The Journal of Clinical Endocrinology & Metabolism, 2012
The 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia. Pregnan... more The 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia. Pregnant women presenting a risk of genetic transmission may be treated with synthetic glucocorticoids such as dexamethasone (DEX) to prevent female fetus virilization. The aim of this study was to assess the potential deleterious effects of DEX exposure on fetal ovarian development. Human fetal ovaries, ranging from 8-11 weeks after fertilization, were harvested from material available after legally induced abortions. They were cultured in the absence or presence of DEX (2, 10, or 50 μm) over 14 d, and histological analyses were performed. The glucocorticoid receptor NR3C1 was present and the signaling pathway active in the fetal ovary as demonstrated by the expression of NR3C1 target genes, such as PLZF and FKBP5, in response to DEX exposure. DEX decreased germ cell density at the 10 and 50 μm doses. Exposure to DEX, even at the highest dose, did not change oogonial proliferation as monitored by 5-bromo-2'-deoxyuridine incorporation and significantly increased the apoptotic rate, detected with cleaved caspase 3 staining. Interestingly, the expression of the prosurvival gene KIT was significantly decreased in the presence of DEX during the course of the culture. We have demonstrated for the first time that in vitro exposure to high doses of DEX impairs human fetal oogenesis through an increase in apoptosis. These data are of high importance, and additional epidemiological studies are required to investigate the female fertility of those women who have been exposed to DEX during fetal life.
Journal de Gynécologie Obstétrique et Biologie de la Reproduction, 2007
Le rinçage folliculaire lors des prélèvements ovocytaires double le taux de grossesses en féconda... more Le rinçage folliculaire lors des prélèvements ovocytaires double le taux de grossesses en fécondation in vitro en cycle semi-naturel
We identified three doublesex and mab-3-related transcription factors (DMRT) that were sexually d... more We identified three doublesex and mab-3-related transcription factors (DMRT) that were sexually differentially expressed in human fetal gonads and present in the ovaries at the time of meiotic initiation. These were also identified in murine embryonic female germ cells. Among these, we focused on DMRTA2 (DMRT5), whose function is unknown in the developing gonads, and clarified its role in human female fetal germ cells, using an original xenograft model. Early human fetal ovaries (8-11 weeks post-fertilization) were grafted into nude mice. Grafted ovaries developed normally, with no apparent overt changes, when compared with ungrafted ovaries at equivalent developmental stages. Appropriate germ cell density, mitotic/meiotic transition, markers of meiotic progression and follicle formation were evident. Four weeks after grafting, mice were treated with siRNA, specifically targeting human DMRTA2 mRNA. DMRTA2 inhibition triggered an increase in undifferentiated FUT4-positive germ cells ...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2011
Although female cancer incidence may be on rise, antineoplastic regimens have become more success... more Although female cancer incidence may be on rise, antineoplastic regimens have become more successful. As a result, an increasing number of women with cancer survive to endure the long-term consequences of chemotherapy. One of the most important long-term consequences of cancers treatments in young female is premature ovarian failure and infertility. Because of the increasing survival rates, many of these young women are seeking methods to preserve their fertility. Currently, embryo/oocytes cryoporeservation obtained after ovarian stimulation appears to provide the best fertility preservation option. However, patients may not have sufficient time to undergo ovarian stimulation prior to chemotherapy and/or the hormones used in ovarian stimulation are contra-indicated for estrogen-dependant tumors. In vitro maturation of oocytes (IVM) has been suggested to avoid ovarian stimulation and time requirement in patients with cancer, and can be combined with ovarian tissue cryobanking. In thi...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2011
