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Polyovular follicles associated with human in vitro fertilization*
FERTILITY AND STERILITY
Vol. 49, No. 3, March 1988
Copyright o 1988 The American Fertility Society
Printed in U.S.A.
Polyovular follicles associated with human in vitro fertilization*
Pramila V. Dandekar, M.Sc.t Mary C. Martin, M.D. Robert H. Glass, M.D. Department of Obstetrics, Gynecology and Reproductive Sciences, University of California, San Francisco, San Francisco, California
Polyovular follicles were found in 61 (24%) of 251laparoscopies performed for in vitro fertilization. Of 898 follicles that contained at least one oocyte, 76 (8%) were polyovular. When the oocytes from polyovular follicles were assessed by morphologic criteria, it was found that 46 of the follicles contained oocytes that were discordant in maturity. Thus, oocytes can develop at different rates even though they are exposed to the same follicular fluid. It is possible that the microenvironment provided by the cellular investments of oocytes may correlate more precisely with oocyte maturity. Fertil Steril 49:483, 1988
Polyovular follicles have been found in the ovaries of a number of mammalian species, including the human. 1 Sherrer et al., 2 examined histologic specimens of human ovaries and demonstrated that polyovular follicles occur more frequently early in life. In women under the age of 20, the prevalence of "polynuclear" follicles was 58%, whereas similar follicles were found in 13% of women aged 20 to 39, and there were none in the ovaries of women 40 years or older. It has been suggested that gonadotropins can increase the number of recruited polyovular follicles,3·4 and Jones et al. 5 retrieved more than one oocyte from 8% of the oocyte-containing follicles aspirated from women undergoing ovarian stimulation with human menopausal gonadotropins (hMG) for in vitro fertilization (IVF). Received February 16, 1987; revised and accepted October 27, 1987. * Presented in part at the Forty-Second Annual Meeting of The American Fertility Society and the Eighteenth Annual Meeting of The Canadian Fertility and Andrology Society, September 27 to October 2, 1986, Toronto, Ontario, Canada. t Reprint requests: Pramila V. Dandekar, M.Sc., Department of Obstetrics, Gynecology and Reproductive Sciences, M1480, University of California, San Francisco, San Francisco, California 94143. Vol. 49, No.3, March 1988
In our IVF program, 8% of oocyte-containing follicles were polyovular in patients stimulated with combined clomiphene citrate (CC)/hMG regimens.6 Moreover, we noted that multiple oocytes retrieved from the same follicle were often at different stages of maturity, a finding that raises questions concerning the effects of the follicular microenvironment on oocyte development. MATERIALS AND METHODS
Between June 1983 and June 1986, 251laparoscopies were performed for IVF following stimulation with CC and hMG (Pergonal, Serono Laboratories, Inc., Randoph, MA). The most common protocol was 50 mg/day CC for 5 days starting on either day 3 or 5 of the cycle, and 1 ampule hMG (75 IU follicle-stimulating hormone [FSH], 75 IU luteinizing hormone [LH]) for 5 days, starting the same day as CC. The hMG was continued for an additional! to 5 days at a dose of 1 to 3 ampules per day. Ultrasound monitoring and determinations of serum estradiol (E 2 ) levels were performed daily starting on the last day of CC administration. Human chorionic gonadotropin (hCG) 5000 IU was given the evening of the day when the ultrasound showed a lead follicle of at least 16 mm in diameter Dandekar et al.
