Endometrial thickness is predictive of histologic endometrial maturation in women undergoing hormone replacement for ovum donation*

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Vol. 66, No.3, September 1996

Copyright c 1996 American Society for Reproductive Medicine

Printed on acid-free paper in U. S. A

Endometrial thickness is predictive of histologic endometrial maturation in women undergoing hormone replacement for ovum donation* Glen E. Hofmann, M.D., Ph.D.t* Jennifer Thie, M.D.t Richard T. Scott, Jr., M.D.§ Daniel Navot, M.D. II Bethesda Hospital, Cincinnati, Ohio; Saint Barnabas Medical Center, Livingston, New Jersey; New York Medical College, Valhalla, New York

Objective: To determine if ultrasonographic endometrial pattern or thickness is predictive of histologic endometrial maturation in women undergoing hormone replacement for ovum donation. Design: Ultrasonographic endometrial thickness and pattern were determined and compared with histologic assessment of endometrial maturation. Patients: Forty-six women underwent 52 preparatory cycles for ovum donation. Transvaginal ultrasound (US) was performed after 14 days of E2 replacement and, after 12 days of P, an endometrial biopsy was performed. In 12 cycles, a continuous dose of 2 mg/d E2 was administered. In cycles with out-of-phase biopsies (dated earlier than day 24) and in the last 34 cycles, all women received an escalating dose of E2 before initiation of P. Additionally, the 46 women underwent 55 ETs with USs performed on cycle day 15. Results: Six women had abnormal biopsies in their first preparatory cycle on the continuous E2 protocol, which normalized with the escalating protocol. All other women had normal biopsies. Women with abnormal biopsies had significantly thinner endometrium (::56 mm) but similar endometrial patterns compared with women with normal biopsies. In women having US in preparatory and transfer cycles, there were no differences in endometrial thickness or pattern between examinations. Conclusions: Endometrial thickness 2:: 7 mm in hormone replacement cycles predicts in phase endometrial histology and can replace the endometrial biopsy. Fertil Steril® 1996;66:380-3 Key Words: Endometrial ultrasound, endometrial histology, endometrial maturation, ovum donation

Current protocols for ovum donation require synchronization of the ovum donor and recipient (1-8). Typically, the recipient undergoes pituitary downReceived March 1, 1996; revised and accepted April 29, 1996.

* Presented in part at the 51st Annual Meeting ofthe American Society for Reproductive Medicine, Seattle, Washington, October 7 to 12, 1995. t Department of Obstetrics and Gynecology, Bethesda Hospital. :j: Reprint requests: Glen E. Hofmann, M.D., Ph.D., Bethesda Center for Reproductive Health and Fertility, Bethesda Hospital, 619 Oak Street, Cincinnati, Ohio 45206 (FAX: 513-569-6386). § Department of Obstetrics and Gynecology, Saint Barnabas Medical Center. II Department of Obstetrics and Gynecology, New York Medical College. 380

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regulation with a GnRH-agonist followed by hormone replacement for endometrial preparation using a variety of hormone replacement protocols. Most ovum donation programs perform a preparatory cycle before the ovum donation cycle to ensure appropriate endometrial maturation with the designated protocol, before cycling the donor. The method of assessment of endometrial maturation traditionally has been through sampling of the endometrium either on normalized cycle day 21 and/or day 28 (35, 7-9). There currently is no consensus as to which day of evaluation and which method of assessment may offer the best evaluation of endometrial maturation and, in particular, uterine receptivity for embryo implantation. Because of this uncertainty, Fertility and Sterility®

the significantly different endometrial morphology noted in hormone replacement cycles (glandularstromal dyssynchrony) (4, 5) and the variability of interpretation of endometrial biopsies (both because ofthe dyssynchrony and the variability of reading by trained examiners) (10), another modality to assess endometrial maturation would be helpful. With the continuing advances being made with high-resolution endovaginal ultrasonography, we examined the possibility that a late follicular endovaginal ultrasound (US) evaluating endometrial thickness and pattern might be able to predict endometrial histology with minimal variation, minimal discomfort (no P support either by suppository or injection and no biopsy), and at a reduced cost. Thus, we retrospectively reviewed ovum recipients presenting to the Bethesda Center for Reproductive Health and Fertility to determine if a late follicular US reliably would predict endometrial histology from a biopsy. MATERIALS AND METHODS