Analyzing the results and validating the procedure of testicular sperm extraction (TESE) performe... more Analyzing the results and validating the procedure of testicular sperm extraction (TESE) performed on the day of oocyte retrieval in non obstructive azoospermia (NOA) patients. Sixty TESE were performed on the day of oocyte retrieval (dOR), in 52 NOA men. Patients were sorted into three groups according to the results of the surgical procedure: 1: sperm recovery with possible sperm freezing (n=20); 2: sperm recovery without freezing (n=27); 3: "negative" biopsy (n=13). ICSI outcomes in the two groups with sperm recovery were compared to those of ICSI performed with frozen-thawed sperm obtained from TESE performed (n=13). The rate of positive sperm retrieval was 78%. While the overall clinical pregnancy rate was 50%, no difference in the fertilization, implantation and clinical pregnancy rates was found in the two groups with positive sperm retrieval as compared to frozen-thawed sperm group. Twelve pregnancies were obtained in patients without further sperm cryopreservation...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2009
To evaluate quality control impact, in assisted reproductive medecine, in view of the stability o... more To evaluate quality control impact, in assisted reproductive medecine, in view of the stability of results. Prospective collection of all IVF cycles at the center of AMP at Antoine-Béclère hospital (Clamart), as well as pregnancy outcomes, from 2002 till 2006. 44.3% in the clinical pregnancy rate and 38.0% in the delivery rate per oocyte retrieval. These rates are stable during the five years studied. The rate of multiple pregnancy is of 30.7%, decreasing, with the decrease in number of transferred embryos. Results are stable over five years duration. Quality control seems to be necessary.
Journal de gynécologie, obstétrique et biologie de la reproduction, 2007
To evaluate the importance of follicular flushing on semi natural cycle IVF. We have compared pro... more To evaluate the importance of follicular flushing on semi natural cycle IVF. We have compared prospectively the reproductive potential of oocytes obtained from follicular fluid (LF, N = 79) to those obtained from follicular flushing (R, N = 47) in 146 oocyte pick ups. The group LF and R were similar with regard to fertilization rate (79.7 versus 88.1%, respectively), percentage of superior grade embryos (28.8 versus 37.8%) and implantation rate (24.1 versus 44.1%). The practice of follicular flushing on semi natural IVF cycle improves the pregnancy rate. The oocytes obtained by follicular flushing had the same reproductive potential than those obtained on follicular fluid.
In-vitro maturation of oocytes (IVM) is a new IVF technology developed in order to avoid iatrogen... more In-vitro maturation of oocytes (IVM) is a new IVF technology developed in order to avoid iatrogenic complications of standard IVF treatments. This technique is particularly useful in patients suffering from polycystic ovary syndrome (PCOS) who are concerned with the risk of ovarian hyperstimulation syndrome. This technique is nowadays routinely practised in many international centres. However, the efficiency of this technique needs to be improved for a better support of maturation conditions to maximize oocyte developmental competence. In order to improve IVM results, the efficiency of two IVM media was retrospectively compared. Ninety-three PCOS candidates undergoing their first IVM cycle were included in this study, and IVM was conducted with TCM-199 or IVM-Medicult medium. This is the first study comparing two maturation media. Both media resulted in the same results concerning total oocyte maturation, fertilization, early embryo development and pregnancy rates.
To understand the psychological motives that led these couples to take risks involved in genetic ... more To understand the psychological motives that led these couples to take risks involved in genetic transmission and the possibility of another spontaneous pregnancy. To report our psychoanalytic experience to explain why after a request of pre-implantation genetic diagnosis (PGD), some women prefer to be spontaneously pregnant. Between 2000 and 2004, out of 643 requests of PGD, 541 have been admitted, among which 123 spontaneous pregnancies appeared during the process of PGD. Some encounters with the patients have shown some reasons to these spontaneous pregnancies - denial of IVF, ambivalence of this desire for a child, denial of the risk, control of the procreation, family loyalty and fertility affirmation. The resort to PGD is more ambivalent than the thought of it because it means, beyond the request, conflict with transmission, culture, and history origins.
