Science Pulse    Paternal-Free Mice

Creating offspring using only female genes and with no paternal contribution is a common phenomenon in nature and in fact is a method of reproduction employed by almost all plants and animals. Mammals however have not been able to reproduce in this way. This impediment is attributed in large part to a process called genomic imprinting.

Experimentally, when mouse embryos are created using only the DNA from 2 eggs, the resulting fetus is well formed, but only a rudimentary placenta develops and the pregnancy fails. This is because the placenta is created mainly by paternal genes, and without the involvement of a sperm, we can’t get a normal placenta.

But if we have 2 copies of almost every gene (one from Mom and one from Dad), why can’t the maternal genes make a placenta? Biologists think that it’s a conflict of interest for Mom’s genes to make the placenta. Since the placenta in many ways is a parasite that fights for Mom’s resources, Mom’s placental genes are deliberately inactivated or switched off and it’s left to Dad to make the placenta. This process of deliberately inactivating a set of genes from one parent, so that the other parent’s genes are left to do the work is called imprinting. These genes carry with them a history of their origin because they are endowed at conception with a maternal or paternal imprint.

One negative consequence of imprinting is that when an imprinted gene is defective or otherwise does not work, the inactive, but perfectly good gene from the other parent can’t be called upon to help out. Diseases like Prader-Willi syndrome and Angelman’s syndrome which have variable physical, mental and behavioral effects on afflicted individuals are caused by defective imprinted genes.

So what happens when an embryo is created using 2 sperm and without maternal DNA? In this instance, as would be expected, the placenta is normal and fully formed, but the fetus is typically deformed and most notably lacks a head. It could be said that without a Mom, mammals lose their heads.

In the Nature paper, the mice without a father were created after exhaustive attempts: 2 live born from 457 reconstructed eggs. And the researchers used a trick to get around the imprinting issue. To make each embryo they used a mature (ovulated) egg and an immature egg from a newborn mouse in which the genomic imprint had not been established (imprinting occurs as eggs grow and mature). This allowed them to overcome the absence of the paternal imprinted genes since there were few or no imprinted (and therefore inactive) genes in the DNA from the immature eggs. The process was not very efficient in creating live offspring, but one of the resulting mice reproduced normally after reaching adulthood. The second mouse was used in tests to determine its DNA normalcy.
- Joe Conaghan, PhD, HCCLD
Dr. Joe Conaghan, PFC's ART Laboratory Director, teaches human reproduction, trains fellow embryologists for licensure and is an inspector for CAP, the licensing body for IVF Laboratories in the United States. His background in research includes involvement in the first PGD on human embryos. He directs PFC's team of all board certified embryologists. His high standards, excellent patient care and extensive experience brings national recognition to our laboratory.

Personal Odyssey    PGD: An Alternative

It had always been our plan to have more than one child. However, as we began to discuss the possibility of having a second baby, we both realized that given the physical, emotional and financial costs of being affected by this disorder, we were not comfortable with consciously bringing another child into the world without doing everything we possibly could do to avoid this for any other child.

After two years of extensive pre-conception counseling, we decided in-vitro fertilization (IVF) with pre-implantation genetic diagnosis (PGD) through Pacific Fertility Center would give us the best possible chance of giving birth to an unaffected child.
Embryo Biopsy
A medical technique whereby embryos can be screened for specific genetic defects prior to transfer to the womb, PGD has been performed for over 10 years and has proven to be a most effective method of diagnosing embryos for known genetic mutations. To-date there have been over 2500 PGDs performed around the world resulting in over 1600 children born without the disease for which they were screened. The error rate for PGD is less than two-percent; therefore, PGD would reduce our chance of having an affected child from 25% to less than 2%.

A little over a year ago we began our IVF with PGD Embryo Biopsy cycle. On Day 3 after retrieval, when our embryos were eight-cells or so in size, a single cell was biopsied from each embryo. These cells then were sent to a lab where the single cell from each embryo was tested for the genetic defect in question. We then transferred two embryos pre-determined to be unaffected by the disorder. In October of last year, we welcomed to the world our miraculous bundle of joy, an - unaffected - little boy.
- Name withheld upon request

Ask the Experts    Improving the Odds

Q:
My husband and I have two sons, age 3 and 6, and we would really like to have a girl. Is there anything we can do to make sure our next child is a girl?

A:
From a purely medical point of view, the answer to your question is yes. There are ways to significantly shift the odds of having a child of one gender or another. Gender is conferred on an embryo by whether an X-bearing sperm (for a girl) or a Y-bearing sperm (for a boy) enters the egg. Unfortunately, despite highly publicized claims, there are no proven effective "at home" methods of sperm separation. Nor does timing of intercourse relative to ovulation affect the 50:50 gender ratio. By natural methods, it’s a flip of the coin.

The only commercially available method for sperm separation that appears to be effective is the sperm sorting process available through Microsort.net. This method is still under FDA investigation for safety and efficacy but does appear to do a reasonable job in separating sperm, especially if the desired gender is female.

The company reports a 90% success rate with separating X-bearing sperm and a 73% success rate in separating Y-bearing sperm. With only a couple of hundred babies born thus far, there does not appear to be any increase in birth defects. Because it is still an experimental process, couples wanting to try this will have to travel to either Fairfax, Virginia (Microsort headquarters) or an affiliated clinic in Southern California for fresh sperm insemination as freezing and thawing of sperm reduces these numbers even further.

