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Endometrial receptivity: the age-related decline in pregnancy rates and the effect of ovarian function
FERTILITY AND STERILITY Copyright
©
Vol. 60, No, 2, August 1993
Printed on acid-free paper in U.
1998 The American Fertility Society
s.
A.
Endometrial receptivity: the age-related decline in pregnancy rates and the effect of ovarian function
Yuval Yaron, M,D, Amnon Botchan, M,D, Ami Amit, M,D.
Abraham Kogosowski, M.D. Israel Yovel M.D. Joseph B. Lessing, M.D.*
In Vitro Fertilization/Embryo Transfer Unit, Serlin Maternity Hospital, Tel Aviv Sourasky Medical Center, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
Objective: To assess the effect of age and ovarian function on endometrial receptivity. Design: Retrospective comparison between standard IVF and ovum donation in younger and older patients «40 and :?40 years of age, respectively). Patients: In standard IVF, there were 325 transfer cycles in older patients and 1,103 transfer cycles in younger ones. In ovum donation, there were 236 transfer cycles in older patients and 222 cycles in younger women. Ovum recipients were then redivided into two groups, according to ovarian function: ovarian failure group (219 cycles) and eugonadal group (239 cycles) in patients with retained ovarian function as manifested by regular menstrual cycles and normal gonadotropins. Results: In standard IVF, clinical pregnancy rates (PRs) were significantly lower in older patients (12.9% versus 23.8%, respectively). In ovum donation, clinical PRs were also significantly lower in older patients (21.2% versus 29.3%, respectively). A significantly higher clinical PR (31.1 %) was noted in patients with ovarian failure, compared with both eugonadal patients undergoing ovum donation (19.7%) and standard IVF patients (21.3%). Conclusions: The decrease in endometrial receptivity with age is responsible for the higher rate of implantation failure in older women. Patients with nonfunctioning ovaries do better than eugonadal patients in ovum donation programs. Fertil Steri11993;60:314-8 Key Words: Ovum donation, IVF, endometrial receptivity
The fact that fertility in women declines with age is true for both normal fertile women, as well as for those treated by assisted reproductive techniques. Many IVF units have previously rejected women > 40 years of age because of the expected decline of fertility in these women (1-4). This decline was neither related to the number of oocytes harvested, fertilized, or transferred nor to the serum steroid levels (5). The lower clinical pregnancy rate (PR) in older women undergoing IVF may be attributed to oocyte senescence, or age-related decline in endo-
Received December 8, 1993; revised and accepted April 27, 1993. * Reprint requests: Joseph B. Lessing, M.D., Serlin Maternity Hospital, Department of Obstetrics and Gynecology, Post Office Box 7079, Tel Aviv 61070, Israel.
314
Yaron et al.
Endometrial receptivity and egg donation
metrial receptivity. The question of which of these is more important has been difficult to resolve because in standard IVF, both the oocytes/embryos and the endometrium are from one and the same patient. However, ovum donation is unique in that the oocytes are obtained from another patient and oocyte/embryo quality may be controlled for, allowing evaluation of endometrial receptivity. When comparing IVF with ovum donation, we examined the role of endometrial receptivity in the agerelated decline of fertility. Initially, only patients with ovarian failure were included in ovum donation programs. Recently, however, ovum donation has proved itself to be an essential mode of treatment for other types of infertility: [1] patients with normal menstrual cycles who have repeatedly failed previous IVF Fertility and Sterility
treatments; [2) those who do not respond to hMG; or [3) individuals with a genetic trait precluding use of their own oocytes. Unlike those with ovarian failure, the endometrium in these patients is subject to the influence of endogenous ovarian steroids. Thus, a comparison between such patients and those with ovarian failure allows for assessment of endometrial receptivity with regard to ovarian function. MATERIALS AND METHODS
Between January 1986 and December 1992, 579 patients underwent 1,428 transfer cycles in standard IVF. We included only those patients treated with hMG alone. A standard protocol was used: on the 3rd day of the menstrual cycle, three ampules of hMG (Pergonal; Teva Pharmaceutical Industries Limited, Petah Tikva, Israel) were given daily. This daily dose was continued until at least two follicles with a mean diameter of 17 mm were seen on ultrasound (US), and the E 2 level was ~800 pg/mL (conversion factor to SI unit, 3.671). At this stage, 10,000 IU of hCG (Chorigon; Teva Pharmaceutical Industries Limited) was administered. Oocyte retrieval was scheduled 35 hours after hCG administration, using an US-guided transvaginal approach. Aspirated oocytes were inseminated and cultured in Ham's F -10 medium (KA 128 NB; Flow Laboratories, Irvine, Scotland), supplemented with 10% to 20% heat-inactivated patients' serum. Embryo transfer was performed 48 to 72 hours after retrieval. A maximum of four embryos was transferred, and the rest was cryopreserved. Results were compared with those of 208 