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Potential enhancement of endometrial receptivity in cycles using controlled ovarian hyperstimulation with antiprogestins: A hypothesis
FERTILITY Copyright
AND
STERILITY~
’ 1997 American
V&67,
Society
for Reproductive
Medicine
Printed
Ko 2, February
on acid-free
paper
1997
m LT. S. A.
Potential enhancement of endometrial receptivity in cycles using controlled ovarian hyperstimulation with antiprogestins: a hypothesis Richard J. Paulson, M.D.*? Mark V. Sauer, M.D.++ Rogerio A. Lobo, M.D.+$ Diuision School
of Reproductive of Medicine,
Endocrinology
Los Angeles,
and Infertility,
Department
of Obstetrics
and
Gynecology,
University
of Southern
California
California
Objective: To manipulate the luteal endometrial progression by the use of antiprogestins. Design: Prospective controlled clinical trial. Setting: The IVF program of the University of Southern California School of Medicine, Los Angeles, California. Patient(s): Thirteen oocyte donors and 20 oocyte recipients. Intervention(s): Controlled ovarian hyperstimulation of oocyte donors, administration of two doses of 2.5 mg of RU486 to the study group, and endometrial biopsies. Main Outcome Measure(s): Serum Ez and P levels, histologic dating of the endometrium, endometrial ultrastructure by scanning electron microscopy. Result(s): No difference in serum Ez or P levels was noted after RU486 administration. The histologic dating was advanced in oocyte donors as compared with recipients undergoing artificial cycles but returned to normal (in phase) after RU486. Pinopods were noted in all recipient biopsies and in donors treated with RU486 but in only one of four biopsies in donor controls. Conclusion(s): Cycles with controlled ovarian hyperstimulation are associated with high early luteal P levels and advanced endometrial histology. Low doses of RU486 may correct the precocious luteinization and restore endometrial receptivity. Fertil Steril@ 1997;67:321-5 Key Words:
Endometrial
receptivity,
antiprogestins,
We previously have proposed (1) that embryo implantation after assisted reproductive cycles is dependent on three factors: embryo quality, endometrial receptivity, and transfer efficiency. In a matched retrospective analysis of data from our institution, we demonstrated that endometrial receptivity to embryo implantation in cycles using controlled ovarian hyperstimulation (COH) is impaired, as compared with artificial cycles in oocyte recipients
Received May 9, 1996; revised and accepted September 25, 1996. * Reprint requests: Richard J. Paulson, M.D., Department of Obstetrics and Gynecology, Women’s and Children’s Hospital, Room lM2, 1240 North Mission Road, Los Angeles, California 90033 (FAX: 213-226-2850). t Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, University of Southern California School of Medicine. $ Present address: Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons, New York, New York. Vol.
67, No. 2, February
1997
RU486,
implantation,
pinopods
(2). Previous studies have shown that cycles using COH are associated with advanced endometrial histology during the luteal phase (3) as well as premature disappearance of endometrial pinopods as determined by scanning electron microscopy (4). The presence of these pinopods is thought to be an ultrastructural marker of the implantation window. It previously has been noted, during cycles using COH with a combination of GnRH agonists and exogenous gonadotropins, that elevated serum P levels on the day of hCG administration arle associated with decreased pregnancy success (5, 6). Whereas this decrease has been noted during IVl? cycles, this effect is notably absent in cycles of oocyte donation (7). This combination of observations strongly suggests that the adverse effect caused by a premature elevation of P is not due to a diminishment of oocyte (and, consequently, embryo) quality. ELasedon the foregoing information, it is our hypothesis that endometrial receptivity in cycles using COH is decreased by premature endometrial luteinization and a prePaulson
et al.
