1) Egg/embryo Quality
2) Receptivity of the Uterus
3) Embryo Transfer (The “Rate Limiting” Technical Step)
Each of these factors are outlined in detail below or linked to prior posts that address them.
1. EGG/EMBRYO QUALITY
There are a number of factors that affect egg and embryo quality. These include:
A. Age and Ovarian Reserve
B. Protocol for Controlled Ovarian Stimulation (COH)
C. Sperm Quality
D. Embryo Selection
A. Age and Ovarian Reserve
Maturation (ripening) is the final step in biological development that a system or organism must undergo in order to prepare for optimal function. Ultimately the functional efficiency of the system and/or organism will be predicated upon its state of readiness to undergo such maturational fine tuning. A poorly developed system/organism can thus never attain optimal functionality. The same concept applies with regard to the development and maturation of a woman’s eggs. Since it is predominantly the egg rather than the sperm that determines the potential of an embryo (the fertilized egg) to develop into a healthy baby (embryo competence), it is little wonder that egg development/maturation is a major determinant of human reproductive performance.
A woman is born with all the eggs she will ever produce. She uses them up through her reproductive life and when they are gone, her reproductive potential ends. It is interesting that a woman’s eggs are already stored in her ovaries 6 months before she is born. In fact, she starts losing her eggs at a furious rate from the get-go such that by the time she is born, more than half of her eggs have already been re-absorbed.
This process of egg attrition continues after birth but at a slower rate. By the time the woman reaches puberty and begins to menstruate (the menarche) and then ovulate, her egg population has dropped from more than 5 million (at 3-4 months post-conception) to less than 1 million on average. The number of eggs present at the time of puberty when the woman’s reproductive potential is launched is genetically determined.
Each immature egg stored in the “ovarian repository” is enveloped by a thin layer of cells. Batches of these so called primordial follicles are sequentially recruited to embark upon what represents a 4-month developmental journey, during which time they undergo complex and magnificent developmental changes designed to prepare them for “service” in a single, designated ovulation cycle. During this recruitment journey many primordial follicles succumb to a process of culling. Those that ultimately survive this 4-month recruitment process each develop a central collection of fluid, lined by a few layers of hormonally active granulosa cells and measure a few millimeters in diameter. At this point they are referred to as antral follicles.
The number of antral follicles available for hormonally induced growth in the very next ovulation cycle (recruitment potential) starts off at quite a large number (as many as 20-50 and sometimes even more) but then declines with advancing age as the primordial follicle population in the ovaries progressively declines with more and more usage. The recruited antral follicles grow in response to stimulation by follicle stimulating hormone (FSH), a hormone produced by the pituitary gland (a cherry-sized structure located just above the roof of the mouth, which hangs from the base of the brain). FSH stimulates the cells lining the inner wall of the follicle (the granulosa cells) which start to produce the female hormone, estrogen in ever increasing amounts. Estrogen makes the lining of the uterus (endometrium) thicken in preparation for ovulation.
Within the first 5 or 6 days of the cycle, one (sometimes two), antral follicles start to grow faster than other members of the cohort, differentiating from them. About five days into the cycle, these selected follicle(s) , i.e the dominant one(s), continue enlarging while all the others begin a downward spiral and ultimately die off within a week or so. Once the selected follicle(s) attain a certain size (approximately 18-20 mm in diameter) and their estrogen output reaches a crescendo, a sudden surge of another pituitary hormone called Luteinizing Hormone (LH) occurs, the follicles rupture and the eggs are released. Ovulation has now taken place.
Under the influence of LH, the granulosa cells in the ovary undergo transformation (luteinization) and now, in addition to estrogen they produce progesterone. The combined effect of estrogen and progesterone causes secretion in the hitherto developed endometrial glands, which further prepares the endometrium to receive the fertilized egg (embryo) about 5-6 days after ovulation.
With every 4 month recruitment journey the number ovarian primordial follicles (eggs) that a woman has available declines and the fewer antral follicles she will have available use in any cycle. Over a period of two decades or so, the progressive erosion in the number of ovarian primordial follicles/eggs is accompanied by such a profound reduction in the woman’s recruitment potential that the pituitary gland, in a frantic attempt to restore recruitment, increases its output of FSH. This manifests in rising blood FSH concentrations.
At a certain point, a reciprocal rise in basal LH follows. The rising basal FSH and LH level signals the onset of the climacteric, a period of roughly 6-8 years in the woman’s reproductive career, during which time the number of primordial follicles (and eggs) in the ovaries, having declined below a critical threshold, triggers certain profound changes. These include (but are not necessarily limited to) hot flashes, ovulation becoming irregular and dysfunctional, menstrual cycles becoming more and more irregular and increasingly less frequent, and a decline in egg quality. This is accompanied by reduced fertility an increased risk of miscarriages. Ultimately the follicle recruitment process trickles to a halt, ovulation and menstruation cease altogether, and menopause has arrived.