Polyovular follicles
483
and at least two other follicles 14 mm or greater in diameter, and the E 2 level was >600 pg/ml. Laparoscopy was performed approximately 35 hours after the hCG injection. Follicles were aspirated with a 12-gauge, 23-cm long, teflon-lined needle connected to a wall suction set at 120 mm Hg. Every effort was made to aspirate follicles individually. If the surgeon believed that two follicles might have been entered with the puncture, the retrieval was not listed as polyovular, even if two or more oocytes were retrieved. The volume of fluid removed from each follicle was measured. Our procedures for oocyte culture, sperm preparation and insemination, and embryo culture have been described previously. 7 The maturity of recovered oocytes was assessed immediately following retrieval and again 5 to 6 hours later under a dissecting microscope using the following criteria: (1) mature, well-dispersed cumulus and corona layers; (2) intermediate, dispersed cumulus but a tight corona; and (3) immature, tight, and compact corona and cumulus cell layers. Mature oocytes were inseminated 5 to 6 hours, and immature oocytes 24 to 33 hours after recovery. Intermediate oocytes were inseminated between 7 and 13 hours following retrieval. The presence of two pronuclei and two polar bodies approximately 18 hours after insemination was accepted as evidence of fertilization. Embryos were transferred to the uterus 48 to 54 hours after laparoscopy. Embryo development at this time ranged from the pronuclear to the 6-cell stage, depending on when the oocytes had been inseminated and the intrinsic developmental potential of the embryos.
RESULTS Polyovular follicles were found in 61 (24%) of the 251 laparoscopies. Of the 898 aspirated follicles Table 1 Fertilization of Concordant Oocytes from Polyovular Follicles Follicles(%) with both, one, or no oocytes fertilized Maturity Mature• Intermediateb Immature•
No. of follicles
Both
7 15 5
40 0
71
One
None
29 47 60
0 13 40
• Mature, well-dispersed cumulus and corona layers. b Intermediate, dispersed cumulus but a tight corona layer. • Immature, tight and compact corona and cumulus cell layers. 484
Dandekar et al. Polyovular follicles
Table 2 Fertilization of Discordant Oocytes from Polyovular Follicles Follicles(%) with both, one, or no oocytes fertilized Maturity Mature• Intermediateb Mature Immature• Intermediate Immature
No. of follicles
Both
One
None
29
62
31d
7
9
33
56•
11
8
25
50'
25
• Mature, well-dispersed cumulus and corona layers. b Intermediate, dispersed cumulus but a tight corona layer. • Immature, tight and compact corona and cumulus cell, layers. d 78% of the fertilized oocytes were mature. • 100% of the fertilized oocytes were mature. I 100% of the fertilized oocytes were intermediate.
that contained an oocyte, 76 (8%) were polyovular. There was more than one follicle with two oocytes in 11 laparoscopies, and, in five cases, there were follicles that contained three or four oocytes. Three polyovular follicles contained both a viable oocyte and a degenerated oocyte. Of the remaining 73 polyovular follicles, 27 (37%) had oocytes that were concordant in terms of maturity (Table 1), whereas 46 (63%) had oocytes that were discordant (Table 2). Despite having more than one oocyte, 26% of the polyovular follicles contained 2 ml or less of follicular fluid. This was similar to the volume in 38% of uniovular follicles aspirated from the same women (Table 3). Twenty-eight percent of polyovular follicles contained more than 5 ml of follicular fluid, compared with 19% of uniovular follicles (Table 3). These differences are not statistically significant. The percentage of oocytes fertilized from polyovular follicles was similar to that seen with oocytes of comparable maturity from uniovular follicles (Table 4). Not surprisingly, fertilization of mature oocytes was superior to that of intermeTable 3 Follicular Fluid Volume Distribution in Polyovular and Uniovular Follicles Volume (ml)
Polyovular Uniovular
No. of follicles
<2 %
%
%
76 164
26" 38
46 43b
28 19b
2-5
>5
• Not significantly different from uniovular follicles. b Not significantly different from polyovular follicles. Fertility and Sterility
