Essay Submitted by

1
Anti-Mullerian Hormone (AMH) in Female
Reproduction (Part II)

• Essay Submitted by
• Mohamed D. Mansy
• Specialist of Obstetrics and Gynecology
• Ministry of Health Population (MOHP) Port Said
• 2009

2

• Under supervision of
• Prof. Dr Mahmoud Farouk Midan
• Professor and Head of
• Obstetrics and Gynecology Department
• Faculty of medicine, Al-Azhar university,
  Damietta.
• Dr. Khattab Abd Elhalem Omar Khattab
• Assist. Professor of Obstetrics and Gynecology
• Faculty of medicine, Al-Azhar university,
  Damietta.
• Dr. Rashed Mohamed Rashed
• lecturer in Obstetrics and Gynecology
• Faculty of medicine, Al-Azhar university,
  Damietta.

3
Review of Literature
AMH as a Marker of Ovarian Reserve in Patients
Undergoing Assisted Reproductive Technology (ART)
4

• In women undergoing treatment for infertility,
  ovarian aging is characterized by decreased
  ovarian responsiveness to exogenous gonadotropin
  administration and poor pregnancy outcome.

5

• On the one hand, correct identification of poor
  responders by assessment of their ovarian reserve
  before entering an in vitro fertilization (IVF)
  program is important.

6

• On the other hand, assessment of the ovarian
  reserve may also benefit patients that would
  generally be excluded from IVF programs because
  of advanced age.

7

• Several studies have shown that AMH is an
  excellent marker to determine ovarian
  responsiveness also in an IVF program.

8

• Hormone measurements in the early follicular
  phase (day 3 of spontaneous cycle),
  retrospectively or in a group of unselected
  patients, revealed that AMH levels are lower in
  patients with poor ovarian response than in women
  with normal response.
• )Seifer, et al., 2002 and vanRooij et al., 2002)

9

• ovarian responsiveness being defined as the
  number of oocytes retrieved, or as cancellation
  due to impaired or absent follicular growth.

10

• AMH serum levels were shown to be highly
  correlated with the number of antral follicles
  before treatment and number of oocytes retrieved
  upon ovarian stimulation
• (vanRooij, et al., 2002).

11

• serum AMH levels had a better predictive value
  than serum levels of FSH, inhibin B and E2, and
  that the predictive values for AMH and AFC were
  almost identical (ROCAUC 0.85 vs 0.86)

12

• To achieve a reliable predictive outcome, one
  single hormone measurement for AMH seems
  sufficient
• (Fanchin, et al., 2005a).
• The absence of regulation of AMH by gonadotropins
  was shown in both rodents and man.
• AMH acts as a paracrine rather than a systemic
  factor.

13

• Treatment of IVF patients with a single, high
  dose of gonadotropin-releasing hormone (GnRH)
  agonist, resulting in a rise of endogenous FSH
  and LH, does not affect AMH serum levels
• (vanRooij, et al., 2002).

14

• Similarly, in conditions where FSH levels are
  suppressed, such as pregnancy, AMH levels remain
  constant
• (LaMarca, et al., 2005a)
• Thus, AMH is not influenced by the gonadotropic
  status and reflects only the follicle population.

15

• However, from a clinical point of view, poor
  responders should be identified before treatment
  therefore, it is more useful to determine serum
  AMH levels during a spontaneous cycle.

16

• Throughout the controlled ovarian
  hyperstimulation protocol, serum AMH levels
  correlated well with the decrease in number of
  small antral follicles (12 mm) (Fanchin, et al.,
  2003a) reflecting the complete conversion of
  small antral follicles into large antral
  follicles in response to FSH stimulation.

17

• Indeed, no correlation with the number of growing
  follicles (gt 12 mm) was observed, in line with
  the low expression of AMH in these follicles
• (Weenen, et al., 2004).

18

• In the days following hCG treatment, AMH serum
  levels initially declined, possibly as a result
  of the luteinization of granulosa cells upon hCG
  treatment that also causes a decline in E2
  levels.