Antiprogestins
and
endometrial
receptiuity
321
Estradlol
(1000
1
pg/mL)
/IA
OV
/
/ 0
/
2
4
Days
Progesterone
/ 7
post-HCG
(ng/mL)
ing exogenous E, and P stimulation of their endometria during mock cycles in anticipation of subsequent transfer cycles. Four of the oocyte donors represented the study group. They received a total of two doses of 2.5 mg RU486 orally, given immediately after follicle aspiration and on the following day. A second group of four donors served as the control group and received no RU486. The day of hCG administration was designated arbitrarily as day 0. Both groups of donors underwent phlebotomy for serum measurements of Ez and P on days 0,2,4, and 7. On day 7 after hCG, an endometrial biopsy was performed on all subjects. The biopsies were analyzed by standard histologic criteria for dating by a single pathologist and by scanning electron microscopy for the presence of pinopods. The remaining five oocyte donors had endometrial biopsies performed on day 9 after hCG. These biopsies were analyzed by histologic dating only. All 20 potential recipients had endometrial biopsies performed on the 7th day of P stimulation. Because recipients are synchronized to donors by beginning P on the day after hCG administration to the donors, these endometria were synchronous with those obtained from the donors on the 7th day after hCG. All 20 specimens were analyzed histologically and four were selected randomly for analysis by scanning electron microscopy in a manner identical to that of the do:nor and control groups. Statistical analysis was by a:nalysis of variance and Fisher’s exact test, as appr’opriate. RESULTS
Days
post-HCG
Figure 1 (A), Serum Ez levels in controls and study (RU486) groups. W, RU486; q , controls. CB), Serum P levels in controls and study (RU486) groups. n , RLJ486; q , controls.
mature appearance of the implantation window caused by a combination of premature secretion and supraphysiologic levels of P in the early luteal phase. The purpose of the present study was to investigate the possibility of restoring normal endometrial receptivity in cycles using COH by inhibiting P action on the endometrium. MATERIALS
AND
MJXTHODS
This study was approved by our Institutional Review Board and informed consent was obtained from all subjects. We studied 13 oocyte donors who underwent a standard regimen of COH with a combination of leuprolide acetate (Lupron; TAP Pharmaceuticals, North Chicago, IL), hMG, and hCG, and 20 prospective recipients of oocyte donation who were undergo322
Paulson
et al.
Antiprogestins
Serum E2 and P levels in the study and control groups of oocyte donors are depicted in Figure 1A and B. Estradiol levels were slightly higher in the study group, with peak values on the day of hCG of 3,929 + 871 pg/mL (conversion factor to SI unit, 3.671; mean -C SD) as compared with ;3,529 ? 837 pg/ mL in the controls. The slight trend toward higher values in the study group persisted throughout the cycles, but there were no statistically significant differences noted between the two groups. There were also no statistically significant differences in P levels between the two groups. The results of the histologic dating in the four subgroups of subjects are depicted in Table 1. Biopsies
and
endometrial
receptivity
Table
1
Endometrial Type
Histology
Results
of cycle
Donors (day 9) Controls (day 7) Study (RU486) group Recipients (day 7)
Histologic
(day
7)
dating
Advanced In phase-advanced In phase In phase
Fen’ility
and
Sterility@
obtained on day 9 after hCG in oocyte donors were all classified as advanced. The control group biopsies, obtained 2 days earlier on day 7 after hCG, demonstrated variable histology, with two specimens described as advanced and two specimens described as in phase. Biopsies obtained from potential recipients demonstrated a similar variability, despite the precise timing of P administration. Biopsies obtained in the study group exposed to low doses of RU486 had the most consistent in-phase pattern of all the groups. The results of the scanning electron microscopy analysis are summarized in Table 2. All of the biopsy specimens in the recipients demonstrated pinopods. However, biopsies obtained from the hyperstimulated control cycles demonstrated pinopods in only one of four specimens. The study group, which was exposed to RU486, demonstrated pinopods in all four of the specimens. The comparison of the control group to the other two groups was statistically significant (P < 0.05). Figure 2A, B, and C show representative sections of these biopsies. DISCUSSION
There is little disagreement that there is such a thing as endometrial receptivity. It generally is accepted that the most viable of embryos will not, or cannot, implant upon an endometrium that is not receptive to implantation. The use of the intrauterine device contraception as well as postcoital contraception methodology is based on this principle. More specifically, in the human, as in several other mammalian species, there appears to be a “window of implantation” during which implantation may occur. Several markers of this window of implantation have been proposed. These include endometrial histology, the production of certain proteins by the endometrium (8, 91, and the presence of pinopods as observed by scanning electron microscopy (4). The precise function of the pinopods is not clear. Previous reports have suggested that these cytoplasmic projections are involved in the pinocytosis of uterine fluids and macromolecules (10). Because the appearance of the pinopods is limited to 24 to 48 hours at the approximate time of blastocyst implantation (ll), their presence is thought to indicate the “open-
Table
2
Results Type
of Scanning
Electron
of biopsy
Microscopy Pinopods
Recipients Control group Study group (RU486) * P < 0.05, Vol.
