ART

Question 1

Define infertility

Answer 1

Infertility defined as the failure to conceive after 1 year of regular unprotected intercourse.

Question 2

Describe the causes of the infertility

Answer 2

• Mechanical blockage to egg and sperm meeting
• Failure of gamete production or release
• Failure of fertilisation/implantation & miscarraige
• Unknown/unexplained

Question 3

Mechanical blockage to egg and sperm meeting- examples

Answer 3

Infection/occlusion of vas deferens or uterine tubes.
- Previous ligation for sterilisation.
- Endometriosis. (Endometriosis in the uterine tubes will block them. Chlamydia can also cause blockage of the uterine tubes)
- Congenital defects (congenital absence of the vas deferens)

Question 4

Failure of gamete production or release- examples

Answer 4

Anovulation, maternal age, PCOS.

Azoospermia, asthenozoospermia, teratozoospermia.

Question 5

Failure of fertilisation/implantation & miscarraige

Answer 5

Genetic factors.
Endometrial receptivity, maternal age.

Question 6

What is vasectomy?

Answer 6

Ligation of the vas deferens

Question 7

What is endometrioisis?

Answer 7

cell from the lining of the uterus find their way into other places in the pelvis. They will implant and respond the high E2 levels and proliferate.

Question 8

Process of ART- assisted reproductive technology?

Answer 8

Start by Inducing ovulation with exogenous hormones.
By-passing the uterine tube (IVF).
Can make the egg and the sperm meet in a dish
Direct collection of sperm from the testis/epididymis.
Aspirate sperm
Direct insertion of the sperm into the egg (ICSI).
Donor gametes.
Donor eggs and donor sperm
Usually is a combination of the above.

Question 9

What are the ways of inducing ovultion?

Answer 9

• Administering gonadotrophins (hormones)
• Removing negative feedback

Question 10

Exogenous gonadotrophins are used to do what?

Answer 10

Used to treat women who are anovulatory or who have oligo/amenorrhoea.
The aim is to induce single dominant follicle.
Daily injections…monitor by ultrasound during the cycle.

Question 11

How do exogenous gonadotrophins induce ovulation?

Answer 11

In the normal menstrual cycle there are oocytes that are recruited into the cycle by the inter cycle rise in FHS.
As they start to grow they produce estrogen. As they produce estrogen, there is negative feedback on the hypothalamus and pituitary, reducing FSH.
The reduction in FSH causes most of the oocytes to undergo apoptosis as they are FSH dependant. But the most advanced which has the most FSH receptors and started to induce LH receptors survives the fall in FSH and will go onto become the dominant follicle.
BUT sometimes the follicles are not producing enough estrogen to cause ovulation. If you were to administer exogenous gonadotrophins (mainly FSH) you can kickstart the follicle development, causing the selection of the dominant follicle.
You need to monitor this by ultrasound as if you give too much FSH you don’t get the dominant follicle selection as FSH will remain high.
Can be made via recombinant technology but also by collection from old woman’s urine and purifying. Post menopausal women have more FSH in their urine as they have low estrogen levels, so less negative feedback.

Question 12

Why does removing negative feedback do?

Answer 12

Normally at the end of the cycle there is the death of CL and progesterone levels fall. This fall in progesterone levels results in a rise in FSH, starting the new cycle.
So if you remove the feedback estrogen is placing on the cycle you will get a natural rise in FSH

Gonadotrophin levels may be normal, but are not cyclical.
Inter-cycle rise in FSH relies on death of the corpus luteum.
ie. fall in levels of progesterone and estradiol.
There is no corpus luteum in the absence of ovulation
Cannot reduce progesterone as there has not been a corpus luteum to make any.
There are follicles in the ovary making estradiol so we can remove the negative feedback of this.

Question 13

In what situations would it be good to induce ovulation by removing negative feedback?

Answer 13

You have a bit of FSH production, but not enough to stimulate the gonads
A few follicles producing estrogen, but not enough to cause ovulation.

So if you can remove the feedback from estrogen, we will release the negative feedback on the hypothalamus and pituitary and endogenous FHS will therefore rise.

