometrial pattern on day of HCG administration Change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration Oestradiol level on HCG day P-value on HCG day Number of oocytes retrieved Number of embryos
Rb
P-valuec
−0.047 −0.097 0.057 −0.002 0.150 −0.228 0.007
0.000 0.002 NS NS 0.000 0.003 NS
0.000 −0.197 −0.020 0.046
NS NS NS 0.010
NS, not statistically significant. a Clinical pregnancy served as dependent variable; Method = enter. b Partial coefficients. c P-values. R values were considered statistically non-significant unless P < 0.05.
The day of stimulation within each cycle on which the endometrial characteristics were measured varies between studies. Some investigators measured them on the day of HCG administration (Basir et al., 2002; Bassil, 2001; Puerto et al., 2003; Sharara et al., 1999), oocyte retrieval (Bassil, 2001; Gonen and Casper, 1990; Sharara et al., 1999) or embryo transfer (Bassil, 2001; Kovacs et al., 2003; Puerto et al., 2003; Raga et al., 1999), and after pituitary suppression, on the sixth day of gonadotrophin stimulation, and on the day of HCG administration (McWilliams and Frattarelli, 2007), which makes it difficult to compare between studies. The endometrial thickness on day 3 of gonadotrophin stimulation can be seen as the basal status when menstrual bleeding has just stopped, and the endometrial thickness on the day of HCG administration is often used to document adequate endometrial development. So, we used the measurement taken at these two defined points. Recently, Al-Ghamdi et al. (2008) demonstrated a significant increase in pregnancy rate as endometrial thickness
increased in a 2464-cycle cohort study. McWilliams and Frattarelli (2007) confirmed this result, reporting that pregnant women had significantly greater endometrial thickness on day 6 of gonadotrophin stimulation and on the day of HCG administration, and had a greater change in endometrial thickness compared with non-pregnant women. In reviewing our data, pregnant women have a significantly but slightly thinner endometrial thickness on day 3 of gonadotrophin stimulation than that of non-pregnant women (6.03 ± 1.55 mm versus 6.23 ± 1.7 2 mm, P = 0.008), but the difference is not clinically significant, with the results falling within the range of measurement error. A significant difference was observed in endometrial thickness on the day of HCG administration, and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration between pregnant women and non-pregnant women. This finding indicates that adequate endometrial development is favourable for improved pregnancy rate. We postulate that the endometrium of non-pregnant women may be
296
J Zhao et al. ROC Curve 1.0 Source of the curve The endometrial thickness on day 3 of stimulation. The endometrial thickness on day of HCG administration.
0.8
Sensitivity
The change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration.
0.6
Reference line.
0.4
0.2
0.0 0.0
0.2
0.4
0.6
0.8
1.0
1 - Specificity Diagonal segments are produced by ties.
Figure 2 Receiver operator characteristic curve of endometrial thickness parameters (endometrial thickness on the day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration). The area under the curve was 0.428 for endometrial thickness on the day 3 of gonadotrophin stimulation, 0.596 for endometrial thickness on the day of HCG administration, 0.606 for the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration. Diagonal segments are produced by ties.