control
67, No. 2, February
present
4of4 1 of4* 4of4
group
versus
1997
recipients
and study
group.
Figure 2 (A), Representative section of biopsy (obtained from a prospective recipient undergoing a cycle of exogenous Ez and P. Abundant pinopods are demonstrated. (B), Biopqy obtained from an oocyte donor; note absence of pinopods. (C), Biopsy from oocyte donor treated with RU486. Pinopods are once again noted.
ing” of the window of implantation (4). The surface of the pinopods also may contain receptors or harbor high concentrations of adhesion molecu.les (9). However, the precise driving force behind a generation of a receptive endometrium and the opening of the window of implantation is not clear. Whereas a variety of substances are produced by Paulson
et al.
Antiprogestins
and
endometrial
reeeptiuity
323
the endometrium around the time of aspiration, the success of oocyte donation to agonadal women presents a powerful argument that estrogen and P are all that is needed to drive the endometrium and establish endometrial receptivity. Whereas other substances may be involved in the communication between the embryo and the endometrium and in fact may be critical to implantation, their production necessarily must be secondary to that of estrogen and P. Furthermore, because many programs use oral estrogen replacement, the precise type of estrogen used (Ea, estrone, estrone sulfate) also must not be critical. If estrogen and P are all that is needed, then the next logical question must be whether varying quantities of either steroid or variable timing of administration may play a role in the generation of an inhospitable or, alternately, receptive endometrium. We previously have demonstrated that COH inhibits endometrial receptivity of the subsequent luteal phase. The precise reason for this effect is not clear. Initially, it was tempting to recollect the use of diethylstilbestrol (DES) for postcoital contraception and draw a parallel between the high levels of Ez during COH and the use of exogenous DES to block implantation. However, studies examining high Ez levels during assisted reproductive techniques cycles have not demonstrated a decrease in embryo implantation rates with increasing Ez levels (12). These data typically are confounded by the observation that high E2 levels are associated with a better ovarian response and thus a potential increase in oocyte quality. However, whereas inadequate levels of E2 may result in an endometrium that is too thin to support nidation (13), it does not appear that excessive E2 levels per se inhibit implantation. By contrast, P levels above a certain threshold are known to induce a predictable cascade of luteal events in an endometrium that is primed adequately with estrogen. Because P is present in low levels throughout the menstrual cycle (note that, in absolute terms, P levels are always higher than E2 levels), it is clear that a “P threshold” must exist below which no luteinization takes place. Once this threshold is exceeded, luteinization proceeds. It is not yet clear if higher levels of P can cause a more rapid luteal progression (14) or if there exists only a single P threshold. Previous reports that have demonstrated a decreased pregnancy rate in cycles with elevated P levels on the day of hCG administration (5, 6) have used variable levels of P as this “threshold.” However, it is not clear if this threshold already was exceeded on the day of hCG or if the elevation of P on the day of hCG was simply a marker for exceeding the P threshold at another time. As indicated above, it seems clear that the elevated P levels primarily are related to a decrease in endometrial 324
Paulson
et al.
