Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
The Impact of Embryo Quality on Pregnancy Outcome in Elderly Women Undergoing in Vitro Fertilization-Embryo Transfer (IVF-ET) DOV FELDBERG, 1 JACOB FARHI, 1 DOV DICKER, 1 JACOB ASHKENAZI, 1 MICHAL SHELEF, l and JACK A. GOLDMAN 1'2
Most infertility programs, especially IVF-ET units, limit the admission to women aged under 38 (1-3). Nevertheless, in many Western countries a change in birth habits is observed in the past decade or two. Furthermore, one is aware of demands for infertility treatment by women aged 40 and over, including IVF therapy (4). Edwards et al. (5,6) claim that since fertilization rates are low in this age group, while implantation processes are rare and defective, they are considered part of the unexplained infertility group, until biological factors of the aging reproductive system are more accurately defined. It is well to remember that at the present time reconstructive surgery of the fallopian tubes in patients over 40 with a severe mechanical factor is no longer considered indicated in view of low rates of success in this age group (7). Thus, the in vitro fertilization alternative must be offered as another therapeutic process, as long as gonadotropins are within normal limits. Moreover, there is no doubt that nowadays the "elderly" woman with infertility problems requires a thorough diagnostic and therapeutic workup. There is little information on the subject of patients over 40 as compared to a group of younger patients treated in an IVF-ET program (8,9). Data from our own unit (10) on the subject of ovulation induction in an elderly female population, using a specific protocol including pFSH + hMG for preparation to IVF, indicate impressive achievements in pregnancy rates; yet first-trimester abortion rates were exceptionally high in women over 40 when compared to a population of younger women treated with an identical protocol for ovulation induction. Fishel and associates (l l) also reported rather
Submitted: October 16, 1989 Accepted: May 15, 1990
Most IVF-ET units limit the procedure to women below age 38. Nevertheless, demands for infertility treatment, including I V F therapy, are more frequent nowadays. We compared 46 cycles for ovulation induction for I V F in 46 women aged40 or more (Group I) to 51 induced cycles for this procedure in younger women o f mean age 30.2 years (Group II). Cancellation rates due to early luteinization or ovulation were significantly higher in group I than in group H (28.2 and 17.6%, respectively) (P < 0.001). Also, significantly higher abortion rates were observed in older women (62.5%), in comparison to the younger control group, (25%), (P < 0.001). It is suggested that the high abortion rate considered to be due to genetic factors in older women, may possibly also be due to the aging uterine environment. Furthermore, while embryos with fragmentations may often produce clinical pregnancies in the young, the aging uterus in the elderly woman does not encourage the development o f clinical pregnancies in such embryos. Consequently the higher abortion rate in the elderly woman. KEY WORDS: in vitro fertilization; elderly women; embryo quality, pregnancy outcome.
INTRODUCTION One of the controversial issues in the management of infertility is the problem of the woman's age. Sherman Institute of Fertility, Department of ObstetricsGynecology, Golda Meir Medical Center (Hasharon Hospital), Petah-Tikva, and Tel-Aviv University Medical School, TelAviv, Israel. 2 To whom correspondence should be addressed at Department of Obstetrics-Gynecology, Golda Meir Medical Center, (Hasharon Hospital), Petah Tikva, Israel, 49372.
257
0740-7769/90/1000-0257506.00/0 9 1990PlenumPublishingCorporation
258
FELDBERG ETAL.
low "take-home baby" rates resulting from those "elderly" pregnancies as compared to a younger age group, while Johnston (1) emphasizes the occurrence of autosomal dominant diseases in offspring of these aged women, often associated with sperm defects of an aged husband. The present publication reports clinical and laboratory data of pregnancies achieved in women older than 40 years following an IVF-ET procedure, compared to a control group of younger women. We have attempted to concentrate on the possible causative factors for the large number of abortions in this group of "elderly" women. MATERIALS AND METHODS Between July and December 1987, 46 women aged over 40 years (group I) were treated in 46 cycles of ovulation induction for the purpose of IVF. Mean age was 40.6 -+ 0.5 years, with a range of 40-43 years. During the same period of time and under identical conditions, 51 patients (group II) of mean age 30.2 +-- 2.7 years, with a range of 27-33.6 years, were treated during 51 ovulation induction cycles. Clinical data of the study group and the control group are listed in Table I. The mean duration of infertility, the different indications for IVF treatment, and the number of previous cycles of therapy were identical in both groups. The ovulation induction protocol (12) consisted of the administration of 3-4 ampoules of pure follicle stimulating hormone (pFSH) (Metrodin, Serono Laboratories, Randolph, MA) daily for 3 days, starting on the third cycle day, followed by 3-4 ampoules daily of human menopausal gonadotropin Table I. Clinical Data of Study and Control Groups Data Mean duration infertility (years; M --- SD) Tubal infertility Unexplained infertility Male infertility Endometriosis Cervical factor Previous IVF cycles* 1 2 />3
*P, NS.