Table 5
Classification of results with discriminant analysis.a,b Predicted group membership
Clinical outcome
Original
Count %
Cross-validatedc
Count %
Non-pregnancy Pregnancy Non-pregnancy Pregnancy Non-pregnancy Pregnancy Non-pregnancy Pregnancy
Non-pregnancy
pregnancy
560 435 60.7 43.1 557 438 60.3 43.4
363 575 39.3 56.9 366 572 39.7 56.6
Total
923 1010 100.0 100.0 923 1010 100.0 100.0
a
58.7% of original grouped cases correctly classified. 58.4% of cross-validated grouped cases correctly classified. c Cross-validation is performed only for those cases in the analysis. In cross validation, each case is classified by the functions derived from all cases other than that case. b
associated with pathological abnormalities, such as squamous cell metaplasia or lacking a normal proliferative response to the rising oestradiol level. In a binary logistic regression model, the endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, and the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration were independent predictive factors for pregnancy. A few studies (Al-Ghamdi et al., 2008; Richter et al., 2007; Weissman et al., 1999), including the present study, have shown a steady and gradual increase in pregnancy rate as endometrial thickness
increases, and do not show a detrimental effect of thicker endometrium on pregnancy rate; however, Weissman et al. 1999 reported a high miscarriage rate with increased endometrial thickness (>14 mm). Most studies (Noyes et al., 2001; Rashidi et al., 2004) have reported a significant difference in pregnancy rate above and below a threshold thickness, but did not show a continuous relationship as reported in this study. In the present study, no threshold values for endometrial thickness were identified. To assess the ability of endometrial thickness to predict pregnancy, ROC curves were used. The area under the curve, calculated as (sensitivity/[1-specificity]), is a measure that
Endometrial characteristics and IVF-ET outcome shows how ‘good’ (AUC close to 1) or ‘bad’ (AUC close to 0.5) a test is. In the present study, the results from ROC curve analysis (Figure 2) suggest that the data analysed in this study cannot establish a true endometrial thickness ‘cut-off’ value, and these endometrial parameters have no predictive value for pregnancy; however, adequate endometrial development is required for pregnancy to occur. In contrast to our results, several investigators, such as Gonen and Casper (1990) and Bassil (2001), showed that endometrial responsiveness and thickness during the IVF stimulation seem to be better prognostic predictors of success. Additionally, some studies (Kovacs et al., 2003; Noyes et al., 1995; Wang et al., 2010) reached the same conclusion as the current report, in which the authors believe that endometrial thickness or morphology has no relationship with IVF–embryo transfer outcome. In addition to endometrial thickness, many published studies have assessed the correlation between endometrial pattern and the outcome of assisted reproductive techniques, with conflicting conclusions (Gonen et al., 1989; Khalifa et al., 1992; Mercé et al., 2008; Noyes et al., 2001; Oliveira et al., 1997; Sharara et al., 1999; Chen et al., 2010; Zaidi et al., 1995). In the present study, we found that women with endometrial pattern A or B on day of HCG administration had a significantly higher clinical pregnancy rate than that of women with pattern C, whereas the clinical pregnancy rates were similar in women with different endometrial patterns on day 3 of stimulation. These data indicate that the basal endometrial pattern on day 3 of gonadotrophin stimulation has no positive or negative correlation with clinical pregnancy rate, and the endometrial pattern on the day of HCG administration is correlated with clinical pregnancy rates. Our data contrast with the results of other authors (Bassil, 2001; Khalifa et al., 1992), who showed similar pregnancy rates between endometrial patterns A and B. The present data, however, were in accordance with the results described by Sher et al. (1991) and Bohrer et al. (1996). The former found a significant difference in pregnancy rates (33% versus 7%) in favour of the triple-line endometrium associated with an endometrial thickness of over 9 mm. The latter concluded that a homogeneous endometrium (pregnancy rate, 2.9%) that fails to convert to the trilaminar pattern (pregnancy rate, 23%) is less receptive to implantation. The contradictory conclusion among these authors may partly be due to the differences in laboratory and stimulation methods, heterogeneity of the various studies, and differences in statistical evaluation. A triple-line pattern at ultrasound examination reflects endometrial proliferation. The presence of such a pattern on the day of HCG injection has been found to be associated with a higher pregnancy rate than is the absence of this pattern. The absence of a triple-line pattern may be a sign of premature secretory changes in the endometrium, and that the time of maximal endometrial receptivity has passed (Bourgain and Devroey, 2003). In addition to independent evaluation of endometrial characteristics on clinical outcome, we also assessed the combined predictive effect of the endometrial characteristics, including endometrial thickness on day 3 of gonadotrophin stimulation, endometrial thickness on the day of HCG administration, the change in endometrial thickness from the third day of gonadotrophin stimulation to the day of HCG administration, endometrial pattern on the day 3 of
297 gonadotrophin stimulation, and endometrial pattern on the day of HCG administration, on the clinical outcome with discriminant analysis. The result indicated that combined endometrial thickness, changes and endometrial pattern could not predict the outcome of IVF–embryo transfer correctly. What we found in the present study was not in accordance with an earlier study (Chen et al., 2010). One earlier study showed that the pathological changes in the endometrium may be a cause of infertility (Ruız-Velasco et al., 1997). A thin endometrium or a homogenous, hyperechogenic endometrium on the day of HCG administration may be associated with pathological abnormalities, which may interfere with normal embryo implantation, and ultrasound examination may not be adequate to identify pathology abnormalities in the endometrium. An important limitation of this study is that it is retrospective in nature. We believe, however, that the results are of interest because similar studies have been published with conflicting results. A well-designed and powered randomized clinical trial will be needed to determine which results, if any, are substantiated.