Antiprogestins
and
endometrial
receptivity
receptivity rather than to oocyte quality (7). Accordingly, it was tempting to speculate that a delay in P action by an exogenous antiprogestin potentially would restore this endometrial receptivity. In this preliminary study, we attempted to look at two markers of the implantation window: endometrial histology and endometrial ultrastructure by scanning electron microscopy. Unfortunately, the supply of RU486 was small, thus limiting the number of subjects who could be studied and diminishing the power of the study. However, it does appear that a total dose of 5 mg of RU486 did have a physiologic effect on the endometrium of these hyperstimulated cycles. Because of the variability of the endometrial histology, no clear differences were observed. However, there did appear to be a significant difference in the presence of the pinopods on the 7th day of P among recipients or on the 7th day after hCG in the donors. If should be noted that even in cycles in which the P levels do not rise before hCG administration, the increase in P values is, substantially higher in stimulated than in natural cycles. For example, in the present study, at the time of follicle aspiration, the P levels were 9.1 ng/mL (conversion factor to SI unit, 3.180) in the study group and 8.2 ng/mL in the control group. These values contrast sharply with values of approximately 1 ng/mL at the time of ovulation in natural cycles (15). In summary, our preliminary study suggests that the antiprogesterone RU486, when afdministered in the early luteal phase stimulated cycles in low doses, has no effect on serum E2 or P levels but may delay the appearance of the implantation window to the time that the window is normally present to allow nidation of the implanting blastocysts. It is clear that our preliminary study awaits confirmation by larger studies, by the confirmation of a dose-response evaluation as well as the establishment of the optimal dose of antiprogestin to ‘be used. When these criteria are established, clinical trials of antiprogestin administration for the purpose of enhancing endometrial receptivity to embryo implantation may be undertaken. REFERENCES 1. Paulson RJ, Sauer MV, Lobo RA. Factors affecting embryo implantation after human in vitro fertilization: a hypothesis. Am J Obstet Gynecol 1990; 163:2020-3. RJ, Sauer MV, Lobo RA. Embryo implantation after 2. Paulson human in vitro fertilization: importance of endometrial receptivity. Fertil Steril 1990;53:870-4. JE, Acosta AA, Hsiu J-G, Jones HW Jr. Advanced 3. Garcia endometrial maturation after ovulation induction with human menopausal gonadotropin human chorionic gonadotropin for in vitro fertilization. Fertil Steril 1984;41:31-5. 4. Psychoyos A, Nikas G. Uterine pinopodes as markers of uterine receptivity. Assist Reprod Rev 1994;4:26-32. W, Sinton E, Schlenker T, Huynh D, Hamilton 5. Schoolcraft Fertility
and
Sterility’B
6.
7.
8.
9.
10.
Vol.
F, Meldrum DR. Lower pregnancy rate with premature luteinization during pituitary suppression with leuprolide acetate. Fertil Steril 1991;55:563-6. Silverberg KM, Burns WN, Olive DL, Riehl RM, Schenken RS. Serum progesterone levels predict success of in vitro fertilization/embryo transfer in patients stimulated with leuprolide acetate and human menopausal gonadotropins. J Clin Endocrinol Metab 1991;73:797-803. Legro RS, Ary BA, Paulson RJ, Stanczyk FZ, Sauer MV. Premature luteinization as detected by elevated serum progesterone is associated with a higher pregnancy rate in donor oocyte in-vitro fertilization. Hum Reprod 1993;8:1506-11. Hegele-Hartung C, Mootz U, Beier HM. Luteal control of endometrial receptivity and its modification by progesterone antagonists. Endocrinology 1992; 131:2446-60. Lessey BA, Castelbaum AJ, Sawin SW, Buck CA, Schinnar R, Bilker W, et al. Aberrant integrin expression in the endometrium of women with endometriosis. J Clin Endocrinol Metab 1994;79:643-9. Martel D, Frydman R, Glissant M, Maggioni C, Roche D,
67, No. 2, February
1997
11.
