In Vitro Fertilization

In Vitro Fertilization David Stanford, M.D.

1

I.

Assisted Reproductive Technologies A.

ARTs involve many variations of the same theme 1. Terms related to ARTs a.

In vitro fertilization (IVF)

b.

Gamete intrafallopian transfer (GIFT)

c.

Pronuclear intrafallopian transfer (PROST)

d.

Zygote intrafallopian transfer (ZIFT)

e.

TEST

f.

Gametes: sperm and oocytes (eggs)

g.

Zygote: fertilized oocyte

h.

Embryo: any fertilized oocyte, but prior to 14 day, sometimes referred to as "preembryo"

2.

The normal sequence of events from ovulation through division a.

Once fertilization has taken place, the genetic material from both parents form visible circles within the oocyte's cytoplasm, which are called pronuclei

b.

This precedes the alignment of genetic material and the first cell division

c.

The disappearance of the pronuclei signals that chromosomal alignment has begun; this is termed syngamy

e.

The pronuclei become visible somewhere around 16 hours after the oocytes are inseminated

f.

The first cell division occurs about 24 hours after insemination and further cell divisions occur about every 12 hours

3.

Any fertilized oocyte is technically an embryo, but, prior to 14 days, they are sometimes referred to as preembryos; fertilized oocytes are also known as "zygotes"

C. Variations of ART 1.

IVF involves a.

Oocyte retrieval

b.

Extracorporeal fertilization

c.

Placement of two-to-six-cell embryos in the uterine cavity 48-54 hours after oocyte retrieval

2.

GIFT involves a.

Oocyte retrieval

b.

Placement of sperm and oocytes into the fallopian tubes

3.

PROST involves a.

Our oldest patient to carry to term was 41.8 years old. I don’t know where these women are coming from. We are not seeing them. As a woman starts out young menstruating the cycle length decreases. Just before she goes into that totally can’t get pregnant stage, there is a decrease in cycle length. Historically, a woman who at a younger age a cycle length of 28 to 30 days who is now 35 or older has a cycle length of 25 to 26 days, it is beginning to get into that stage where she is probably has an age-related oocyte factor. That is a really easy clinical piece of history. If you talk about age-related infertility this is are our data for our donor oocyte recipients. We periodically we have somebody ask us who is like 52 or 54 whether we will do a donor. I don’t know the answer to that. We do it on a case by case basis because there is no real hard fact answers on that, but these woman will get pregnant. From an obstetric standpoint the most important thing is that we do our job in screening. When you look at this woman because they are older the major risk obstetrically has medical complications. There is a higher incidence of gestational diabetes. There is a higher incidence of hypertension, and those will obviously adversely affect the pregnancy outcome, but they are not because of the fact that they are pregnant. Most of this woman has healthy kids, so they can conceive and deliver even though they are in there 50’s. Every so often you will have somebody who is even in there 60’s. They do need to be monitored as you probably would most of your high-risk for gestational diabetes and gestational hypertension. The kids go far now that they do well. It is tough to get to eggs. We have advertisements out nation wide all the time to recruit people. You cannot pay woman for doing it. You can only pay them for their time. They make about $2000 because that is the time requirements we have for them. Waiting lists are from anywhere of 6 months to a year to get a decent match. People try to look at other ways of obtaining oocytes. It would be great if somebody could learn how to freeze eggs, but at the present time you can’t freeze eggs. I just mentioned that because we’ve had cloning recently as one of the scariest things I’ve heard. I honestly didn’t believe you couldn’t clone a human adult tissue because it is too differentiated.

These fertilized oocytes with visible pronuclei are called pronuclear embryos

d.

Two things, which dramatically influence pregnancy rates are, age and number of embryos transferred.

