1 Reproductive Medicine Unit, the Queen Elizabeth Hospital, University of Adelaide, 28 Woodville Road, Woodville, South Australia 5011, Australia, 2 Oxford School of Biological and Molecular Sciences, Oxford Brookes University, Oxford, UK and 3 Prince Henry's Institute of Medical Research, PO Box 5152, Clayton, Victoria 3168, Australia
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Abstract |
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Key words: activin/follistatin/inhibin/PCOS
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Introduction |
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In a recent publication following extensive recruitment of siblings with hyperandrogenaemia compatible with PCOS, a proposal was made that there is a linkage of PCOS to the follistatin gene (Urbanek et al., 1999). Follistatin is a well-known intra-ovarian regulator which binds activin, a protein also found in the ovary and partly responsible for follicular growth (Findlay, 1993
; Schneyer et al., 1996
; Hillier, 1999
). Follistatin also binds other members of the transforming growth factor ß (TGFß) family, including bone morphogenetic proteins (Iemura et al., 1998
). Mice with gene-targeted hyperexpression of the follistatin gene have impaired follicular development (Guo et al., 1998
). Overexpression of follistatin has been associated with the arrest of follicular development, an increase in ovarian androgen development and a decrease in serum FSH concentrationsall cardinal features of PCOS. Follistatin concentrations increase in the early follicular phase, then fall and then have a short surge mid-cycle. It has been noted that in patients with PCOS there is an absence of this mid-cycle surge (Lockwood et al., 1998
).
The human ovary expresses mRNA for all three subunits of activin and inhibin (, ßA, ßB) and follistatin (Burns et al., 1990
; Schwall et al., 1990
; Eramaa et al., 1993
; Roberts et al., 1993
) as well as the proteins for activin and inhibin subunits and follistatin. These proteins are detectable in the circulation with cyclic, specific variations for inhibin and activin A and can be measured in follicular fluid from small and large follicles for all three molecules (Schneyer et al., 1996
; Lambert-Messerlian et al., 1997
; Magoffin and Jakimiuk, 1997
, 1998
; Lau et al., 1999
). Activin A is produced by follicles in culture (Smitz et al., 1998
) and has been proposed to promote proliferation of granulosa cells, enhance FSH receptor expression, decrease LH-induced androgen production, increase pituitary FSH secretion and generally promote follicle growth (Li et al., 1995
; Silva and Knight, 1998
; Smitz et al., 1998
). Follistatin is a binding protein for activin and inhibits most of these actions of activin (Hillier and Miro, 1993
; de Winter et al., 1996
; Hillier, 1999
). Both proteins have effects on oocyte maturity and development, with activin promoting post-fertilization development and follistatin inhibiting this process. In mice where activin receptors have been inactivated by recombinant technology, FSH concentrations are low and animals are infertile. Follistatin knockout animals die early in life (Matzuk et al., 1995
) while overexpressing animals are infertile, have small ovaries and exhibit a block in folliculogenesis between primary and secondary follicles (Guo et al., 1998
).
Data relating to follistatin, activin A and inhibin B concentrations in PCOS are scanty. In this study we show that circulating follistatin concentrations are increased and activin A concentrations decreased in subjects with PCOS while inhibin B values did not alter.
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Materials and methods |
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Statistical analysis was performed after assessing the distribution of the data. Parametric comparisons were performed using Student's t-test; non-parametric comparisons utilized the Mann-Whitney test and Spearman's test of correlation. P < 0.05 was considered significant.
