Anti-Mullerian hormone is a fertility in animals

1. Anti-Mullerian Hormone as a Fertility
Marker in Veterinary Field
Dr. P. M. Kekan
(M.V.Sc., Ph.D.)
Department of Veterinary Physiology
College of Veterinary and Animal Sciences,
Parbhani – 431402

2. Introduction
Causes of
Infertility
Hormonal
Gynecological
Pathological
Nutrional

3. Introduction Contd…
Apart from this there is another cause of
infertility i.e.
AMH
What is AMH ?????

4. Introduction Contd…
Anti-Mullerian Hormone
Alfred Jost named it ‘hormone inhibitrice’
which later became better known as
Mullerian Inhibiting Substance (MIS),
 Mullerian Inhibiting Factor (MIF) or
Anti-Mullerian Hormone (AMH)
(Claes, 2015).

5. Introduction Contd…
Anti-Mullerian hormone, is a glycoprotein of 140
kDa belonging to the transforming growth factor
beta family that is expressed only in the gonads
(Cate et al., 1986)
Presently, AMH is the best endocrine marker of
the ovarian follicular reserve in human, in mouse,
and recently the AMH is also a reliable endocrine
marker of the population of small antral
gonadotropin responsive follicles in the cow
(Rico et al., 2009)

6. Introduction Contd…
 Over the past 10 years, attention
has been focused on AMH in humans
in the context of assisted
reproductive technologies (ART).
 Numerous clinical studies have
shown that, low plasma AMH
concentrations are indicative of
ovarian ageing

7. Secretion of AMH
 Anti-Mullerian hormone is produced by
granulosa cells of all primordial, primary,
secondary follicles, as well as antral follicles up to
4 to 5 mm diameter.
 The function of AMH in females is to regulate or
limit the recruitment of primordial follicles into
folliculogenesis, by reducing the responsiveness
to these follicles to follicle stimulating hormone. .
(Visser et al., 2006)

8. Granulosa cells

9. Granulosa cells

10. Anti-Mullerian as a fertility indicator
in heifers
 The question arises as to how early in development,
AMH can be measured as an indicator of fertility.
 Identification of heifers with low or high fertility at
birth or weaning would be advantageous to producers
for making management decisions.
 If measure of AMH at weaning could predict
subsequent fertility, this would not only reduce
replacement heifer costs, but also identify less fertile
heifers at an age that would allow their marketing as
stocker-feeder cattle at a more optimal time.

11. AMH Concentration in Heifers
• Newberry (2016) observed serum AMH
ranged from 40 to 990 pg/ml, with a mean of
300 pg/ml at weaning.
• Moreover, when heifers were compared by
quartiles, based on serum AMH concentration
at weaning failed to detect any effect of AMH
on subsequent heifer cyclicity at breeding,
estrous response, after synchronization.

12. AMH Concentration in Heifers
Quartile 1 Quartile 2 Quartile 3 Quartile 4
40 to 230
pg/ml
270 to 450
pg/ml
500 to 770
pg/ml
800 to 1730
pg/ml
Comparison of heifers based on the difference
between AMH concentrations at breeding versus
weaning revealed that none of the heifers in the
lowest quartile (Q1) became pregnant after AI,
compared with 80% in the highest quartile

13. AMH Concentration in Heifers
• Results indicated that either AMH concentration at
breeding or the change in AMH from weaning to
breeding can identify beef heifers more likely to
conceive to AI and to conceive early in the breeding
season.
• Rico et al., (2012) reported that the heifers below 150
pg/ml remained non-pregnant, he suggested that the
animals having low AMH concentration below cut-off
level by determining single blood sample should be
culled.
• The cut-off value may differ breed wise (Chachere,
2015).

14. AMH & AFC as a Predictor of embryo
production in cows
• Center et al. (2018) investigated the use of
Anti-Mullerian hormone (AMH) and/or follicle
counts as a predictor of subsequent
superovulatory response and embryo
production in 79 beef cows.
• The distribution of AMH concentrations were
divided into quartiles (Q1 lowest, Q2, Q3 and
Q4 highest).

15. AMH & AFC as a Predictor ---
Quartiles Q1 Q2 Q3 Q4
AMH (ng/ml) 0.013 to 0.068 0.069 to 0.263 0.264 to 0.363 0.364 to 0.898
No of Follicles 13.46 ± 0.91b 14.95 ± 0.98b 16.79 ± 0.94ab 19.33 ± 0.94a
No. of embryos 9.77 ± 1.76b 9.36 ± 1.91b 15.50 ± 1.83ab 20.13 ± 1.83a
abc in a row differ significantly (P < 0.05)
 Donors cow in AMH Q4 had a greater (P < 0.001) numbers of
3 – 5 mm follicles as compared to Q1 and Q2.
 Result confirm that AMH concentration is positively correlated
with number of small AFC (3-5 mm) in ovaries.
 AMH & AFC Might be used to predict superovulatory response in cows

16. AMH in Follicular Fluid
• Liang et al. (2016) determined the AMH
concentration in follicular fluid (FF) recovered
from different size follicles and evaluate whether
the AMH concentration in FF is associated with
the antral follicular count.
• Ovaries of buffaloes were collected at the
slaughter house, and all follicles were counted
and categorized as small (3–5 mm), medium (5–8
mm), and large (>8 mm). They reported that,
AMH concentration in FF significantly decreases
as the size of follicle increases in diameter.

17. AMH During Estrous Cycle
• Souza et al. (2014) Estimated the AMH
concentration during 3 different stages of
estrous cycle.
• They observed that, the AMH concentration at
three different stages of synchronized estrus
was not affected (133.2 + 10.6, 132.7 + 9.7
and 150.9 + 10.6 pg/ml) during proestrus,
estrus and diestrus respectively.