diate and immature oocytes in both categories of follicles. Five women had polyovular follicles in more than one cycle. Interestingly, all the repeat polyovula,r follicles were in the same ovary that had produced the polyovular follicle in the initial cycle. Of the 55 women who had polyovular follicles, 15 were 20 to 29 years of age, 38 were 30 to 40, and 2 were over 40. The percentage of our total population having polyovular follicles in each age group was 47% for ages 20 to 29, 22% for ages 30 to 40, and 9% for over 40. Eleven pregnancies occurred following the 61laparoscopies. Seven patients have delivered, 1 had an unexplained intrauterine demise at 6 months, 1 had an ectopic pregnancy, and 2 had spontaneous abortions. One successful singleton pregnancy followed a transfer of only two embryos, both derived from the same follicle. The pregnancy rate in women with polyovular follicles was 19% per transfer, whereas it was 15% in women with only uniovular follicles. DISCUSSION
Despite the surgeon's perception that only one follicle was entered, there is a possibility that aspirates containing two or more oocytes originated from more than one follicle. Supporting the view that we were dealing with polyovular follicles are the morphologic studies of Sherrer et al., 2 in which 13% of women in the reproductive age group had polyovular follicles. In addition, our prevalence of polyovular follicles (8%) is the same as reported by the group from Norfolk. 5 •8 Moreover, the distribution of volumes of follicular fluid from polyovular follicles in our study was not significantly different from that obtained from follicles containing one oocyte (Table 3). Twenty-six percent of polyovular follicles contained only 2 ml or less of follicular fluid. The distribution of cases with polyovular follicles shows a decreasing incidence with age, which Table 4 Fertilization of Oocytes from Uniovular and Polyovular Follicles from the Same Patients Polyovular
Uniovular Maturity
Total
% Fertilized
Total
% Fertilized
Maturea Intermediateb Immature•
62 68 34
84 74 21
54 69 34
89 67 24
Mature, well-dispersed cumulus and corona layers. Intermediate, dispersed cumulus but tight corona. • Immature, tight and compact corona and cumulus layers.
a
b
Vol. 49, No.3, March 1988
correlates well with the data obtained by Sherrer et al. 2 This suggests that these follicles might be more easily recruited in the younger woman, with or without exogenous stimulation. Approximately two thirds of the polyovular follicles contained oocytes that were judged by morphologic criteria to be discordant in terms of maturity. The criterion used to judge maturity (i.e., assessment of the dispersion of the cumulus and corona cell layers) correlates well with nuclear events in immature eggs. However, with mature and intermediate oocytes, there can be a discrepancy between the morphologic categorization and thecategorization obtained by noting germinal vesicle breakdown and extrusion of the first polar body. 9 The discrepancy becomes minor if polar body formation is evaluated just before sperm insemination. A number of studies have correlated oocyte maturity with bioactive LH, FSH, prolactin, or progesterone in follicular fluid. 10 •11 The overlap in values seen in these studies and our demonstration that oocytes from the same follicle can differ in maturity suggest that these are not exact relationships. It is possible that the microenvironment provided by the cellular investments of the oocyte, as opposed to follicular fluid content, might correlate more precisely with oocyte maturity. 12•13 Polan et aP 2 demonstrated an association between decreasing aromatase activity of granulosa cells and increasing oocyte maturity. The differing maturities of oocytes surrounded by the same follicular fluid also could be explained by the position of the oocyte in the follicle. In the rabbit, if an oocyte of a polyovular follicle is located near the antrum, it develops at the same rate as oocytes from uniovular follicles. However, if the oocyte is located in the mural granulosa, there is retarded development. 14 Unfortunately, the techniques involved in aspirating human follicles for IVF do not allow a determination of the position of the oocyte within the follicle. All of our patients were stimulated with hMG and CC and they had higher levels of serum E 2 than those found in unstimulated cycles. Treatment of monkeys with diethylstilbestrol (DES) was reported to stimulate the formation of polyovular follicles. 16 Unfortunately, the ages of the monkeys were unknown. If monkeys in the control group were older than those in the DES group, the age factor alone might account for the disparity in polyovular follicles. J ones4 suggested that exogenous gonadotropins in women, perhaps by raising Dandekar et al.
