1 University of Bristol Centre for Reproductive Medicine, St Michael's Hospital, Southwell Street and 2 University of Bristol Division of Child Health, Institute of Child Health, Royal Hospital for Sick Children, Bristol BS2 BEG, UK
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: competent oocytes/follicle diameter/monofollicular unstimulated cycles/serum oestradiol
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Evidence of follicular and oocyte competence was taken as normal two-pronucleate fertilization and progressive embryo cleavage (using laboratory methods as previously described) (Hull et al., 1985). As an end-point, implantation might seem to be the obvious choice to determine follicle competence; however, only a minority of embryos implant. Cycles with an unfertilized oocyte or in which an oocyte could not be collected were initially included to look for possible differences in oestradiol and FD from `fertilized' cycles, analysed on the day of onset of the LH surge. These differences were investigated using a two-tailed Student's t-test on the log-transformed data. If no oocyte was collected, this could have been due to technical difficulties encountered in the single follicle aspiration or because the `follicle' was in fact a follicular cyst. If an oocyte was collected but no embryo resulted, this was unlikely to be related to sperm dysfunction as we had controlled for that. It could have been because endogenous FSH or endogenous LH inadequately stimulated the follicle or because the oocyte recovery procedure was inappropriately timed. Therefore, for the definitive study of the normal relationship between oestradiol and FD, only data from the `fertilized' cycles were used because significant differences were found between the `fertilized`, `non-fertilized' and `no oocyte' groups.
Oestradiol was measured as a daily routine (not in batches) using a fluoroimmunoassay (DELFIA; Wallac UK Ltd., Milton Keynes, UK) as previously described (Harlow et al., 1995). The interand intra-assay coefficients of variation were 13.8 and 7.1% (at 603 pmol/l). The coefficient of variation for ultrasound FD measurement was 1.7% at 11 mm and 1.2% at 20 mm. As is usual in clinical practice, estimations of follicle size were rounded up or down to the nearest whole integer. For example, an FD of 16 mm included all measurements from 15.5 to 16.4 mm.
For statistical analysis, oestradiol values were transformed to log base 10 and the relationship to FD was investigated by a `random intercept' model which allowed for variation between subjects, between repeated cycles in the same subject (a maximum of four in this instance), and repeated measurements within cycles (maximum six). In this way, a formula was proposed for the calculation of serum oestradiol from FD as:
![]() |
In this model, u, v and e were assumed to be normally distributed variables, with means equal to 0 and variances that were estimated as part of the modelling process.
There was no significant variation in the slope (b), either across women or across cycles per woman (data not shown).
Model fitting was undertaken using SAS v6.12, PROC MIXED, (SAS Institute, Cary, NC, USA).
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
FD and oestradiol concentration were measured on the day of the onset of the LH surge in all 128 monofollicular cycles and the results are summarized in Table I. The distribution of oestradiol was skewed but normalization of the distribution by log-transformation indicated a coefficient of variation of only 5%, compared with 1013% for FD. Mean FD appeared to be smaller in the `no oocyte' group by 0.8 mm compared with the other groups but the difference was not significant. However, mean oestradiol was significantly lower in the `no oocyte' and `unfertilized' cycles compared with the `fertilized' cycles (P = 0.03 and P = 0.008 respectively). Therefore further analysis was restricted to the `fertilized' group.
|
|
|
![]() |
![]() |
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Failure to collect an oocyte or failure of an oocyte to fertilize may occur for several reasons, as discussed previously. Data from such cycles were excluded, as they were likely to represent sub-optimal follicular development. This is supported by the significantly lower serum oestradiol concentrations that were measured in these cycles.