Forty-six women presenting to the Bethesda Center for Reproductive Health and Fertility as candidates for ovum donation because of Turner's syndrome (n = 1) or the inability to achieve adequate ovarian hyperstimulation with gonadotropins (n = 45) were studied. All underwent appropriate medical, physical, and psychological evaluation before admission into the ovum donation program. All women underwent a preparatory cycle using midluteal pituitary down-regulation with the GnRHagonist leuprolide acetate (0.5 mg SC daily, except the Turner's patient). Initially, all women received E2 replacement with a fixed dose (1 mg twice per day, fixed-dose protocol) of oral micronized E2 (Estrace; Mead Johnson, Evansville, IN) (7). After 14 days of E 2, a pelvic US was performed. Daily P supplementation then was initiated (50 mg P in oil 1M) while continuing the E2 at 1 mg twice per day. After 12 days of P in oil, an endometrial biopsy was performed. If the biopsy was out of phase or after the 12th patient, all patients underwent estrogen replacement with an escalating dose of micronized E2 (escalating protocol, 2 mg/d for 5 days, 4 mg/d for 4 days, 6 mg/d for 4 days) (3-5, 7 -9) before initiation of 1M P in oil. An endometrial biopsy then was performed after decreasing the E2 to 1 mg twice per day and initiating P in oil for 12 days. Transvaginal ultrasonography was performed using an ATL Ultramark 4 (Advanced Technologies Laboratories, Bothell, WA) with a 5-mHz Array IVT, recording maximal endometrial thickness in a sagittal plane in the fundus and endometrial pattern (11). The US findings were recorded, but all clinical decisions were based on the endometrial histology. Vol. 66, No.3, September 1996

An endometrial biopsy was performed on normalized cycle day 27 (day 15 = day 1 of P in oil) with a pipelle. The tissue was Formalin fixed and paraffin embedded and stained with hematoxylin and eosin for histologic dating by one ofthe examiners (G.E.H.) according to the criteria of Noyes et al. (12). An inphase biopsy required evidence of subepithelial decidualization (day 25). The 46 women underwent 52 preparatory cycles with late follicular USs and subsequent endometrial biopsies, and eventually 55 transfer cycles with late follicular USs. Comparisons of endometrial thicknesses and histologic dating were made by unpaired two-tailed Student's t-tests. Comparisons between endometrial thickness and patterns in the same women in a preparatory cycle (in phase) and a transfer cycle were made using a paired t-test. An a error of 0.05 was considered statistically significant. No ETs were made onto endometria previously shown to be out of phase without a subsequent in-phase biopsy. Thus, assessment of the reproductive (implantation) potential of out-of-phase endometrium, which also was abnormally thin «7 mm), was never determined, and no comparisons of pregnancy results were made between patients with endometrial thickness of <7 or 2'=.7 mm. RESULTS

Forty-six women had 52 USs and endometrial biopsies in preparatory cycles. Six women had out-ofphase biopsies when on the fixed dose of E 2, all of which normalized when on the escalating dose. Forty additional women underwent preparatory cycles where the biopsies were in phase (6 with a fixed dose of E2 and 34 with an escalating dose). Endometrial thickness was significantly thinner in women whose biopsy was out of phase (Fig. lA) compared with women whose biopsy was in phase (Fig. IB) (P = 0.000001) initially or after correction with the escalating E2 protocol (P = 0.0001) (Table 1). All women whose biopsy was out of phase on a fixed dose of E2 normalized on the escalating dose (Table 1). All women with out-of-phase biopsies had endometrial thickness < 7 mm (range 5 to 6 mm), whereas all women with in-phase biopsies had endometrial thickness> 7 mm (range 8 to 13). Endometrial pattern (11) was similar between women with in-phase or out-of-phase biopsies and was always pattern 1 or 2, with pattern 1 predominating in >85% of the cycles (Table 1). Additionally, the same 46 women after achieving an in-phase endometrial biopsy underwent 55 ETs with USs performed on day 15 of the transfer cycle. Both the fixed and escalating E2 protocols were used. There were no significant differences in the en domeHofmann et al.

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381

A

B

Figure 1 (A), Hematoxylin and eosin-stained endometrium from women on the continuous estrogen replacement with thin «7 mm) endometrium on US demonstrating an arrest of development (menstrual day 21 ). Note the significant stromal edema, relative lack of spiral arteriole development, and the total lack of periarteriolar or subepithelial decidualization. Magnification, x 200. (B), Hematoxylin and eosin-stained endometrium of the same woman on the escalating estrogen protocol with thick (~ 7 mm) endometrium that is in phase (day 26). Note the lack of stromal edema, the prominent presence of spiral arterioles, and prominent periarteriolar and subepithelial decidualization. Magnification, x 200).

trial thickness or pattern between the preparatory cycle and transfer cycles (P = 0.096), and no woman developed an endometrial thickness < 7 mm in a subsequent evaluation.