Question 14

What are the 2 ways to remove oestrogen feedback and what drugs are used?

Answer 14

1) Block the E2 receptor on the pituitary gonadotroph cells with SERM (selective estrogen receptor modulator) : Clomid/Clomiphene.

2) Stop E2 being made by the gonads using an aromatase inhibitor.
Drugs ending in ‘zole’ eg. Letrozole.
Testosterone -> (via aromatase) Estradiol

Question 15

Outline the IVF cycle

Answer 15

Hypothalamic-pituitary down regulation (GnRH)
Administer exogenous GnRH
To prevent an LH surge and loss of follicles (premature ovulation)
Ovarian stimulation (monitoring follicles)
To obtain as many follicles as possible
hCG trigger
Oocyte retrieval
Fertilisation in vitro
Mix with sperm in a dish
Embryo culture 3 – 5 days
Embryo or Blastocyst Transfer
Pregnancy confirmation
Luteal phase support - Cyclogest (progesterone)

As failure will occur at each stage, we require as many eggs as possible and so hyper-stimulate the ovaries to increase follicle numbers.

Question 16

Outline the IVF cycle

Answer 16

At the beginning of the cycle the CL has died, progesterone levels fall, this has removed the negative feedback on the pituitary and hypothalamus.
There is an intercycle rise of FSH
In the ovaries there will normally be two dozen small antral follicles. These are follicles that 3 months ago left the primordial pool and started to grow independent of LH and FSH.
When they get to the small antral stage, and start to have a fluid filled space inside, they start to express FSH receptors.
Then as soon as the next menstrual cycle starts and FSH rises, they all get recruited into the cycle and start growing.
As they grow they produce estrogen, this estrogen will act via negative feedback to reduce FSH.
The lack of FSH will kill most, but the most advanced one will have the most FSH receptors and also have some LH receptors, it will become the dominant follicle.
It will continue to produce estrogen and this will have a negative affect on FSH, which will lead to atresia.

Question 17

How is a single follicle selected in each cycle- IVF?

Answer 17

IVF is different. Exogenous FSH is given every day
Endogenous FSH is switched off with GnRH – it is given at a constant level, shutting down the axis.
No matter how much estrogen the follicles produce, it has no effect as we are giving FSH exogenously. So all the follicles are selected, we want 10-15. Any more is dangerous. This is monitored via ultrasound.

Question 18

Why do we down-regulate HPG axis using GnRH antagonist or agonist?

Answer 18

prevent an LH surge

Question 19

Downregulate Hypothamo-pituitary gonadal axis using GnRH antagonist or agonist.
Prevents the LH spike and potential loss of the follicles- what is the significance of this?

Answer 19

LH surge causes resumption of meiosis 1 and initiation of meiosis 2.

Question 20

Why do we require a lot of eggs?

Answer 20

As failure will occur at each stage, we require as many eggs as possible and so hyper-stimulate the ovaries to increase follicle numbers.

Question 21

Give FSH by subcutaneous injection every day. Growth of multiple follicles.
Monitor follicle growth with ultrasound until most follicles 12–19mm.
At this point hCG trigger given (GnRH agonist or Kisspeptin may be used).
Why is LH not used?

Answer 21

LH is not used as it has a short half life, hCG is used as it binds to the LH receptor
Will cause meiosis 1 resumption and initiation of meiosis 2

Question 22

Timing of ovarian stimulation?

Answer 22

36 hours allowed for completion of meiosis I and initiation of meiosis II before egg collection.
There is a window of opportunity where the eggs are fertile but they WONT be ovulated, so we can go in and collect them

Question 23

Describe the process of oocyte retrieval

Answer 23

Transvaginal needle goes through the wall of the vagina.
Under ultrasound guidance with a trans vaginal ultrasound probe, we can take the needle right to the follicle and aspirate the follicular fluid inside, and the egg will come out with it.

Collect mature eggs from ovary using transvaginal needle aspiration under ultrasound guidance.