Previous studies relating serum oestradiol to ovarian FD are based on small numbers of observations using transabdominal ultrasound scanning. A close correlation between FD and serum oestradiol concentration was shown but this was based on only 15 cycles studied using transabdominal ultrasound (Hackeloer et al., 1979). Furthermore, oestradiol estimations did not relate to specific FD. Others set arbitrary oestradiol criteria for human chorionic gonadotrophin (HCG) administration in otherwise unstimulated IVF cycles and also administered HCG if follicles showed disproportionate oestradiol:FD ratios (Paulson et al., 1990
). They proposed that higher oestradiol concentrations were required at small follicle diameters (1100 pmol/l at 15 mm) than at larger diameters (735 pmol/l at 20 mm) to justify giving HCG, prior to oocyte recovery but there are no scientific data to support this hypothesis.
In our study, also involving unstimulated IVF, the cycle was not perturbed by any hormonal treatment and oocyte collection was timed by the onset of the endogenous LH surge. The onset of the surge was precisely estimated by 4 hourly measurements. Only monofollicular cycles producing a fertilizable oocyte were used for our purpose of deriving optimal reference criteria for individual follicles. The present study is based on a large number of observations using transvaginal ultrasound scanning, the method of choice for ovarian imaging in most current gynaecological practice (Kossoff et al., 1991). The study was continued in each cycle until the day of onset of the LH surge, as after this oestradiol concentrations fall with the onset of luteinization (Muasher et al., 1990
); also, any practical predictive value would have passed. Transvaginal scanning did not begin until day 9 of the cycle, primarily to ensure the presence of a dominant follicle at the onset of monitoring (van Santbrink et al., 1995
; Fauser and Van Heusden, 1997
).
In cycles in which an oocyte could not be collected or fertilized, the peak FD was not reduced. By contrast, peak serum oestradiol concentrations were significantly reduced, suggesting an association between impaired follicular secretion of oestradiol and defective oocyte maturation. The findings suggest that serum oestradiol may be a better index of follicular functional capacity. Serum oestradiol concentrations rose exponentially with follicular enlargement suggesting progressively enhanced activity of individual granulosa cells.
Other reported attempts to predict serum oestradiol concentrations have not been helpful. Based on 21 conception cycles achieved by ovulation induction (Fink et al., 1982), follicular maturity did not correspond with oestrogen production in one-third of cases, even though a wide range of peak urinary oestrogen output (100600 µmol/24 h) was accepted as being normal. Ultrasound assessment of endometrial thickness or morphology has been advocated as an indirect index of oestrogen state for monitoring exogenous gonadotrophin-stimulated cycles (Zaidi et al., 1995
; Remohi et al., 1997
), but has not proved valuable. It is probably too insensitive for the high oestradiol concentrations expected in gonadotrophin-stimulated cycles.
In this study, we determined means and lower limits of serum oestradiol for follicles of every diameter from 10 to 24 mm and specifically on the day of the LH surge (see Tables I and II). At the onset of the LH surge, the mean FD was 21 mm and lower limit 14.5 mm, and geometric mean oestradiol 1279 pmol/l and lower limit 594 pmol/l. Those oestradiol values are consistent with the findings in Table II
for all follicles of 21 mm diameter, but the lower oestradiol limit was also consistent with the mean for follicles of 1516 mm diameter. Thus there is wide functional variation between follicles of similar size. Nevertheless, our findings provide reliable minimum and mean values of oestradiol to be expected for a follicle of any given diameter that can produce a competent oocyte, and these criteria might reasonably be applied to follicles of mixed sizes within a stimulated cohort as in IVF or ovulation induction treatment. Whether ovulation induction or down-regulation have significant effects on these relationships has yet to be tested in future studies. From these data, we know that for any FD, a particular serum oestradiol concentration can be said to be within expected limits for a competent follicle, that is, one which contains a fertilizable oocyte. However, these data cannot predict whether a follicle of a particular size is likely to contain a competent fertilizable oocyte, and can only give an indication of what might be expected from a particular follicle.
