Projects / Programmes
Gene expression in cumulus cells of ovarian follicles in in vitro fertilization cycles
| Code |
Science |
Field |
Subfield |
| 3.05.00 |
Medical sciences |
Human reproduction |
|
| Code |
Science |
Field |
| B570 |
Biomedical sciences |
Obstetrics, gynaecology, andrology, reproduction, sexuality |
| Code |
Science |
Field |
| 3.05 |
Medical and Health Sciences |
Other medical sciences |
cumulus cells, granulosa cells, biomarkers, oligonucleotide microarrays, in vitro fertilization, GnRH analogues, oocyte quality, blastocyst
Researchers (31)
Organisations (2)
Abstract
Introduction: In vitro fertilization (IVF) procedures have significantly improved pregnancy rate in infertile couples. However, their success is limited. Although most oocytes are capable of fertilization, only half of them develop into embryos and still fewer implant. In order to increase success rates of IVF more than one embryo is being transferred. This, on the other hand, increases the risk of multiple pregnancy with harmful consequences to the mother and the child. To avoid adverse outcomes, single embryo transfer is increasingly being used. This means that for the transfer the embryo with the highest implantation potential that will lead to a successful pregnancy is to be selected from among all available embryos.
A high-quality oocyte plays an essential role in the development of a high-quality embryo. Oocyte selection for fertilization and selection of embryos for transfer is currently based on the evaluation of subjective morphological criteria. As morphological evaluation is not a reliable method for the assessment of oocyte competence and embryo implantation potential, new, non-invasive, objective and reliable indicators of oocyte and embryo quality have been increasingly searched for. Bidirectional communication between the oocyte and granulosa (GC) and cumulus (CC) cells is crucial for the development of competent oocytes. We assume that the differences in intrafollicular processes responsible for the development of high-quality oocytes and embryos with high implantation potential are reflected in gene expression of GC and CC. Thus it would be possible to predict the quality of oocytes and embryos with the gene expression analysis of these cells.
Controlled ovarian stimulation with gonadotropins in combination with gonadotropin-releasing hormone (GnRH) antagonists or agonists is being used in IVF. The reports on pregnancy and live birth rates generated by either protocol are opposing. To date no study whole genome analysis of CC has been made to identify biomarkers, which would be predictive of blastocyst development regarding the GnRH analogue used.
Aim: Using genome wide gene expression analysis of GC and CC we will identify the differential gene expression in these cells between oocytes that will fertilize and those that will not, and between embryos that will implant and those that will not. Identification of reliable biomarkers would reduce the need for multiple embryo transfer without lowering IVF success rates.
Further, in this prospective randomised study, we will identify the biomarkers of blastocyst development regarding the protocol used. Additionally, we aim at finding whether biomarkers of blastocyst development are different regarding the protocol used for ovarian stimulation.
The comparison of CC gene expression in unfertilized oocytes and CC gene expression in oocytes that develop into blastocysts will permit identification of biomarkers of blastocyst development. We assume that biomarkers of blastocyst development will be similar regardless of the GnRH analogue used.
Methods: The CC whole genome gene expression will be analyzed using microarray technique. The gene expression of potential biomarkers will be analysed using real time polymerase chain reaction (qPCR).
Expected results: Based on CC and GC gene expression, biomarkers of oocytes which will fertilize and oocytes which will develop to blastocysts will be identified. These biomarkers will be useful in cases of selective fertilization of a limited number of oocytes or in the decision on which oocytes or embryos are appropriate for transfer or freezing.
We expect that biomarkers in an individual GnRH analogue treatment protocol will be different, but not in the group of biomarkers representative of the oocyte development to the blastocyst stage. This would imply that the GnRH antagonist protocol, which is more patient friendly, does not differ in efficiency from the GnRH agonist protocol at the bio-molecular level.
Significance for science
Functional analyses of differentially expressed genes revealed that enriched biological pathways differ according to the cell type. In granulosa cells (GC), the highly expressed genes represent biological functions as inflammatory and immune response and cell communication. In cumulus cells (CC), the highly expressed genes represent biological functions as multicellular organismal development and signal transduction. Furthermore, in the present study we discovered two genes, that had not previously been described in human GC and CC; PROK2, which was highly expressed in GC, and PNCK which was highly expressed in CC,. PROK2 was a part of the gene network connected with haematological system development and function, and for this reason we presume that this gene could have a role in angiogenesis during follicular development. The central gene of this network was TNF. During mammalian ovulation, TNF causes the apoptosis of ovarian surface epithelium and breakdown of the extracellular matrix in the follicle wall allowing the oocyte to be released from the follicle. The connection of PROK2 with TNF might thus indicate that PROK2 is involved in the process of releasing the cumulus-oocyte complex from the follicle at the time of ovulation. PNCK gene inhibits MAPK signalling pathway, whose activation is crucial for final oocyte maturation and GC luteinization. Based on the results of our study we presume that PNCK in CC maintains inactivation of MAPK pathway after the completion of oocyte maturation. These results have upgraded existing data on differentially expressed genes between GC and CC.
Significance for the country
Regarding the CC gene expression results we cofirmed that protocols of ovarian stimulation either combination with GnRH agonists or GnRH antagonists are comparable. There was no CC gene expression difference regarding between different GnRH analogue used on various oocyte maturation stages. These results supports results of metaanalyses of clinical studies which showed non significant difference in live born babies between different GnRH analogue used.
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