Polyovular follicles
485
estrogen levels, could induce the inclusion of more than one oocyte in a follicle. The mechanism is uncertain, but it has been hypothesized that high hormone levels could stimulate release of hydrolytic and proteolytic enzymes from cytoplasmic processes of interfollicular stromal fibroblasts. 3 Fusion of adjacent follicles then could occur following degradation of intervening connective tissue and depolymerization of the ground substance. Alternatively, hMG may be more successful in recruiting polyovular follicles to develop, so that the relative proportion that progress to maturity instead of regressing may be increased. Simonetti et al., 8 found a slight increase in degenerated oocytes (23.6%) from polyovular follicles compared with oocytes from uniovular follicles (20.3%). Our prevalence of degenerated oocytes in polyovular follicles was only 2%, and this difference could reflect their use of hMG or hMG/FSH protocols, which provide greater numbers of oocytes compared with our hMG/CC protocols, but also yield a greater percentage of degenerated oocytes.5 Both onr group and that of Simonetti et al. 8 found no difference in the fertilizability of oocytes from polyovular follicles compared with those from uniovular follicles when the maturities were comparable. Moreover, in our series, the attainment of a pregnancy following transfer of embryos derived only from a polyovular follicle suggests that these embryos are not at a reproductive disadvantage. REFERENCES 1. Hartman CG: Polynuclear ova and polyovular follicles in the opossum and other mammals with special reference to the problem of fecundity. Am J Anat 37:1, 1926 2. Sherrer CW, Gerson B, Woodruff JD: The incidence and significance of polynuclear follicles. Am J Obstet Gynecol 128:6, 1977 3. Papadaki L: Binovular follicles in the adult human ovary. Fertil Steril 29:342, 1978 4. Jones GS: Induction of ovulation. Ann Rev Med 19:351, 1968
486
Dandekar et al. Polyovular follicles
5. Jones HW Jr, Acosta AA, Andrews MC, Garcia JE, Jones GS, Mayer J, McDowell JS, Rosenwaks Z, Sandow BA, Veeck LL, Wilkes CA: Three years of in vitro fertilization at Norfolk. Fertil Steril42:826, 1984 6. Dandekar P, Martin M, Andreyko J, Marshall L, Taylor R, Dumesic D, Monroe S, Glass R: Polyovular follicles in IVF patients (Abstr 149P). Presented at the forty-second annual meeting of The American Fertility Society and the eighteenth annual meeting of The Canadian Fertility and Andrology Society, Toronto, Ontario, Canada, September 27 to October 2, 1986. Birmingham, The American Fertility Society, 1986, p 53 7. Dandekar PV, Quigley MM: Laboratory setup for human in vitro fertilization. Fertil Steril 42:1, 1984 8. Simonetti S, Chillik CF, Veeck LL, Jones HW: Incidence and outcome of follicles having more than one oocyte in IVF (Abstr 161P). Presented at the forty-second annual meeting of The American Fertility Society and the eighteenth annual meeting of The Canadian Fertility and Andrology Society, Toronto, Ontario, Canada, September 27 to October 2, 1986. Birmingham, The American Fertility Society, 1986, p 57 9. Laufer N, Tarlatzis BC, DeCherney AH, Master JT, Haseltine FP, MacLusky N, Naftolin F: Asynchrony between human cumulus-corona cell complex and oocyte maturation after human menopausal gonadotropin treatment for in vitro fertilization. Fertil Steril 42:366, 1984 10. Laufer N, Botero-Ruiz W, DeCherney AH, Haseltine F, Polan ML, Behrman HR: Gonadotropin and prolactin levels in follicular fluid of human ova successfully fertilized in vitro. J Clin Endocrinol Metab 58:430, 1984 11. Cha K-Y, Barnes RB, Marrs RP, Lobo RA: Correlation of the bioactivity of luteinizing hormone in follicular fluid with oocyte maturity in the spontaneous cycle. Fertil Steril 45:338, 1986 12. Polan ML, Laufer N, Ohkawa R, Botero-Ruiz W, Haseltine FP, DeCherney AH, Behrman HR: The association between granulosa cell aromatase activity and oocyte-corona-cumulus-complex maturity from individual human follicles. J Clin Endocrinol Metab 59:170, 1984 13. Laufer N, DeCherney AH, Haseltine FP, Behrman HR: Steroid secretion by the human egg-corona-cumulus complex in culture. J Clin Endocrinol Metab 58:1153, 1984 14. Bomsel-Helmreich 0: The preovulatory human oocyte and its microenvironment. In Fertilization ofthe Human Egg In Vitro, Edited by HM Beier, HR Lindner. Berlin, SpringerVerlag, 1983, p 19 15. Graham CE, Bradley CF: Polyovular follicles in squirrel monkeys after prolonged diethylstilboestrol treatment. J Reprod Fertil 27:181, 1971
Fertility and Sterility
Vol. 49, No. 3, March 1988
Copyright o 1988 The American Fertility Society
Printed in U.S.A.
