Ovarian Tissue Transplantation Options Explored
By Tommaso Falcone, MD
Over the past few decades, considerable attention has been given to the long-term reproductive function of females undergoing cancer treatment. Many treatments damage the ovaries and induce premature ovarian failure.
There are several options for these patients. Some are routinely available clinically, such as in vitro fertilization (IVF) and embryo freezing. Others, such as oocyte and ovarian tissue freezing, are considered experimental. Oocyte freezing still requires IVF and therefore sufficient time for ovarian stimulation. Furthermore, this approach is not possible in prepubertal children.
Ovarian tissue can be removed surgically and frozen. Usually the cortex is cut into
small pieces and the tissue cryopreserved and subsequently transplanted back.
However, since this is an avascular transplant, ischemic necrosis of the graft occurs
before neovascularization can occur. As an alternative, we proposed removing,
freezing and subsequently thawing and transplanting an ovary with a vascular
pedicle to try to diminish the ischemia. We chose a heterotopic site, the inferior
epigastric vessels that could theoretically provide an easier access. A sheep model
was used. The ovary was removed from its initial site with its vascular pedicle, cryopreserved, thawed and transplanted to the inferior epigastric vessels.
Our first experiments showed successful autotransplantation of frozen/thawed
ovaries with their vascular pedicle. This was manifested by restoration of selected
ovarian function including in vitro maturation of collected oocytes, presence of
follicles from several stages of folliculogenesis and blood vessels expressing
specific markers of vascularization, and proliferation and apoptosis of ovarian
cells. Thus, heterotopic autotransplantation of a whole frozen/thawed ovary
allows for development of preovulatory follicles, oocyte growth, and restoration
of vascularization and cellular function. However, there was some necrosis of the
vessels (above), probably due to the differential freezing characteristics of different
Additional improvements are required to enhance the efficiency of auto transplantation
of frozen/thawed ovaries to produce more oocytes. Freeze-thaw protocols still require further optimization and depletion of the primordial follicle reserve will need to be addressed. But current evidence suggests that whole frozen-thawed ovary transplantation may be successful in humans in the future.
Dr. Falcone is Professor and Chairman of the Ob/Gyn & Women's Health Institute. Physicians may reach him at 216.444.1758 or email@example.com.
New Horizons in Embryo and Oocyte Cryopreservation
By Nina Desai, PhD, HCLD
In vitro fertilization (IVF) is one of the most exciting and dynamic areas in medicine. In the past two decades, tremendous strides have been made with IVF techniques. Clinical pregnancy outcomes have risen dramatically, from 20 percent to almost 60 percent in younger patients. With this increase has come a movement to reduce the number of embryos being created and also to avoid high order multiple pregnancies by reducing the
number of embryos being transferred.
Effective techniques for cryopreservation of embryos and unfertilized oocytes for a future cycle have been the focus of research in laboratories all over the world. Traditional cryopreservation technology has involved the use of cryoprotectant agents, dehydration
of the cell and programmed slow cooling at rates of -0.3 ºC a minute down to -36 ºC, before plunging in liquid nitrogen. The entire process can take as long as two hours. The most frequent cause of damage is intracellular ice crystal formation leading to cellular disruption on thaw. Oocyte cryopreservation has been even more challenging than embryo freezing due to the large volume of the egg and the susceptibility of the meiotic spindle to depolymerization as temperatures are reduced. Disruption of the meiotic spindle and aberrant chromosome segregation during oocyte freezing can lead to an increase in embryos with aneuploidy.
A new cryopreservation technique known as “vitrification” shows tremendous promise and has been adopted by the Cleveland Clinic’s IVF program for cryopreservation of embryos at all stages, as well as oocyte cryopreservation. Vitrification involves ultra rapid freezing of the oocyte/embryo from 37 ºC to -196 ºC at a rate of -20,000 to -30,000 ºC per minute. High concentrations of cryoprotectants and high cooling rates are necessary to transition the embryos into a “glass-like state” to avoid cryoinjury. To achieve such high cooling rates, the oocyte/embryo is frozen in miniscule fluid volumes of less than one microliter.