Unfortunately, after processing, Microsort severely decreases the number of sperm available for insemination. Because sperm counts are so low after sorting, it is usually necessary to do in vitro fertilization with sperm injection (IVF-ICSI) to significantly improve the chances that the sperm will fertilize the egg in the IVF laboratory. PFC is a participating site in the FDA investigation for Microsort and we have used sperm specimens that have been previously Micro-sorted for IVF-ICSI.

Beyond Microsort, the only way to improve the odds of recovering one gender over another to close to 100% is to undergo preimplantation genetic diagnosis (PGD). PGD uses a DNA-binding technique to determine if there are a correct number of chromosomes in the embryo at the time of IVF. To do this, embryos on Day 3 of culture (5-10 cells) undergo a biopsy to remove a single cell. The rest of the embryo remains in culture in the IVF laboratory. The cells from the embryos are analyzed for the correct number of chromosomes. Currently, PFC with its cytogenetic partner, St. Barnabas Medical Center, tests for 9 chromosome pairs, 13, 15, 16, 17, 18, 21, 22, X and Y. This screening is called "aneuploidy screening" and we allow our patients to know and select the gender of their normal embryos for transfer if they so wish.

Although IVF with PGD is the most effective method for gender selection, it is certainly the most expensive and there is no absolute guarantee that the transfer of the screened embryos will result in pregnancy. A PFC physician can best discuss the odds based on the woman’s age and the couple’s history of childbirth.

Many couples undergoing PGD are doing so to screen for specific genetic defects or are specifically undergoing gender selection because of their risks of having a genetic disease that only affects males (X-linked diseases).

On the other hand, PGD for elective gender selection, either for "family balancing" as in your case, or even for having a first child of a particular gender poses difficult ethical issues. Just because we can do this, should we? What will the couple do with normal embryos of the undesired gender? At PFC, we do not encourage PGD for elective gender selection but if a couple is undergoing IVF and wishes to undergo aneuploidy screening for the 9 chromosomes, we do allow them to select to transfer embryos by gender. We encourage them to consider donating excess embryos of the undesired gender for adoption by other couples.

Women or couples interested in this procedure should discuss it with their Reproductive Endocrinologist. At PFC, we also refer our PGD patients for a special genetic counseling session in preparation for this process.
--- Carolyn Givens, MD

Critical Review    Pushing the Limits of Prenatal Portraiture

Seizing on this yearning, a new crop of commercial ultrasound studios has mushroomed all over the country, offering parents a chance to have a first look via an elaborate 3D and even 4D video ultrasound. At least three such businesses are in the Bay Area. Yet new parents contemplating a nonmedical 3D ultrasound simply for novelty or posterity should be fully aware of this technology in a rapidly evolving marketplace.

The safety of common medical ultrasounds is undisputed. For over 35 years, ob-gyns have used 2D ultrasound technology as standard practice to medically diagnose the health of a weeks-old fetus, enjoying an early glimpse of its emerging shape, major organ development, tissue and blood flow and when desired, the gender. The ultrasound repertoire is so common; over 80 million procedures are now performed in the US each year, reports one clinic.

Nevertheless, the Food and Drug Administration (FDA) and the primary medical association that oversees ultrasonography - the American Institute of Ultrasound Medicine (AIUM), have thus far refused to endorse 3D and 4D ultrasounds offered by commercial studios. The concern is less about the technology itself, and more about how it is applied. While the sound wave levels used for a 2D and a 3D/4D are reportedly of the same frequency, (it’s the computer diagnosis that creates the image differentiation), there is more built-in oversight in the medical community performing diagnostic ultrasounds.

For instance, is the person performing the commercial fetal portrait properly trained? Right now, it is up to the 3D studio to make sure that the person controlling the knobs and holding the transducer has undergone the same training standards required for ultrasonography at an ob-gyn office. Professional (non-physician) ultrasound practitioners undergo nearly three years of training, including 12-18 months for didactic and 12-18 months of clinical practice in order to gain the key certification from the American Registry of Diagnostic Medical Sonographers (ARDMS).

Moreover, there is concern that a commercial portrait ultrasound will reveal a developmental problem with the fetus that should be observed and discussed only through a physician/patient relationship. Another concern is that enthusiastic parents will forego a routine medical ultrasound after obtaining an elaborate portraiture one. In response, many commercial ultrasound studios are requiring patients to bring proof of a prior medical diagnostic ultrasound.

Finally, knowing a bit about the technology helps parents make an informed decision. In the medical community, the standard is to expose the fetus to the lowest possible exposure level for the shortest amount of time, usually 10 minutes or so. Because frequent ultrasonography at higher levels can produce a heating effect in bone and tissue, the aim is to minimize exposure. Yet some commercial fetal portrait studios offer deluxe packages involving a 45 minute video ultrasound.

A spokesperson from 3DBabyVu insists that the potential for physical damage to the fetus via a wrong decimal level setting is literally and virtually not possible, at least with the standard GE Voluson machines, which provide a cap to the frequency level. Yet he admitted that the same machines have two other settings for cardiac mode and vascular mode to examine more robust adult tissue. If patients choose to purchase a dynamic 3D or 4D image package offered by one of these enterprising studios, we strongly recommend that you learn as much as possible and even consult with your ob-gyn if you are at all confused. Also, it is best to confirm that the sonographer at the commercial studio is ARDM certified. Because the practice of fetal portraiture imaging is self-regulated, it is the patient’s responsibility to be aware of current research and be as informed as possible prior to using this new technology.
--- Eldon Schriock, MD

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