Polyovular follicles associated with human in vitro fertilization*
Pramila V. Dandekar, M.Sc.t Mary C. Martin, M.D. Robert H. Glass, M.D. Department of Obstetrics, Gynecology and Reproductive Sciences, University of California, San Francisco, San Francisco, California
Polyovular follicles were found in 61 (24%) of 251laparoscopies performed for in vitro fertilization. Of 898 follicles that contained at least one oocyte, 76 (8%) were polyovular. When the oocytes from polyovular follicles were assessed by morphologic criteria, it was found that 46 of the follicles contained oocytes that were discordant in maturity. Thus, oocytes can develop at different rates even though they are exposed to the same follicular fluid. It is possible that the microenvironment provided by the cellular investments of oocytes may correlate more precisely with oocyte maturity. Fertil Steril 49:483, 1988
Polyovular follicles have been found in the ovaries of a number of mammalian species, including the human. 1 Sherrer et al., 2 examined histologic specimens of human ovaries and demonstrated that polyovular follicles occur more frequently early in life. In women under the age of 20, the prevalence of "polynuclear" follicles was 58%, whereas similar follicles were found in 13% of women aged 20 to 39, and there were none in the ovaries of women 40 years or older. It has been suggested that gonadotropins can increase the number of recruited polyovular follicles,3·4 and Jones et al. 5 retrieved more than one oocyte from 8% of the oocyte-containing follicles aspirated from women undergoing ovarian stimulation with human menopausal gonadotropins (hMG) for in vitro fertilization (IVF). Received February 16, 1987; revised and accepted October 27, 1987. * Presented in part at the Forty-Second Annual Meeting of The American Fertility Society and the Eighteenth Annual Meeting of The Canadian Fertility and Andrology Society, September 27 to October 2, 1986, Toronto, Ontario, Canada. t Reprint requests: Pramila V. Dandekar, M.Sc., Department of Obstetrics, Gynecology and Reproductive Sciences, M1480, University of California, San Francisco, San Francisco, California 94143. Vol. 49, No.3, March 1988
In our IVF program, 8% of oocyte-containing follicles were polyovular in patients stimulated with combined clomiphene citrate (CC)/hMG regimens.6 Moreover, we noted that multiple oocytes retrieved from the same follicle were often at different stages of maturity, a finding that raises questions concerning the effects of the follicular microenvironment on oocyte development. MATERIALS AND METHODS
Between June 1983 and June 1986, 251laparoscopies were performed for IVF following stimulation with CC and hMG (Pergonal, Serono Laboratories, Inc., Randoph, MA). The most common protocol was 50 mg/day CC for 5 days starting on either day 3 or 5 of the cycle, and 1 ampule hMG (75 IU follicle-stimulating hormone [FSH], 75 IU luteinizing hormone [LH]) for 5 days, starting the same day as CC. The hMG was continued for an additional! to 5 days at a dose of 1 to 3 ampules per day. Ultrasound monitoring and determinations of serum estradiol (E 2 ) levels were performed daily starting on the last day of CC administration. Human chorionic gonadotropin (hCG) 5000 IU was given the evening of the day when the ultrasound showed a lead follicle of at least 16 mm in diameter Dandekar et al.
Polyovular follicles
483
and at least two other follicles 14 mm or greater in diameter, and the E 2 level was >600 pg/ml. Laparoscopy was performed approximately 35 hours after the hCG injection. Follicles were aspirated with a 12-gauge, 23-cm long, teflon-lined needle connected to a wall suction set at 120 mm Hg. Every effort was made to aspirate follicles individually. If the surgeon believed that two follicles might have been entered with the puncture, the retrieval was not listed as polyovular, even if two or more oocytes were retrieved. The volume of fluid removed from each follicle was measured. Our procedures for oocyte culture, sperm preparation and insemination, and embryo culture have been described previously. 7 The maturity of recovered oocytes was assessed immediately following retrieval and again 5 to 6 hours later under a dissecting microscope using the following criteria: (1) mature, well-dispersed cumulus and corona layers; (2) intermediate, dispersed cumulus but a tight corona; and (3) immature, tight, and compact corona and cumulus cell layers. Mature oocytes were inseminated 5 to 6 hours, and immature oocytes 24 to 33 hours after recovery. Intermediate oocytes were inseminated between 7 and 13 hours following retrieval. The presence of two pronuclei and two polar bodies approximately 18 hours after insemination was accepted as evidence of fertilization. Embryos were transferred to the uterus 48 to 54 hours after laparoscopy. Embryo development at this time ranged from the pronuclear to the 6-cell stage, depending on when the oocytes had been inseminated and the intrinsic developmental potential of the embryos.