Couples with a risk of transmitting X-linked diseases included in a preimplantation genetic diagn... more Couples with a risk of transmitting X-linked diseases included in a preimplantation genetic diagnosis (PGD) center need early and rapid fetal sex determination during pregnancy in two situations. The first situation corresponds to control of embryo sexing after PGD, the second one being that of couples in PGD program having a spontaneous pregnancy. Determination of fetal sex can be achieved by karyotyping using invasive procedures such as chorionic villus sampling (CVS), amniocentesis or cordocentesis and by non-invasive procedures such as ultrasound (US) examination. CVS is the earliest invasive procedure for fetal sex determination and molecular analysis of X-linked genetic disorders during the first trimester but it is associated with a risk of fetal loss. US allows reliable fetal sex determination only during the second trimester. Recently, reliable non-invasive fetal sex determination was realized by using SRY gene amplification in maternal serum. We report the prospective use ...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2004
Assisted hatching (AH) has been proposed to improve the implantation rate in patients with poor p... more Assisted hatching (AH) has been proposed to improve the implantation rate in patients with poor prognosis for pregnancy. This systematic review of randomized controlled trials addresses primary outcomes of clinical pregnancy and embryo implantation. Several different methods for AH have been introduced over the years but comparative studies are lacking. AH using laser technology is probably the best technique. Assisted hatching increases the pregnancy and implantation rates significantly in patients with a poor prognosis undergoing IVF or ICSI, particularly in older women. More randomized trials are required to confirm any positive effects on the take-home-baby rate and to compare the different techniques.
Morphologie : bulletin de l'Association des anatomistes, 1998
The extended use of Fish with centromeric probes in many cytogenetic laboratories is often impair... more The extended use of Fish with centromeric probes in many cytogenetic laboratories is often impaired by the cost of this technique. Polymerase Chain Reaction (PCR) constitutes a simple way to generate and label such centromeric probes at low cost. Two types of human DNA source can be used: 1--Somatic hybrid cell lines containing a unique human chromosome. The specific amplification of the human subset of alphoid DNA is realised with a primer pair specific for the consensus region of human alpha satellite sequence. 2--Total Human DNA. This time, a primer pair specific for the alpha satellite DNA of the chromosome of interest must be designed. These probes, labelled during the PCR reaction by direct incorporation of modified dUTP, are actually widely used in our laboratory, alone or mixed with other probes (chromosome painting or locus specific probes).
a Unité d'assistance médicale à la procréation, service de gynécologie-obstétrique (Pr Frydman), ... more a Unité d'assistance médicale à la procréation, service de gynécologie-obstétrique (Pr Frydman), 157, rue de la Porte-de-Trivaux,
ABSTRACT Cryopreservation of human oocytes might provide an alternative approach to freezing supe... more ABSTRACT Cryopreservation of human oocytes might provide an alternative approach to freezing supernumerary embryos obtained during IVF. This process, performed on immature denuded prophase 1 mouse oocytes, was investigated. We first investigated the capacity of frozen, immature, murine oocytes to continue in vitro maturation after thawing. We then evaluated the risk to offspring from chromosomal damage by cytogenetical and cytological (spindle) analysis. Finally, we attempted to determine the reasons for and the stage of maturation failure. A total of 700 immature oocytes was frozen, 629 (90%) were recovered intact after thawing, and 53% extruded the first polar body, versus 74% for the control group. Freezing was not accompanied by an increase in aneuploidy in maturing oocytes (18 and 15% for thawed and control oocytes, respectively). Consequently, the first meiotic division occurred normally, without an increase in nondisjunction. Spindle analysis demonstrated only a few abnormalities (15 and 2% for thawed and control oocytes, respectively) incompatible with further development. Oocytes arrested during in vitro maturation were mainly at the metaphase I stage (64 and 76% for thawed and control oocytes, respectively). Whereas 17% of thawed oocytes were blocked before the formation of the first meiotic spindle, this never occurred in the control group. Immature murine oocytes can withstand cryo-preservation, which is encouraging for future human application of this technique.