Conclusion The endometrium, with adequate growth, triple-line on HCG day, or both, seems to be favourable for pregnancy. The endometrial thickness and the change, however, are not prognostically useful in predicting the occurrence of pregnancy, and the combined endometrial characteristics cannot predict the clinical outcome correctly.
Acknowledgement The authors thank everyone in the department of Reproductive Medicine in the Centre South University Xiangya Hospital for their scientific advice and encouragement.
References Al-Ghamdi, A., Coskun, S., Al-Hassan, S., AL-Rejjal, R., Awartani, K., 2008. The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET) outcome. Reprod. Biol. Endocrinol. 6, 37. Basir, G.S., O, W.S., So, W.W., Ng, E.H., Ho, P.C., 2002. Evaluation of cycle-to-cycle variation of endometrial responsiveness using transvaginal sonography in women undergoing assisted reproduction. Ultrasound Obstet. Gynecol. 19, 4844– 4889. Bassil, S., 2001. Changes in endometrial thickness, width, length and pattern in predicting pregnancy outcome during ovarian stimulation in in vitro fertilization. Ultrasound Obstet. Gynecol. 18, 258– 263. Bohrer, M.K., Hock, D.L., Rhoads, G.G., Kemmann, E., 1996. Sonographic assessment of endometrial pattern and thickness in patients treated with human menopausal gonadotropins. Fertil. Steril. 66, 2442–2447. Bourgain, C., Devroey, P., 2003. The endometrium in stimulated cycles for IVF. Hum. Reprod. Update 9, 515–522. Chen, S.-L., Wu, F.-R., Luo, C., Chen, X., Shi, X.-Y., Zheng, H.-Y., Ni, Y.-P., 2010. Combined analysis of endometrial thickness and
298 pattern in predicting outcome of in vitro fertilization and embryo transfer: a retrospective cohort study. Reprod. Biol. Endocrinol. 8, 30. De Geyter, C., Schmitter, M., De Geyter, M., Nieschlag, E., Holzgreve, W., Schneider, H.P., 2000. Prospective evaluation of the ultrasound appearance of the endometrium in a cohort of 1,186 infertile women. Fertil. Steril. 73, 106–113. Dickey, R.P., Olar, T.T., Curole, D.N., Taylor, S.N., Rye, P.H., 1992. Endometrial pattern and thickness associated with pregnancy outcome after assisted reproduction technologies. Hum. Reprod. 7, 418–421. Dietterich, C., Check, J.H., Choe, J.K., Nazari, A., Lurie, D., 2002. Increased endometrial thickness on the day of human chorionic gonadotropin injection does not adversely affect pregnancy or implantation rates following in vitro fertilization-embryo transfer. Fertil. Steril. 77, 7817–7886. Fanchin, R., 2001. Assessing uterine receptivity in 2001. Ann. N. Y. Acad. Sci. 943, 185–202. Gonen, Y., Casper, R.F., 1990. Prediction of implantation by the sonographic appearance of the endometrium during controlled ovarian stimulation for in vitro fertilization (IVF). J. In Vitro Fert. Embryo Transf. 7, 146–152. Gonen, Y., Casper, R.F., Jacobson, W., Blankier, J., 1989. Endometrial thickness and growth during ovarian stimulation: a possible predictor of implantation in in vitro fertilization. Fertil. Steril. 52, 446–450. Khalifa, E., Brzyski, R.G., Oehninger, S., Acosta, A.A., Muasher, S.J., 1992. Sonographic appearance of the endometrium: the predictive value for the outcome of in-vitro fertilization in stimulated cycles. Hum. Reprod. 7, 6776–6780. Kovacs, P., Matyas, S., Boda, K., Kaali, S.G., 2003. The effect of endometrial thickness on IVF/ICSI outcome. Hum. Reprod. 18, 2337– 2341. McWilliams, G.D., Frattarelli, J.L., 2007. Changes in measured endometrial thickness predict in vitro fertilization success. Fertil. Steril. 88, 74–81. Mercé, L.T., Barco, M.J., Bau, S., Troyano, J., 2008. Are endometrial parameters by three-dimensional ultrasound and power Doppler angiography related to in vitro fertilization/embryo transfer outcome? Fertil. Steril. 