12.
13.
14. 15.
Paulson
Psychoyos A. Scanning electron microscopy of postovulatory human endometrium in spontaneous cycles and cycles stimulated by hormone treatment. J Endocrinoll987; 114:319-24. Nikas G, Drakakis P, Loutradis D, Mara-Skoufari C, Koumantakis E, Michalas S, et al. Uterine pinopods as markers of the ‘nidation window’ in cycling women receiving exogenous oestradiol and progesterone. Hum Reprod 1995; 10:1208-13. Chenette PE, Sauer MY, Paulson RJ. Very high serum estradiol levels are not detrimental to clinical outcome of in vitro fertilization. Fertil Steril 1990;54:858-63. Check JH, Nowroozi K, Choe J, Dietterich C. Influence of endometrial thickness and echo patterns on pregnancy rates during in vitro fertilization. Fertil Steril 1991;56:1173-5. Meldrum DR. Female reproductive aging-ovarian and uterine factors. Fertil Steril 1993;59:1-5. Lobo RA. The menstrual cycle. In: Mishell DR Jr, Davajan V, Lobo RA, editors. Infertility, contraception and reproductive endocrinology. 3rd edition. Cambridge (MA): 13lackwell Scientific Publications, 1991:104-24.
et al.
Antiprogestins
and
endometrial
rteceptiuity
325
AND
STERILITY~
’ 1997 American
V&67,
Society
for Reproductive
Medicine
Printed
Ko 2, February
on acid-free
paper
1997
m LT. S. A.
Potential enhancement of endometrial receptivity in cycles using controlled ovarian hyperstimulation with antiprogestins: a hypothesis Richard J. Paulson, M.D.*? Mark V. Sauer, M.D.++ Rogerio A. Lobo, M.D.+$ Diuision School
of Reproductive of Medicine,
Endocrinology
Los Angeles,
and Infertility,
Department
of Obstetrics
and
Gynecology,
University
of Southern
California
California
Objective: To manipulate the luteal endometrial progression by the use of antiprogestins. Design: Prospective controlled clinical trial. Setting: The IVF program of the University of Southern California School of Medicine, Los Angeles, California. Patient(s): Thirteen oocyte donors and 20 oocyte recipients. Intervention(s): Controlled ovarian hyperstimulation of oocyte donors, administration of two doses of 2.5 mg of RU486 to the study group, and endometrial biopsies. Main Outcome Measure(s): Serum Ez and P levels, histologic dating of the endometrium, endometrial ultrastructure by scanning electron microscopy. Result(s): No difference in serum Ez or P levels was noted after RU486 administration. The histologic dating was advanced in oocyte donors as compared with recipients undergoing artificial cycles but returned to normal (in phase) after RU486. Pinopods were noted in all recipient biopsies and in donors treated with RU486 but in only one of four biopsies in donor controls. Conclusion(s): Cycles with controlled ovarian hyperstimulation are associated with high early luteal P levels and advanced endometrial histology. Low doses of RU486 may correct the precocious luteinization and restore endometrial receptivity. Fertil Steril@ 1997;67:321-5 Key Words:
Endometrial
receptivity,
antiprogestins,
We previously have proposed (1) that embryo implantation after assisted reproductive cycles is dependent on three factors: embryo quality, endometrial receptivity, and transfer efficiency. In a matched retrospective analysis of data from our institution, we demonstrated that endometrial receptivity to embryo implantation in cycles using controlled ovarian hyperstimulation (COH) is impaired, as compared with artificial cycles in oocyte recipients
Received May 9, 1996; revised and accepted September 25, 1996. * Reprint requests: Richard J. Paulson, M.D., Department of Obstetrics and Gynecology, Women’s and Children’s Hospital, Room lM2, 1240 North Mission Road, Los Angeles, California 90033 (FAX: 213-226-2850). t Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, University of Southern California School of Medicine. $ Present address: Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons, New York, New York. Vol.
