GDF9 and BMP 15 are secreted by oocyte and these regulates the function of cumulus cells thus helps in enhancing the oocyte developmental competence and embryo development.

1. Muzamil Ahmad
M.V.Sc. Student
Animal Biotechnology

2.  Recent
reproductive technologies have opened up
possibilities in accelerating genetic gain and
enhancing production potential in livestock .

3.  These







technologies include
Artificial insemination (AI).
Multiple ovulation and embryo transfer
(MOET).
In vitro production of embryos.
Animal Cloning.
Estrus synchronization.
Marker assisted selection ,and
Transfer of genes between animals.
(transgenesis).

4.  In
vitro production of embryos (IVEP)has emerged as a
reliable alternative method to conventional ovarian
super stimulation methods

Important research tool for animal embryology.

5.  Previous
studies have clearly demonstrated that, while
the intrinsic quality of the oocyte determines the
proportion of oocytes developing to blastocysts.
 The
post-fertilization culture environment has the
biggest influence on blastocyst quality.
[Russell et al., 2006]
 However, the exact influence of culture conditions
during critical events is still unknown.

6. Animal
cloning
Transgenic
animal
production
IVM
In vitro
embryo
production
IVF

7.  Oocyte
in vitro maturation (IVM) is a
reproductive technology that enables
mature oocytes to be generated ex vivo.
 IVM
involves removal of cumulus-oocyte-complexs
(COCs) and culturing them into standard culturing
medium for 24h upto meiosis II.

8.  The
efficiency of IVM technologies or assisted
reproductive technologies (ARTs) is limited by


The oocyte developmental competence ,and
Subsequent embryo development as compared to in
vivo.

9. a) Meiotic Maturation
b) Fertilization
c) Preimplantation development
d) Development to term /successful pregnancy

10.  Oocytes
gradually and sequentially acquire
developmental competence during the course of
folliculogenesis as the oocyte grows and its
companion somatic cells differentiate.
(Eppig et al., 1994).

11. (1) The origin of the oocyte.
(2) Follicle health.
(3) Hormonal stimulation.
(Lonergan et al., 1994)
(Blondin et al.,1995)
(Sirard et al., 2006)
(4) Communication between the oocyte and its surrounding
cumulus cells (CCs)
(Krisher, 2004)

12.  Previously
it was thought that oocyte is a passive
recipient of Cumulus Cell function.
 But
now it is evidenced oocyte is a key regulator of
its developmental competence by regulating its micro
environment via the secretion of paracrine molecules
like GDF9 , BMP15 etc.

13.  Oocyte
plays active role throughout folliculogenesis
via secretion of paracrine factors ( like GDF9 &
BMP15) that maintain an appropriate
microenvironment for the acquisition of its
developmental competence.
(Dong et al., 1996; Eppig et al., 1997;Gilchrist et al., 2004).

14.  Falck
and colleagues demonstrated in 1959 that intact
rabbit preovulatory follicles do not luteinize when
transplanted into the anterior chamber of the eye.

In contrast to oocyte-free explants of either the
follicle wall or granulosa cells that do undergo
morphological luteinization.
(Falck 1959).

15.  Subsequently,
Nalbandov and colleagues
confirmed this work using rabbit dominant
follicles in situ, showing that removal of the
oocyte caused spontaneous luteinization of
granulosa and theca cells and elevated secretion
of progesterone to levels produced by corpora
lutea.
( Nalbandov 1970).

16.  It
is known that bidirectional interactions occurs
between oocyte and cumulus cells.
 Bidirectional
interaction occurs via; gap junction and
paracrine signaling.
 Helps
in growth and development of both oocyte and
cumulus cells.
( Eppig et al.,1997,2002;Matzuk et al., 2002)

18.  Growth
differentiation factor 9 (GDF9)and Bone
morphogenetic protein 15 (BMP15) are two closely
related members of the Transforming Growth Factorβ (TGF-β ) superfamily.
 TGF-β
superfamily ; large family of structurally
related proteins.
 Controls
proliferation, differentiation, development
etc in variety of cells/tissues.