89, 1111–1117. Noyes, N., Liu, H.C., Sultan, K., Schattman, G., Rosenwaks, Z., 1995. Endometrial thickness appears to be a significant factor in embryo implantation in in-vitro fertilization. Hum. Reprod. 10, 919– 922. Noyes, N., Hampton, B.S., Berkeley, A., Licciardi, F., Grifo, J., Krey, L., 2001. Factors useful in predicting the success of oocyte donation: a 3-year retrospective analysis. Fertil. Steril. 76, 92–97. Oliveira, J.B., Baruffi, R.L., Mauri, A.L., Petersen, C.G., Borges, M.C., Franco, J.G., 1997. Endometrial ultrasonography as a predictor of pregnancy in an in-vitro fertilization programme after ovarian stimulation and gonadotrophin-releasing hormone and gonadotrophins. Hum. Reprod. 12, 25152–25518. Puerto, B., Creus, M., Carmona, F., Civico, S., Vanrell, J.A., Balasch, J., 2003. Ultrasonography as a predictor of embryo
J Zhao et al. implantation after in vitro fertilization: a controlled study. Fertil. Steril. 79, 1015–1022. Raga, F., Bonilla-Musoles, F., Casan, E.M., Klein, O., Bonilla, F., 1999. Assessment of endometrial volume by three-dimensional ultrasound prior to embryo transfer: clues to endometrial receptivity. Hum. Reprod. 14, 28512–28854. Rashidi, B.H., Sadeghi, M., Jafarabadi, M., Nejad, E.S.T., 2004. Relationships between pregnancy rates following in vitro fertilization or intracytoplasmic sperm injection and endometrial thickness and pattern. Eur. J. Obstet. Gynecol. Reprod. Biol. 120, 179– 184. Richter, K.S., Bugge, K.R., Bromer, J.G., Levy, M.J., 2007. Relationship between endometrial thickness and embryo implantation, based on 1,294 cycles of in vitro fertilization with transfer of two blastocyst-stage embryos. Fertil. Steril. 87, 535–539. Rinaldi, L., Lisi, F., Floccari, A., Lisi, R., Pepe, G., Fishel, S., 1996. Endometrial thickness as a predictor of pregnancy after in-vitro fertilization but not after intracytoplasmic sperm injection. Hum. Reprod. 11, 15381–15541. Ruız-Velasco, V., Alfani, G.G., Sanchez, L.P., Marcela, V.A., 1997. Endometrial pathology and infertility. Fertil. Steril. 67, 6876– 6892. Sharara, F.I., Lim, J., McClamrock, H.D., 1999. Endometrial pattern on the day of oocyte retrieval is more predictive of implantation success than the pattern or thickness on the day of hCG administration. J. Assist. Reprod. Genet. 16, 523– 528. Sher, G., Herbert, C., Maassarani, G., Jacobs, M.H., 1991. Assessment of the late proliferative phase endometrium by ultrasonography in patients undergoing in-vitro fertilization and embryo transfer (IVF/ET). Hum. Reprod. 6, 2322–2337. Wang, L., Qiao, J., Li, R., Zhen, X., Liu, Z., 2010. Role of endometrial blood flow assessment with color Doppler energy in predicting pregnancy outcome of IVF-ET cycles. Reprod. Biol. Endocrinol. 8, 122. Weissman, A., Gotlieb, L., Casper, R.F., 1999. The detrimental effect of increased endometrial thickness on implantation and pregnancy rates and outcome in an in vitro fertilization program. Fertil. Steril. 71, 147–149. Yuval, Y., Lipitiz, S., Dor, J., Achiron, R., 1999. The relationships between endometrial thickness, and blood flow and pregnancy rates in in-vitro fertilization. Hum. Reprod. 14, 1067–1071. Zaidi, J., Campbell, S., Pittrof, R., Tan, S.L., 1995. Endometrial thickness, morphology, vascular penetration and velocimetry in predicting implantation in an in vitro fertilization program. Ultrasound Obstet. Gynecol. 6, 1911–1998.
Declaration: The authors report no financial or commercial conflicts of interest.
Received 22 February 2014; refereed 11 May 2014; accepted 21 May 2014.