Cleveland Clinic’s IVF laboratory uses a novel carrier called the “cryoloop” for vitrification. The cryoloop is a fine nylon loop measuring no larger than the eye of a sewing needle. The loop is dipped in cryoprotectant and the embryo/oocyte is placed on the fluid film (<1ul) using a micropipette. The cryoloop is then immediately immersed directly in liquid nitrogen instantaneously freezing the sample.
Vitrification has had a tremendous impact on our embryo cryopreservation program. We were one of the first laboratories in the United States to successfully apply this methodology for cryopreservation of eight-cell embryos (Desai et al, 2007). To date, the post-thaw survival rate has been 95 percent. We have achieved a clinical pregnancy rate of 45 percent (based on fetal heart by ultrasound). Vitrification technology has completely replaced conventional methods for freezing in our laboratory.
The existence of excess cryopreserved embryos, together with steady improvements in clinical outcomes, often present an ethical dilemma for patients who become pregnant and no longer need their frozen embryos. Many have difficulties consenting to
embryo disposal or donation for a variety of reasons, including religious beliefs. Oocyte cryopreservation has the potential to resolve some of these issues. While vitrification appears to have increased the efficacy of the procedure, oocyte freezing is still in its infancy. This new technology is very appealing and offers women the ability to bank their gametes in much the same way as men bank sperm. Cleveland Clinic now offers oocyte cryopreservation on an experimental basis to a select group of patients who may benefit from this procedure, including cancer patients who can freeze oocytes before undergoing radiation and chemotherapy.
The process of oocyte cryopreservation is much more difficult than embryo freezing because of the large volume of the egg and the susceptibility of meiotic spindle disruption during the freezing process, resulting in embryos with chromosomal abnormalities. Using the cryoloop, Cleveland Clinic’s IVF laboratory has been able to successfully freeze oocytes in fluid volumes of less than one microliter.
Dr. Desai is the Director of IVF and Clinical Research. For more information about oocyte cryopreservation, contact her at 216.839.2907, or firstname.lastname@example.org.
New In Vitro Fertilization Director Named
Cynthia Austin, MD, has been selected to direct Cleveland Clinic’s In Vitro Fertilization Program. Dr. Austin joined the Beachwood Fertility Center staff in 2000 after serving as co-director of University Hospitals’ in vitro program.
Dr. Austin is a graduate of Boston University and George Washington University Medical School. During postgraduate training at MetroHealth Medical Center at University Hospitals, she focused on infertility and reproductive endocrinology. Throughout her career, she has earned a reputation for her welcoming and thorough approach to couples struggling to achieve pregnancy. She takes pride in individualizing her treatment of her patients.
“The right approach depends on many things, including the patient’s age; overall health; medical history; extent of the disorder and the course it is expected to take; tolerance for specific medications, procedures or therapies; and most important, the couples’ personal preferences,” she says. “This makes a detailed evaluation and consultation before any procedure of the utmost importance.” Sometimes, even the most comprehensive evaluation won’t reveal a single factor. But Dr. Austin says that even when the cause isn’t clear, effective treatments are available.
The American Society of Reproductive Medicine reports that 85 percent to 90 percent of cases of infertility are successfully treated with conventional therapies, such as drug therapy or surgical repair. The vast majority of surgical procedures used to address infertility can now be performed on an outpatient basis either through a laparoscopic or hysteroscopic approach. Both types of endoscopic procedures offer success rates as good or better than open surgery and result in reduced costs, quicker recovery, fewer complications, less pain and better cosmetic results.
For those patients who are unable to conceive with these methods, reproductive technology, such as IVF, may be the solution. One of the strengths of Cleveland Clinic’s In Vitro Fertilization Program is that in addition to state-of-the-art technology, the center provides a caring environment where the emphasis is on accessibility, personal attention and emotional support.