patients who underwent a total of 458 ovum donation cycles. These included patients with ovarian failure (primary or secondary) as well as eugonadal patients who were enrolled because of failure of previous treatments or because of a genetic trait precluding use of their own oocytes. Eugonadal patients were defined based on a history of regular menstrual cycles in the presence of normal gonadotropins (FSH < 20 mIU/mL, LH < 20 mIU/mL [conversion factor to SI unit, 1.00). According to the regulations of the Israeli Ministry of Health, oocyte donation may only be done anonymously by patients undergoing IVF themselves. Oocyte donors were therefore young (~32 years old), healthy women undergoing IVF, who willingly consented to donate oocytes when there was an excess thereof. Patients on standby for oocyte donation, regardless of ovarian function, were treated with 2 mg E2 valerate (Progynon; Schering AG, Berlin, Germany) three times Vol. 60, No.2, August 1993
a day for endometrial preparation. The duration of the treatment with E2 varied in accordance with the availability of the oocytes. Beginning on the day when oocytes for donation were aspirated, a daily dose of 100 mg P in oil (Geston; Paines and Byrne Limited, Greenford, Middlesex, United Kingdom) was administered. On the same day, at least five donated oocytes were inseminated with sperm from the recipient's spouse. In cases of male factor, donor sperm was used. Embryo transfer was carried out 48 to 72 hours later at a four- to eight-cell stage. Twelve days later, the serum ~-subunit of hCG levels were determined, and if pregnancy was confirmed, the daily doses of E2 valerate and P were continued until 14 weeks of gestation. Ultrasound was performed at 6 and 8 weeks' gestation to confirm clinical pregnancy. The proportion of cycles using donor sperm did not differ significantly among all groups. Embryo quality was assessed by gross morphological features. Only embryos with equal-size blastomeres and no, or minimal, fragmentation were transferred (grades 1 and 2, respectively) (6). Results were analyzed statistically by x 2 test and Student's t-test. Logistic regression analysis was used to assess independent effects of age and ovarian function on clinical PRs. RESULTS
Patients were first divided according to age. In standard IVF, there were 325 transfer cycles in patients ~ 40 years of age, and 1,103 transfer cycles in patients < 40 years (Table 1). The mean numbers of embryos transferred were 3.4 ± 1.2 and 3.1 ± 1.4, respectively; the mean numbers of blastomeres/ embryo were 5.2 ± 1.8 and 5.7 ± 1.6, respectively. There was no difference in the embryo quality when comparing both groups (Grade 1 embryos 78% and 82%, respectively,). The clinical PRs/transfer were significantly lower in older patients (12.9% versus 23.8%, P = 0.0001, respectively). The rates of
Table 1 Results of Treatment in Patients Undergoing Standrd IVF
Transfer cycles Pregnancies* Miscarriagest
Age < 40
Age:2: 40
P value
1,103 262 (23.8) 37 (14.1)
325 42 (12.9) 13 (30.9)
0.0001 0.0065
* Numbers in parentheses are clinical PRs. t Numbers in parentheses are miscarriage rates.
YarOB et al.
Endometrial receptivity and egg donation
315
Table 2 Results of Treatment in All Patients Undergoing Egg Donation
Transfer cycles Pregnancies* Miscarriagest
~
Age < 40
Age
222 65 (29.3) 8 (12.3)
236 50 (21.2) 9 (18.0)
40
0.043 0.56
miscarriage were significantly higher in older patients (30.9% versus 14.1%, P = 0.0065, respectively). In ovum donation, there were 236 cycles in patients age 40 years or above and 222 cycles in patients < 40 years (both in eugonadal patients and in those with ovarian failure). The mean numbers of embryos transferred were 3.3 ± 1.5 and 3.0 ± 1.2, respectively. The mean numbers of blastomeres/ embryo were 5.6 ± 1.7 and 5.5 ± 1.8, respectively. There was no difference in the embryo quality when comparing both groups (grade 1 embryos 79% and 84%, respectively). Clinical PRs/transfer were significantly lower in older patients (21.2% versus 29.3%, P = 0.043, respectively). Unlike patients undergoing standard IVF, miscarriage rates did not differ significantly in either group (Table 2). Patients were then redivided according to treatment and ovarian function (Table 3). Recipients were redivided into two groups: ovarian failure group, 219 cycles in patients diagnosed as having either primary or secondary ovarian failure, and eugonadal group, 239 cycles in patients with retained ovarian function. These were compared with the results of 1,428 transfer cycles in standard IVF. The mean number of embryos transferred was 3.1 ± 1.3, 3.2 ± 0.9, and 2.9 ± 1.1, respectively. The mean numbers of blastomeres/embryo were 5.2 ± 1.9, 5.3 ± 1.2, and 5.8 ± 1.8, respectively. There was no difference in the embryo quality when comparing both groups (grade 1 embryos 82%, 80%, and 77% respectively). A significantly higher PR was noted in patients with ovarian failure (31.1 %, P < 0.005), compared with both eugonadal patients undergoing ovum donation (19.7%) and standard IVF patients (21.3%). Miscarriage rates did not differ significantly in all groups. Moreover, logistic regression analysis was also used to analyze the effect of each variable (age, ovarian function) while controlling for the other. Independent effects for age (Wald x2 = 4.28, Yaron et al.
= =
0.039) and ovarian function (Wald 0.025) on PRs were observed.
x2
=
5.04;
Pvalue
* Numbers in parentheses are clinical PRs. t Numbers in parentheses are miscarriage rates.
316
P P
Endometrial receptivity and egg donation
DISCUSSION
With modern means of ovulation induction and laboratory techniques, oocyte retrieval and ET may be achieved in most patients undergoing IVF. Pregnancy rates, however, are still disappointingly low (22.8%/transfer) (7). In older patients, conception rates are even lower. Cumulative conception rates after five treatment cycles were 54 % in women between 20 and 34 years of age, but only 20.2% for those age 40 years and above (4). We too found a significant decline in clinical PRs in women 40 years or older undergoing IVF (Table 1). Navot et al. (8) claim that poor oocyte quality, rather than implantation failure, is the leading cause of age-related decline in female fertility. Higher PRs were found in patients 40 years or older who received donated oocytes, when compared with previous standard IVF treatments with their own oocytes. The authors attributed this finding to a better quality of the donated oocytes/embryos, rather than to endometrial receptivity. However, they did not consider the fact that a significantly' higher amount of embryos were transferred in donation cycles as compared with standard IVF cycles in the same patients (4.5 versus 1.0), precluding any comparison between the treatments on grounds of oocyte or embryo quality. Others believe that with age, endometrial receptivity plays a more important role in the decline in PRs. It has been claimed that an age-related decrease in uterine perfusion might be the cause for the decline in fertility with age (9). Indeed, in aging animals, marked uterine and endometrial changes
Table 3 of Age
Results of Treamtent in All Groups, Regardless
Oocyte donation
Transfer cycles Pregnancies* Miscarriagest
Standard IVF
Eugonadal
Ovarian failure
1,428 304 (21.3) 50 (16.4)
239 47 (19.7) 8 (17.0)
219 68 (31.1) 9 (13.2)
* Numbers in parentheses are clinical PRs. Pregnancy rates in patients with ovarian failure are significnatly greater than both eugonadal and standard IVF patients (P < 0.005, x 2 test). t Numbers in parentheses are miscarriage rates. No significant difference exists among the groups. Fertility and Sterility
were detected. These included an increased collagen content, a reduced number of stromal cells (10), and fewer E2 receptors on endometrial cell surface (11). However, no such direct evidence is available in humans. Moreover, Sauer et al. (12) stated that endometrial-biopsy specimens from women receiving hormone replacement therapy are histologically indistinguishable from that of normal endometrium, regardless of the patient's age. Furthermore, the authors found no statistically significant differences in implantation and ongoing PRs, when comparing older with younger women. The number of cycles in each group was low, perhaps accounting for a lack of significant difference. Conversely, our results show that clinical PRs in ovum donation were higher in patients < 40 years, compared with those 40 years or older (Table 2). Similar findings were previously reported (13, 14). Because oocytes donated to both groups of recipients were obtained from young patients, the difference in clinical PRs cannot be attributed to oocyte quality. This supports the contention that a decrease in endometrial receptivity with age is responsible for a higher rate of implantation failure in older women. With standard IVF, the need for controlled ovarian hyperstimulation (CO H) results in supraphysiological levels of E 2, known to have an interceptive effect on the endometrium in animals and in humans (15-18). On the other hand, oocyte donation programs alleviate the need for COH, and the use of simple hormone replacement therapy creates a more physiological endometrial environment. Thus, when such unstimulated patients receive donated oocytes, higher PRs are achieved when compared with standard IVF (19-21), GIFT (22), and with frozen-thawed embryos (13). Undoubtedly, in our study patients with ovarian failure receiving donated oocytes performed better than standard IVF patients (Table 3). However, eugonadal patients undergoing ovum donation did not fare as well as those with ovarian failure and had clinical PRs that did not differ statistically from standard IVF patients. This may be the result of adverse effects of endogenous ovarian hormones on the endometrium in eugonadal patients. An alternative explanation for the higher PR in patients with ovarian failure compared with eugonadal patients might be related to higher gonadotropin levels in the former. This may possibly be mediated by a beneficial effect of gonadotropins on the endometrium because LH receptors were found in the endometrium of animals (23) and humans (Reshef E, Lei ZM, Chegini N, Rao Ch V, Luborsky JL, abstract). Still another Vol. 60, No.2, August 1993
explanation may lie in the fact that the group of eugonadal patients was somewhat heterogeneous, including patients who had failed previous standard IVF treatments and possibly having an as-yet unrecognized endometrial factor. Similar results were reported by Edwards et al. (3) in patients undergoing ovum donation. They too found that clinical PRs were higher in previously amenorrhoeic women than in eugonadal women, irrespective of age and number of embryos transferred. If, indeed, patients with non functioning ovaries do better in ovum donation programs, perhaps pretreatment with a GnRH analogue that creates medical hypophysectomy may improve success rates in eugonadal patients undergoing ET (24). Our results show that miscarriage rates are significantly higher in standard IVF patients who were 40 years or older, compared with younger patients (Table 1). This may either be the result of poor oocyte quality in older women or due to age-related endometrial factors. However, in patients undergoing ovum donation, no such difference was noted (Table 2). This supports the contention that poor oocyte quality, rather than a decline in receptivity, is responsible for the higher miscarriage rates in older women. This is also in agreement with the high rate of chromosomally abnormal ova known to be present in older women (25). In conclusion, clinical PRs decline with age as a result of reduced endometrial receptivity. Miscarriage rates increase with age, mainly because of oocyte senescence. Patients with ovarian failure do better in ovum donation than patients with functioning ovaries. REFERENCES 1. Edwards RG, Fishel SB, Cohen J, Fehilly CB, Purdy JM,
2.
3.
4.
5.
Slater JM, et al. Factors influencing the success of in vitro fertilization for alleviating human infertility. J In Vitro Fert Embryo Transf 1984;1:3-23. Sharma V, Riddle A, Mason BA, Pampiglione J, Campbell S. An analysis of factors influencing the establishment of a clinical pregnancy in an ultrasound-based ambulatory in vitro fertilization program. Fertil Steril 1988;49:468-78. Edwards RG, Morcos S, Macnamee M, Balmaceda JP, Walters DE, Asch R. High fecundity of amenorrhoeic women in embryo-transfer programmes. Lancet 1991; 338:292-4. Tan SL, Roystone P, Campbell S, Jacobs HS, Betts J, Mason B, Edwards RG. Cumulative conception and livebirth rates after in-vitro fertilization. Lancet 1992;339:1390-4. Romeu A, Muasher SJ, Acosta AA, Veeck LL, Diaz J, Jones GS, et al. Results of in vitro fertilization attempts in women 40 years of age and older: the Norfolk experience. Fertil Steril 1987;47:130-6.
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6. Veeck LL. Oocyte assessment and biological performance. Ann NY Acad Sci 1989;541:259-74. 7. Medical Research International, Society for Assisted Reproductive Technology (SART), The American Fertility Society. In vitro fertilization-embryo transfer (lVF-ET) in the United States: 1990 results from the IVF-ET Registry. Fertil Steril 1992;57:15-24. 8. Navot D, Bergh PA, Anne Williams M, John Garrisi G, Guzman I, Sandler B, et al. Poor oocyte quality rather than implantation failure as a cause of age-related decline in female fertility. Lancet 1991;337:1375-7. 9. Goswamy RK, Williams G, Steptoe PC. Decreased uterine perfusion: a cause of infertility. Hum Reprod 1988;3:955-9. 10. Craig SS, Jollie WP. Age changes in density of endometrial stromal cells of the rat. Exp Gerontol 1985;20:93-7. 11. Han Z, Kokkonen GC, Roth GS. Effect of aging on populations of estrogen receptor-containing cells in the rat uterus. Exp Cell Res 1989;180:234-42. 12. Sauer MV, Paulson RJ, Lobo RA. A preliminary report on oocyte donation extending reproductive potential to women over 40. N Engl J Med 1990;323:1157-60. 13. de Ziegler D, Frydman R. Different implantation rates after transfers of cryopreserved embryos originating from donated oocytes or from regular in vitro fertilization. Fertil Steril 1990;54:682-8. 14. Levran D, Ben-Shlomo I, Dor J, Ben-Rafael Z, Nebel L, Mashiach S. Aging of endometrium and oocytes: observations on conception and abortion rates in an egg donation model. Fertil Steril1991;56:1091-4. 15. Gidley-Baird AA, O'Neill C, Sinosich MJ, Porter RN, Pike IL, Saunders DM. Failure of implantation in human in vitro fertilization and embryo transfer patients: the effects of altered progesterone/estrogen ratios in humans and mice. Fertil Steril 1986;45:69-74. 16. Martel D, Frydman R, Glissant M, Maggioni C, Roche D, Psychoyos A. Scanning electron microscopy of postovula-
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17.
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tory human endometrium in spontaneous cycles and cycles stimulated by hormone treatment. J Endocrinol 1987;114: 319-24. Forman RG, Eychenne B, Nessmann C, Frydman R, Robel P. Assessing the early luteal phase in in vitro fertilization cycles: relationships between plasma steroids, endometrial receptors, and endometrial histology. Fertil Steril 1989;51: 310-6. Fossum GT, Davidson A, Paulson RJ. Ovarian hyperstimulation inhibits embryo implantation in the mouse. J In Vitro Fert Embryo Transf 1989;6:7-10. Navot D, Laufer N, Kopolovic J, Rabinowitz R, Birkenfeld A, Lewin A, et al. Artificially induced endometrial cycles and establishment of pregnancies in the absence of ovaries. N Engl J Med 1986;314:806-11. Paulson RJ, Sauer MV, Lobo RA. Embryo implantation after human in vitro fertilization: importance of endometrial receptivity. Fertil Steril 1990;53:870-4. Sauer MV, Paulson RJ, Macaso TM, Francis MM, Lobo RA. Oocyte and pre-embryo donation to women with ovarian failure: an extended clinical trial. Fertil Steril 1991;55:39-43. Asch RH, Balmaceda JP, Ord T, Borrero C, Cefalu E, Gastaldi C, et al. Oocyte donation and gamete intrafallopian transfer in premature ovarian failure. Fertil Steril 1988;49:263-7. Ziecik AJ, Stanchev PD, Tilton JE. Evidence for the presence of luteinizing hormone/human chorionic gonadotropin-binding sites in the porcine uterus. Endocrinology 1986;119:1159-63. Meldrum DR, Wisot A, Hamilton F, Gutlay-Yeo AL, Man B, Huynh D. Artificial agonadism and hormone replacement for oocyte donation. Fertil Steril1989;52:509-11. Riesch HA, Weiss NS, Clarke EA, Miller AB. Risk factors for spontaneous abortion and its recurrence. Am J Epidemiol 1988;128:420-30.
Fertility and Sterility
©
Vol. 60, No, 2, August 1993
Printed on acid-free paper in U.
1998 The American Fertility Society
s.
A.
Endometrial receptivity: the age-related decline in pregnancy rates and the effect of ovarian function
Yuval Yaron, M,D, Amnon Botchan, M,D, Ami Amit, M,D.
Abraham Kogosowski, M.D. Israel Yovel M.D. Joseph B. Lessing, M.D.*
In Vitro Fertilization/Embryo Transfer Unit, Serlin Maternity Hospital, Tel Aviv Sourasky Medical Center, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
Objective: To assess the effect of age and ovarian function on endometrial receptivity. Design: Retrospective comparison between standard IVF and ovum donation in younger and older patients «40 and :?40 years of age, respectively). Patients: In standard IVF, there were 325 transfer cycles in older patients and 1,103 transfer cycles in younger ones. In ovum donation, there were 236 transfer cycles in older patients and 222 cycles in younger women. Ovum recipients were then redivided into two groups, according to ovarian function: ovarian failure group (219 cycles) and eugonadal group (239 cycles) in patients with retained ovarian function as manifested by regular menstrual cycles and normal gonadotropins. Results: In standard IVF, clinical pregnancy rates (PRs) were significantly lower in older patients (12.9% versus 23.8%, respectively). In ovum donation, clinical PRs were also significantly lower in older patients (21.2% versus 29.3%, respectively). A significantly higher clinical PR (31.1 %) was noted in patients with ovarian failure, compared with both eugonadal patients undergoing ovum donation (19.7%) and standard IVF patients (21.3%). Conclusions: The decrease in endometrial receptivity with age is responsible for the higher rate of implantation failure in older women. Patients with nonfunctioning ovaries do better than eugonadal patients in ovum donation programs. Fertil Steri11993;60:314-8 Key Words: Ovum donation, IVF, endometrial receptivity
The fact that fertility in women declines with age is true for both normal fertile women, as well as for those treated by assisted reproductive techniques. Many IVF units have previously rejected women > 40 years of age because of the expected decline of fertility in these women (1-4). This decline was neither related to the number of oocytes harvested, fertilized, or transferred nor to the serum steroid levels (5). The lower clinical pregnancy rate (PR) in older women undergoing IVF may be attributed to oocyte senescence, or age-related decline in endo-
Received December 8, 1993; revised and accepted April 27, 1993. * Reprint requests: Joseph B. Lessing, M.D., Serlin Maternity Hospital, Department of Obstetrics and Gynecology, Post Office Box 7079, Tel Aviv 61070, Israel.
314
Yaron et al.
Endometrial receptivity and egg donation
metrial receptivity. The question of which of these is more important has been difficult to resolve because in standard IVF, both the oocytes/embryos and the endometrium are from one and the same patient. However, ovum donation is unique in that the oocytes are obtained from another patient and oocyte/embryo quality may be controlled for, allowing evaluation of endometrial receptivity. When comparing IVF with ovum donation, we examined the role of endometrial receptivity in the agerelated decline of fertility. Initially, only patients with ovarian failure were included in ovum donation programs. Recently, however, ovum donation has proved itself to be an essential mode of treatment for other types of infertility: [1] patients with normal menstrual cycles who have repeatedly failed previous IVF Fertility and Sterility
treatments; [2) those who do not respond to hMG; or [3) individuals with a genetic trait precluding use of their own oocytes. Unlike those with ovarian failure, the endometrium in these patients is subject to the influence of endogenous ovarian steroids. Thus, a comparison between such patients and those with ovarian failure allows for assessment of endometrial receptivity with regard to ovarian function. MATERIALS AND METHODS
Between January 1986 and December 1992, 579 patients underwent 1,428 transfer cycles in standard IVF. We included only those patients treated with hMG alone. A standard protocol was used: on the 3rd day of the menstrual cycle, three ampules of hMG (Pergonal; Teva Pharmaceutical Industries Limited, Petah Tikva, Israel) were given daily. This daily dose was continued until at least two follicles with a mean diameter of 17 mm were seen on ultrasound (US), and the E 2 level was ~800 pg/mL (conversion factor to SI unit, 3.671). At this stage, 10,000 IU of hCG (Chorigon; Teva Pharmaceutical Industries Limited) was administered. Oocyte retrieval was scheduled 35 hours after hCG administration, using an US-guided transvaginal approach. Aspirated oocytes were inseminated and cultured in Ham's F -10 medium (KA 128 NB; Flow Laboratories, Irvine, Scotland), supplemented with 10% to 20% heat-inactivated patients' serum. Embryo transfer was performed 48 to 72 hours after retrieval. A maximum of four embryos was transferred, and the rest was cryopreserved. Results were compared with those of 208 patients who underwent a total of 458 ovum donation cycles. These included patients with ovarian failure (primary or secondary) as well as eugonadal patients who were enrolled because of failure of previous treatments or because of a genetic trait precluding use of their own oocytes. Eugonadal patients were defined based on a history of regular menstrual cycles in the presence of normal gonadotropins (FSH < 20 mIU/mL, LH < 20 mIU/mL [conversion factor to SI unit, 1.00). According to the regulations of the Israeli Ministry of Health, oocyte donation may only be done anonymously by patients undergoing IVF themselves. Oocyte donors were therefore young (~32 years old), healthy women undergoing IVF, who willingly consented to donate oocytes when there was an excess thereof. Patients on standby for oocyte donation, regardless of ovarian function, were treated with 2 mg E2 valerate (Progynon; Schering AG, Berlin, Germany) three times Vol. 60, No.2, August 1993
a day for endometrial preparation. The duration of the treatment with E2 varied in accordance with the availability of the oocytes. Beginning on the day when oocytes for donation were aspirated, a daily dose of 100 mg P in oil (Geston; Paines and Byrne Limited, Greenford, Middlesex, United Kingdom) was administered. On the same day, at least five donated oocytes were inseminated with sperm from the recipient's spouse. In cases of male factor, donor sperm was used. Embryo transfer was carried out 48 to 72 hours later at a four- to eight-cell stage. Twelve days later, the serum ~-subunit of hCG levels were determined, and if pregnancy was confirmed, the daily doses of E2 valerate and P were continued until 14 weeks of gestation. Ultrasound was performed at 6 and 8 weeks' gestation to confirm clinical pregnancy. The proportion of cycles using donor sperm did not differ significantly among all groups. Embryo quality was assessed by gross morphological features. Only embryos with equal-size blastomeres and no, or minimal, fragmentation were transferred (grades 1 and 2, respectively) (6). Results were analyzed statistically by x 2 test and Student's t-test. Logistic regression analysis was used to assess independent effects of age and ovarian function on clinical PRs. RESULTS
Patients were first divided according to age. In standard IVF, there were 325 transfer cycles in patients ~ 40 years of age, and 1,103 transfer cycles in patients < 40 years (Table 1). The mean numbers of embryos transferred were 3.4 ± 1.2 and 3.1 ± 1.4, respectively; the mean numbers of blastomeres/ embryo were 5.2 ± 1.8 and 5.7 ± 1.6, respectively. There was no difference in the embryo quality when comparing both groups (Grade 1 embryos 78% and 82%, respectively,). The clinical PRs/transfer were significantly lower in older patients (12.9% versus 23.8%, P = 0.0001, respectively). The rates of
Table 1 Results of Treatment in Patients Undergoing Standrd IVF
Transfer cycles Pregnancies* Miscarriagest
Age < 40
Age:2: 40
P value
1,103 262 (23.8) 37 (14.1)
325 42 (12.9) 13 (30.9)
0.0001 0.0065
* Numbers in parentheses are clinical PRs. t Numbers in parentheses are miscarriage rates.
YarOB et al.
Endometrial receptivity and egg donation
315
Table 2 Results of Treatment in All Patients Undergoing Egg Donation
Transfer cycles Pregnancies* Miscarriagest
~
Age < 40
Age
222 65 (29.3) 8 (12.3)
236 50 (21.2) 9 (18.0)
40
0.043 0.56
miscarriage were significantly higher in older patients (30.9% versus 14.1%, P = 0.0065, respectively). In ovum donation, there were 236 cycles in patients age 40 years or above and 222 cycles in patients < 40 years (both in eugonadal patients and in those with ovarian failure). The mean numbers of embryos transferred were 3.3 ± 1.5 and 3.0 ± 1.2, respectively. The mean numbers of blastomeres/ embryo were 5.6 ± 1.7 and 5.5 ± 1.8, respectively. There was no difference in the embryo quality when comparing both groups (grade 1 embryos 79% and 84%, respectively). Clinical PRs/transfer were significantly lower in older patients (21.2% versus 29.3%, P = 0.043, respectively). Unlike patients undergoing standard IVF, miscarriage rates did not differ significantly in either group (Table 2). Patients were then redivided according to treatment and ovarian function (Table 3). Recipients were redivided into two groups: ovarian failure group, 219 cycles in patients diagnosed as having either primary or secondary ovarian failure, and eugonadal group, 239 cycles in patients with retained ovarian function. These were compared with the results of 1,428 transfer cycles in standard IVF. The mean number of embryos transferred was 3.1 ± 1.3, 3.2 ± 0.9, and 2.9 ± 1.1, respectively. The mean numbers of blastomeres/embryo were 5.2 ± 1.9, 5.3 ± 1.2, and 5.8 ± 1.8, respectively. There was no difference in the embryo quality when comparing both groups (grade 1 embryos 82%, 80%, and 77% respectively). A significantly higher PR was noted in patients with ovarian failure (31.1 %, P < 0.005), compared with both eugonadal patients undergoing ovum donation (19.7%) and standard IVF patients (21.3%). Miscarriage rates did not differ significantly in all groups. Moreover, logistic regression analysis was also used to analyze the effect of each variable (age, ovarian function) while controlling for the other. Independent effects for age (Wald x2 = 4.28, Yaron et al.
= =
0.039) and ovarian function (Wald 0.025) on PRs were observed.
x2
=
5.04;
Pvalue
* Numbers in parentheses are clinical PRs. t Numbers in parentheses are miscarriage rates.
316
P P
Endometrial receptivity and egg donation
DISCUSSION
With modern means of ovulation induction and laboratory techniques, oocyte retrieval and ET may be achieved in most patients undergoing IVF. Pregnancy rates, however, are still disappointingly low (22.8%/transfer) (7). In older patients, conception rates are even lower. Cumulative conception rates after five treatment cycles were 54 % in women between 20 and 34 years of age, but only 20.2% for those age 40 years and above (4). We too found a significant decline in clinical PRs in women 40 years or older undergoing IVF (Table 1). Navot et al. (8) claim that poor oocyte quality, rather than implantation failure, is the leading cause of age-related decline in female fertility. Higher PRs were found in patients 40 years or older who received donated oocytes, when compared with previous standard IVF treatments with their own oocytes. The authors attributed this finding to a better quality of the donated oocytes/embryos, rather than to endometrial receptivity. However, they did not consider the fact that a significantly' higher amount of embryos were transferred in donation cycles as compared with standard IVF cycles in the same patients (4.5 versus 1.0), precluding any comparison between the treatments on grounds of oocyte or embryo quality. Others believe that with age, endometrial receptivity plays a more important role in the decline in PRs. It has been claimed that an age-related decrease in uterine perfusion might be the cause for the decline in fertility with age (9). Indeed, in aging animals, marked uterine and endometrial changes
Table 3 of Age
Results of Treamtent in All Groups, Regardless
Oocyte donation
Transfer cycles Pregnancies* Miscarriagest
Standard IVF
Eugonadal
Ovarian failure
1,428 304 (21.3) 50 (16.4)
239 47 (19.7) 8 (17.0)
219 68 (31.1) 9 (13.2)
* Numbers in parentheses are clinical PRs. Pregnancy rates in patients with ovarian failure are significnatly greater than both eugonadal and standard IVF patients (P < 0.005, x 2 test). t Numbers in parentheses are miscarriage rates. No significant difference exists among the groups. Fertility and Sterility
were detected. These included an increased collagen content, a reduced number of stromal cells (10), and fewer E2 receptors on endometrial cell surface (11). However, no such direct evidence is available in humans. Moreover, Sauer et al. (12) stated that endometrial-biopsy specimens from women receiving hormone replacement therapy are histologically indistinguishable from that of normal endometrium, regardless of the patient's age. Furthermore, the authors found no statistically significant differences in implantation and ongoing PRs, when comparing older with younger women. The number of cycles in each group was low, perhaps accounting for a lack of significant difference. Conversely, our results show that clinical PRs in ovum donation were higher in patients < 40 years, compared with those 40 years or older (Table 2). Similar findings were previously reported (13, 14). Because oocytes donated to both groups of recipients were obtained from young patients, the difference in clinical PRs cannot be attributed to oocyte quality. This supports the contention that a decrease in endometrial receptivity with age is responsible for a higher rate of implantation failure in older women. With standard IVF, the need for controlled ovarian hyperstimulation (CO H) results in supraphysiological levels of E 2, known to have an interceptive effect on the endometrium in animals and in humans (15-18). On the other hand, oocyte donation programs alleviate the need for COH, and the use of simple hormone replacement therapy creates a more physiological endometrial environment. Thus, when such unstimulated patients receive donated oocytes, higher PRs are achieved when compared with standard IVF (19-21), GIFT (22), and with frozen-thawed embryos (13). Undoubtedly, in our study patients with ovarian failure receiving donated oocytes performed better than standard IVF patients (Table 3). However, eugonadal patients undergoing ovum donation did not fare as well as those with ovarian failure and had clinical PRs that did not differ statistically from standard IVF patients. This may be the result of adverse effects of endogenous ovarian hormones on the endometrium in eugonadal patients. An alternative explanation for the higher PR in patients with ovarian failure compared with eugonadal patients might be related to higher gonadotropin levels in the former. This may possibly be mediated by a beneficial effect of gonadotropins on the endometrium because LH receptors were found in the endometrium of animals (23) and humans (Reshef E, Lei ZM, Chegini N, Rao Ch V, Luborsky JL, abstract). Still another Vol. 60, No.2, August 1993
explanation may lie in the fact that the group of eugonadal patients was somewhat heterogeneous, including patients who had failed previous standard IVF treatments and possibly having an as-yet unrecognized endometrial factor. Similar results were reported by Edwards et al. (3) in patients undergoing ovum donation. They too found that clinical PRs were higher in previously amenorrhoeic women than in eugonadal women, irrespective of age and number of embryos transferred. If, indeed, patients with non functioning ovaries do better in ovum donation programs, perhaps pretreatment with a GnRH analogue that creates medical hypophysectomy may improve success rates in eugonadal patients undergoing ET (24). Our results show that miscarriage rates are significantly higher in standard IVF patients who were 40 years or older, compared with younger patients (Table 1). This may either be the result of poor oocyte quality in older women or due to age-related endometrial factors. However, in patients undergoing ovum donation, no such difference was noted (Table 2). This supports the contention that poor oocyte quality, rather than a decline in receptivity, is responsible for the higher miscarriage rates in older women. This is also in agreement with the high rate of chromosomally abnormal ova known to be present in older women (25). In conclusion, clinical PRs decline with age as a result of reduced endometrial receptivity. Miscarriage rates increase with age, mainly because of oocyte senescence. Patients with ovarian failure do better in ovum donation than patients with functioning ovaries. REFERENCES 1. Edwards RG, Fishel SB, Cohen J, Fehilly CB, Purdy JM,
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