Empty Follicle Syndrome

The Empty Follicle Syndrome (EFS) is a frustrating condition in which no oocytes (eggs) are retrieved at IVF, even though ultrasound and estradiol measurements showed the presence of many potential follicles. The mechanism responsible for EFS remains obscure. Many hypotheses have been put forward, but none truly explain this syndrome. The most likely cause of EFS is ovarian ageing, as many patients who suffer from EFS are also poor responders. If an EFS cycle does occur during your treatment, please make sure you discuss it thoroughly with your fertility physician. EFS is an infrequent event and has been estimated to occur in between 2 - 7% of IVF cycles. However, the overall risk of recurrence in later IVF cycles is 20% and the risk of recurrence is higher as the age of the patient increases.

To date, Empty Follicle Syndrome (EFS) has only been reported in GnRH agonist down-regulated IVF cycles. Some cases have been successfully treated by changing the batch, or by repeating the dose of hCG. A case of EFS was reported recently on PubMed in both GnRH antagonist and GnRH agonist down-regulated IVF cycles when final oocyte maturation was triggered using urinary hCG (u-hCG). Failure to retrieve oocytes occurred, despite administration of a further dose of u-hCG from a different batch and a delayed repeated oocyte recovery performed in the second GnRH agonist down-regulated cycle. A successful oocyte recovery cycle was achieved after triggering of an endogenous gonadotrophin surge using GnRH agonist in an antagonist down-regulated cycle. Nine oocytes were readily retrieved from 10 follicles, at 36 h after GnRH agonist administration, and eight of these fertilized normally. Two good quality embryos were used for fresh transfer and four were cryopreserved for future use. The authors concluded that EFS can occur in GnRH antagonist down-regulated IVF cycles, and can be successfully treated by triggering a natural gonadotrophin surge using GnRH agonist in the absence of any response to previous treatment methods. This represents a novel therapeutic modality for this uncommon but frustrating condition.

Another paper reports experience with three IVF cycles in which no oocytes were collected. In all cases, an additional IVF cycle was performed. The ovarian stimulation protocol, ultrasound and hormonal surveillance methods, human chorionic gonadotrophin timing and oocyte retrieval technique were similar in all patients. The assessment of additional cycles demonstrated a poor response in terms of oocyte quality, since the number of mature oocytes was low despite the high number of oocytes collected. Thus, the data suggest that in these patients, EFS should be considered as a borderline form of poor response to ovarian stimulation. If this is confirmed, EFS should be a recurrent event and an empty cycle could be a good predictor that a subsequent stimulated cycle will be an unfavourable.

A group from UK recently reported five cases in which no oocytes were retrieved after standard ovarian stimulation for in-vitro fertilization (IVF), and in which it was found that mistakes had been made at the time of human chorionic gonadotrophin (HCG) administration. In all five cases, oocyte retrieval was achieved after injecting HCG, when necessary, and reprogramming aspiration 24–36 h later. A mean of 7 ± 3.2 MII oocytes were recovered per patient and 3.2 ± 0.8 embryos were transferred. Three clinical pregnancies were obtained, and four healthy infants were born. In their program, these were the only cases of empty follicle syndrome (EFS) that appeared over a total of 1118 cycles, and were all explained by human error in the administration of HCG. Our experience too shows that human error could be considered a significant factor in the aetiology of empty follicle syndrome, and that EFS may be in part avoided by taking simple preventive measures.

A novel method of rescuing empty follicle syndrome (EFS) was recently published and provides evidence that it is a drug-related problem rather than a clinical dysfunction. In a preliminary study the authors from UK established that in EFS the serum beta-human chorionic gonadotrophin (beta-HCG) concentrations 36 h after HCG administration never exceeded 10 mIU/ ml. beta-HCG concentrations were thus used to confirm EFS when oocytes were not retrieved from one ovary after controlled ovarian hyperstimulation. The procedure was suspended leaving intact all follicles in the second, ovary. After confirmation of EFS, a second HCG from a different batch was administered and 36 h later mature oocytes were retrieved from the intact ovary, suggesting a fault with the HCG previously administered. Three patients have been treated in this way. In the first case, four out of five mature eggs were fertilized after intracytoplasmic sperm injection (ICSI) resulting in the transfer of three top grade (grade 1) embryos. In the second case all seven mature oocytes fertilized after in-vitro fertilization (IVF) and three grade 1 embryos were transferred resulting in a twin pregnancy, now delivered. In the third case, five out of nine oocytes were fertilized after ICSI and one out of the three treated with high insemination concentration IVF fertilized, resulting in the transfer of three ICSI embryos.

We suspected hCG batch-to-batch consistency a couple of times with our own cases, but could never confirm the above. Would love other clinics to post their experiences here.

Weekend Punjabi Puttar Joke

Bill Gates was looking for a new Chairman for Microsoft Europe and
organized an enormous session to interview a suitable candidate.

5000 hopeful candidates assemble in a large room. One candidate is Santa
Singh.

Bill Gates thanks all the candidates for coming and then asks all those who
do not know JAVA to leave.

2000 people leave the room.

Santa says to himself, "I do not know JAVA, but I have nothing to lose if I
stay. I'll give it a try".


Bill Gates asks the candidates who have never had experience of managing
more than 100 people to leave.

Another 2000 people leave the room.

Santa says to himself "I have never managed anybody by myself, but I have
nothing to lose if I stay. What can happen to me?" So he stays.


Then Bill Gates asks candidates who do not have management diplomas to
leave.

500 people leave the room.

Santa says to himself, "I left school at 15, but what have I got to lose?"
So he stays in the room.


Lastly, Bill Gates asks the candidates who do not speak Serbo - Croat to
leave.

498 people leave the room.

Santa says to himself, "I do not speak a singlr word of Serbo - Croat, but
what do I have to lose?"
So he stays and finds himself with one other candidate.


Everyone else has gone. Bill Gates now joins them and says
"Apparently you are the only two candidates who speak Serbo - Croat, so I'd
now like to hear you have a conversation together in that language."

Calmly, Santa turns to the other candidate and says "Hor Paape, ki haal
chaal?"

The other candidate answers "O vadiya veere, tu sunaa". Bolo Ta Ra Ra Ra:)

Tubal Transfer Procedures

The Tubal Embryo Transfer (TET) procedure involves the transfer of embryos that are more advanced in development than those in ZIFT i.e. cleaved embryos. The aim is to achieve fertilization in-vitro (in the laboratory) and then replace the cleaved embryos in the Fallopian tube, the normal site for fertilization. It is suitable only for women who have at least one healthy Fallopian tube. TET is carried out as a day-case procedure under a general anesthetic, using laparoscopy(see picture). You may experience some abdominal discomfort and pain for a few days after the procedure, but painkillers can relieve this. The main groups of patients selected for TET procedure are women who are keen to have gamete intra-Fallopian transfer (GIFT) but where there is doubt about the chance of fertilization, and women who encountered difficulties in previous embryo transfers through the cervix. The success rate for TET is in the region of 25-40% per embryo transfer, but varies depending on many factors such as the woman's age and the number of embryos transferred, etc.

The ZIFT procedure involves the transfer of embryos (zygotes) into the Fallopian tube 24 hours after fertilization. The aim is to achieve fertilization in-vitro (in the laboratory) and then replace the early embryos in the Fallopian tube, the normal site for fertilization. ZIFT offers the best of both IVF and GIFT procedures. It is suitable only for women who have at least one healthy Fallopian tube. ZIFT is carried out as a day-case procedure under a general anesthetic, using laparoscopy. You may experience some abdominal discomfort and pain for a few days after the procedure, but painkillers can relieve this. The main groups of patients selected for ZIFT procedure are women who are keen to have gamete intra-Fallopian transfer (GIFT) but where there is doubt about the chance of fertilization, and women who encountered difficulties in previous embryo transfers through the cervix. If the quality of the embryo's in a preceding IVF cycle was poor, some fertility specialists will advise the patient to have ZIFT rather than IVF.

Gamete Intra-Fallopian Transfer (GIFT) is a procedure in which the eggs are mixed with sperm and the mixture is then injected into the Fallopian tube. It is suitable for women who have at least one healthy Fallopian tube. The aim of GIFT is to allow fertilization to occur in the right place and implantation at the right time. There are selected groups of patients to whom gamete intra-Fallopian transfer (GIFT) is recommended:
a)Couples with unexplained infertility.
b)Women with minimal endometriosis.
c)Men with infertility problem (not severe).
d)Couples who have had failed donor insemination.
e)Where transcervical embryo transfer is impossible and the patient has one healthy tube.

The procedure is carried out as a day case surgery under a general anaesthetic, and involves making three small cuts in the abdomen. Fertility drugs are used to stimulate the ovaries, monitoring is also carried out as with IVF treatment. Eggs may be collected either by a vaginal ultrasound scan or laparoscopically using a fine needle and gentle suction.Once the eggs are collected, they are examined under the microscope for quality. The best eggs are then mixed with washed and prepared sperm in a small volume of culture medium. The end of the fallopian tube is grasped gently and a guided fine canula is passed through the fimbrial end of the tube(see picture). The mixture of the eggs and sperm is then deposited into the Fallopian tube. In most cases, three or four eggs are usually transferred per cycle. Any spare eggs that are collected may be fertilized in-vitro and the resulting embryos are frozen (if they are of good quality to freeze) for later embryo replacement. After the GIFT procedure, the patient is usually given hormonal supplements, in the form of tablets, pessaries, gel or injection to help the implantation of embryos.The success rate after GIFT is similar to that of in-vitro fertilization (IVF) treatment. Some specialists claim a higher success rate with GIFT compared to IVF.

More recently, attempts have been made to simplify the procedure by collecting the eggs by vaginal ultrasound and injecting the egg and sperm mixture into the tube via the cervix using a special catheter. The procedure is usually performed under ultrasound guidance. The reported pregnancy rates with this method are less than that with conventional laparoscopy GIFT.

Last week, we did a big Tubal Transfer Procedures Workshop at the Prince Aly Khan Hospital IVF Center (www.testtubebabyclinic.com) where we performed the above procedures on 9 patients. We have made a teaching DVD of the same to teach post-graduates which will be released at the World Congress on Fallopian Tubes to be held at Kolkata from 18-21 October 2007. You can access the meeting homepage on www.sisab.net/tubes2007

Frozen Embryo Transfer

Embryo freezing is a well-established form of assisted conception treatment. An increasing number of IVF clinics worldwide are now able to freeze spare embryos for later transfer. The first frozen embryo baby was born in 1984. Embryo cryopreservation allows multiple embryo transfers from a single egg collection and improves the chances of livebirth. Amongst the advantages of embryo cryopreservation are maximizing the potential for conception for IVF and prevent wastage of viable normal spare embryos. Perhaps this is the most important advantage of cryopreservation. Approximately 50% of women may have spare embryos available for freezing. In some clinics, the pregnancy and live birth rates with frozen-thawed embryo transfer is as high as those achieved with fresh embryo transfer. Freezing all embryos for subsequent transfer may be advised for women who are at a high risk of developing severe ovarian hyperstimulation syndrome following ovarian stimulation for in-vitro fertilization (IVF).

Embryo Freezing is also resorted to when embryo implantation may be compromised in cases such as the presence of endometrial polyps, poor endometrial development, break through bleeding near the time of embryo transfer or illness as also difficulty encountered at fresh embryo transfer e.g. cervical stenosis (inability to pass through the cervical canal because the cervix is narrowed or scarred, etc). Cryopreservation of embryos is very important to be incorporated in the egg donation programs. It is not always possible to synchronize the recipient’s cycle with that of the egg donor. In some countries, it is mandatory to freeze all embryos created from donated eggs, quarantined for a period of six months and until the donor have a repeat negative screening tests. As a result of successful cryopreservation programs, frozen embryos have also become available for donation to infertile couples.

Embryos can be frozen at any stage (pronucleate, early cleaved and blastocyst) if they are of good quality. Embryos are stored in batches of one or more embryos depending on the number of embryos that are likely to be transferred into the uterus at a later date. Embryos are mixed with a cryoprotectant fluid (to protect embryos from damage during freezing process). Then, the mixture is put either in a plastic straw or a glass ampoule and stored in liquid nitrogen at a very low temperature –196°C using a specialized programmable machine. Thawing of embryo involves removing the embryos from the liquid nitrogen, thaw at room temperature, remove the cryoprotectant fluid and mix the embryo in a special culture media. The mixture is then kept in the incubator ready for transfer.

If the embryos were frozen at cleaved stage or blastocyst, they can be thawed and replaced in the same day. However, if they were frozen at the two-pronucleate stage, then they are thawed on the day before and cultured overnight to allow them to divide and are replaced when they become 2-4 cell embryo (s). Different countries have different regulation concerning the length of time that embryos can be frozen. In the United Kingdom, embryos can be stored for a maximum of 10 years. India is yet to define the upper limit of storage.

The first step to transfer embryos from one centre to another is to write to the centre which has the couple's embryos stored in order to release the embryos. Consent from both partners is required. The couple will need to take responsibility of the embryos once they leave the center. The embryos are transferred using a small liquid nitrogen container specially designed.
Not all embryos survive the freezing and thawing process. In a good freezing program, a survival rate of 75-80% should be expected. Therefore, it may be necessary to thaw out several embryos to get two or three good embryos to replace. Damage of embryos does occur as a result of freezing, not during the storage but during the cooling and thawing process. It is important that both the couple and the clinic to keep in contact with regard their frozen embryos. Most IVF clinics will send an annual reminder letter to patients who have frozen embryos stored.

The letter usually offer a couple five choices:

Continue storage for a further period of time.
Arrange for embryo transfer treatment cycle before the expiry date.
Allow embryos to perish by interfering with the thawing process.
Donate embryos to ethically approved research.
Donate embryos to infertile couple.

Frozen/thawed embryos may be transferred into the uterus in a natural cycle, a hormone replacement cycle or a stimulated cycle. In general, the three methods have similar pregnancy and live birth rates. A Natural cycle is usually recommended in young women with regular menstrual cycles and ovulation. It involves serial ultrasound scans to check the development of the follicle and endometrium, blood tests to check the levels of hormone LH, estrogen and progesterone. Embryo transfer is usually performed about 3-4 days after the LH surge (2-3 days after ovulation). The woman is given no drugs until the day of embryo transfer. On the day of embryo transfer, the woman may start a course of progesterone pessaries or tablets to support the luteal phase. Natural cycles have the advantages of a naturally prepared endometrium and reduced cost. The disadvantages of natural cycle frozen embryo transfer is the risk of failure of ovulation. Also, the date of ovulation can not be predicted.

Hormone replacement cycle with or without GnRh agonist is usually recommended for older women, woman without ovaries or non-functioning ovaries, women with irregular infrequent menstrual cycles or ovulation. It involves giving estrogen in the form of tablets or skin batches and later adds progesterone in the form of tablets, pessaries, gel or injection. Different IVF clinics have different protocols for giving these medications and in some women GnRh agonists may be given in addition to hormone replacement to "switch off" any hormone production by the ovaries which may interfere with the treatment. After embryo transfer, both estrogen and progesterone are continued until the pregnancy test. In the test is positive, the woman should continue the medication for a further 8-10 weeks. Hormone replacement cycle allows accurate programng the date of embryo transfer and ensures that the endometrium is adequately prepared to receive the embryos.

Stimulated cycle is where fertility drugs such as clomifene tablets or FSH injection is given aiming to produce one or two follicles. When the follicle is mature and the endometrium developed satisfactorily, hCG injection is given to induce ovulation. Embryo transfer is usually performed 2-3 days after the ovulation. This regimen is usually recommended for women do not ovulate regularly and did not respond to hormone replacement treatment in a previous cycle.

The success rates depend on many factors; mainly the woman’s age and number of embryos transferred. The outcome of pregnancies resulted from frozen embryo transfer is similar to fresh embryo transfer in the incidence of biochemical pregnancy, blighted ovum, early and late miscarriage, ectopic pregnancy, preterm deliveries and term deliveries. To date there is no evidence that babies born after frozen embryo transfer have any increased incidence of congenital abnormality.

There are several ethical and moral issues surrounding the embryo freezing process. These include the following:

Fate of the stored embryos on the death of couple - 'orphaned' embryos.
Ownership of the embryos if the couple divorce.
Safety of embryo freezing.
Concern that the length of time embryos have been kept in storage might have a detrimental effect on the outcome of frozen embryo transfer and possible increase in fetal abnormalities. However, no long-term studies have been carried out since the age of the oldest child born as a result of frozen embryo transfer is only 14 years. In addition, there is no evidence that extended storage is detrimental to the outcome of treatment.

Trans-rectal Electro-ejaculation (TREE)

Trans-rectal electroejaculation (TREE) technique is used to collect semen from men who are unable to ejaculate. The technique has been used in animals since 1930s and in man since 1948 onwards. The aim of trans-rectal electroejaculation (TREE) is to directly stimulate the seminal vesicles. There are selected groups of patients for whom trans-rectal electroejaculation (TREE) is recommended. The majority of spinal cord injuries are among young men. About 5% of these men are likely to achieve pregnancies with their partners without assistance. The loss of fertility in these men is due to a decline of sperm quality, impotence and ejaculatory failure. About 50% of them may able to achieve ejaculation with the use of penile vibrators. Men who are unable to ejaculate by other means, should be offered rectal electroejaculation. Sexual dysfunction is common in insulin dependent diabetics. Trans-rectal electroejaculation (TREE) may be advised for men who are unable to achieve erection and ejaculation. Men with multiple sclerosis who are unable to achieve erection and ejaculation will also benefit from TREE. Men who are impotent and or unable to ejaculate because of psychosexual problems may be offered rectal electroejaculation after failed psychosexual treatment.

The procedure is performed as a day case surgery usually under a short general anesthesia. However, in men with spinal cord injury (SCI) it may be performed without anesthetics (although about 5% of these men may require a general anesthesia. The procedure takes about 15 minutes. The man usually lies on his right side. A metal or plastic tube (proctoscope) is lubricated and inserted into his rectum to inspect the lining of the rectum (see picture). A special probe is inserted into the rectum, the probe is attached to an electric stimulation power unit through which electric stimulation is delivered(see picture). The power is increased gradually until ejaculation occurs. At the same time an assistant will apply massage to the prostate gland and milk the urethra to obtain the ejaculate. At the end of the procedure, another proctoscope is inserted to check for any burn injury in the rectum caused by the hot rectal probe. Like any surgical procedure there are a few potential problems. Autonomic hypereflexia (very high blood pressure) is a potentially severe complication to any spinal cord injured man. Because the normal defenses which protect against a sustained high blood pressure during orgasm are lost. This can have serious consequences such as brain hemorrhage. For this reason, SCI men are usually given pills to lower their blood pressure before they undergo trans-rectal electroejaculation (TREE). Other problems that may occur include burn injury and complications associated with the general anesthesia.

After sperm preparation the fresh sperm is then used for treatment such as insemination, IVF or ICSI depending upon the sperm count and quality. Alternatively the semen sample is frozen for future use. Treatment options will depend upon the sperm count and quality. Other factors such as the fertility of the female partner and the duration of infertility, etc. will also influence the treatment options available. The options include the following:

Intrauterine insemination (IUI)

Intrauterine insemination (IUI) with washed and prepared sperm. This will be advised if the sperm problem is mild and the female partner is young, has regular menstrual cycles and ovulation, and has healthy patent Fallopian tubes. A pregnancy rate of 5-10% per treatment cycle is expected, depending on many factors such as the woman’s age and treatment cycle number, etc.

IVF (In Vitro Fertilization)

In-vitro fertilization (IVF) is usually advised if the female partner is 35 years or older, has blocked or damaged Fallopian tubes or severe endometriosis and the male partners sperm count or quality is not suitable for IUI.

ICSI (Intra-cytoplasmic Sperm Injection)

Intracytoplasmic sperm injection (ICSI) is advised when there is a severe sperm problem, i.e. sperm is not suitable for standard IVF treatment.

Recent advances in assisted conception such as IVF and ICSI have made it possible to achieve pregnancies and live births with ejaculate containing just a few sperm.

All about hCG testing

The American College of Obstetricians and Gynecologists (ACOG) has issued advice on recognizing and avoiding false-positive human chorionic gonadotropin (hCG) test results. In its latest Committee Opinion report, the college acknowledges that "clinically significant false positive hCG test results are rare," citing one study in which five out of 162 women studied had evidence of serum assay interference sufficient to provide misleading results. But the college points out that such results, if unrecognized, can lead to significant and unnecessary clinical intervention.

Reviewing the causes of false-positive hCG results, the ACOG reports that some individuals have circulating factors in their serum that interact with the hCG antibody. The most common of these are heterophilic antibodies—human antibodies that can target animal-derived antigens used in immunoassays. People who have worked as animal laboratory technicians or in veterinary facilities, or who have grown up on farms, are more likely than others to have developed heterophilic antibodies, the college says.

What is a misleading serum test result? The ACOG states: "If results are misleading, they usually are seen with values below 1,000 mIU/mL." To rule out the presence of heterophilic antibodies or other interfering substances, it lists four methods:
1.A urine test: heterophilic antibodies are not present in urine.
2.Rerunning the assay with serial dilutions of serum: lack of linearity confirms assay interference.
3.Preabsorbing serum: some laboratories can pre-absorb serum to remove heterophilic antibodies before the assay is performed.
4.Using another assay: some assays may recognize the aberrant, non-active forms of hCG that some individuals naturally produce. "Repeating the hCG measurement in a different assay system can best detect this problem," says the college.

Concluding, it states: "Caution should be exercised whenever clinical findings and laboratory results are discordant. Although false-positive hCG results are rare, if unrecognized they may lead to unwarranted clinical interventions for conditions such as persistent trophoblastic disease. The physician must judge whether the risks of waiting for confirmation of results outweighs the risks of failing to take immediate action."

Venter Sci-Fi

Craig Venter, the scientist who controversially commercialized the efforts of the Human Genome Project, is claiming to have constructed a synthetic chromosome with his research team, giving rise to an artificial life form. Mr. Venter's institute is expected to make an announcement concerning the development in the coming weeks. If true, the potential applications are simply awe-inspiring. Organisms could be "created" for a plethora of uses. Potential applications include, but are not limited to; medical treatments, environment protection, energy production and, unfortunately, biological weapons of vast sophistication. Speaking about the research Mr. Venter said:

"[this is] a very important philosophical step in the history of our species. We are going from reading our genetic code to the ability to write it. That gives us the hypothetical ability to do things never contemplated before".

The team was able to construct an artificial chromosome of 381 genes and the DNA sequence they have pieced together is based upon the bacterium Mycoplasma genitalium (see picture). The original bacterium had a fifth of its DNA removed and was able to live successfully with the synthetic chromosome in place. The man-made organism, dubbed Mycoplasma laboratorium, still relied upon the intracellular machinery already present to carry out tasks such as replication and metabolism, and in this respect the entity is not an entirely new life form. Though the potential may be phenomenal, at the moment all there is to go on is a scientist's word, and in science facts are the only language of proof. We will be waiting for an official announcement with baited breath.