RESULTS Polyovular follicles were found in 61 (24%) of the 251 laparoscopies. Of the 898 aspirated follicles Table 1 Fertilization of Concordant Oocytes from Polyovular Follicles Follicles(%) with both, one, or no oocytes fertilized Maturity Mature• Intermediateb Immature•
No. of follicles
Both
7 15 5
40 0
71
One
None
29 47 60
0 13 40
• Mature, well-dispersed cumulus and corona layers. b Intermediate, dispersed cumulus but a tight corona layer. • Immature, tight and compact corona and cumulus cell layers. 484
Dandekar et al. Polyovular follicles
Table 2 Fertilization of Discordant Oocytes from Polyovular Follicles Follicles(%) with both, one, or no oocytes fertilized Maturity Mature• Intermediateb Mature Immature• Intermediate Immature
No. of follicles
Both
One
None
29
62
31d
7
9
33
56•
11
8
25
50'
25
• Mature, well-dispersed cumulus and corona layers. b Intermediate, dispersed cumulus but a tight corona layer. • Immature, tight and compact corona and cumulus cell, layers. d 78% of the fertilized oocytes were mature. • 100% of the fertilized oocytes were mature. I 100% of the fertilized oocytes were intermediate.
that contained an oocyte, 76 (8%) were polyovular. There was more than one follicle with two oocytes in 11 laparoscopies, and, in five cases, there were follicles that contained three or four oocytes. Three polyovular follicles contained both a viable oocyte and a degenerated oocyte. Of the remaining 73 polyovular follicles, 27 (37%) had oocytes that were concordant in terms of maturity (Table 1), whereas 46 (63%) had oocytes that were discordant (Table 2). Despite having more than one oocyte, 26% of the polyovular follicles contained 2 ml or less of follicular fluid. This was similar to the volume in 38% of uniovular follicles aspirated from the same women (Table 3). Twenty-eight percent of polyovular follicles contained more than 5 ml of follicular fluid, compared with 19% of uniovular follicles (Table 3). These differences are not statistically significant. The percentage of oocytes fertilized from polyovular follicles was similar to that seen with oocytes of comparable maturity from uniovular follicles (Table 4). Not surprisingly, fertilization of mature oocytes was superior to that of intermeTable 3 Follicular Fluid Volume Distribution in Polyovular and Uniovular Follicles Volume (ml)
Polyovular Uniovular
No. of follicles
<2 %
%
%
76 164
26" 38
46 43b
28 19b
2-5
>5
• Not significantly different from uniovular follicles. b Not significantly different from polyovular follicles. Fertility and Sterility
diate and immature oocytes in both categories of follicles. Five women had polyovular follicles in more than one cycle. Interestingly, all the repeat polyovula,r follicles were in the same ovary that had produced the polyovular follicle in the initial cycle. Of the 55 women who had polyovular follicles, 15 were 20 to 29 years of age, 38 were 30 to 40, and 2 were over 40. The percentage of our total population having polyovular follicles in each age group was 47% for ages 20 to 29, 22% for ages 30 to 40, and 9% for over 40. Eleven pregnancies occurred following the 61laparoscopies. Seven patients have delivered, 1 had an unexplained intrauterine demise at 6 months, 1 had an ectopic pregnancy, and 2 had spontaneous abortions. One successful singleton pregnancy followed a transfer of only two embryos, both derived from the same follicle. The pregnancy rate in women with polyovular follicles was 19% per transfer, whereas it was 15% in women with only uniovular follicles. DISCUSSION