19

• During the midluteal phase, AMH serum levels
  slightly increased, probably as a result of the
  presence of newly developed, small antral
  follicles
• (Fanchin, et al., 2005b).

20

• All combined these studies strongly support a
  role of serum AMH level as a marker for ovarian
  responsiveness.
• However, the application of AMH to predict
  ongoing pregnancy seems limited, although day 3
  serum AMH levels are higher in patients that
  become pregnant after IVF treatment than in those
  who do not.
• (Hazout, et al., 2004).

21

• Therefore, it is likely that the quantitative
  aspect of AMH as a marker of the ovarian reserve
  has contributed predominantly to the association
  with pregnancy outcome.

22

• Indeed, other studies did not observe a
  predictive value of AMH serum levels for ongoing
  pregnancy after IVF treatment
• (Penarrubia, et al., 2005).
• Moreover, the decrease in AMH in FSH-treated
  women might be the result of a growth stimulation
  by FSH of the follicles that enlarge, thereby
  losing their AMH expression.

23

• Small follicles (8-12 mm in diameter) secreted
  AMH at levels that were approximately three times
  as high as those of large follicles (16-20 mm in
  diameter)
• (Fanchin, et al., 2005c).
• Serum AMH levels correlated strongly with the
  AFC, the number of follicles retrieved, age,
  inhibin B and FSH.

24

• AMH assessment was shown to predict ovarian
  reserve with a sensitivity of 80 and a
  specificity of 85 (Tremellen, et al., 2005).
• AMH levels have also been shown to be 10-fold
  lower in the cancelled cycles compared with
  patients who had a completed IVF cycle. In about
  75 of cancelled cycles, AMH levels were below
  the detection limit (Muttukrishna, et al., 2004).

25

• For the first time, clinicians may have a
  reliable serum marker of ovarian response that
  can be measured independently of the day of the
  menstrual cycle
• (LaMarca, et al., 2007).

26

• Recently, Raziehi, et al., (2008) concluded
  that, it appears that AMH serum levels are
  associated with ovarian response in ART cycles
  and can be served as a novel marker for ovarian
  reserve.
• Riggs, et al., (2008) cocluded that
  Anti-Müllerian hormone correlates better than
  age, follicle-stimulating hormone, luteinizing
  hormone, inhibin B, and E2 with the number of
  retrieved oocytes.

27

• AMH is a predictor of ovarian response and
  suitable for screening. Levels 1.26 ng/ml are
  highly predictive of reduced ovarian reserve and
  should be confirmed by a second line antral
  follicle count. Measurement of AMH supports
  clinical decisions, but alone it is not a
  suitable predictor of IVF success
• (Gnoth, et al., 2008).

28

• The AFC and AMH are the most significant
  predictors of poor response to ovarian
  stimulation during ART. The AMH and AFC, either
  alone or in combination, demonstrate a similar
  predictive power but are not predictive of
  nonconception, which is dependent on the woman's
  age
• (Jayaprakasan , et al., 2008).

29

• The AMH and inhibin B levels on the day of oocyte
  retrieval are correlated to reproductive outcome
• (Wunder, et al., 2008).

30
Review of Literature
AMH as a Marker of Ovarian Dysfunction
31

• In the human, follicle development to the antral
  stage continues throughout life until depletion
  of follicles at the menopause, even in the
  presence of conditions under which endogenous
  gonadotrophin release is substantially
  diminished.
• (Richardson and Nelson, 1990)

32

• AMH serum levels have been measured in women
  affected by hypergonadotrophic amenorrhoea
  (premature ovarian failure, POF) or by
  hypogonadotrophic amenorrhoea (functional
  hypothalamic amenorrhoea). AMH serum levels have
  been found to be normal in women with
  hypogonadotrophic amenorrhoea and to be
  undetectable in 83 of women with
  hypergonadotrophic amenorrhoea (POF). The
  remaining 17 of patients had very low AMH serum
  levels.