18. AMH During Estrous Cycle
• Pfeiffer et al. (2014) determined AMH at
estrus during synchronized and a natural
bovine estrous cycle.
• The mean concentration of AMH did not differ
between natural (0.0543 ± 0.0076 ng/ml) or
synchronized (0.0428 ± 0.0076 ng/ml) estrous
cycle.

19. AMH During Estrous Cycle
• Batista et al. (2016) evaluated the association
between AMH and AFC in Bos taurus and Bos
indicus.
• The experiment was conducted on cyclic heifers and
calves of 2 to 4 months.
• They reported positive correlation between plasma
AMH and AFC in calves and concluded that calves
had higher plasma AMH concentration than cyclic
heifers, whereas, the AMH concentration remained
steady throughout the estrous cycle.

20. AMH During Estrous Cycle
• Almeida et al. (2011) determined the concentrations
of AMH in the blood of mares during estrous cycle,
and pregnancy. Blood samples were collected daily
from 6 mares during estrous cycle and samples from
6 pregnant mares monthly throughout gestation
period.
• The AMH concentration of cyclic mare was 0.96 ±
0.08 ng/ml, which was higher than pregnant mares
(0.72 ± 0.05 ng/ml).
• There was no effect of cycle stage and month of
gestation on serum AMH concentration.

21. AMH During Estrous Cycle
Monniaux et al. 2013

22. AMH During Estrous Cycle
• The high AMH concentration on the day of estrus and
then decreasing on day 3 and 7 of estrous cycle may be
due to inhibition of AMH production by FSH at
granulosa cell level.
• The non significant decrease in AMH concentration
following estrus is not associated with concomitant
changes in the numbers of follicles detected by ovarian
ultrasonography, but it could result from the inhibiting
action of FSH upon AMH production by granulosa cells
of the AMH secreting follicles in the basal follicular
growth stage. (Rico et al., 2011)

23. AMH and Pregnancy/Fertility
• There is a individual variation in AMH
concentration.
• There is positive association between AMH and
fertility.
• There is positive correlation between AMH and
AFC (3-5 mm) and fertility.
• Cows with low AMH had greater risk of
pregnancy loss between day 30 and 65 of
gestation than cows with intermediate or high
AMH. Ireland et al., (2011)

24. Correlation of AMH, AFC and
Pregnancy
• Rico et al. (2011) studied the regulation of
anti-mullerian production in 22 Holstein cows,
which were divided into two groups according
to low and high ovulatory response.
• Ultrasonography was performed three times
weekly from day of estrus till next estrus and
also endocrine and follicular parameters were
compared throughout estrous cycle in both
the groups.

25. Correlation of AMH, AFC and
Pregnancy
• They observed that the AMH concentration
was strongly correlated with average number
of follicles with diameter between 3 - 5 mm.
• The average AMH concentration was higher
(P< 0.01) in the high ovulatory response group
(94.8 6 + 11.2 pg/ml) than in the low
ovulatory group (40.7 6 + 14.8 pg/ml).
• The pregnancy rate was higher in the group
with high AMH and AFC (3-5 mm).

26. Therefore, it is concluded that the AMH test can be done
at any stage of estrous cycle, as there is no fluctuation in
AMH concentration during estrous cycle.

27. Breed wise variation in AMH & AFC
Breed Murrah Gyr Holstein
AMH
(ng/ml)
0.18 ± 0.03 0.60 ± 0.12 0.24 ± 0.06
AFP 25.6 ± 2.1 60.0 ± 12.2 35.9 ± 6.8
 Baldrighi et al. (2014) evaluated plasma
concentrations of Anti-Mullerian hormone and the
ovarian antral follicle population in cyclic heifers of
Murrah, Holstein and Gyr.
The cut-off value of AMH is 0.24 , 0.60 and 0.18 for
Holstein, Gyr and Murrah buffaloes, respectively .

28. AMH as a marker for diagnosis of
Granulosa-cell tumors in Mares
• Almeida et al. (2011) determined the
concentrations of AMH in the blood of cyclic
mares (n=6), pregnant mares (n=6) and in the
mares of granulosa-cell tumors (n=11).
• Granulosa-cell tumors in mares was confirmed
by histopathology.

29. AMH as a marker for diagnosing GCT in
Mares
Cycle Mares
(n=6)
Pregnant Mares
(n=6)
GCT Mares
(n=11)
0.96 ± 0.08
ng/ml
0.72 ± 0.05
ng/ml
1901.4 ± 1144.6
ng/ml
In granulosa-cell tumor mares, serum concentrations
of AMH were higher than those in cyclic and pregnant
mares.

30. Influence of Gender in AMH of Asian &
African Elephants
Species Gender Mean (ng/ml)
Asian Female (n=18) 0.19 ± 0.02
African Female (n=41) 0.17 ± 0.04
Asian Male (n=20) 40.27 ± 3.18
African Male (n=18) 31.01 ± 4.22
 There is no significant difference between Asian and African
elephants within the sex.
 But significantly (P<0.05) higher AMH is recorded in male
elephants as compared to female

31. Estimation of Anti - Mullerian
Hormone by ELISA
• Serum AMH concentration can be evaluated
by using bovine AMH ELISA kit, AL - 114
(Ansh Labs, Webster, TX, USA).
• The cost of kit is Rs. 40,000/-
• The cost of each sample will be approximately
Rs. 500/-

32. Future Research
To develop the RAPID CARD for
instant determination of
approximate AMH concentration in
blood.

33. Conclusions
• In cattle, circulating AMH concentration can
help field veterinarians to predict antral
follicular population (AFP) in the ovaries.
• One time determination of AMH
concentration will help to predict the
reproductive performance of the animals at
the heifer stage.
• It will help to select the high potential animals
for assisted reproductive technologies.