Question 24

IVF vs IUI

Question 25

Describe the sperm retrieval

Answer 25

There is a density gradient fluid that is denser at the bottom than top
The semen is then taken and then layered on top of the density gradient.
It is then centrifuged in a process called density centrifugation.
The most mature sperm are dense and will be at the bottom of the tube.
The dead sperm are less dense are therefore higher up.

In IVF there are a series of droplets of media underneath mineral oil, where the sperm and the eggs will be mixed.

Question 26

Process of IVF- protocol

Answer 26

Can see egg with cumulus and lots of sperm around it, ready to be fertilised.

Control factors such as nutrients, acidity, humidity, temperature, gas composition of air, and exposure to light.

The sperm and the egg are incubated together at a ratio of about 75,000:1.
Duration of this co-incubation traditionally 16 – 18 hours . Approximately 65% of the eggs will fertilize.

Question 27

Embryo Culture

Answer 27

When the egg divides in meiosis 1 it divides asymmetrically
The egg contains all the cellular machinery for the early embryo, the sperm just brings DNA.
Therefore it does divide in half, there is production of a polar body.
After meiosis 1, there is 1 polar body visible.
After meiosis 2 which happens at fertilisation, there is a second polar body.
So 2 polar bodies = 2 meiotic divisions

Can see two pro nuclei. One has all the male DNA in it one has all the female DNA in it.
Thy move towards each other and fuse = diploid -> mitosis

After 3 days you see 6-8 cells
At 5 days = blastocyst stage = around 100 cells
This has a better implantation rate.

Question 28

How is the embryo transferred?

Answer 28

Embryo transferred to the patient’s uterus through catheter, which goes through the vagina and cervix into the uterus, usually under ultrasound guidance.

Single embryo transfer is the norm in order to avoid multiple pregnancies, though 2 – 3 may be transferred in women over 40 or who have had repeated implantation failure.

Question 29

Success rate

Answer 29

Around 6.5 million children born worldwide to date with IVF.
As you get older it is less successful.

Question 30

Intracytoplasmic sperm injection - ICSI

Answer 30

Microscope has two arms, on the ends are two ultra fine pipettes. These are connected to two controllers to give movement of the two pipettes and suck and blow.
Used in low sperm count, low motility or repeated fertilisation failure.
Single sperm used so can collect sperm by needle aspiration from epididymis or testis.
Inject sperm into the egg. The sperm may not even be swimming

The egg is being held with a holding pipette, and one sperm is injected into the egg

Question 31

Is ICSI safe?

Answer 31

The fertilisation rate is 90%
Natural means of sperm selection is bypassed.
Some evidence of increased genetic damage, but equivocal.
Other defects 9.9% compared with 5% of non-ICSI.
Patients may be being pushed to ICSI as higher fertilisation rates. In 2013 there were more ICSI cycles than IVF for the first time.
Biggest risk with infertility treatment is still multiple pregnancy.

Question 32

Gamete donation

Answer 32

Sperm
Very common…..who for?
Same sex couple
Asospermic males
Freezing (cryopreservation) is essential for donor sperm…why?
As it needs to be quarantined for 6m to make sure that the donor has no diseases.
Reasonable function after thawing.
Change in the law may reduce the number of UK donors.

Eggs
Who for?
Women who have PCOS, fertility problems
World-wide shortage of donor eggs.
Need to go through an IVF cycle to access eggs.
Eggs can now be cryopreserved by vitrification. (frozen)
Can be a waiting list/expensive.

Question 33

Donor identity and anonymity

Answer 33

Since 2005 donors also have right to access information about themselves held by the HFEA.

1) Whether their donation has been successful.
2) The number of children born as a result of their donation
3) The sex and year of birth of any children born.

Children born from donations in the UK have the right to ask the donors identity once they are over 18.

Question 34

HFEA

Answer 34

All in vitro assisted reproductive procedures in UK require licence from Human Fertilisation and Embryology Authority.
Paid for by fee added to each procedure and each procedure is documented and reported.
Premises/staff/procedures and paperwork all inspected at short notice.
League tables of success rates are published. How reliable are these?

Question 35

What is vitrification?

Question 36

Access to infertility treatment?

Answer 36

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