33

• AMH measurement could provide information on
  ovarian reserve when women with secondary
  amenorrhoea are investigated and initial follicle
  recruitment is not abolished in hypogonadotrophic
  hypogonadism.
• (LaMarca, et al., 2006)

34

• Premature ovarian failure (POF) is generally
  irreversible. However, developing follicles up to
  the antral stage are reported in POF and
  anti-Müllerian hormone (AMH) might be a good
  indicator of follicular presence.

35

• The dysfunction of AMH production in the antral
  follicles of POF women observed in their series
  of ovarian biopsies suggests the possibility of
  the presence of a defect in antral follicular
  development, particularly concerning the
  granulosa cells.

36

• A decreased antral granulosa cell AMH production
  could alter the mechanism of follicular
  recruitment in these patients. AMH could be
  useful in discriminating POF patients with a
  sizable follicular population from patients with
  no or few ovarian follicles, with a lesser
  potential for ovulation and pregnancy
• (Méduri, et al., 2007)

37

• PCOS is one of the most common endocrine
  disorders in women of reproductive age
• (Franks, 1995)
• It is characterized by anovulation manifested as
  oligo- or amenorrhea, elevated levels of
  circulating androgens, and polycystic ovaries as
  visualized by ultrasound. The diagnosis is based
  on the presence of at least two of the described
  characteristics, as defined by the Rotterdam
  Consensus (2004)

38

• AMH production is reported to increase in women
  with polycystic ovary syndrome (PCOS) compared to
  controls
• (Fallat, et al., 1997, Cook, et al., 2002 and
  Laven, et al., 2004)
• This may be the result of aberrant activities of
  the granulosa cells in the polycystic ovaries
• (Mulders, et al., 2004)
• AMH measurement can offer a high specificity and
  sensitivity (92 and 67, respectively) as a
  marker for PCOS.

39

• On this basis it has been proposed that in
  situations where accurate ultrasonography data
  are not available, AMH could be used instead of
  the follicle count as a diagnostic criterion for
  PCOS.
• (Pigny, et al., 2006)

40

• AMH levels in amenorrhoeic is higher than in
  oligomenorrhoeic women with PCOS, which could
  indicate a role for AMH in the pathogenesis of
  PCOS-related anovulation.
• Alternatively, high AMH values could reflect a
  more evident impairment in follicular development
  and granulosa cell function in the ovaries of
  amenorrhoeic than oligomenorrhoeic PCOS women.
• (LaMarca, et al., 2004b)

41

• Anovulation in PCOS could be the result of
  excessive pituitary LH secretion and inhibin
  production by multiple, small follicles that
  inhibit FSH secretory dynamics.
• Increased ovarian AMH production may exert a
  paracrine negative control on follicle growth,
  sufficient to prevent selection of a dominant
  follicle.

42

• Pellatt, et al. (2007) concluded The reduction of
  AMH in follicles greater than 9mm from normal
  ovaries appears to be an important requirement
  for the selection of the dominant follicle.
• AMH production per GC was 75 times higher in
  anovPCOs, compared with normal ovaries. This
  increase in AMH may contribute to failure of
  follicle growth and ovulation seen in polycystic
  ovary syndrome.

43

• It is important to bear in mind that these raised
  levels could be misleading if used as a marker of
  ovarian reserve. The overproduction of AMH could
  have important implications for the mechanism of
  anovulation in these women.

44

• The reduction in serum level of AMH in PCOS
  patients was lower than in the controls. The
  authors concluded that this may indicate a
  sustained reproductive lifespan in these
  patients.
• (Cook, et al., 2002)

45

• On histological examination, polycystic ovaries
  exhibit the same number of primordial follicles,
  whereas the number of developing follicles is
  doubled compared with normal ovaries
• (Webber, et al., 2003)
• Hence, it may be proposed that the process of
  ovarian ageing is delayed in women with PCOS as
  high AMH levels may inhibit the primordial
  follicle pool depletion.
• (Cook, et al., 2002 )

46

• Serum AMH level can be used as a marker for
  the number of growing follicles. Besides being a
  marker for a diminishing follicle pool, serum AMH
  level can also serve as a marker in ovarian
  pathophysiology, in which the antral follicle
  pool is enlarged.