The Journal of Clinical Endocrinology & Metabolism, 2012
The 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia. Pregnan... more The 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia. Pregnant women presenting a risk of genetic transmission may be treated with synthetic glucocorticoids such as dexamethasone (DEX) to prevent female fetus virilization. The aim of this study was to assess the potential deleterious effects of DEX exposure on fetal ovarian development. Human fetal ovaries, ranging from 8-11 weeks after fertilization, were harvested from material available after legally induced abortions. They were cultured in the absence or presence of DEX (2, 10, or 50 μm) over 14 d, and histological analyses were performed. The glucocorticoid receptor NR3C1 was present and the signaling pathway active in the fetal ovary as demonstrated by the expression of NR3C1 target genes, such as PLZF and FKBP5, in response to DEX exposure. DEX decreased germ cell density at the 10 and 50 μm doses. Exposure to DEX, even at the highest dose, did not change oogonial proliferation as monitored by 5-bromo-2'-deoxyuridine incorporation and significantly increased the apoptotic rate, detected with cleaved caspase 3 staining. Interestingly, the expression of the prosurvival gene KIT was significantly decreased in the presence of DEX during the course of the culture. We have demonstrated for the first time that in vitro exposure to high doses of DEX impairs human fetal oogenesis through an increase in apoptosis. These data are of high importance, and additional epidemiological studies are required to investigate the female fertility of those women who have been exposed to DEX during fetal life.
Journal de Gynécologie Obstétrique et Biologie de la Reproduction, 2007
Le rinçage folliculaire lors des prélèvements ovocytaires double le taux de grossesses en féconda... more Le rinçage folliculaire lors des prélèvements ovocytaires double le taux de grossesses en fécondation in vitro en cycle semi-naturel
We identified three doublesex and mab-3-related transcription factors (DMRT) that were sexually d... more We identified three doublesex and mab-3-related transcription factors (DMRT) that were sexually differentially expressed in human fetal gonads and present in the ovaries at the time of meiotic initiation. These were also identified in murine embryonic female germ cells. Among these, we focused on DMRTA2 (DMRT5), whose function is unknown in the developing gonads, and clarified its role in human female fetal germ cells, using an original xenograft model. Early human fetal ovaries (8-11 weeks post-fertilization) were grafted into nude mice. Grafted ovaries developed normally, with no apparent overt changes, when compared with ungrafted ovaries at equivalent developmental stages. Appropriate germ cell density, mitotic/meiotic transition, markers of meiotic progression and follicle formation were evident. Four weeks after grafting, mice were treated with siRNA, specifically targeting human DMRTA2 mRNA. DMRTA2 inhibition triggered an increase in undifferentiated FUT4-positive germ cells ...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2011
Although female cancer incidence may be on rise, antineoplastic regimens have become more success... more Although female cancer incidence may be on rise, antineoplastic regimens have become more successful. As a result, an increasing number of women with cancer survive to endure the long-term consequences of chemotherapy. One of the most important long-term consequences of cancers treatments in young female is premature ovarian failure and infertility. Because of the increasing survival rates, many of these young women are seeking methods to preserve their fertility. Currently, embryo/oocytes cryoporeservation obtained after ovarian stimulation appears to provide the best fertility preservation option. However, patients may not have sufficient time to undergo ovarian stimulation prior to chemotherapy and/or the hormones used in ovarian stimulation are contra-indicated for estrogen-dependant tumors. In vitro maturation of oocytes (IVM) has been suggested to avoid ovarian stimulation and time requirement in patients with cancer, and can be combined with ovarian tissue cryobanking. In thi...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2011