A few additional pieces of information are needed here:
- The ovary has two well defined hormone producing “compartments”: The first, the follicle, houses the egg and has an inner lining of granulosa cells which in response to FSH produce estrogen. The second is the stroma or theca, connective tissue that surrounds the follicles and produces male hormones (predominantly testosterone).
- Testosterone is the building block from which granulosa cells manufacture estrogen. It is carried in “bucket brigade” fashion from the stroma to the follicle. LH promotes production of testosterone by the stroma.
- Some LH is essential. In fact, neither follicle growth with estrogen production, nor proper egg development and maturation is possible without some delivery of testosterone to the follicle granulosa cells. However, too much LH, by causing overproduction of stromal (thecal) male hormones can have a detrimental effect on follicle growth (as evidenced by a decline in estrogen production) and with egg development, which in turn ultimately interferes with egg maturation. In such cases, when (and if) the eggs fertilize, the resulting embryo(s) are much more likely to have an irregular chromosome number (aneuploidy) and thus be “incompetent”.
- There are situations such as advanced age (above 40 years), diminishing ovarian reserve, and polycystic ovarian syndrome (PCOS) which are all associated with a sustained elevation in LH as well as an overgrowth of ovarian stroma (stromal hyperplasia or hyperthecosis). In such cases, the overgrowth of ovarian stroma/theca serves as an enlarged target for LH activity and thus will over-produce testosterone with adverse effects on egg development and embryo quality.
Simply stated, the potential for a woman’s eggs to undergo orderly development and maturation, successful fertilization and subsequent progression to competent embryos while genetically determined is also profoundly influenced by ovarian hormonal influences which can have an adverse influence on follicle development, egg quality and ultimately on embryo competence.
It follows that protocols for controlled ovarian hyperstimulation (COH) must strike a balance between optimizing follicle growth and development, and avoidance of excessive ovarian testosterone production. We do have access to medications that act as gonadotropin releasing hormone agonists-GnRHa (e.g. Lupron, Nafarelin, Synarel, Buserelin etc) and GnRH-antagonists (Cetrotide, Orgalutron, Ganirelix etc.), by which it is possible to pharmacologically regulate pituitary release of LH. We also have pure DNA recombinant FSH (Folistim, Puregon and Gonal F), and pure recombinant LH preparations (Luveris). By prescribing customized protocols of ovarian stimulation, it is now possible to avoid over-exposure to testosterone in women at risk (e.g. those with diminished ovarian reserve, older women and those who have PCOS) and in the process improve egg/embryo quality.
So, why do some still persist in using fertility drugs that contain high concentrations of LH such as Menopur and Repronex in such cases, and why use protocols that promote LH over-exposure (Flare protocols)?
There is an unfortunate tendency on the part of many IVF practitioners to place the blame for “poor quality embryos” on the embryology laboratory. The truth of the matter is that while the IVF embryology laboratory plays a pivotal role in achieving optimal fertilization and embryo quality, no embryologist, regardless of expertise, can produce competent embryos from aneuploid eggs (thosse with an irregular number of chromosomes---see below.
It is important to recognize that the chromosomal configuration of the embryo that determines its “competence.” A “competent" embryo is one that is euploid (i.e. has all its 46 chromosomes), and upon being transferred to a receptive uterus is most likely to propagate a viable pregnancy. Embryos that are aneuploid (an irregular number of chromosomes) are incapable of propagating “competent” embryos. The fact is that less than 40% of embryos resulting from fertilization of even young women’s eggs are capable of making a healthy baby. As women get older, this gets progressively worse, such that by her mid-40’s the number drops to less than 10%.
The above serves to explain the progressive decline in IVF birthrates that starts in the mid thirties, and greatly accelerates after age 40. For those women whose age and/or degree of ovarian resistance makes having a baby with their own eggs unappealing or unlikely, ovum donation (using eggs from a young donor - usually compatible and anonymous) is an excellent option. In fact, IVF-ovum donation is one of the most successful methods of achieving pregnancy, regardless of the woman’s age.
B. Individualizing Controlled Ovarian Hyperstimulation (COH) Protocols
Protocols for controlled ovarian hyperstimulation (COH) should be geared toward optimizing follicle and egg development and avoiding over exposure to ovarian androgens (mainly testosterone). The fulfillment of these objectives requires an individualized approach to COH and that the administration of human chorionic gonadotropin (hCG) or recombinant luteinizing hormone (LHr) to “trigger” ovulation, be timed precisely.
It is common practice to administer gonadotropin releasing hormone (GnRH) agonists (GnRHa such as Lupron, Buserelin, Nafarelin,Synarel; and GnRH-antagonists (e.g. Orgalutron, Ganirelix, Cetrotide). GnRH- antagonists block pituitary LH release within a few hours of the initial administration. GnRHa act by causing an initial outpouring and then, over 4-5 days, a depletion of Pituitary FSH and LH.