47

• In a study by Eldar-Geva, et al,. (2005),
  categorization of PCOS women by presence or
  absence of hyperandrogenism showed that AMH
  levels were significantly different between
  groups. Both groups had increased AMH levels
  compared to control women, but levels in women
  with PCO and hyperandrogenism were even further
  elevated.
• These results suggest that the nonvisible pool of
  follicles may be further increased in the
  presence of increased androgen levels.

48

• The increased insulin levels in some PCOS women
  can, in part, account for the hyperandrogenism,
  because insulin acts synergistically with LH to
  enhance androgen production by theca cells.
• However, serum AMH levels do not seem to
  correlate with BMI and insulin levels
• (Fleming, et al., 2005).

49

• In contrast, in a small study, LaMarca, et
  al., (2004b) observed a positive correlation
  between serum AMH levels and the Homeostatic
  model assessment (HOMA) index .

50

• In a study of obese PCOS women, metformin
  treatment suppressed androstenedione levels and
  ovulation rate although androgen levels were
  still above the upper limit of the normal range.
• Metformin administration also resulted in a small
  but significant reduction of serum AMH levels
  after 8 months of treatment, whereas the follicle
  number did not change significantly.
• (Fleming, et al., 2005)

51

• In a smaller study, metformin treatment for 6
  months also decreased serum AMH levels only
  slightly, and levels remained strongly elevated
  compared to controls.
• (Piltonen, et al., 2005)

52

• Future challenges comprise the availability of
  a well standardized assay and the development of
  AMH agonists and antagonists as possible tools to
  manipulate ovarian function for contraception or
  ovarian longevity.
• (Broekmans, et al., 2008)

53
Review of Literature
AMH as a Marker of Granulosa Cell Tumours (GCTs)
54

• AMH expression is limited to ovarian granulosa
  cells, hence it has been proposed to use AMH as a
  marker of granulosa cell tumours (GCTs).
• It has been shown that AMH serum levels are
  increased in women affected by GCTs.
• (Rey, et al., 2000)

55

• The mean AMH level was 190.3 ng/ml (range 2-1124
  ng/ml) in patients with GCTs in one study.
• (Lane, et al., 1999)
• In patients in whom serial measurements were made
  following initial tumour resection, the length of
  time that an elevation in AMH levels preceded
  clinical detection of a recurrence was 16 months.
  
• (Long, et al., 2000)

56

• AMH seems to be superior to alpha-inhibin and
  oestradiol in the follow-up of GCT. It is largely
  recognized that AMH and alpha-inhibin exhibit a
  higher degree of sensitivity than oestradiol in
  progressive GCT. Indeed, oestradiol production by
  GCT is widely variable
• (Lappohn, et al., 1989)

57

• In the follow-up, AMH seems to be a marker of
  comparable value to alpha-inhibin. However, AMH
  is elevated only in GCT, while inhibin may be
  elevated in different types of cancer
• (Healy, et al., 1993)

58
Based on the published studies it is possible to
elaborate the following recommendations for the
use of AMH as a marker of GCT

• High serum AMH levels are found in 7693 of
  patients with GCT.
• Serum AMH levels should normalize within few days
  following surgery.
• Persistent AMH levels are indicative of residual
  disease.
• When bilateral ovariectomy is performed AMH must
  be undetectable in serum. Even very low levels
  may be suggestive of residual disease.
• A post-operative rise in serum AMH levels may
  precede the clinical recurrence. Hence, the
  clinical examination and imaging should be
  anticipated.
• AMH levels should be obtained every 6 months for
  at least 5 years after the surgery. However, it
  should not be ignored that a late recurrence (up
  to 30 years after initial treatment) has been
  reported in up to 33 of patients (Stenwig, et
  al., 1979).
• Recurrences in juvenile GCTs occur within a year
  of initial diagnosis at a mean of 56 months.
  Hence, in juvenile GCT, AMH serum levels should
  be determined every month following the surgery.