67, No. 2, February
1997
RU486,
implantation,
pinopods
(2). Previous studies have shown that cycles using COH are associated with advanced endometrial histology during the luteal phase (3) as well as premature disappearance of endometrial pinopods as determined by scanning electron microscopy (4). The presence of these pinopods is thought to be an ultrastructural marker of the implantation window. It previously has been noted, during cycles using COH with a combination of GnRH agonists and exogenous gonadotropins, that elevated serum P levels on the day of hCG administration arle associated with decreased pregnancy success (5, 6). Whereas this decrease has been noted during IVl? cycles, this effect is notably absent in cycles of oocyte donation (7). This combination of observations strongly suggests that the adverse effect caused by a premature elevation of P is not due to a diminishment of oocyte (and, consequently, embryo) quality. ELasedon the foregoing information, it is our hypothesis that endometrial receptivity in cycles using COH is decreased by premature endometrial luteinization and a prePaulson
et al.
Antiprogestins
and
endometrial
receptiuity
321
Estradlol
(1000
1
pg/mL)
/IA
OV
/
/ 0
/
2
4
Days
Progesterone
/ 7
post-HCG
(ng/mL)
ing exogenous E, and P stimulation of their endometria during mock cycles in anticipation of subsequent transfer cycles. Four of the oocyte donors represented the study group. They received a total of two doses of 2.5 mg RU486 orally, given immediately after follicle aspiration and on the following day. A second group of four donors served as the control group and received no RU486. The day of hCG administration was designated arbitrarily as day 0. Both groups of donors underwent phlebotomy for serum measurements of Ez and P on days 0,2,4, and 7. On day 7 after hCG, an endometrial biopsy was performed on all subjects. The biopsies were analyzed by standard histologic criteria for dating by a single pathologist and by scanning electron microscopy for the presence of pinopods. The remaining five oocyte donors had endometrial biopsies performed on day 9 after hCG. These biopsies were analyzed by histologic dating only. All 20 potential recipients had endometrial biopsies performed on the 7th day of P stimulation. Because recipients are synchronized to donors by beginning P on the day after hCG administration to the donors, these endometria were synchronous with those obtained from the donors on the 7th day after hCG. All 20 specimens were analyzed histologically and four were selected randomly for analysis by scanning electron microscopy in a manner identical to that of the do:nor and control groups. Statistical analysis was by a:nalysis of variance and Fisher’s exact test, as appr’opriate. RESULTS
Days
post-HCG
Figure 1 (A), Serum Ez levels in controls and study (RU486) groups. W, RU486; q , controls. CB), Serum P levels in controls and study (RU486) groups. n , RLJ486; q , controls.
mature appearance of the implantation window caused by a combination of premature secretion and supraphysiologic levels of P in the early luteal phase. The purpose of the present study was to investigate the possibility of restoring normal endometrial receptivity in cycles using COH by inhibiting P action on the endometrium. MATERIALS
AND
MJXTHODS
This study was approved by our Institutional Review Board and informed consent was obtained from all subjects. We studied 13 oocyte donors who underwent a standard regimen of COH with a combination of leuprolide acetate (Lupron; TAP Pharmaceuticals, North Chicago, IL), hMG, and hCG, and 20 prospective recipients of oocyte donation who were undergo322
Paulson
et al.