Despite the surgeon's perception that only one follicle was entered, there is a possibility that aspirates containing two or more oocytes originated from more than one follicle. Supporting the view that we were dealing with polyovular follicles are the morphologic studies of Sherrer et al., 2 in which 13% of women in the reproductive age group had polyovular follicles. In addition, our prevalence of polyovular follicles (8%) is the same as reported by the group from Norfolk. 5 •8 Moreover, the distribution of volumes of follicular fluid from polyovular follicles in our study was not significantly different from that obtained from follicles containing one oocyte (Table 3). Twenty-six percent of polyovular follicles contained only 2 ml or less of follicular fluid. The distribution of cases with polyovular follicles shows a decreasing incidence with age, which Table 4 Fertilization of Oocytes from Uniovular and Polyovular Follicles from the Same Patients Polyovular
Uniovular Maturity
Total
% Fertilized
Total
% Fertilized
Maturea Intermediateb Immature•
62 68 34
84 74 21
54 69 34
89 67 24
Mature, well-dispersed cumulus and corona layers. Intermediate, dispersed cumulus but tight corona. • Immature, tight and compact corona and cumulus layers.
a
b
Vol. 49, No.3, March 1988
correlates well with the data obtained by Sherrer et al. 2 This suggests that these follicles might be more easily recruited in the younger woman, with or without exogenous stimulation. Approximately two thirds of the polyovular follicles contained oocytes that were judged by morphologic criteria to be discordant in terms of maturity. The criterion used to judge maturity (i.e., assessment of the dispersion of the cumulus and corona cell layers) correlates well with nuclear events in immature eggs. However, with mature and intermediate oocytes, there can be a discrepancy between the morphologic categorization and thecategorization obtained by noting germinal vesicle breakdown and extrusion of the first polar body. 9 The discrepancy becomes minor if polar body formation is evaluated just before sperm insemination. A number of studies have correlated oocyte maturity with bioactive LH, FSH, prolactin, or progesterone in follicular fluid. 10 •11 The overlap in values seen in these studies and our demonstration that oocytes from the same follicle can differ in maturity suggest that these are not exact relationships. It is possible that the microenvironment provided by the cellular investments of the oocyte, as opposed to follicular fluid content, might correlate more precisely with oocyte maturity. 12•13 Polan et aP 2 demonstrated an association between decreasing aromatase activity of granulosa cells and increasing oocyte maturity. The differing maturities of oocytes surrounded by the same follicular fluid also could be explained by the position of the oocyte in the follicle. In the rabbit, if an oocyte of a polyovular follicle is located near the antrum, it develops at the same rate as oocytes from uniovular follicles. However, if the oocyte is located in the mural granulosa, there is retarded development. 14 Unfortunately, the techniques involved in aspirating human follicles for IVF do not allow a determination of the position of the oocyte within the follicle. All of our patients were stimulated with hMG and CC and they had higher levels of serum E 2 than those found in unstimulated cycles. Treatment of monkeys with diethylstilbestrol (DES) was reported to stimulate the formation of polyovular follicles. 16 Unfortunately, the ages of the monkeys were unknown. If monkeys in the control group were older than those in the DES group, the age factor alone might account for the disparity in polyovular follicles. J ones4 suggested that exogenous gonadotropins in women, perhaps by raising Dandekar et al.