19.  GDF9
& BMP15 are expressed and translated in
oocytes .
 Synthesized
as preproproteins, consisting of a signal
peptide, a large proregion and a mature region.
( Shimasaki et al., 2004)

20. Properties :
 Lack of 4th cysteine residue (otherwise conserved in
other members of TGF-β superfamily)

 Function
mostly as homodimer/and or sometimes as
heteriodimers.
 Required for female fertility: as mutations
(homozygus) leads to infertility due to block in
follicullogenisis at early stage.

21. 
GDF9 and BMP 15 have recently been shown to
signal through known TGF-β superfamily receptors
to activate the SMAD intracellular cascade

22.  Two
pathways BMP pathway and TGFβ/activin
pathway which respectively activate SMAD1/5/8 and
SMAD2/3 messengers
 BMP15
utilizes BMP pathway to activate
SMAD1/5/8
 GDF9
utilizes SMAD2/3

24. Proliferation of Granulosa cells
Differentiation of Granulosa cells
Prevention of apoptosis
Inhibition of Luteinization
Cumulus cell expansion
Metabolism

25. GDF 9 and BMP15 stimulates granulosa cell
proliferation and DNA synthesis.
Expression of Ccnd2 mRNA cell cycle
transcript >>>>>cyclin d 2 protein of cell
cycle
 Assessed by uptake of [H3]thymidine

(Glichrist et al.,2006)

26. Oocyte secreted factors i-e,GDF9 & BMP15
differentiates the granulosa cells into mural cells
(MGCs) and cumulus cells (CCs) at antral stage of
follicle.

MGCs under influence of FSH

CCs
under influence of OSFs

(Hussein et.al.,2005;Diaz et al.,2007)

27.  BMP15
prevent Cumulus Cell apoptosis by
maintaining a localized gradient of antiapoptotic factors within the COC
(Hussein et al., 2005)

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Expression of anti-apoptotic gene Bcl2
Suppression of pro-apoptotic protein Bax

28. 
Suppression of LH receptor (LHCGR)
( Eppig et al.,1997)

Inhibition of FSH induced progestrone production

29. 
Two signaling pathways:(i) Gonadotrophin or epidermal growth factor (EGF)
stimulation and
(ii) Paracrine signals of oocyte (GDF9 &BMP15)

Synthesise extracellular matrix (ECM)

Hyaluronan (HA) major component

tumor necrosis factor alpha induced protein 6 (TNFAIP6)

Pentraxin 3(PTX3)
(Russell et al., 2006)

30. 
Prior to the LH surge, cumulus cells require GDF-9 to
support the metabolic cascades such as glycolysis and
sterol biosynthesis
(Sugiura et al. 2005).

Evidence for this comes from findings of reduced
cholesterol synthesis from acetate in cumulus-oocyte
complexes of Gdf9 − /−mutant mice.
(Sugiura et al. 2005).

33. IVF
IVF
Blastocyst
Inner cell
mass
Blastocyst
ICM
Hussein et al.,2005..

34. Treatment
No.of oocytes
used
% Cleavage Rate
% Blastocycst
from cleaved
oocytes
CONTROL
205
86.5 ±3.2
41.0 ±0.9
GDF9
191
88.1 ±2.0
49.5 ±3.9
BMP15
189
88.7 ±4.2
57.5 ±2.4
GDF9+BMP15
184
88.9 ±2.9
55.1 ±4.5
293H
187
80.4 ±2.2
27.1 ±3.1
(Hussein et al.,2005)

35.  It
is now clear that oocyte regulates the
cumulus cell functions like differentiation,
apoptosis, expansion, luteinization
 And
cumulus cells provide microenvironment
to oocytes for development
 In
order to increase the efficiency of ARTs
the OSFs (GDF9 & BMP15) may be utilized in
culture media