59

• Several authors have also hypothesized a role for
  recombinant AMH in the treatment of epithelial
  ovarian cancer
• Based on the physiological inhibiting role of AMH
  on the mullerian ducts, researchers have
  demonstrated that AMH inhibits epithelial ovarian
  cancer cell in vitro.
• (Stephen, et al., 2001, 2002)

60

• The administration of recombinant AMH in
  immunosuppressed mice with human epithelial
  ovarian cancer implants has been followed by
  reduced dimensions of the graft (Stephen et al.,
  2002). This of course will be the basis for
  future research aiming to investigate the
  possible clinical role of AMH as neo-adjuvant or
  most probably adjuvant therapy for ovarian cancer
  
• (LaMarca and Volpe, 2007)

61
Table (4) Modifications in Markers of Ovarian
Function (Oestradiol, Inhibin B and AMH) During a
Woman's Life and in Some Pathological Conditions.
62
Summary and Conclusions
63
Summary

• Anti-Müllerian hormone (AMH) is a dimeric
  glycoprotein, a member of the transforming growth
  factor (TGF) superfamily. It is produced
  exclusively in the gonads, and is involved in the
  regulation of follicular growth and development.
  In the ovary AMH is produced by the granulosa
  cells of early developing follicles and seems to
  be able to inhibit the initiation of primordial
  follicle growth and FSH-induced follicle growth.
  A clearer understanding of its role in ovarian
  physiology may help clinicians to find a role for
  AMH measurement in the field of reproductive
  medicine.

64

• As AMH is largely expressed throughout
  folliculogenesis, from the primary follicular
  stage towards the antral stage, serum levels of
  AMH may represent both the quantity and quality
  of the ovarian follicle pool. Compared to other
  ovarian tests, AMH seems to be the best marker
  reflecting the decline of reproductive age. AMH
  measurement could be useful in the prediction of
  the menopausal transition. It could also be used
  to predict poor ovarian response and possibly the
  prognosis of in vitro fertilization (IVF) cycles.

65

• AMH has been shown to be a good surrogate marker
  for polycystic ovary syndrome (PCOS).
• Also, its use as a marker for granulosa cell
  tumours has been proposed.
• The administration of recombinant AMH in
  immunosuppressed mice with human epithelial
  ovarian cancer implants has been followed by
  reduced dimensions of the graft. This of course
  will be the basis for future research aiming to
  investigate the possible clinical role of AMH as
  neo-adjuvant or most probably adjuvant therapy
  for ovarian cancer.

66
Conclusions

• AMH is a very sensitive indicator of ovarian
  ageing and for ovarian response to controlled
  ovarian stimulation. Compared with the other
  ovarian tests, AMH seems to be the best marker
  for reflecting the oocyte/follicle pool. Also,
  serum levels of AMH are a good candidate for
  inclusion in standard diagnostic procedures to
  assess ovarian dysfunctions, such as premature
  ovarian failure.

67

• AMH can be measured at any time during the
  cycle, which is a great advantage in clinical
  practice. However, complementary studies in
  various clinical situations are necessary to
  attest the superiority of this compared to the
  other markers.

68

• There seems to be little doubt that research
  on AMH will continue in the years to come. A
  clearer understanding of its role in ovarian
  physiology may help clinicians to find a role for
  AMH measurement in the field of reproductive
  medicine.

69
Recommendations
70

• In order to determine whether serum AMH level
  has prognostic value, additional prospective
  studies in a normal population are necessary to
  provide definite proof for this concept.

71

• The positive correlation between serum AMH
  levels and number of antral follicles is also
  observed in women with PCOS. The elevated levels
  of AMH in these women strongly suggest that serum
  AMH levels may also be used in the diagnosis of
  PCOS.

72

• The difference in serum levels of AMH
  between subgroups of PCOS women suggests that AMH
  might also be used to establish a
  subclassification of this heterogeneous syndrome.
  However, more studies, preferably prospective,
  with thoroughly analyzed patient cohorts are
  necessary to define cutoff values. In addition,
  studies are necessary to determine whether serum
  AMH levels are also indicative of improved
  ovarian function upon treatment of PCOS women.

73

• Genetic studies of well-defined population
  cohorts would also provide more knowledge about
  the role of AMH in ovarian physiology.

74
thank you
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