Polyovular follicles
485
estrogen levels, could induce the inclusion of more than one oocyte in a follicle. The mechanism is uncertain, but it has been hypothesized that high hormone levels could stimulate release of hydrolytic and proteolytic enzymes from cytoplasmic processes of interfollicular stromal fibroblasts. 3 Fusion of adjacent follicles then could occur following degradation of intervening connective tissue and depolymerization of the ground substance. Alternatively, hMG may be more successful in recruiting polyovular follicles to develop, so that the relative proportion that progress to maturity instead of regressing may be increased. Simonetti et al., 8 found a slight increase in degenerated oocytes (23.6%) from polyovular follicles compared with oocytes from uniovular follicles (20.3%). Our prevalence of degenerated oocytes in polyovular follicles was only 2%, and this difference could reflect their use of hMG or hMG/FSH protocols, which provide greater numbers of oocytes compared with our hMG/CC protocols, but also yield a greater percentage of degenerated oocytes.5 Both onr group and that of Simonetti et al. 8 found no difference in the fertilizability of oocytes from polyovular follicles compared with those from uniovular follicles when the maturities were comparable. Moreover, in our series, the attainment of a pregnancy following transfer of embryos derived only from a polyovular follicle suggests that these embryos are not at a reproductive disadvantage. REFERENCES 1. Hartman CG: Polynuclear ova and polyovular follicles in the opossum and other mammals with special reference to the problem of fecundity. Am J Anat 37:1, 1926 2. Sherrer CW, Gerson B, Woodruff JD: The incidence and significance of polynuclear follicles. Am J Obstet Gynecol 128:6, 1977 3. Papadaki L: Binovular follicles in the adult human ovary. Fertil Steril 29:342, 1978 4. Jones GS: Induction of ovulation. Ann Rev Med 19:351, 1968
486
Dandekar et al. Polyovular follicles
5. Jones HW Jr, Acosta AA, Andrews MC, Garcia JE, Jones GS, Mayer J, McDowell JS, Rosenwaks Z, Sandow BA, Veeck LL, Wilkes CA: Three years of in vitro fertilization at Norfolk. Fertil Steril42:826, 1984 6. Dandekar P, Martin M, Andreyko J, Marshall L, Taylor R, Dumesic D, Monroe S, Glass R: Polyovular follicles in IVF patients (Abstr 149P). Presented at the forty-second annual meeting of The American Fertility Society and the eighteenth annual meeting of The Canadian Fertility and Andrology Society, Toronto, Ontario, Canada, September 27 to October 2, 1986. Birmingham, The American Fertility Society, 1986, p 53 7. Dandekar PV, Quigley MM: Laboratory setup for human in vitro fertilization. Fertil Steril 42:1, 1984 8. Simonetti S, Chillik CF, Veeck LL, Jones HW: Incidence and outcome of follicles having more than one oocyte in IVF (Abstr 161P). Presented at the forty-second annual meeting of The American Fertility Society and the eighteenth annual meeting of The Canadian Fertility and Andrology Society, Toronto, Ontario, Canada, September 27 to October 2, 1986. Birmingham, The American Fertility Society, 1986, p 57 9. Laufer N, Tarlatzis BC, DeCherney AH, Master JT, Haseltine FP, MacLusky N, Naftolin F: Asynchrony between human cumulus-corona cell complex and oocyte maturation after human menopausal gonadotropin treatment for in vitro fertilization. Fertil Steril 42:366, 1984 10. Laufer N, Botero-Ruiz W, DeCherney AH, Haseltine F, Polan ML, Behrman HR: Gonadotropin and prolactin levels in follicular fluid of human ova successfully fertilized in vitro. J Clin Endocrinol Metab 58:430, 1984 11. Cha K-Y, Barnes RB, Marrs RP, Lobo RA: Correlation of the bioactivity of luteinizing hormone in follicular fluid with oocyte maturity in the spontaneous cycle. Fertil Steril 45:338, 1986 12. Polan ML, Laufer N, Ohkawa R, Botero-Ruiz W, Haseltine FP, DeCherney AH, Behrman HR: The association between granulosa cell aromatase activity and oocyte-corona-cumulus-complex maturity from individual human follicles. J Clin Endocrinol Metab 59:170, 1984 13. Laufer N, DeCherney AH, Haseltine FP, Behrman HR: Steroid secretion by the human egg-corona-cumulus complex in culture. J Clin Endocrinol Metab 58:1153, 1984 14. Bomsel-Helmreich 0: The preovulatory human oocyte and its microenvironment. In Fertilization ofthe Human Egg In Vitro, Edited by HM Beier, HR Lindner. Berlin, SpringerVerlag, 1983, p 19 15. Graham CE, Bradley CF: Polyovular follicles in squirrel monkeys after prolonged diethylstilboestrol treatment. J Reprod Fertil 27:181, 1971
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