Antiprogestins
Serum E2 and P levels in the study and control groups of oocyte donors are depicted in Figure 1A and B. Estradiol levels were slightly higher in the study group, with peak values on the day of hCG of 3,929 + 871 pg/mL (conversion factor to SI unit, 3.671; mean -C SD) as compared with ;3,529 ? 837 pg/ mL in the controls. The slight trend toward higher values in the study group persisted throughout the cycles, but there were no statistically significant differences noted between the two groups. There were also no statistically significant differences in P levels between the two groups. The results of the histologic dating in the four subgroups of subjects are depicted in Table 1. Biopsies
and
endometrial
receptivity
Table
1
Endometrial Type
Histology
Results
of cycle
Donors (day 9) Controls (day 7) Study (RU486) group Recipients (day 7)
Histologic
(day
7)
dating
Advanced In phase-advanced In phase In phase
Fen’ility
and
Sterility@
obtained on day 9 after hCG in oocyte donors were all classified as advanced. The control group biopsies, obtained 2 days earlier on day 7 after hCG, demonstrated variable histology, with two specimens described as advanced and two specimens described as in phase. Biopsies obtained from potential recipients demonstrated a similar variability, despite the precise timing of P administration. Biopsies obtained in the study group exposed to low doses of RU486 had the most consistent in-phase pattern of all the groups. The results of the scanning electron microscopy analysis are summarized in Table 2. All of the biopsy specimens in the recipients demonstrated pinopods. However, biopsies obtained from the hyperstimulated control cycles demonstrated pinopods in only one of four specimens. The study group, which was exposed to RU486, demonstrated pinopods in all four of the specimens. The comparison of the control group to the other two groups was statistically significant (P < 0.05). Figure 2A, B, and C show representative sections of these biopsies. DISCUSSION
There is little disagreement that there is such a thing as endometrial receptivity. It generally is accepted that the most viable of embryos will not, or cannot, implant upon an endometrium that is not receptive to implantation. The use of the intrauterine device contraception as well as postcoital contraception methodology is based on this principle. More specifically, in the human, as in several other mammalian species, there appears to be a “window of implantation” during which implantation may occur. Several markers of this window of implantation have been proposed. These include endometrial histology, the production of certain proteins by the endometrium (8, 91, and the presence of pinopods as observed by scanning electron microscopy (4). The precise function of the pinopods is not clear. Previous reports have suggested that these cytoplasmic projections are involved in the pinocytosis of uterine fluids and macromolecules (10). Because the appearance of the pinopods is limited to 24 to 48 hours at the approximate time of blastocyst implantation (ll), their presence is thought to indicate the “open-
Table
2
Results Type
of Scanning
Electron
of biopsy
Microscopy Pinopods
Recipients Control group Study group (RU486) * P < 0.05, Vol.
control
67, No. 2, February
present
4of4 1 of4* 4of4
group
versus
1997
recipients
and study
group.
Figure 2 (A), Representative section of biopsy (obtained from a prospective recipient undergoing a cycle of exogenous Ez and P. Abundant pinopods are demonstrated. (B), Biopqy obtained from an oocyte donor; note absence of pinopods. (C), Biopsy from oocyte donor treated with RU486. Pinopods are once again noted.
ing” of the window of implantation (4). The surface of the pinopods also may contain receptors or harbor high concentrations of adhesion molecu.les (9). However, the precise driving force behind a generation of a receptive endometrium and the opening of the window of implantation is not clear. Whereas a variety of substances are produced by Paulson
et al.