individual patients was minimal. Endometrial pattern did not correlate with histologic findings. The best method for assessing endometrial adequacy for implantation remains elusive. Endometrial biopsy has long been the "gold standard." However, exactly when during the cycle the biopsy should be performed remains controversial. Traditionally, assessment of the endometrium for ovum recipients was to sample the endometrium on normalized cycle days 21 and 28 in two different cycles (4). The endometrial biopsy often is difficult to interpret, with the same examiners differing in the dating on repetitive readings (10), with fertile women occasionally having out-of-phase endometrial biopsies (13), and women with completely normal menstrual cycles having out-of-phase endometrial biopsies (14). Additionally, some authors believe a more exact assessment of endometrial receptivity would require a day 21 endometrial biopsy with assessment of the a y f33 integrin (15). Thus, when and how to assess the endometrium is not agreed upon. We chose to eval ua te the endometrium with transvaginal US, which was evaluated against the endometrial biopsy. Criterion for a normal biopsy was evidence of luteal transformation as evidenced by exhausted glands, prominence of spiral arterioles, and, in particular, sheets of subepithelial decidualized stromal cells. This last criteria is used in the original description of endometrial dating (12) to date the latest development of the endometrium before neutrophil infiltration and menstruation and is detected easily with essentially no chance for misinterpretation. Thus, although the potential weakness of the "gold standard" is acknowledged, we have attempted to use it as unambiguously as possible. Previous studies have examined the role of ultrasonography of the endometrium in women undergoing hormone replacement cycles for ovum donation (8, 16-18). One study examined the endometrial thickness on cycle day 21 in conjunction with histol-

Table 1

Comparison of Ultrasound and Histologic Dating

DISCUSSION

Assessment of endometrial maturation in preparation for recipient ovum donation cycles traditionally has been done with the endometrial biopsy, and assessment of endometrial maturation by the traditional criteria of Noyes et al. (12). This study demonstrates that a late follicular transvaginal US assessing endometrial thickness correlates accurately with the histologic findings in 100% of the patients who underwent both biopsy and US, and that between-cycle variability in endometrial thickness for 382

Hofmann et al.

Ultrasound and endometrial maturation

Out-of-phase biopsyt (n = 6) Corrected in-phase biopsy (n = 6)

In-phase biopsy (n

=

40)

Thickness*

Pattern 1

mm

%

Histologic dating*

.4

87

22.3 ± .8

9.8 ± 1.9 9.8 ± 1.4

100

26.0 ± .7 25.6 ± .7

5.8 ±

94

* Values are means ± SD. t Out-of-phase biopsies were significantly thinner than corrected biopsies (P = 0.0001) and in-phase biopsies (P = 0.000001), had similar endometrial patterns (P > 0.05), and were significantly lagging in histologic maturation compared with corrected biopsies in the same woman and in the women with in-phase biopsies (P = 0.0001).

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ogy and hormone receptor levels in the endometrium (8). Ultrasonography was similar in women of all ages undergoing hormone replacement. Another study examined endometrial thickness in recipients at the time of ET; the results suggested that the thickness correlated with pregnancy outcome with few, if any, pregnancies occurring with a thickness < 7.5 mm (16). Endometrial biopsies were not performed in this study. Another study examined endometrial thickness and pattern in recipient women on hormone replacement who were known to have normal biopsies (17). Endometrial thickness did not influence pregnancy outcome, but pattern did. Most of the pregnancies in this study (17) occurred in women with endometrial thickness> 7 mm. A final study examining endometrium ultrasonographically included evaluation of uterine artery impedance (18). No differences were found in pregnancy rates based on endometrial thickness or uterine artery impedance. Endometrial pattern was found to be predictive of outcome. This study does not comment as to whether the endometrium was assessed histologically in a preparatory cycle. The benefits of transvaginal US are the ease of performance, immediate knowledge of the results, reduced cost to the patient, lack of 12 days of 1M P in oil required for endometrial maturation in the absence of corpus luteum function (oral progestin or suppositories can be used to induce menses), minimal discomfort from the US, and no painful endometrial biopsy. Patient acceptance of the US compared with the traditional methodology is very high. As further knowledge of the mechanisms of endometrial receptivity evolves, particularly regarding integrins, the role of US in assessing endometrial maturation and receptivity must be re-evaluated. Our data support the finding that when a late follicular US demonstrates an endometrial thickness 2: 7 mm, the late luteal endometrial biopsy will always be in phase. With biopsies with endometrial thickness < 7 mm, a late luteal biopsy may be indicated. Studies that have measured pregnancy outcome and the ultrasonographic appearance of the endometrium (16, 17) support the role of endometrial thickness in assessing uterine receptivity.

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