Dr. Austin currently is investigating new treatment alternatives for polycystic ovary syndrome (PCOS) and is developing strategies to help women with PCOS make lifestyle changes to impact their overall wellness.
She also has turned her attention to teaching, for which she has found a passion. After raising three children of her own, she is happy to share her insight into what makes her young students tick. The innovative curriculum of the Lerner College of Medicine fits her goal of training self-reliant, independent thinkers.
To refer a patient to Dr. Austin for evaluation or treatment, please call 216.444.6601.
Infertility Program Enhanced by State-of-the-art Labs
The IVF Laboratory and associated research laboratories at Cleveland Clinic’s Fertility Center create a unique environment for providing the highest level of clinical care and development of cutting-edge technology to improve clinical outcomes. The labs encompass approximately 1200 square feet and contain state-of-the art equipment for micromanipulation, cryopreservation, cell fusion, and image analysis of gametes and embryos. The clinical laboratory has 16 incubators eight microscope stations, five laminar flow hoods for gamete handling, a laser for assisted hatching and three micromanipulation stations for ICSI or PGD. The culture environment of the embryo is carefully regulated through a specially designed air system.
The newest equipment added to the clinical laboratory has been the Spindleview System, which allows real-time imaging of the meiotic spindle of living oocytes. Using advanced polarized light imaging technology, the embryologist is able to visualize the position and organization of the meiotic spindle assembly in oocytes before and after freezing. This allows the embryologist to avoid disrupting the oocyte spindle during intracytoplasmic sperm injection (ICSI), thus reducing the risk of aneuploid embryos. More recently, this imaging system has been applied by our laboratory to critically assess protocols for egg freezing. The strong link between the research and clinical labs at the Cleveland Clinic Fertility Center has been instrumental in developing “high tech”
solutions to improve clinical IVF.
For the past 30 years, our state-of-the-art Andrology Laboratory and Reproductive
Tissue Bank has been providing the highest quality services to couples being
evaluated for infertility. The laboratory features state-of-the-art testing using the
latest, most effective diagnostic tools and analyses. The laboratory is certified by
CLIA in the state of Ohio and accredited since 1994 by the College of American
Pathologists (CAP) Reproductive Laboratory Program.
The laboratory offers comprehensive semen analysis with a computerized system that allows for the quality of the semen to be thoroughly studied. Other tests include direct and indirect immunobead testing to detect antisperm antibodies in serum, sperm or seminal plasma. Special stains can be utilized for the presence of sperm in azoospermic
specimen. A post-vasectomy screen is available for those patients who recently have had a vasectomy.
Services also include the processing of sperm for intrauterine insemination;
providing frozen donor semen for artificial insemination; and sperm freezing for
patients prior to cancer treatment or vasectomy.
Now On Bookshelves
Cleveland Clinic Gynecologists Collaborate on New Text
Basic, Advanced, and Robotic Laparoscopic Surgery: Female Pelvic Surgery Video Atlas Series
Tommaso Falcone, MD, FRCSC, FACOG
Vice Chair, Office of Professional Staff Affairs
Professor and Chair, Ob/Gyn & Women’s Health Institute
Jeffrey M. Goldberg, MD
Section Head, Reproductive Endocrinology and Infertility
Ob/Gyn & Women’s Health Institute
This book and accompanying videos demonstrate the most important aspects of laparoscopy including anatomy, instrumentation and principles of laparoscopic suturing.
Hours of video footage, detailed discussion and illustrations provide a clear description of the most effective laparoscopic procedures, how to manage complications and more.
Treatment Guide Available
Cleveland Clinic has created a comprehensive guide to the treatment of infertility, which is available online.
This free downloadable guide includes:
- The causes of infertility
- When couples should seek help
- Available treatment options
- A directory of Cleveland Clinic physicians and locations providing care
If your patients are interested in comprehensive information on infertility, they can access it at clevelandclinic.org/infertilityguide.