Oocyte And Somatic Cell Communication Flashcards
What happens to the transzonal processes when they are exposed to the LH surge?
One of the first things that happens when exposed to the LH surge is that these transzonal processes cease to exist. The oocyte then is essentially on its own.
What 3 factors secreted by the oocyte are thought to be important for growth and differentiation of the follicle?
1) . GDF-9
2) . BMP-15
3) . BMP-6
GDF-9 is expressed by all oocytes at al stages and clearly has different roles.
GDF-9 and BMP-15 are expressed specifically by the oocyte.
BMP-6 is not expressed specifically by the oocyte. It is expressed by the cumulus cell layer and the granulosa cell layers within the follicles. The oocyte is the only structure within the follicle that actually expresses the mRNA for BMP-6 so it appears that it produces quite a lot of the BMP-6 which is then taken up within the somatic cells.
What are the main 4 somatic cell factors and what cells express them?
1) . Anti-Müllerian hormone (AMH) - expressed from somatic cells most highly during the Gonadotropin-responsive stages of follicle development. AMH is used as a marker for the ovarian reserve on assisted reproduction. Because granulosa cells secrete AMH we can use AMH levels as an index of how well a woman is likely to response to ovarian stimulation.
2) . Inhibin alpha - expressed highly in the granulosa cell layer.
3) . Inhibin Beta A - expressed in the granulosa cell layer.
4) . Follistatin - very highly expressed by the granulosa somatic cells in a number of species. Follistatin is an antagonist of a number of regulatory systems. It is thought of primarily as an Inhibin antagonist, but also binds to the BMP proteins.
Describe BMP receptor signal transduction.
BMP receptors are composed of two subunits. A type 2 receptor that binds the BMP ligand and then once receptor-ligand interaction has occurred then the type 1 receptor binds. This then activated smads. Smad 4 is the co-smad. This complex then translocates to the nucleus and affects gene transcription.
In the granulosa cells (and a little bit of the oocytes) we have the expression of the BMP receptor 1A (BMPR1A) and BMPR1B (both type 1 receptors). The granulosa cells within these follicles also express BMPR2.
These receptors are specific for a number of local factors that we have been talking about. Therefore, as we find the ligand and the receptors it is likely that they are doing something important.
What evidence do we have of the importance of these local factors such as BMP-15?
The best evidence we have for the importance of these local factors is in animals with mutations.
Natural knockouts for BMP15 were found to be infertile and the ovary was completely inactive.
It was found with one copy of the BMP-15 mutation they would have an increased ovulation rate.
It is clear that BMP-15 is an important local factor that affects both the early phases of follicle development and likely the terminal phases.
What information can knockout mice give us about the the importance of somatic cell factors such as AMH?
Knockout mice can also give us information about the importance of local factors such as the components of the inter follicular cascade.
AMH knockout mice showed more growing follicles in prepubertal animals, but in adult individuals the number of primordial follicles was lower in knockout individuals, but they still had an increased number of growing follicles.
The conclusion from this was that AMH was probably one of the factors controlling the initiation of follicle development and when you take it away you have high rates of initiation and more growing follicles. This means that as the rate of initiation is higher the primordial reserve gets depleted and you have less primordial follicles in the adult.
AMH in sheep appears to dictate the rate at which follicles progress through the gonadotrophin-responsive stages.
How is primordial follicle initiation controlled?
The mechanism of action is unknown.
Potential initiators of primordial follicles include the TGF-beta family (GDF-9, BMP-15, AMH), Ckit produced by granulosa cells, somatic EGF and somatic Insulin/IGF.
There are probably a number of systems that control this important process with a lot of built in redundancy.
How do we know what stages gonadotrophins effect early follicle development?
We know what effects gonadotrophins have at each stage mainly from the results of hypo-gonadotrophin and hyper-gonadotrophin graft experiments.
In the 2 months after the after procedure hyper-gonadotrophic grafts had grown right up to the antral phase, but in the hypo-gonadotrophic grafts we only saw follicles up to the secondary phase.
This indicates that gonadotrophin is effecting the latter stages of development of the follicle in the later stages of folliculogenesis.
After 4-month post graft however there is no difference between the two groups.
This provides evidence that the early stages of follicular development are gonadotrophin-independent (as we still get growth up to the secondary phase in both groups), the stages after the secondary phase up until the early antral phase are gonadotrophin responsive but not dependent (we still get development of the follicle through these stages in both groups but it is slower without gonadotrophins).
Gonadotrophin levels also have an effect on follicular morphology. We see fewer granulosa cells dividing in the hyper gonadotrophic follicles.
Hormone levels are also very different on hypergonadotrophic and hypogonadotrophic animals. Levels of Inhibin A and oestrodiol remain very low in hypo graft animals.
It seems likely that preantral follicles secrete something (Inhibin A or activin) that positively feeds back on the early stages of follicle development. This fits in with the fact that there is a good correlation between the numbers of growing follicles and the numbers of primordial follicles in the ovaries of most species.
The study concluded that:
- FSH does not affect the rate of primordial follicle activation.
- FSH is not an absolute requirement for pre-antral follicle development .
- FSH can affect the rate of the latter stages of pre-antral development.
- further evidence of local feedback loops controlling the early stages of pre-antral follicle development (activin?).
What evidence do we have of positive feedback loops in the early stages of follicle development?
It seems likely that preantral follicles secrete something (Inhibin A or activin) that positively feeds back on the early stages of follicle development. This fits in with the fact that there is a good correlation between the numbers of growing follicles and the numbers of primordial follicles in the ovaries of most species.
What are the advantages of in vitro growth and maturation of follicles?
Advantages of in-vitro growth:
- supersede existing ART technology.
- overcomes the problem of reintroduction of malignant tissue.
- applicable to individuals at a range of ages.
- great flexibility in timing and tissue use.
What are some of the problems with in-vitro follicle maturation?
The big problem is that it is very technically demanding, especially as it must result in oocytes that can give rise to normal individuals. It is very high risk.
The main problem with growing relatively large structures on vitro is the issues with getting gas into the centre.
What are the most promising methods of on-vitro follicle growth and maturation?
Within cortical patches ultra thin section of approximately 100um seem to be the most successful method.
Another way is to isolate the follicles, but this can be very difficult to do. People have done it and culture system for isolated follicles do exist (e.g. Plastic or in-situ culture for primordial follicles). Culture systems have also been published for pre-antral follicles.
What method of in-vitro follicle growth seems to be most promising?
There has been some success with isolated follicle growth with primordial and preantral follicles in mice
Cortical strip ex plants show high rates of initiation.
Ultra thin sections show tertiary follicles.
It is probable that a multi stage follicle culture system is necessary for in vitro growth.
In-situ culture»_space;> isolated follicle culture 1»_space;> isolated follicle culture 2»_space;> In-vitro maturation.
In-vitro culture is proving hard though. It is a matter of defining the right signals for each stage of maturation.
Describe how the somatic cells and the oocyte communicate with each other.
There is 2-way communication between the oocyte and the somatic cells that surround it.
One of the ways that the oocytes communicate with the surrounding somatic cells is via trans-zonal processes that project out from the granulosa cells , go through the zona pellucida and interact with the oocyte. This is the main mechanism whereby we have transfer of nutrients and inhibitory signal to keep the oocyte in meiotic arrest. We also have transfer of local factors from the oocyte to the somatic cells to influence their rate of proliferation. These transzonal processes are very important.