Analyzing the results and validating the procedure of testicular sperm extraction (TESE) performe... more Analyzing the results and validating the procedure of testicular sperm extraction (TESE) performed on the day of oocyte retrieval in non obstructive azoospermia (NOA) patients. Sixty TESE were performed on the day of oocyte retrieval (dOR), in 52 NOA men. Patients were sorted into three groups according to the results of the surgical procedure: 1: sperm recovery with possible sperm freezing (n=20); 2: sperm recovery without freezing (n=27); 3: "negative" biopsy (n=13). ICSI outcomes in the two groups with sperm recovery were compared to those of ICSI performed with frozen-thawed sperm obtained from TESE performed (n=13). The rate of positive sperm retrieval was 78%. While the overall clinical pregnancy rate was 50%, no difference in the fertilization, implantation and clinical pregnancy rates was found in the two groups with positive sperm retrieval as compared to frozen-thawed sperm group. Twelve pregnancies were obtained in patients without further sperm cryopreservation...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2009
To evaluate quality control impact, in assisted reproductive medecine, in view of the stability o... more To evaluate quality control impact, in assisted reproductive medecine, in view of the stability of results. Prospective collection of all IVF cycles at the center of AMP at Antoine-Béclère hospital (Clamart), as well as pregnancy outcomes, from 2002 till 2006. 44.3% in the clinical pregnancy rate and 38.0% in the delivery rate per oocyte retrieval. These rates are stable during the five years studied. The rate of multiple pregnancy is of 30.7%, decreasing, with the decrease in number of transferred embryos. Results are stable over five years duration. Quality control seems to be necessary.
Journal de gynécologie, obstétrique et biologie de la reproduction, 2007
To evaluate the importance of follicular flushing on semi natural cycle IVF. We have compared pro... more To evaluate the importance of follicular flushing on semi natural cycle IVF. We have compared prospectively the reproductive potential of oocytes obtained from follicular fluid (LF, N = 79) to those obtained from follicular flushing (R, N = 47) in 146 oocyte pick ups. The group LF and R were similar with regard to fertilization rate (79.7 versus 88.1%, respectively), percentage of superior grade embryos (28.8 versus 37.8%) and implantation rate (24.1 versus 44.1%). The practice of follicular flushing on semi natural IVF cycle improves the pregnancy rate. The oocytes obtained by follicular flushing had the same reproductive potential than those obtained on follicular fluid.
In-vitro maturation of oocytes (IVM) is a new IVF technology developed in order to avoid iatrogen... more In-vitro maturation of oocytes (IVM) is a new IVF technology developed in order to avoid iatrogenic complications of standard IVF treatments. This technique is particularly useful in patients suffering from polycystic ovary syndrome (PCOS) who are concerned with the risk of ovarian hyperstimulation syndrome. This technique is nowadays routinely practised in many international centres. However, the efficiency of this technique needs to be improved for a better support of maturation conditions to maximize oocyte developmental competence. In order to improve IVM results, the efficiency of two IVM media was retrospectively compared. Ninety-three PCOS candidates undergoing their first IVM cycle were included in this study, and IVM was conducted with TCM-199 or IVM-Medicult medium. This is the first study comparing two maturation media. Both media resulted in the same results concerning total oocyte maturation, fertilization, early embryo development and pregnancy rates.
To understand the psychological motives that led these couples to take risks involved in genetic ... more To understand the psychological motives that led these couples to take risks involved in genetic transmission and the possibility of another spontaneous pregnancy. To report our psychoanalytic experience to explain why after a request of pre-implantation genetic diagnosis (PGD), some women prefer to be spontaneously pregnant. Between 2000 and 2004, out of 643 requests of PGD, 541 have been admitted, among which 123 spontaneous pregnancies appeared during the process of PGD. Some encounters with the patients have shown some reasons to these spontaneous pregnancies - denial of IVF, ambivalence of this desire for a child, denial of the risk, control of the procreation, family loyalty and fertility affirmation. The resort to PGD is more ambivalent than the thought of it because it means, beyond the request, conflict with transmission, culture, and history origins.
