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P De Sutter

Publications and source records attributed to P De Sutter.

At least 55 records · Page 3Linked to original sources
PubMed1997

Triple colour fluorescent in-situ hybridization for chromosomes X,Y and 1 on spare human embryos.

The potential for implantation of human embryos obtained by in-vitro fertilization is presumably determined to a large extent by their chromosomal constitution but cytogenetic analysis of preimplantation embryos has been hampered by a number of practical and technical problems. With the advent of fluorescent in-situ hybridization (FISH) a practical method for numerical chromosomal analysis has become available. A limited amount of data has been obtained with FISH on human embryos using probes binding to chromosomes X, Y, 16, 18 and 13/21 combined or for chromosomes X and Y or 1 and 17. It was our purpose to extend these data by the combined analysis of chromosomes X, Y and 1 in spare human embryos. A short fluorescent in-situ hybridization procedure involving the simultaneous use of three deoxyribonucleic acid probes detected with red, green, and a mixture of red and green was used to determine chromosomal abnormalities in 116 spare embryos with a poor morphological score and/or displaying one or more multinucleated blastomeres. The majority of the embryos was obtained by intracytoplasmic sperm injection. Less than half of the embryos (n = 54) were diploid and only 39 of them were uniformly XY11 or XX11; two embryos showed a non-disjunction and 13 embryos were aneuploid. Of the remainder, 22 were mosaic, nine were either haploid, triploid or tetraploid and 12 embryos were classified as chaotic. The latter pattern was particularly frequent in multinucleated blastomeres. Our data are comparable with those obtained with FISH using other chromosomal probes and confirm that the majority of preimplantation embryos carry a numerical chromosomal defect. Aneuploidy for chromosome 1 does not appear to be more common in preimplantation embryos than is reported for other chromosomes. Although the high incidence of chromosomal anomalies is presumably biased by the fact that only embryos with a poor morphological score were analysed, it nevertheless indicates that natural selection is the foremost reason for the low implantation rates of human preimplantation embryos in in-vitro fertilization (IVF) programmes.

Chromosome Aberrations↗
PubMed1997

Transport-in-vitro fertilization/intracellular sperm injection: a prospective randomized study.

We performed a prospective randomized clinical trial to investigate whether long distance oocyte transport prior to an intracytoplasmic sperm injection (ICSI) procedure influences fertilization rates, embryo quality and/or embryo implantation rates. After informed consent, 100 infertile couples booked for ICSI treatment were randomized into two groups. In group 1 (n = 50), patients were stimulated and monitored in Rotterdam (The Netherlands), and oocyte retrieval, ICSI procedure and embryo transfer took place in Gent (Belgium). In group 2 (n = 50), patients were stimulated, monitored and punctured in Rotterdam and the oocytes were transported in their follicular fluid in an isothermic transport box to Gent, where the ICSI procedure and the embryo transfer took place. In both groups the stimulation and monitoring regimen, puncture technique, laboratory conditions and transfer policy were identical. In both groups, the number of fertilized oocytes (7.13 +/- 0.65 versus 5.53 +/- 0.60, P = 0.08), the number of transferred embryos (2.36 +/- 0.09 versus 2.40 +/- 0.11, P = 0.87) and the embryo implantation rate [20/113 (17.7%) versus 19/103 (18.4%), P = 0.89] was similar. In group 1, the number of retrieved oocytes was higher (10.83 +/- 0.95 versus 8.44 +/- 0.93, P = 0.05). The total score of the embryos obtained (18.90 +/- 1.73 versus 12.64 +/- 1.26, P = 0.01), the number of good quality embryos (4.63 +/- 0.49 versus 2.98 +/- 0.38, P = 0.02), the mean score of the transferred embryos (3.32 +/- 0.11 versus 2.94 +/- 0.13, P = 0.05) and the number of embryos available for cryopreservation (2.70 +/- 0.45 versus 1.48 +/- 0.38, P = 0.03) were significantly higher in group 1. Therefore, long distance transport of oocytes prior to ICSI does not affect oocyte fertilization and embryo implantation rates, although a negative effect on embryo quality cannot be excluded.

Adult↗
PubMed1997

Disintegration of chromosomes in dead sperm cells as revealed by injection into mouse oocytes.

Intracytoplasmic sperm injection of immotile (dead) ejaculated human spermatozoa has been carried out by several centres for the treatment of infertility caused by severe asthenozoospermia (necrozoospermia). No healthy pregnancies have been reported as yet, suggesting irreversible damage to sperm DNA, centrioles and/or other important structures. We investigated this hypothesis by injection of immotile human spermatozoa obtained from several male infertility patients into mouse oocytes and analysis of the oocyte activation rate and sperm chromosome integrity. