Science Pulse    Controversy of Immunology

It is easy for patients to scapegoat the body's immune system when apparently healthy looking embryos fail to implant, or a pregnancy is thwarted for unknown reasons. Autoimmune factors related to recurrent pregnancy loss (RPL) have been fairly well studied, resulting in treatment methods that are relatively standard.

On the other hand, implicating the immune system for repeated IVF failures represents an area of medicine that can be subject to abuse. Practitioners of reproductive endocrinology in the U.S. as well as Great Britain consider this one of the more controversial topics in their field.

There is no shortage of data analyzing the role of immunology in reproductive success or failure. A number of comprehensive studies in the mid- to late1990s were undertaken to identify a potential cause and effect relationship between abnormal immune test results associated with reproductive failure. Yet the tests reached the same conclusions; the most rigorous studies failed to provide a correlation.

Today many years after most reproductive endocrinologists might have thought the topic was put to rest, women with greater research capabilities on the internet who actively seek answers for their fertility problems continue to come across offers of clinical immunological investigations and treatments that lack true scientific basis. As pointed out by the Royal College of Obstetricians and Gynecologists, "Praying to Artemis of Ephesus, a goddess associated with fertility, might be as useful as undergoing some of the fertility tests offered on the Internet".

The market for potential abuse is considerable, given that nearly 15% of American couples suffer from infertility, 10% of whom suffer from unexplained infertility. Additionally 2% of childbearing women may experience recurrent pregnancy loss or RPL, (generally defined as three or more consecutive pregnancy losses before 20 weeks gestation) and as many as 60% of such RPL will demonstrate no cause. Despite the costs, the lack of scientific evidence and the majority of skeptical practicing reproductive endocrinologists, people nevertheless seek treatment for purported immune imbalances.

Many such patients might be genuinely mixed up, finding it difficult to distinguish potential immunological causes of RPL from failed implantation following IVF procedures. Indeed, there is some evidence that RPL can occur as a result of an imbalance of some immune factors. But RPL is very different than a failed pregnancy at the implantation stage. If patients are given the impression that studies will support unproven treatments, it is understandable that frustrated patients may agree to experimental treatments.

What is clear, however, among the majority of practicing reproductive endocrinologists is the myriad of studies conducted in the 1990s demonstrated a sound scientific approach to the question, and no causal relationship was found. The American Society for Reproductive Medicine (ASRM) also summarized the literature and published an opinion paper concluded scientific evidence is not sufficient to suggest immune therapies are valuable for IVF. The Royal College concurs: “It is clear that the advice given on many sites is strongly influenced by the personal prejudices of doctors practicing non-evidence based medicine. Much of the data they provide has never been exposed to the rigorous scrutiny of peer review. The couples are emotionally vulnerable, and there is currently no scientific evidence to justify the use of these tests and treatments.” Nevertheless, a highly visible, albeit controversial industry exists, comprised of just a handful of practicing physicians and associated laboratories offering panels of immunological tests and subsequent treatment.

Patients are rarely informed that there is no standardized testing methodology among laboratories, so the interpretation of test results (normal, borderline, or abnormal) is frequently inconsistent. If and when therapies are administered, many are designed to modify the immune system (i.e. glucocorticoid treatment, intravenous immunoglobulin, and peripheral leukocyte immunization) or to compensate for the suspected effects of the immune defect (i.e. heparin or aspirin to reduce thrombophilia from thrombogenic autoantibodies).

Given the lack of strong scientific proof of meaningful associations between abnormal immune testing and adverse reproductive outcome, combined with the poor quality of the standards of such tests, PFC physicians maintain a packet of information for our patients who inquire about potential immunological causes to their infertility.

In the next issue of Fertility Flash, we will provide a follow-up article that will include more detailed descriptions of the tests that have been conducted as well as an introductory description of what is considered viable immunological tests and treatments for RPL, versus those that are considered more controversial for repeated IVF failure.
Eldon Schriock, MD

Dr. Eldon Schriock along with PFC's medical team is continually evaluating the latest research. Our patients' welfare is PFC's first priority. With this in mind, be assured we do not include new technologies and treatments unless they are backed with solid, evidenced-based research.

Conception Health    Keeping Egg Freezing in Perspective

Almost 20 years ago, a paper in a British medical journal Lancet announced the arrival of a new technology: Oocyte Cryopreservation (Chen, C., 1986, Vol 1, Page 884). What was initially thought to be a landmark paper turned out to be the poster child for the procedure, as Chen himself and many others were unable to repeat the process with consistency. Although it is difficult to open any magazine today without reading about this wonderful new technology, less than 1% of eggs that have been frozen and thawed have resulted in live born infants.

We have learned much about the freezing of human oocytes over the years, yet despite a massive and consistent effort by the scientific community, a reliable method to freeze eggs with the same success as embryos and sperm remains elusive.

Our ability to freeze any cell depends on many factors, but most significantly on how much water the cell contains. Because water expands in volume as it turns to ice, cells must be dehydrated prior to freezing to prevent the cell from rupturing. The addition of a cryoprotectant, which does not expand upon freezing, can greatly reduce the risk of cell rupture.

Scientists have been freezing and thawing sperm with good success for over 100 years. In many ways, sperm are ideal for freezing as they exist as individual cells, they are the smallest human cells and they contain very little water. It is thought that sperm can be stored perhaps indefinitely after being added to a solution of cryoprotectant, and then frozen to minus 1960C.

In contrast to the sperm, the oocyte is the largest human cell and it contains much more water. The oocyte is also much more sensitive and is very intolerant of the chemical and physical stresses that are created during freezing and thawing. Further, the availability of oocytes is much more limited. When an oocyte is ovulated, or retrieved from the ovary during an IVF cycle, ideally it is ready to be fertilized by a single sperm. In anticipation of fertilization, the oocyte prepares to discard half of its DNA - a process called meiosis. Any changes in the physical or chemical environment around the oocyte can disrupt meiosis, leading to an oocyte with too much or too little DNA. Even after we overcome the hurdles of sensitivity and cell water content, there are other obstacles to freezing and thawing oocytes successfully.

In scientific literature, most papers that report success with egg freezing involve very few patients and therefore even fewer pregnancies and deliveries. Porcu et al., 1997, Tucker et al., 1998 and Young et al., 1998 are typical examples of papers that report successful deliveries from just one patient's frozen oocytes. Between them, these authors froze 34 eggs, of which 15 survived thawing. In larger studies, Porcu et al., 2000 and Fabbri et al., 2001 were able to obtain large numbers of oocytes for freezing (1502 and 1769 respectively), resulting in overall survival after freezing at just over 50% for both studies. Just over half of the oocytes that survived freezing fertilized, and about half of these made good quality embryos. Yet the number of babies delivered reported by Porcu was low (9 births plus 7 ongoing pregnancies). Fabbri reported only fertilization and embryo development rates as a measure of success in his study and has not yet reported on pregnancies and births.

Wider application and success with oocyte freezing depends on continued improvements with the technology and on careful selection of oocytes to freeze. While many researchers are continuing to improve the freezing process, much of the success so far has been with the use of good quality or young oocytes. In the Porcu study, most of the oocytes were collected from young women who would presumably have good quality oocytes. We would expect results to be worse if the eggs were from older women, although no such studies have been undertaken. • Despite all the hype, oocyte freezing will fall short of mainstream therapy in the near future until new technologies improve the process. Oocyte cryopreservation may be an especially disappointing prospect for older women. With this in mind, this year PFC will take part in a large scale study involving Japanese IVF centers and other US centers on an alternative technology called vitrification. This involves an ultra-rapid freezing process that we hope will allow more oocytes to be frozen before they are compromised by the effects of the physical and chemical stresses indicative of typical slow freezing methods. Vitrification has shown good success with human oocytes and embryos in recent Japanese studies.
Joe Conaghan, PhD

Ask the Experts    Metformin & PCOS Treatment

A.
Metformin (brand name: Glucophage) is indeed an FDA-approved drug for type 2 diabetes. It is also a promising new treatment in the portfolio of ovulation induction medications for women with polycystic ovary syndrome (PCOS).

Many women with PCOS suffer from insulin resistance (high blood insulin levels), a problem that is thought to possibly impede ovulation and elevate male hormone levels.

By way of background, PCOS is experienced by as many as 10 percent of women of reproductive age. An inability to ovulate normally and problems associated with an overproduction of male type hormone are typical findings in women diagnosed with PCOS. The “polycystic” aspect can be seen in the ovaries via ultrasound, which reveals a large multitude of tiny follicular cysts instead of a smaller group of well-defined emerging follicles preparing for ovulation.

Many women with PCOS respond well to clomiphene citrate (brand name: Clomid), which stimulates increased blood levels of FSH (follicle stimulating hormone) and LH (luteinizing hormone) to induce the growth of a follicle and eventual ovulation. Approximately 70% of patients treated with clomiphene citrate will ovulate and 40% will conceive, the majority within three to six ovulatory cycles.

A small fraction of patients who see no improvement from clomiphene treatment alone are good candidates for metformin, or a combination of clomiphene and metformin. Offering metformin provides such women with an alternative oral medication before being directed to the injectable stimulation medications. As an insulin-sensitizing medication, metformin decreases insulin levels, which is thought to help restore the normal ovarian hormone profile (reduces male hormone), thus allowing for spontaneous growth of a follicle and ovulation to occur. Alternatively, metformin enables the patient to become more sensitive to clomiphene. It is important to note that of those patients who do not ovulate on clomiphene alone, most benefit by the combination of metformin with clomiphene.

Metformin and other insulin-sensitizing medications may offer other benefits for women with PCOS, who are reported to be three times more prone to early pregnancy loss compared to ovulatory women. In several reports involving as yet small populations of PCOS patients, the use of these drugs appears to significantly reduce the rate of early miscarriage. One must approach this news with caution, however, until prospective controlled trials on this topic are conducted.
Carl Herbert, MD

Personal Odyssey    From Spot and Elvis to PFC

After many summers assisting vets in Atlanta and realizing that maybe I wanted to do something that involved research, I decided during my Junior year to go to Graduate School, but then the dilemma followed, grad studies in what? My undergrad professor suggested that I study a topic from a class that I really enjoyed learning, something that really sparked my interest. That's when I remembered how much I enjoyed my Animal Reproduction class, where we learned all about hormones and how they effectively operate the reproductive system.

In 1993, after graduating from UGA with a B.S., I began my Master's program in Reproductive Physiology. The first day I was asked to report to the UGA farm. I knew I was going to have some type of research animal but I was hoping for something small, cute and furry. As I pulled into the UGA Farm parking lot, everyone was assembled in front of the Swine Barn.

Yes, my research animal was to be a pig. Being from Atlanta, Georgia, a major metropolitan city and only briefly being introduced to pigs during undergrad I was a bit worried about how I was going to handle such large animals that weighed 300 lbs or more with very sharp teeth. Then there was the smell. Over several weeks I quickly realized that pigs were the most friendly and smartest animals I had ever worked with and that I had it made over the grad students working with goats and cows. Not only could I teach my pigs to come when their names were called but also to learn how to stand still for certain procedures, including ultrasounds. They were very happy every morning to leave their pig huts to go on a daily heat-check (ovulation) walk with me through the female pens, informing me in their own language who was ready for breeding. Usually the presence of a male causes a female in heat to stand stiff and flag their ears up away from their heads. Now aren't you glad we have ovulation predictor kits for humans?

My 2 years at UGA passed very quickly, I soon found myself asking the same question again, what's next? My professors urged me to continue on for a PhD after which they promised I would have a world of opportunities at hand to choose from. So in 1995, I said goodbye to UGA and my most favorite boars, Spot and Elvis and headed north to North Carolina State University, where I again would work with the most abundant domestic animal in the state. My professor, Dr. Flowers, was the Pig Expert, who knew everything to know about pigs, especially their reproductive behavior and systems.

At NCSU I thought I would be just looking at hormones in blood samples. Yet Dr. Flowers was more interested in predictors of male fertility in boars. Male fertility research involved collecting another fluid. How in the world to obtain a sample from a pig? My first week was spent in the boar housing facility where I learned how to collect my first semen samples from boars. Lets just say that it is a very scary and trusting event, perched down underneath a huge 400 lbs animal for 30 minutes or more. After my "initiation" into the pig repro group, I learned all about how to study sperm binding, and egg penetration and all of the other semen analysis techniques. All I had to do was to come up with a major research topic and find a way to execute it. I decided that I would look at sperm parameters and by using IVF and fertilization results as my endpoint, I could determine what semen parameters really mattered for boars. So over the next four years, I made trips to the local abattoir to obtain pig ovaries and to the boar house, making many male pigs happy every morning.

Finally in 1999, I graduated with a PhD in Reproductive Physiology. My professors sent me on my way, although disappointed that I did not want to become an Animal Science or Reproductive Physiology professor. I loved teaching reproduction and physiology to students while in grad school but I didn't want the pressure of obtaining grants and the publish or perish regime. I had decided that I liked IVF and wanted to do more in that field, but this time, I wanted to help people in some capacity. I began to investigate the human IVF field and sent out my CV. Dr. Joe Conaghan of PFC offered me a position.

So in 2000, I moved to San Francisco, 3,000 miles away from everyone and anything I had every really known. It was both scary and exciting. I was trained in all aspects of Human Embryology, Andrology, and Endocrinology and now work among many other embryologists to give our patients and their embryos the best care possible. We have a great lab team, made up of diverse, experienced, well educated, dedicated, and caring individuals. Each of us is Board Certified (ABB) and takes on the responsibility to know and learn the latest technologies as they arise.

Last April, I took the last High Complexity Laboratory Director (HCLD) exam and received certification as a Lab Director. But there is always something new to learn and challenging IVF cases to keep me on my toes. Working at PFC as an embryologist is a very detail orientated job, which involves a great deal of careful microscopic work, but the end result makes it all worth while, and the diverse patient cases and challenges make it all exciting all over again each day. I truly love being in the lab and assisting patients with their reproductive care. You can see it in my smile, the next time you hear me say, "Hello, my name is Jean and I am here to do your name check, can I have your first and last name?" But I especially enjoy being asked questions about your embryos. Then I get to teach all over again.
Jean Popwell, PhD, HCLD

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-- Best regards from all of us at Pacific Fertility Center.