Antiprogestins
and
endometrial
reeeptiuity
323
the endometrium around the time of aspiration, the success of oocyte donation to agonadal women presents a powerful argument that estrogen and P are all that is needed to drive the endometrium and establish endometrial receptivity. Whereas other substances may be involved in the communication between the embryo and the endometrium and in fact may be critical to implantation, their production necessarily must be secondary to that of estrogen and P. Furthermore, because many programs use oral estrogen replacement, the precise type of estrogen used (Ea, estrone, estrone sulfate) also must not be critical. If estrogen and P are all that is needed, then the next logical question must be whether varying quantities of either steroid or variable timing of administration may play a role in the generation of an inhospitable or, alternately, receptive endometrium. We previously have demonstrated that COH inhibits endometrial receptivity of the subsequent luteal phase. The precise reason for this effect is not clear. Initially, it was tempting to recollect the use of diethylstilbestrol (DES) for postcoital contraception and draw a parallel between the high levels of Ez during COH and the use of exogenous DES to block implantation. However, studies examining high Ez levels during assisted reproductive techniques cycles have not demonstrated a decrease in embryo implantation rates with increasing Ez levels (12). These data typically are confounded by the observation that high E2 levels are associated with a better ovarian response and thus a potential increase in oocyte quality. However, whereas inadequate levels of E2 may result in an endometrium that is too thin to support nidation (13), it does not appear that excessive E2 levels per se inhibit implantation. By contrast, P levels above a certain threshold are known to induce a predictable cascade of luteal events in an endometrium that is primed adequately with estrogen. Because P is present in low levels throughout the menstrual cycle (note that, in absolute terms, P levels are always higher than E2 levels), it is clear that a “P threshold” must exist below which no luteinization takes place. Once this threshold is exceeded, luteinization proceeds. It is not yet clear if higher levels of P can cause a more rapid luteal progression (14) or if there exists only a single P threshold. Previous reports that have demonstrated a decreased pregnancy rate in cycles with elevated P levels on the day of hCG administration (5, 6) have used variable levels of P as this “threshold.” However, it is not clear if this threshold already was exceeded on the day of hCG or if the elevation of P on the day of hCG was simply a marker for exceeding the P threshold at another time. As indicated above, it seems clear that the elevated P levels primarily are related to a decrease in endometrial 324
Paulson
et al.
Antiprogestins
and
endometrial
receptivity
receptivity rather than to oocyte quality (7). Accordingly, it was tempting to speculate that a delay in P action by an exogenous antiprogestin potentially would restore this endometrial receptivity. In this preliminary study, we attempted to look at two markers of the implantation window: endometrial histology and endometrial ultrastructure by scanning electron microscopy. Unfortunately, the supply of RU486 was small, thus limiting the number of subjects who could be studied and diminishing the power of the study. However, it does appear that a total dose of 5 mg of RU486 did have a physiologic effect on the endometrium of these hyperstimulated cycles. Because of the variability of the endometrial histology, no clear differences were observed. However, there did appear to be a significant difference in the presence of the pinopods on the 7th day of P among recipients or on the 7th day after hCG in the donors. If should be noted that even in cycles in which the P levels do not rise before hCG administration, the increase in P values is, substantially higher in stimulated than in natural cycles. For example, in the present study, at the time of follicle aspiration, the P levels were 9.1 ng/mL (conversion factor to SI unit, 3.180) in the study group and 8.2 ng/mL in the control group. These values contrast sharply with values of approximately 1 ng/mL at the time of ovulation in natural cycles (15). In summary, our preliminary study suggests that the antiprogesterone RU486, when afdministered in the early luteal phase stimulated cycles in low doses, has no effect on serum E2 or P levels but may delay the appearance of the implantation window to the time that the window is normally present to allow nidation of the implanting blastocysts. It is clear that our preliminary study awaits confirmation by larger studies, by the confirmation of a dose-response evaluation as well as the establishment of the optimal dose of antiprogestin to ‘be used. When these criteria are established, clinical trials of antiprogestin administration for the purpose of enhancing endometrial receptivity to embryo implantation may be undertaken. REFERENCES 1. Paulson RJ, Sauer MV, Lobo RA. Factors affecting embryo implantation after human in vitro fertilization: a hypothesis. Am J Obstet Gynecol 1990; 163:2020-3. RJ, Sauer MV, Lobo RA. Embryo implantation after 2. Paulson human in vitro fertilization: importance of endometrial receptivity. Fertil Steril 1990;53:870-4. JE, Acosta AA, Hsiu J-G, Jones HW Jr. Advanced 3. Garcia endometrial maturation after ovulation induction with human menopausal gonadotropin human chorionic gonadotropin for in vitro fertilization. Fertil Steril 1984;41:31-5. 4. Psychoyos A, Nikas G. Uterine pinopodes as markers of uterine receptivity. Assist Reprod Rev 1994;4:26-32. W, Sinton E, Schlenker T, Huynh D, Hamilton 5. Schoolcraft Fertility
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