Oocyte retrieval

The pregnancy rates go up dramatically based upon the number of embryos that you transfer and then it goes down again. That is because usually these patients either that is all they had, or those are your PCO patients. We know that although PCO patients can be pregnant with IVF they tend to have somewhat lower pregnancy rates. There are some PCO patients for reasons unknown to anybody that have problems with recurrent miscarriages. We can’t figure out how to fix it. The problem with it is the more embryos you transfer the higher the triplet rates, so it is always a trade off. The largest cost is from the complications of multi-gestation. They can get 100’s of cycles of IVF for one bad triplet or quadruplet pregnancy. In this country we are still financially driven in capital systems that people want to get pregnant. If I know I get more people pregnant by putting more embryos then that is the temptation. However we start coming out with recommendations that we limit that number of embryos in this country to three or four. The triplet rate will be at about 5%, and there are some programs that are running triplets rates up to 30%. I think what we are looking to here is to start using frozen embryos more and more. They can be kept up to 2,012 years. They certainly can be kept for longer than 10 years because we have some. We switched the stage of which we freeze them. Some of our cycles we sent to do a freeze. If we have a hyperstimulation patient who has lot of eggs we will freeze them all to avoid hyperstimulation. If you get 16 eggs you will probably get 10 to 12 good embryos and of that we like to transfer 4 to 6. The average number that was frozen was eight. During the same time period we thawed 162. Some of these embryos had been frozen years ago and some of them had only been frozen recently. Their time of freezing was quite variable. If we have had a chance to freeze them in our new facility, and we’ve done the freeze, the thaw, and the transfer we are running at about 40%. Right now we are having trouble showing a difference between our fresh and our frozen transfer rate. It might be if that is true that we will go to frozen embryo transfers and limiting it to two because there is no symbiotic effect. It isn’t like if I put four back two of those are going to help the other two survive. Each embryo has its own individual rate of being able to survive. If only put two back you will have a lower overall success. It costs $3000 to have an embryo transfer. It is not that cheap. We are stating immature human oocyte. You have to take them out before they have selected the dominant follicle, and that is usually somewhere around day 10. You learn what she does, and you bring her in the next month. If you have a decent ultrasound machine you can see these. They get to be about 8-12 mm. Once it is more than a 12mm follicle it is dominant and you can’t use it anymore. You can usually get anywhere from four to eight eggs. The problem is these eggs are still very adherent in the follicle. If you think about is when a follicle matures just before ovulation you have this egg sitting on this little pedestal. The egg sort of floats free right before she gets ready to ovulate. If you are going to stick a needle in there it is really easy to get the egg out. Regular IVF is easy. Now you are doing this at a time when this egg is still plastered into the granulosa. Instead of using a sharp needle use a blunt needle and

2

4.

b.

Extracorporeal fertilization

c.

Transfer of pronuclear embryos to the fallopian tubes

IF/TEST involves a.

Oocyte retrieval

b.

Extracorporeal fertilization

c.

Transfer of two-to-six-cell embryos (zygotes) into the fallopian tubes

The last thing that we are starting to do now has to do with colorful Doppler’s. There is some preliminary evidence to suggest that the blood flow can be related functionally to the quality of eggs or to the quality of endometrium.

D. IVF 1.

Ovulation induction

2.

Oocyte retrieval

essentially do suction D and C. The retrieval are more difficult in terms of you may not get the eggs. We are using cold culture. You just wait for them to kick out their first polar body and then you know that they have gone to that stage. When they have kicked out their first polar body you can see this under phase contrast. You supplement it with estrogen and progesterone. If we can even get a 20% pregnancy rate that would be great because there is no drugs. If you have taken away that you have take away $3000 per cycle of the cost. For people who always hyperstimulate you have taken away the stimulation. Those are the two groups that we targeted. Oocyte donors this would be ideal for. She is not exposed to any drugs. It reduces the cost. You have really taken away most of the major complications and injections to doing donor oocytes. That is the reason why people want to do it. Whether it works or not remains to be seen.

a.

Ultrasound-guided

b.

Laparoscopically-guided

3.

Fertilization in the laboratory

4.

Embryo transfer intrauterine

E. GIFT 1.

Ovulation induction

2.

Oocyte retrieval must be done laparoscopically

3.

Oocytes and sperm placed intratubal; fertilization not documented

F.

ZIFT/PROST 1.

Ovulation induction

2.

Oocyte retrieval done by ultrasound

3.

Fertilization done in the laboratory

4.

Embryo transfer; requires laparoscopy a.

PROST: pronuclear embryo transferred intratubal

b.

ZIFT: two-to-six-cell embryo transferred intratubal

Table 1 Assisted Reproductive Technologies IVF

GIFT

ZIFT

Ovulation induction

+

+

+

Ultrasound-guided retrieval

+

-

+

Laparoscopy

+

+

+ = (?)

Embryo transfer

+ = IU

-

+ = IT

Requires normal tubes

-

+

+

Assesses fertilization

+

-

+

(?) = laparoscopy only if fertilization occurs IU = intrauterine IT = intratubal ZIFT is the same as PROST

Table 2 Indications for ART IVF

GIFT

ZIFT

3

Disease

+

-

Tubal disease

+

+ (*)

+ (*)

Endometriosis

+

+

+

Polycystic ovarian disease

+

+

+

Perimenopausal ovulation dysfunction -

-

-

Antisperm antibodies

+

(?) -

+

Proven fertility

+

+

+

No proven fertility

+

-

+

Azoospermia

-

-

Etiology unknown

+

+ (#)

Male factor: + motile sperm

+

* = must have normal fallopian tubes # = GIFT will not document fertilization, because some unexplained fertility is due to a lack of fertilization, GIFT may not be the method of choice

II.

Pregnancy Rates A.

The concept of cumulative pregnancy rates and monthly fecundity are important to evaluate the relative success of any therapy 1.

Monthly fecundity is simple a.

Definition: the number of women conceiving in a given month is divided by the number of women trying to conceive in a month

4