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Results |
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Discussion |
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Several studies have examined the patterns of inhibin and activin in follicular fluid from subjects with PCOS. Erickson et al. were the first to report that there was no difference in the follicular fluid concentrations of follistatin between normal and PCOS follicles (Erickson et al., 1995). They also observed no difference between atretic and healthy follicles. Examination of follicular fluid from small ovaries (Magoffin and Jakimiuk, 1998
) found no differences in the activin A and follistatin concentrations between controls and PCOS in size-matched follicles, whereas inhibin B concentrations were higher in dominant follicles from control ovaries. The advantage of both these studies was the use of unstimulated ovaries. Another study indicated that follicular aspirates from women undergoing IVF treatment had normal concentrations of follistatin although the rate of increase in follistatin concentrations from small to large follicles was less than expected in PCOS (Lambert-Messerlian et al., 1997
). The implications of this observation are clouded by the fact that aspirates were obtained after ovarian stimulation and administration of human chorionic gonadotrophin. The origins of circulating follistatin and activin are unclear, especially as follistatin has been shown to remain constant across the menstrual cycle and is not altered by administration of GnRH antagonist which acutely lowers FSH and LH concentrations (Kettel et al., 1996
).
Liao et al. recently produced preliminary results in follistatin gene mutations in PCOS (Liao et al., 2000). Although they did not find a mutation in the coding region of the Chinese population, there remains the possibly that there may be changes in follistatin in other ethnic populations or indeed changes in the regulation of the follistatin gene.
Inhibin B has been measured in follicular fluid and blood from women with PCOS. Two studies have shown higher concentrations in the circulation (Anderson et al., 1998; Lockwood et al., 1998
), although their populations of no more than 10 subjects makes a comparison difficult. This higher concentration has not been confirmed in follicular fluid (Lambert-Messerlian et al., 1997
; Magoffin and Jakimiuk, 1998
). In our study using a much larger population size, we were not able to confirm any changes in the peripheral blood using the identical assays in the studies of Anderson or Lockwood (Anderson et al., 1998
; Lockwood et al., 1998
).
The current study implicates follistatin and activin in the pathophysiology of PCOS in that follistatin is higher and activin is lower in the circulation of women with this condition. The study has a large number of well-characterized patients with PCOS clearly distinguished from the control group. All control subjects had blood taken in the early follicular phase at a time that follicles are similar in size to those that characterize the PCO ovary. However, the PCOS subjects were heavier yet slightly younger than control women, raising the issue that comparisons may be affected by these parameters. It has been found that there was no relationship of activin A and age in women even though there was a transient rise at about the time of menopause (Loria et al., 1998). Inhibin B probably does not change with age until menopause is imminent (Burger et al., 1999
) and changes cannot be explained on the basis of age. Multivariate analysis showed no relationship between age and any of the analytes measured. There is no significant association between follistatin and BMI; however, activin A shows a significant association with BMI which is negatively correlated in the control group and positively correlated in the PCOS women. It is well established that PCOS women are often overweight with an increase in their BMI. We have shown that the larger PCOS women have a positive correlation with activin A concentrations. This is in contrast to the control group whose activin A concentrations were negatively correlated with increasing BMI. The follistatin concentrations in the PCOS group were nearly twice as high as for the control group. It is postulated that the higher follistatin concentrations have a much greater effect on reducing the activity of activin A, thus reducing the FSH effect from the pituitary with a consequential effect on ovulation rate. This is the opposite effect to that of age, where activin A increases as women become older and the FSH concentration in blood increases (Reame et al., 1998
).
Follicles in PCOS are arrested at 810 mm in diameter and a variety of intrafollicular regulators have been implicated, including epidermal growth factor, insulin-like growth factor (IGF) and TGFß (Hillier and Miro, 1993; Findlay, 1994
; Hillier, 1999
). Overexpression of follistatin could be one mechanism whereby follicular growth is impeded. The previous studies that failed to show any difference in follicular fluid concentrations of follistatin could be explained by thecal production of follistatin and the rich blood supply in this area of the ovary leading to rapid clearance into the blood. The lack of difference in inhibin B does not accord with two previous studies in this area (Lambert-Messerlian et al., 1997
; Magoffin and Jakimiuk, 1998
). More investigations are needed before the true situation with inhibin B is resolved.
The current study raises the possibility that follistatin from the ovary or other tissues could inhibit follicle development in women prone to PCOS. Given the information relating abnormalities in other growth factors, including IGF and insulin, follistatin and activin regulation may be areas for diagnosis and pharmacological intervention in this condition.
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Acknowledgements |
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Notes |
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References |
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Submitted on September 13, 2000; accepted on January 4, 2001.