Our data from apparently competent pre-ovulatory follicles show considerable variance for FD and oestradiol, FD more so than oestradiol. It appears from these data that precise assessment of follicle maturity can only be obtained through considering both size and endocrine function. However, reference to oestradiol concentrations as an index of demonstrable follicle maturity (Table I) shows that an FD of 21 mm on average or 15 mm minimum is required. The clinical value of these data is to provide definition of what normal endocrine output is from a follicle which provides an oocyte capable of fertilization. Adjustment will need to be made for inter-laboratory differences but the assay systems used are widely available commercially, together with correction factors from assay to assay.
![]() |
Acknowledgments |
---|
![]() |
Notes |
---|
* After a short illness, Michael Hull, Professor of Reproductive Medicine and Surgery in the Division of Obstetrics and Gynaecology, University of Bristol, died on 22 November, 1999, aged 60 years. He went to Bristol from London in 1976 as Consultant Senior Lecturer in Obstetrics and Gynaecology with a developing reputation in clinical reproductive endocrinology from his time in London and elsewhere. His contributions to knowledge and practice of reproductive medicine, particularly in all aspects of male and female infertility, brought international renown to his unit. He was appointed to a Personal Chair in 1989. He brought strong ethical principles to his research and clinical practice and campaigned locally and nationally for public understanding and funding of infertility services.
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Fink, R.S., Bowes, L.P., Mackintosh, C.E. et al. (1982) The value of ultrasound for monitoring ovarian responses to gonadotrophin stimulant therapy. Br. J. Obstet. Gynaecol., 89, 856861.[ISI][Medline]
Hackeloer, B.J., Fleming, R., Robinson, H.P. et al. (1979) Correlation of ultrasonic and endocrinologic assessment of human follicular development. Am. J. Obstet. Gynecol., 135, 122128.[ISI][Medline]
Harlow, C.R., Cahill, D.J., Maile, L.A. et al. (1995) Time-dependent effects of transforming growth factor alpha on aromatase activity in human granulosa cells. Hum. Reprod., 10, 25542559.[Abstract]
Hull, M.G.R., Joyce, D.N., McLeod, F.N. et al. (1985) An economic and ethical way to introduce in-vitro fertilization to infertility practice, and findings related to post-coital sperm/mucus penetration in isolated tubal, `cervical' and unexplained infertility. Ann. N. Y. Acad. Sci., 442, 318323.[ISI][Medline]
Kossoff, G., Griffiths, K.A. and Dixon, C.E. (1991) Is the quality of transvaginal images superior to transabdominal ones under matched conditions? Ultrasound Obstet. Gynaecol., 1, 2935.[ISI][Medline]
Muasher, S.J., Kruithoff, C., Webster, S. et al. (1990) Natural cycle in-vitro fertilization: a simplified treatment method. Fertil. Steril., 53, 826827.
Paulson, R.J., Sauer, M.V., Francis, M.M. et al. (1990) In vitro fertilization in unstimulated cycles: a clinical trial using hCG for timing of follicle aspiration. Obstet. Gynecol., 52, 785791.
Remohi, J., Ardiles, G., Garcia-Velasco, J.A. et al. (1997) Endometrial thickness and serum oestradiol concentrations as predictors of outcome in oocyte donation. Hum. Reprod., 12, 22712276.[Abstract]
Testart, J., Frydman, R., Feinstein, M.C. et al. (1981) Interpretation of plasma luteinizing hormone assay for the collection of mature oocytes from women: definition of a luteinizing hormone surge initiating rise. Fertil. Steril., 36, 5054.[ISI][Medline]
van Santbrink, E.J., Hop, W.C., van Dessel, T.J. et al. (1995) Decremental follicle-stimulating hormone and dominant follicle development during the normal menstrual cycle. Fertil. Steril., 64, 3743.[ISI][Medline]
Zaidi, J., Campbell, S., Pittrof, R. and Tan, S.L. (1995) Endometrial thickness, morphology, vascular penetration and velocimetry in predicting implantation in an in vitro fertilization program. Ultrasound Obstet. Gynecol., 6, 191198.[ISI][Medline]
Submitted on February 24, 2000; accepted on May 22, 2000.