Couples with a risk of transmitting X-linked diseases included in a preimplantation genetic diagn... more Couples with a risk of transmitting X-linked diseases included in a preimplantation genetic diagnosis (PGD) center need early and rapid fetal sex determination during pregnancy in two situations. The first situation corresponds to control of embryo sexing after PGD, the second one being that of couples in PGD program having a spontaneous pregnancy. Determination of fetal sex can be achieved by karyotyping using invasive procedures such as chorionic villus sampling (CVS), amniocentesis or cordocentesis and by non-invasive procedures such as ultrasound (US) examination. CVS is the earliest invasive procedure for fetal sex determination and molecular analysis of X-linked genetic disorders during the first trimester but it is associated with a risk of fetal loss. US allows reliable fetal sex determination only during the second trimester. Recently, reliable non-invasive fetal sex determination was realized by using SRY gene amplification in maternal serum. We report the prospective use ...
Journal de gynécologie, obstétrique et biologie de la reproduction, 2004
Assisted hatching (AH) has been proposed to improve the implantation rate in patients with poor p... more Assisted hatching (AH) has been proposed to improve the implantation rate in patients with poor prognosis for pregnancy. This systematic review of randomized controlled trials addresses primary outcomes of clinical pregnancy and embryo implantation. Several different methods for AH have been introduced over the years but comparative studies are lacking. AH using laser technology is probably the best technique. Assisted hatching increases the pregnancy and implantation rates significantly in patients with a poor prognosis undergoing IVF or ICSI, particularly in older women. More randomized trials are required to confirm any positive effects on the take-home-baby rate and to compare the different techniques.
Morphologie : bulletin de l'Association des anatomistes, 1998
The extended use of Fish with centromeric probes in many cytogenetic laboratories is often impair... more The extended use of Fish with centromeric probes in many cytogenetic laboratories is often impaired by the cost of this technique. Polymerase Chain Reaction (PCR) constitutes a simple way to generate and label such centromeric probes at low cost. Two types of human DNA source can be used: 1--Somatic hybrid cell lines containing a unique human chromosome. The specific amplification of the human subset of alphoid DNA is realised with a primer pair specific for the consensus region of human alpha satellite sequence. 2--Total Human DNA. This time, a primer pair specific for the alpha satellite DNA of the chromosome of interest must be designed. These probes, labelled during the PCR reaction by direct incorporation of modified dUTP, are actually widely used in our laboratory, alone or mixed with other probes (chromosome painting or locus specific probes).
a Unité d'assistance médicale à la procréation, service de gynécologie-obstétrique (Pr Frydman), ... more a Unité d'assistance médicale à la procréation, service de gynécologie-obstétrique (Pr Frydman), 157, rue de la Porte-de-Trivaux,
ABSTRACT Cryopreservation of human oocytes might provide an alternative approach to freezing supe... more ABSTRACT Cryopreservation of human oocytes might provide an alternative approach to freezing supernumerary embryos obtained during IVF. This process, performed on immature denuded prophase 1 mouse oocytes, was investigated. We first investigated the capacity of frozen, immature, murine oocytes to continue in vitro maturation after thawing. We then evaluated the risk to offspring from chromosomal damage by cytogenetical and cytological (spindle) analysis. Finally, we attempted to determine the reasons for and the stage of maturation failure. A total of 700 immature oocytes was frozen, 629 (90%) were recovered intact after thawing, and 53% extruded the first polar body, versus 74% for the control group. Freezing was not accompanied by an increase in aneuploidy in maturing oocytes (18 and 15% for thawed and control oocytes, respectively). Consequently, the first meiotic division occurred normally, without an increase in nondisjunction. Spindle analysis demonstrated only a few abnormalities (15 and 2% for thawed and control oocytes, respectively) incompatible with further development. Oocytes arrested during in vitro maturation were mainly at the metaphase I stage (64 and 76% for thawed and control oocytes, respectively). Whereas 17% of thawed oocytes were blocked before the formation of the first meiotic spindle, this never occurred in the control group. Immature murine oocytes can withstand cryo-preservation, which is encouraging for future human application of this technique.
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Papers by N. Frydman