Group I (n = 46)
Group II (n = 51)
8.9 -+ 1.3 25 (54.3%) 13 (28.3%) 5 (10.9%) 2 (4.3%) 1 (2.2%)
7.4 --- 2.6 34 (66.7%) 10 (19.7%) 5 (9.8%) 1 (1.9%) 1 (1.9%)
21 (45.6%) 13 (28.3%) 12 (26.1%)
16 (31.3%) 21 (41.2%) 14 (27.5%)
(hMG) (Pergonal, Serono Laboratories, Randolph, MA) according to individual response in each case, for as long as necessary. Daily monitoring of the development of ovarian follicles was started on the sixth day of the menstrual cycle and included plasma E2 and progesterone levels, as well as ultrasound estimation of the size of follicles. Following the presence of at least two follicles over 16 mm in diameter, E 2 levels above 400 pg/ml, and progesterone lower than 1.5 ng/ml, human chorionic gonadotropin (hCG) (Chorigon, Teva, Cfar-Saba, Israel), 10,000 units, was given. Ovum pickup was done by means of vaginal ultrasound under paracervical anesthesia (13). The morphology of the oocytes was described according to oocytec-corona--cumulus structure of the classification by Laufer et al. (14), and that of the embryos according to their pronuclear structure, polyspermic fertilization, regularity of blastomeres, and degree of fragmentation (15). All patients were given support in the luteal phase by means of progesterone suppositories, 25 mg twice daily, and serum beta hCG was done 10 days after embryo transfer. Clinical pregnancy was defined as a pregnancy in which a sac, embryo, and fetal heart were demonstrated by ultrasound. Statistical evaluation was done by means of Student's t test and chi-square analysis.
RESULTS There was a significant difference in cancellation rates between the study group (28.2%) and the control group (17.6%) (P < 0.001). The reasons for cancellations were early ovulation or luteinization and refractory response of the ovaries to increasing levels of menotropins. No statistically significant differences were observed between the number of follicles present in response to ovulation induction, as well as between the average number of oocytes asTable II. Results of IVF in Study and Control Groups Data
Group I
Group II
No. inductions No. ovum pickups Cancellation rate* No. follicles aspirated No. oocytes No. oocytes/pickup (M -+ SD)
46 33 28.2% 132 114 3.5 --- 0.9
51 42 17.6% 186 141 3.4 --+ 1.1
* P < 0.001.
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
259
EMBRYO QUALITY AND PREGNANCY OUTCOME IN ELDERLY WOMEN
pirated, in the two groups (Table II). In no case in either group were no oocytes aspirates. No statistically significant differences were found between the morphological forms of oocyte fertilization rates or pregnancy rates in the study group of older women and those in the control group of young women (Tables III and IV). In order to define the etiological factors of abortions the morphological structure of embryos was evaluated. Findings such as pronuclear structure, polyspermic fertilizations, and the symmetry of blastomeres were compared in pregnancies and abortions. The polyspermic fertilization rates were 7.9% in the study group and 3.2% in the control group, a statistically significant difference. In all other variables no statistically significant difference was considered to be associated with the morphological structure of blastomeres and the pregnancy rates in the two groups (Table V). When the pregnancy rate achieved with the transfer of fragmented versus nonfragmented embryos in the study group was examined (Table VI), no statistically significant difference was found. Nevertheless, of all embryos transferred, only I of 9 embryos with fragmentations achieved pregnancy (11.2%), while 7 of 22 transferred embryos without fragmentation achieved pregnancy (32.7%). Identical findings were observed in the control group (Table VII). In the control group 31.2% of fragmented embryos resulted in pregnancy, as compared to
Table III. Distribution of Fertilized and Nonfertilized Oocytes* in Group I According to Their Morphology and Establishment of Pregnancy Pregnancy +, Fertilization
Pregnancy - , Fertilization
Total, Fertilization
Oocyte classification~
+
-
+
-
+
-
PSD PSF PMD PMF PLD PLF DSD DSF DMD DMF DLD DLF
4 3 1 3 2 5 2 2 0 3 0 0
2 3 3 1 1 2 2 1 7 1 1 0
3 3 6 3 6 1 1 0 6 3 3 3
3 3 4 4 1 1 2 1 3 2 0 2
7 6 7 6 8 6 3 2 6 6 3 3
4 6 7 6 2 3 4 2 10 4 1 2
Description of oocytes: P, pale; D, dark; S, small; M, medium; L, large; De, dense; F, diffuse. * P , NS.
Table IV. Distribution of Fertilized and Nonfertilized Oocytes* in Group II According to Their Morphology and Establishment of Pregnancy Pregnancy +, Fertilization
Pregnancy - , Fertilization
Total, Fertilization
Oocyte classificationa
+
-
+
-
+
-
PSDe PSF PMDe PMF PLDe PLF DSDe DSF DMDe DMF DLDe DLF
5 3 4 3 2 4 8 2 2 6 4 2
2 2 2 1 1 4 4 2 1 6 2 1
6 4 2 2 2 8 6 0 4 4 2 2
2 2 1 1 1 4 4 2 2 2 3 2
11 7 6 5 4 12 14 2 6 10 6 4
4 4 3 2 2 8 8 4 3 8 5 3
* P, NS. a Description of oocytes: P, pale; D, dark; S, small; M, medium; L, large; De, dense; F, diffuse.
29.1% in embryo transfer of nonfragmented conceptuses. DISCUSSION The outcome of 32 treatment cycles in 24 women aged 40 or more were compared to 38 cycles in 31 patients aged in their thirties. Therapy in both groups was given during the same period and by the same medical and laboratory team. No statistically significant differences were obvious as far as indications for IVF or previous therapeutic trials. The protocol for induction of ovulation was identical in both groups. Cancellation rates in the study group were significantly higher than in the control group, 28.2% as compared to 17.6%, respectively, but were acceptTable V. Fertilization and Pregnancy Rates in Study and Control Groups Data No. oocytes fertilized* No. transfers*
Group I
Group II
(n = 46)
(n = 51)
79 (68%) 31
108 (71.8)% 40
2.5 • 0.4 8
2.9 • 0.3 12
25.8% 5/8 (62.5%)
30% 3/12 (25%)
No. embryos per transfer* (M • SD) No. clinical pregnancies*
Pregnancy rates per transfer* Abortion rate**
*P, NS. ** T < 0.001.
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
260
FELDBERG ETAL.
Table VI. Pregnancy Rate in Embryos with and Without Fragmentation in the Study Group With fragmentation
Without fragmentation
+
1
-
8
7 15 22 7/22 (32.7%)
Pregnancy
Total Pregnancy rate*
9 1/9 (11.2%)
*P, NS.
able when compared to cancellations reported in the literature in IVF-treated younger population groups (16). In only three cases the cause for cancellation was refractory ovarian response, while in all other cases treatment cycles were cancelled because of early luteinization or ovulation. These findings differ from those of Kerin et al. (17), who reported poor ovarian response to induction in women aged 40 years or more. Gindoff et al. (8) nevertheless reported normal ovarian reaction to induction of ovulation with menotropins in an older age group. It is interesting to note that in the patients of the study group who reacted normally to ovarian stimulation, no difference was observed in the endocrine and follicular reaction as compared to those in the control group of younger patients. This fact is obvious in that the number of oocytes aspirated at ovum pickup, the morphologic characteristics, fertilization rates, and cell division were similar in both groups. Similar findings were reported by Romeo and associates (9), who studied a group of 29 "elderly" women, treated with IVF-ET at the Norfolk Center. In our group no significant difference was registered in the mean number of transferred embryos and in the pregnancy rates, which were similar, namely, 31.8% in the control group. The most dominant finding was the very high abortion rate in the study group of older women (62.5%) as compared to 25% in the control group. Other groups observed identical, high abortion Table VII. Pregnancy Rate in Embryos with and Without Fragmentation in the Control Group Pregnancy
With fragmentation
Without fragmentation
+ Total Pregnancy rate*
5 11 16 5/16 (31.2%)
7 17 24 7/24 (29.1%)
*P, NS.
rates in patients aged 40 or more, when treated with IVF-ET (9,18). Trounson et al. (19) attempted to find a correlation between the electron microscopic structure of embryos and the prediction of pregnancy, though without success. We compared the morphologic picture of oocytes according to the classification of Laufer et al. (14) dealing with fertilization rates in the study and control groups. From findings in Tables IV and V it is clearly evident that no correlation may be found between the morphology of oocytes resulting in pregnancy and that of oocytes that did not achieve pregnancy. In our study group polyspermic fertilizations were observed (7.8%) twice as often as in the control group (3.2%). It is possible that a hint may be found here: although oocytes in an older age group are microscopically and morphologically similar to those of the younger age group, they nevertheless demonstrate a functional defect of the zona pellucida, making possible polyspermic fertilization in a high percentage of cases. Possibly this is one cause for more frequent genetic aberrations in embryos of older women. In attempting to compare the morphology of embryos with form and symmetry of blastomeres in different phases of division, no statistically significant difference was found between the two groups. In focusing on a specific structural defect in the development of the embryo, i.e., fragmentations, we evaluated all embryos transferred with and without fragmentation in both groups of women. Although not statistically significant, a clear difference was established, as in the study group only 11.2% of fragmented embryos resulted in pregnancies, while 32.7% of nonfragmented e m b r y o s achieved clinical pregnancies. This finding was not registered in the control group, in which 31.2% of fragmented embryos, as compared to 29.1% of nonfragmented embryos, resuited in clinical pregnancies. We believe that these findings have not been reported this far. We suggest that the high rate of abortions, considered by most authors (8,9) to be due to genetic factors in older women, may possibly be explained, at least in part, by the uterine factor or environment in the uterine cavity. Undoubtedly an embryo with fragmentations possesses the potential to result in a smaller number of pregnancies than an embryo without this phenomenon. Moreover, while the young uterine environment allows the development of these " w e a k " embryos to produce clinical pregnancies, the aging
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
EMBRYO QUALITY AND PREGNANCY OUTCOME IN ELDERLY WOMEN
uterus in elderly women does not encourage it and a small number of pregnancies is registered in the implantation of embryos with fragmentations. Thus, the same uterine environment--endocrine as well as histological--which is "insufficient" due to advanced age, is unable to supply the requirements for the development of pregnancy as in younger age, resulting in high abortion rates. This has been found in animal experiments (18) in which it has been demonstrated that, with aging, the uterus becomes more fibrotic, the blood supply is poor, the thickness of the endometrium is reduced, and its capacity to produce prostaglandins declines. Nevertheless, the conclusion that the uterine factor is etiologically related to increased early pregnancy wastage in elderly women is currently challenged by data on embryo donation in menopausal women. Asch et al. (20) reported a high pregnancy rate in elderly women in his embryo donor program suggestive of ovarian and not uterine factor in those patients. In view of the above, it seems to us that the aging woman is entitled to IVF-ET treatment according to the usual indications, identical to those which apply to the younger patient population, with the hope of achieving the same pregnancy rates as in the latter group. Naturally, the "elderly" women should be informed of the current chances and higher abortion rates and warned of the need for chorionic villous samplings or amniocentesis in pregnancy. Further studies are necessary to evaluate if the tendency observed in our small group of patients will have statistical support in a large number of women studied.
4.
5.
6. 7.
8. 9.
10. 11.
12.
13.
14.
15.
16.
17.
REFERENCES 18. 1. Johnston WIH, Oke K, Speirs A, et al.: Patient selection for in vitro fertilization: Physical and psychological aspects. Ann NY Acad Sci 1985;442:490-503 2. Utian WH, Goldfarb JM, Sheean LA: Implementation of an in vitro fertilization program. J Vitro Fert Embryo Transfer 1984;1:72-75
19.
20.
3. Laufer N, DeCherney AH, Haseltine FP, et aL: Human in vitro fertilization employing individualized ovulation induc-
Journal o f in Vitro Fertilization a n d E m b r y o Transfer, VoL 7, N o . 5, 1990
261
tion by human menopausal gonadotropins. J Vitro Fert Embryo Transfer 1984;1:56-62 Quigley MM, Wolf DP: Human in vitro fertilization and embryo transfer at the University of Texas, Houston. J Vitro Fert Embryo Transfer 1984;1:29-33 Edwards RG, Fishel SB, Cohen J, et al.: Factors influencing the success of in vitro fertilization for alleviating human infertility. J Vitro Fert Embryo Transfer 1984;1:3-23 Edwards RG: In vitro fertilization and embryo replacement: Opening lecture. Ann NY Acad Sci 1985;442:1-22 Wood C, McMaster R, Rennie G, et al.: Factors influencing pregnancy rates following in vitro fertilization and embryo transfer. Fertil Steril 1985;43:245-250 Gindoff PR, Jewelewicz R: Reproductive potential in the older woman. Fertil Steril 1986;46:989-1001 Romeo A, Muasher SJ, Acosta AA, et al.: Results of in vitro fertilization attempts in women 40 years of age and older: The Norfolk experience. Fertil Steril 1987;47:130-136 Feldberg D, Ashkenazi J, Dicker D, et al.: IVF-ET in elderly women. Personal Communication, 1987 Fishel SB, Cohen J, Fehilly G, et al.: Factors influencing human embryonic development in vitro. Ann NY Acad Sci 1985;442:342-356 Feldberg D, Goldman JA, Shelef M, et al.: Comparison of a fixed and dynamic protocol for embryo transfer in an IVF/ ET program. Hum Reprod 1988;3:747-750 Feldberg D, Goldman JA, Ashkenazi J, et al.: Transvaginal oocyte retrieval controlled by vaginal probe for in vitro fertilization: A comparative study. J Ultrasound Med 1988;7: 339-343 Laufer N, DeCherney AH, Tarlatzis BC, et al.: The association between preovulatory serum 17-beta estradiol pattern and conception in human menopausal gonadotropin--human choronic gonadotropin stimulation. Fertil Steril 1986;46:7376 Veeck LL, Wortham JWE Jr, Witmyer J, et al.: Maturation and fertilization of morphologically immature human oocytes in a program of in vitro fertilization. Fertil Steril 1983 ;39:594-598 Jones HW, Jr, Acosta A, Andrews MC, et al.: The importance of the follicular phase to success and failure in in vitro fertilization. Fertil Steril 1983;40:317-321 Kerin JF, Warnes GM, Quinn P, et al.: In vitro fertilization and embryo transfer program. J Vitro Fert Embryo Transfer 1984;1:63-71 Gosden RG: Maternal age: A major factor affecting the prospects and outcome of pregnancy. Ann NY Acad Sci 1984; 442:45-57 Trounson A, Sathananthan AH: The application of electron microscopy in the evaluation of two to four cell human embryos cultured in vitro for embryo transfer. J Vitro Fert Embryo Transfer 1984;1:153-165 Asch R, Balmaceda JP, Ord T, et al.: Oocyte donation and gamete intrafallopian transfer in premature ovarian failure. Fertil Steril 1988;49:263-267
The Impact of Embryo Quality on Pregnancy Outcome in Elderly Women Undergoing in Vitro Fertilization-Embryo Transfer (IVF-ET) DOV FELDBERG, 1 JACOB FARHI, 1 DOV DICKER, 1 JACOB ASHKENAZI, 1 MICHAL SHELEF, l and JACK A. GOLDMAN 1'2
Most infertility programs, especially IVF-ET units, limit the admission to women aged under 38 (1-3). Nevertheless, in many Western countries a change in birth habits is observed in the past decade or two. Furthermore, one is aware of demands for infertility treatment by women aged 40 and over, including IVF therapy (4). Edwards et al. (5,6) claim that since fertilization rates are low in this age group, while implantation processes are rare and defective, they are considered part of the unexplained infertility group, until biological factors of the aging reproductive system are more accurately defined. It is well to remember that at the present time reconstructive surgery of the fallopian tubes in patients over 40 with a severe mechanical factor is no longer considered indicated in view of low rates of success in this age group (7). Thus, the in vitro fertilization alternative must be offered as another therapeutic process, as long as gonadotropins are within normal limits. Moreover, there is no doubt that nowadays the "elderly" woman with infertility problems requires a thorough diagnostic and therapeutic workup. There is little information on the subject of patients over 40 as compared to a group of younger patients treated in an IVF-ET program (8,9). Data from our own unit (10) on the subject of ovulation induction in an elderly female population, using a specific protocol including pFSH + hMG for preparation to IVF, indicate impressive achievements in pregnancy rates; yet first-trimester abortion rates were exceptionally high in women over 40 when compared to a population of younger women treated with an identical protocol for ovulation induction. Fishel and associates (l l) also reported rather
Submitted: October 16, 1989 Accepted: May 15, 1990
Most IVF-ET units limit the procedure to women below age 38. Nevertheless, demands for infertility treatment, including I V F therapy, are more frequent nowadays. We compared 46 cycles for ovulation induction for I V F in 46 women aged40 or more (Group I) to 51 induced cycles for this procedure in younger women o f mean age 30.2 years (Group II). Cancellation rates due to early luteinization or ovulation were significantly higher in group I than in group H (28.2 and 17.6%, respectively) (P < 0.001). Also, significantly higher abortion rates were observed in older women (62.5%), in comparison to the younger control group, (25%), (P < 0.001). It is suggested that the high abortion rate considered to be due to genetic factors in older women, may possibly also be due to the aging uterine environment. Furthermore, while embryos with fragmentations may often produce clinical pregnancies in the young, the aging uterus in the elderly woman does not encourage the development o f clinical pregnancies in such embryos. Consequently the higher abortion rate in the elderly woman. KEY WORDS: in vitro fertilization; elderly women; embryo quality, pregnancy outcome.
INTRODUCTION One of the controversial issues in the management of infertility is the problem of the woman's age. Sherman Institute of Fertility, Department of ObstetricsGynecology, Golda Meir Medical Center (Hasharon Hospital), Petah-Tikva, and Tel-Aviv University Medical School, TelAviv, Israel. 2 To whom correspondence should be addressed at Department of Obstetrics-Gynecology, Golda Meir Medical Center, (Hasharon Hospital), Petah Tikva, Israel, 49372.
257
0740-7769/90/1000-0257506.00/0 9 1990PlenumPublishingCorporation
258
FELDBERG ETAL.
low "take-home baby" rates resulting from those "elderly" pregnancies as compared to a younger age group, while Johnston (1) emphasizes the occurrence of autosomal dominant diseases in offspring of these aged women, often associated with sperm defects of an aged husband. The present publication reports clinical and laboratory data of pregnancies achieved in women older than 40 years following an IVF-ET procedure, compared to a control group of younger women. We have attempted to concentrate on the possible causative factors for the large number of abortions in this group of "elderly" women. MATERIALS AND METHODS Between July and December 1987, 46 women aged over 40 years (group I) were treated in 46 cycles of ovulation induction for the purpose of IVF. Mean age was 40.6 -+ 0.5 years, with a range of 40-43 years. During the same period of time and under identical conditions, 51 patients (group II) of mean age 30.2 +-- 2.7 years, with a range of 27-33.6 years, were treated during 51 ovulation induction cycles. Clinical data of the study group and the control group are listed in Table I. The mean duration of infertility, the different indications for IVF treatment, and the number of previous cycles of therapy were identical in both groups. The ovulation induction protocol (12) consisted of the administration of 3-4 ampoules of pure follicle stimulating hormone (pFSH) (Metrodin, Serono Laboratories, Randolph, MA) daily for 3 days, starting on the third cycle day, followed by 3-4 ampoules daily of human menopausal gonadotropin Table I. Clinical Data of Study and Control Groups Data Mean duration infertility (years; M --- SD) Tubal infertility Unexplained infertility Male infertility Endometriosis Cervical factor Previous IVF cycles* 1 2 />3
*P, NS.
Group I (n = 46)
Group II (n = 51)
8.9 -+ 1.3 25 (54.3%) 13 (28.3%) 5 (10.9%) 2 (4.3%) 1 (2.2%)
7.4 --- 2.6 34 (66.7%) 10 (19.7%) 5 (9.8%) 1 (1.9%) 1 (1.9%)
21 (45.6%) 13 (28.3%) 12 (26.1%)
16 (31.3%) 21 (41.2%) 14 (27.5%)
(hMG) (Pergonal, Serono Laboratories, Randolph, MA) according to individual response in each case, for as long as necessary. Daily monitoring of the development of ovarian follicles was started on the sixth day of the menstrual cycle and included plasma E2 and progesterone levels, as well as ultrasound estimation of the size of follicles. Following the presence of at least two follicles over 16 mm in diameter, E 2 levels above 400 pg/ml, and progesterone lower than 1.5 ng/ml, human chorionic gonadotropin (hCG) (Chorigon, Teva, Cfar-Saba, Israel), 10,000 units, was given. Ovum pickup was done by means of vaginal ultrasound under paracervical anesthesia (13). The morphology of the oocytes was described according to oocytec-corona--cumulus structure of the classification by Laufer et al. (14), and that of the embryos according to their pronuclear structure, polyspermic fertilization, regularity of blastomeres, and degree of fragmentation (15). All patients were given support in the luteal phase by means of progesterone suppositories, 25 mg twice daily, and serum beta hCG was done 10 days after embryo transfer. Clinical pregnancy was defined as a pregnancy in which a sac, embryo, and fetal heart were demonstrated by ultrasound. Statistical evaluation was done by means of Student's t test and chi-square analysis.
RESULTS There was a significant difference in cancellation rates between the study group (28.2%) and the control group (17.6%) (P < 0.001). The reasons for cancellations were early ovulation or luteinization and refractory response of the ovaries to increasing levels of menotropins. No statistically significant differences were observed between the number of follicles present in response to ovulation induction, as well as between the average number of oocytes asTable II. Results of IVF in Study and Control Groups Data
Group I
Group II
No. inductions No. ovum pickups Cancellation rate* No. follicles aspirated No. oocytes No. oocytes/pickup (M -+ SD)
46 33 28.2% 132 114 3.5 --- 0.9
51 42 17.6% 186 141 3.4 --+ 1.1
* P < 0.001.
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
259
EMBRYO QUALITY AND PREGNANCY OUTCOME IN ELDERLY WOMEN
pirated, in the two groups (Table II). In no case in either group were no oocytes aspirates. No statistically significant differences were found between the morphological forms of oocyte fertilization rates or pregnancy rates in the study group of older women and those in the control group of young women (Tables III and IV). In order to define the etiological factors of abortions the morphological structure of embryos was evaluated. Findings such as pronuclear structure, polyspermic fertilizations, and the symmetry of blastomeres were compared in pregnancies and abortions. The polyspermic fertilization rates were 7.9% in the study group and 3.2% in the control group, a statistically significant difference. In all other variables no statistically significant difference was considered to be associated with the morphological structure of blastomeres and the pregnancy rates in the two groups (Table V). When the pregnancy rate achieved with the transfer of fragmented versus nonfragmented embryos in the study group was examined (Table VI), no statistically significant difference was found. Nevertheless, of all embryos transferred, only I of 9 embryos with fragmentations achieved pregnancy (11.2%), while 7 of 22 transferred embryos without fragmentation achieved pregnancy (32.7%). Identical findings were observed in the control group (Table VII). In the control group 31.2% of fragmented embryos resulted in pregnancy, as compared to
Table III. Distribution of Fertilized and Nonfertilized Oocytes* in Group I According to Their Morphology and Establishment of Pregnancy Pregnancy +, Fertilization
Pregnancy - , Fertilization
Total, Fertilization
Oocyte classification~
+
-
+
-
+
-
PSD PSF PMD PMF PLD PLF DSD DSF DMD DMF DLD DLF
4 3 1 3 2 5 2 2 0 3 0 0
2 3 3 1 1 2 2 1 7 1 1 0
3 3 6 3 6 1 1 0 6 3 3 3
3 3 4 4 1 1 2 1 3 2 0 2
7 6 7 6 8 6 3 2 6 6 3 3
4 6 7 6 2 3 4 2 10 4 1 2
Description of oocytes: P, pale; D, dark; S, small; M, medium; L, large; De, dense; F, diffuse. * P , NS.
Table IV. Distribution of Fertilized and Nonfertilized Oocytes* in Group II According to Their Morphology and Establishment of Pregnancy Pregnancy +, Fertilization
Pregnancy - , Fertilization
Total, Fertilization
Oocyte classificationa
+
-
+
-
+
-
PSDe PSF PMDe PMF PLDe PLF DSDe DSF DMDe DMF DLDe DLF
5 3 4 3 2 4 8 2 2 6 4 2
2 2 2 1 1 4 4 2 1 6 2 1
6 4 2 2 2 8 6 0 4 4 2 2
2 2 1 1 1 4 4 2 2 2 3 2
11 7 6 5 4 12 14 2 6 10 6 4
4 4 3 2 2 8 8 4 3 8 5 3
* P, NS. a Description of oocytes: P, pale; D, dark; S, small; M, medium; L, large; De, dense; F, diffuse.
29.1% in embryo transfer of nonfragmented conceptuses. DISCUSSION The outcome of 32 treatment cycles in 24 women aged 40 or more were compared to 38 cycles in 31 patients aged in their thirties. Therapy in both groups was given during the same period and by the same medical and laboratory team. No statistically significant differences were obvious as far as indications for IVF or previous therapeutic trials. The protocol for induction of ovulation was identical in both groups. Cancellation rates in the study group were significantly higher than in the control group, 28.2% as compared to 17.6%, respectively, but were acceptTable V. Fertilization and Pregnancy Rates in Study and Control Groups Data No. oocytes fertilized* No. transfers*
Group I
Group II
(n = 46)
(n = 51)
79 (68%) 31
108 (71.8)% 40
2.5 • 0.4 8
2.9 • 0.3 12
25.8% 5/8 (62.5%)
30% 3/12 (25%)
No. embryos per transfer* (M • SD) No. clinical pregnancies*
Pregnancy rates per transfer* Abortion rate**
*P, NS. ** T < 0.001.
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
260
FELDBERG ETAL.
Table VI. Pregnancy Rate in Embryos with and Without Fragmentation in the Study Group With fragmentation
Without fragmentation
+
1
-
8
7 15 22 7/22 (32.7%)
Pregnancy
Total Pregnancy rate*
9 1/9 (11.2%)
*P, NS.
able when compared to cancellations reported in the literature in IVF-treated younger population groups (16). In only three cases the cause for cancellation was refractory ovarian response, while in all other cases treatment cycles were cancelled because of early luteinization or ovulation. These findings differ from those of Kerin et al. (17), who reported poor ovarian response to induction in women aged 40 years or more. Gindoff et al. (8) nevertheless reported normal ovarian reaction to induction of ovulation with menotropins in an older age group. It is interesting to note that in the patients of the study group who reacted normally to ovarian stimulation, no difference was observed in the endocrine and follicular reaction as compared to those in the control group of younger patients. This fact is obvious in that the number of oocytes aspirated at ovum pickup, the morphologic characteristics, fertilization rates, and cell division were similar in both groups. Similar findings were reported by Romeo and associates (9), who studied a group of 29 "elderly" women, treated with IVF-ET at the Norfolk Center. In our group no significant difference was registered in the mean number of transferred embryos and in the pregnancy rates, which were similar, namely, 31.8% in the control group. The most dominant finding was the very high abortion rate in the study group of older women (62.5%) as compared to 25% in the control group. Other groups observed identical, high abortion Table VII. Pregnancy Rate in Embryos with and Without Fragmentation in the Control Group Pregnancy
With fragmentation
Without fragmentation
+ Total Pregnancy rate*
5 11 16 5/16 (31.2%)
7 17 24 7/24 (29.1%)
*P, NS.
rates in patients aged 40 or more, when treated with IVF-ET (9,18). Trounson et al. (19) attempted to find a correlation between the electron microscopic structure of embryos and the prediction of pregnancy, though without success. We compared the morphologic picture of oocytes according to the classification of Laufer et al. (14) dealing with fertilization rates in the study and control groups. From findings in Tables IV and V it is clearly evident that no correlation may be found between the morphology of oocytes resulting in pregnancy and that of oocytes that did not achieve pregnancy. In our study group polyspermic fertilizations were observed (7.8%) twice as often as in the control group (3.2%). It is possible that a hint may be found here: although oocytes in an older age group are microscopically and morphologically similar to those of the younger age group, they nevertheless demonstrate a functional defect of the zona pellucida, making possible polyspermic fertilization in a high percentage of cases. Possibly this is one cause for more frequent genetic aberrations in embryos of older women. In attempting to compare the morphology of embryos with form and symmetry of blastomeres in different phases of division, no statistically significant difference was found between the two groups. In focusing on a specific structural defect in the development of the embryo, i.e., fragmentations, we evaluated all embryos transferred with and without fragmentation in both groups of women. Although not statistically significant, a clear difference was established, as in the study group only 11.2% of fragmented embryos resulted in pregnancies, while 32.7% of nonfragmented e m b r y o s achieved clinical pregnancies. This finding was not registered in the control group, in which 31.2% of fragmented embryos, as compared to 29.1% of nonfragmented embryos, resuited in clinical pregnancies. We believe that these findings have not been reported this far. We suggest that the high rate of abortions, considered by most authors (8,9) to be due to genetic factors in older women, may possibly be explained, at least in part, by the uterine factor or environment in the uterine cavity. Undoubtedly an embryo with fragmentations possesses the potential to result in a smaller number of pregnancies than an embryo without this phenomenon. Moreover, while the young uterine environment allows the development of these " w e a k " embryos to produce clinical pregnancies, the aging
Journal of in Vitro Fertilization and Embryo Transfer, Vol. 7, No. 5, 1990
EMBRYO QUALITY AND PREGNANCY OUTCOME IN ELDERLY WOMEN
uterus in elderly women does not encourage it and a small number of pregnancies is registered in the implantation of embryos with fragmentations. Thus, the same uterine environment--endocrine as well as histological--which is "insufficient" due to advanced age, is unable to supply the requirements for the development of pregnancy as in younger age, resulting in high abortion rates. This has been found in animal experiments (18) in which it has been demonstrated that, with aging, the uterus becomes more fibrotic, the blood supply is poor, the thickness of the endometrium is reduced, and its capacity to produce prostaglandins declines. Nevertheless, the conclusion that the uterine factor is etiologically related to increased early pregnancy wastage in elderly women is currently challenged by data on embryo donation in menopausal women. Asch et al. (20) reported a high pregnancy rate in elderly women in his embryo donor program suggestive of ovarian and not uterine factor in those patients. In view of the above, it seems to us that the aging woman is entitled to IVF-ET treatment according to the usual indications, identical to those which apply to the younger patient population, with the hope of achieving the same pregnancy rates as in the latter group. Naturally, the "elderly" women should be informed of the current chances and higher abortion rates and warned of the need for chorionic villous samplings or amniocentesis in pregnancy. Further studies are necessary to evaluate if the tendency observed in our small group of patients will have statistical support in a large number of women studied.
4.
5.
6. 7.
8. 9.
10. 11.
12.
13.
14.
15.
16.
17.
REFERENCES 18. 1. Johnston WIH, Oke K, Speirs A, et al.: Patient selection for in vitro fertilization: Physical and psychological aspects. Ann NY Acad Sci 1985;442:490-503 2. Utian WH, Goldfarb JM, Sheean LA: Implementation of an in vitro fertilization program. J Vitro Fert Embryo Transfer 1984;1:72-75
19.
20.
3. Laufer N, DeCherney AH, Haseltine FP, et aL: Human in vitro fertilization employing individualized ovulation induc-
Journal o f in Vitro Fertilization a n d E m b r y o Transfer, VoL 7, N o . 5, 1990
261
tion by human menopausal gonadotropins. J Vitro Fert Embryo Transfer 1984;1:56-62 Quigley MM, Wolf DP: Human in vitro fertilization and embryo transfer at the University of Texas, Houston. J Vitro Fert Embryo Transfer 1984;1:29-33 Edwards RG, Fishel SB, Cohen J, et al.: Factors influencing the success of in vitro fertilization for alleviating human infertility. J Vitro Fert Embryo Transfer 1984;1:3-23 Edwards RG: In vitro fertilization and embryo replacement: Opening lecture. Ann NY Acad Sci 1985;442:1-22 Wood C, McMaster R, Rennie G, et al.: Factors influencing pregnancy rates following in vitro fertilization and embryo transfer. Fertil Steril 1985;43:245-250 Gindoff PR, Jewelewicz R: Reproductive potential in the older woman. Fertil Steril 1986;46:989-1001 Romeo A, Muasher SJ, Acosta AA, et al.: Results of in vitro fertilization attempts in women 40 years of age and older: The Norfolk experience. Fertil Steril 1987;47:130-136 Feldberg D, Ashkenazi J, Dicker D, et al.: IVF-ET in elderly women. Personal Communication, 1987 Fishel SB, Cohen J, Fehilly G, et al.: Factors influencing human embryonic development in vitro. Ann NY Acad Sci 1985;442:342-356 Feldberg D, Goldman JA, Shelef M, et al.: Comparison of a fixed and dynamic protocol for embryo transfer in an IVF/ ET program. Hum Reprod 1988;3:747-750 Feldberg D, Goldman JA, Ashkenazi J, et al.: Transvaginal oocyte retrieval controlled by vaginal probe for in vitro fertilization: A comparative study. J Ultrasound Med 1988;7: 339-343 Laufer N, DeCherney AH, Tarlatzis BC, et al.: The association between preovulatory serum 17-beta estradiol pattern and conception in human menopausal gonadotropin--human choronic gonadotropin stimulation. Fertil Steril 1986;46:7376 Veeck LL, Wortham JWE Jr, Witmyer J, et al.: Maturation and fertilization of morphologically immature human oocytes in a program of in vitro fertilization. Fertil Steril 1983 ;39:594-598 Jones HW, Jr, Acosta A, Andrews MC, et al.: The importance of the follicular phase to success and failure in in vitro fertilization. Fertil Steril 1983;40:317-321 Kerin JF, Warnes GM, Quinn P, et al.: In vitro fertilization and embryo transfer program. J Vitro Fert Embryo Transfer 1984;1:63-71 Gosden RG: Maternal age: A major factor affecting the prospects and outcome of pregnancy. Ann NY Acad Sci 1984; 442:45-57 Trounson A, Sathananthan AH: The application of electron microscopy in the evaluation of two to four cell human embryos cultured in vitro for embryo transfer. J Vitro Fert Embryo Transfer 1984;1:153-165 Asch R, Balmaceda JP, Ord T, et al.: Oocyte donation and gamete intrafallopian transfer in premature ovarian failure. Fertil Steril 1988;49:263-267