See Previous Post on GnRH agonist/antagonist Protocols for Detailed Information
C. Sperm Quality
While about 90% of embryo chromosome aberrations result from egg rather than sperm problems, few would argue that since fertilization is the center pin of IVF success, sperm quality is a vital factor. Nevertheless, the advent of intracytoplasmic sperm injection (ICSI) has all but removed male infertility as an impediment to IVF success.
The introduction of ICSI has made it possible to fertilize eggs with sperm derived from men with the severest degrees of male infertility and in the process to achieve pregnancy rates as high, if not higher than that which can be achieved through conventional IVF performed in cases of non--male factor related infertility. The performance of ICSI in cases of "male factor infertility " has been shown to marginally increase the risk of certain embryo chromosome deletions (leading to a slight increase in early miscarriages) as well as the potential for a resulting male offspring to have male infertility in later life, there is no evidence of any significant increase in the incidence of serious birth defects attributable to the ICSI procedure itself. More relevant is the fact that when ICSI is performed for indications other than male infertility there is NO reported increase
in the risk of subsequent embryo chromosome deletions, miscarriages or in the incidence of subsequent male factor infertility in the offspring.
The Sperm Chromatin Structure Assay or SCSA (aka Sperm DNA Integrity Assay or SDIA) About 12-15% of conventional IVF is associated with unanticipated absent or poor fertilization. This has led many to conclude that male infertility may be an “occult phenomenon” in some men. In fact the SCSA/SDIA has demonstrated that DNA damage may be present in sperm from men with both normal and abnormal semen analyses and that male infertility is equally prevalent in such cases.
D. Evaluating Embryo Quality & Selecting the Best Embryos for ET
No other factor in the IVF process influences success as directly as choosing the “right” embryo for transfer to the uterus. This is due mainly to the fact that aneuploidy of the embryo, is responsible for the majority of IVF failures. Aneuploidy generally results in either: 1) the failure of the embryo to develop to a stage capable of attaching to the uterine wall, 2) miscarriage after implantation, or 3) a chromosomal birth defect such as Down syndrome. The selection of one or more competent embryos for transfer is thus central to IVF success.
Unfortunately, most methods currently used to select the best embryos for transfer are relatively inconsistent – yielding on average less than a 20% pregnancy rate per embryo transferred. This fact is responsible for the tendency to transfer a large number of embryos during an IVF cycle in hopes of attaining a pregnancy. Unfortunately, this can lead to high-order multiple pregnancies which carry substantial risks to both the mother and the babies.
See previous post on CGH Embryo Selection for more detailed information.
2. Uterine Receptivity
I often refer to the conception process in terms of a “seed/soil relationship". Just as a plant can’t grow and thrive without first assuring that both seed and soil are good, neither can a pregnancy be successful without both the seed (embryo) and the soil (uterine environment) being ideal. The uterine factors are just as critical to the equation as the quality of the embryo. There are a number of factors that contribute to the receptivity of the uterus including the contour of the uterine cavity, thickness of the uterine lining, and immunologic factors. Each of these are outlined in my prior posts on uterine receptivity and immunologic implantation failure linked here.
3. The Embryo Transfer : A Rate Limiting Factor
Undoubtedly, embryo transfer is a rate limiting step in IVF. It takes confidence, dexterity, skill and gentility to do a good transfer. Of all the procedures in ART this is arguably the most difficult to teach. It is a true “ART” and many women fail to conceive simply because this procedure is not performed optimally. From a technical perspective, embryo transfer is the most critical phase of the entire process.
The use of ultrasound guidance to optimally place the embryos in the uterus has become an integral part of the ET process. Successful clearance of all other hurdles means nothing if there the embryos are damaged, misplaced, dislodged or if they reflux into the cervix, vagina or fallopian tube(s). Symptoms such as uterine cramping or bleeding during, or immediately after or the procedure are indicators of a poor transfer. A problem-free process is so important that we grade all transfers performed at SIRM on the basis of patient comfort, technical difficulty, and the number of attempts required to achieve the successful transfer of all allotted embryos. Embryo transfer is usually performed 72-144 hours after egg retrieval, but the actual time may depend on whether cleaved embryos or blastocysts are being transferred. Embryos that have not attained at least a 7-cell state of cleavage within 72 hrs of egg retrieval are much less likely to progress to blastocysts and implant successfully into the uterine lining.
See my prior post on Embryo Transfer
Embryo Cryopreservation: At SIRM we vitrify (ultra-rapid freezing) leftover, blastocysts Cryoepreserved blastocysts are thawed and the surviving ones are transferred on the same day while in isolated cases where we have to process day-3 embryos , we thaw and then culture them for an additional 2-3 days to the blastocyst stage before transferring them to the uterus.
In the pursuit of optimizing outcome with IVF, the clinician has a profound responsibility to try and enhance the environment for implantation, so as to maximize the chance of pregnancy and enhance the quality of the life after birth.
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