Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2001-03-13
2003-11-18
Chin, Christopher L. (Department: 1641)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S007100, C435S007920, C435S007940, C435S091500, C436S501000, C436S806000, C436S811000
Reexamination Certificate
active
06649344
ABSTRACT:
BACKGROUND OF THE INVENTION
The normal ovulating woman will recruit approx. 300 immature oocytes (ova) for each menstrual cycle. Normally, during a process of apoptosis all but one oocyte will die before ovulation. Conventional in vitro fertilisation (IVF), treatment for special cases of severe male and female infertility, is based on retrieval of mature human ova followed by fertilisation of the mature oocytes with spermatozoa. The recruitment of human mature oocytes is accomplished by several complicated forms of hormone treatment regimens, often with discomfort or risk for the women involved. These hormone treatment regimens will especially become a problem in the future, as IVF is increasingly offered to perfectly normal women in these programs due to their husbands' poor sperm quality. Furthermore, this type of treatment will normally provide a pregnancy rate of 20% per started cycle.
Because of the risk, discomfort and cost of the hormonal stimulation several other approaches have been tried during the years. In animals in vitro maturation (IVM) has become an efficient method for producing oocytes for IVF, but until now recorded success rates for clinical human IVM have been low (Cha, Trounson, Barnes, Russel). One of the most simple ways to avoid hormonal stimulation has been not to stimulate with hormones at all. This treatment regimen, however, has only provided a limited numbers of pregnancies and the calculated pregnancy rate is never beyond 5% per started cycle.
SUMMARY OF THE INVENTION
The present invention relates to the ability of a test system to predict the timing and development of specific healthy, activated prophase ova to develop further in vitro up to MF-II without exogenous hormonal treatment of the woman, and hereby avoid the degeneration of the majority of the ova, even though the woman is not treated with hormones. In contrast to all other previous hormonal assays always focusing on later stages of ova development, usually ova in the MF-II stage, the measurements aim at the timing of aspiration of non-fertilisable ova, which can mature in vitro to be fertilised. By this method pregnancy rates of more than 15% are achieved.
DETAILED DISCLOSURE OF THE INVENTION
Scientific Background
The nuclear development of the ova are arrested in the diplotene phase as germinal vesicles. This is referred to as the dicytate stage. This stage is characterised by highly diffuse chromosomes, the DNA of which has little affinity for such nuclear strains as Feulgen's reagent. The chromosomes of ova in the dicytate stage bear lateral projections in the form of branches and loops which actively replicate ribonucleic acid (RNA). They closely resemble the “Lampbrush” chromosomes, which are found almost universally in eggs of lower vertebrates and some invertebrates. The RNA may act as the messenger directing protein synthesis within the ova itself. Circumstantial evidence from studies of ova in frogs and toads suggest that some of the RNA may act as the early organiser of mammalian development. Whatever the outcome of the additional studies required to elucidate these complex problems, the dicytate stage should clearly not be referred to as a resting phase, since the oocytes show a high degree of metabolic and synthetic activity at a time when the follicular envelope consist of only a few flattened epithelia cells.
Human oocyte/folicular development is a controlled mechanism both in time and biological processes. 99% of all ova are arrested in the meiotic prophase (that is the dicytate stage) even before birth. From this pool a couple of hundred ova are selected and activated every month (the menstrual cycle) for further development that is resumption of the meiosis though germinal vesicle breakdown (GVB) and Metaphase I (MF-I) to Metaphase II (MF-II) and growth of the follicle.
The reproductive state of the human female is cyclic with a complex interaction between the hypothalamus, anterior pituitary and the ovaries eventually leading to the process of ovulation. The cycle is repeated with an average period of 28-30 days (range 25-35 days). The first phase, menstruation, last 4-5 days. The first day of a cycle is the first day of the first phase, that is the first day of menstrual bleeding. The second follicular phase of the ovary corresponds to the proliferative phase of the endometrium and lasts 10-16 days (i.e. highly variable). Then follows an ovulatory phase (36 hours) and finally a luteal phase which corresponds to the secretory phase of the endometrium and is usually constant at 14 days. Three components critical to the understanding of the menstrual cycle:
1. hypothalamic GnRH control of FSH/LH release,
2. ovarian follicular development to ovulation and subsequent corpus luteum formation and
3. the feedback control of FSH/LH secretion by ovarian hormones.
During the second half of the reproductive cycle, the corpus luteum develops and secretes both oestrogen and progesterone. Oestrogen continues to promote the proliferative activity in the endometrium. Progesterone on the other hand causes the endometrial glands to become distended with secretory products which include glycogen (important for the developing embryo should implantation occur). Endometrial blood flow increases and the spiral arteries become coiled and twisted. The second half of the cycle is called the luteal phase (ovaries) or the secretory phase (uterus). If implantation does not occur then the corpus luteum regresses, there is a rapid fall in the secretion of oestrogen and progesterone, the endometrium undergoes shrinkage due to extracellular fluid loss, the spiral arteries constrict, the endometrial blood flow decreases with cell death and destruction of blood vessels. These chances eventually lead to a phase of menstrual bleeding where all but the basal layer of the endometrium is lost. The first day of the menstrual bleeding is, for practical reasons the first day of the menstrual cyclus.
Ova maturation is the final stage of ova development that prepares for fertilisation and embryo development. It can be divided into two general processes: nuclear maturation and cytoplasmic maturation. Nuclear maturation is defined as the resumption of meiosis and progression to MF-II while cytoplasmic maturation is defined as the extragenomic changes that prepare the egg for activation, pronuclear formation, and early embryogenesis.
The follicle surrounding the ova in the dicytate stage comprises a single layer of flattened granulosa cells. The signs of further development of the ova and follicle involves further multiplication in the number of granulosa cells and also the passage of fluid into spaces between the granulosa cells. As the quantity of fluid increases, the cavities that it occupies increases in size and become confluent to form an antrum. The follicle is now said to be of the Graafian type. With the further expansion of the antrum, the ova occupies a position at one side of the follicle and is surrounded by two or more layers of granulosa cells. The innermost layer of these cells becomes columnar in the shape and constitutes the corona radiata which, as the innermost part of the cumulus oophorus, persists around the egg for a period after ovulation.
The nuclear maturation proceeds in parallel with the follicular development Thus, GVB and MF-I ova are observed during the formation of the antrum that is in the antral and pre-antral follicle. By the time the antrum is fully developed the meiotic division is completed and the first polar body extrusion is seen.
A few hundred ova reach this stage during each menstrual cycle. However, due to presently unexplained factors, the majority of these enter atresia, wherein the ova undergo an apoptotic process and die. The dissolution of the innermost part of the granulosa cells whilst the egg is still in the follicle, or precociously after its departure, is a sure sign that degenerative changes are occurring that will result in the death of the ova. Only a few, normally only 1, will proceed the development.
Technical Background of IVF
If the woman rece
Lindenberg Svend
Mikkelsen Anne Lis
Birch & Stewart Kolasch & Birch, LLP
Chin Christopher L.
Do Pensee T.
Medi-Cult A/S
LandOfFree
Assay to indicate the presence of non-fertilizable ova does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Assay to indicate the presence of non-fertilizable ova, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Assay to indicate the presence of non-fertilizable ova will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3173923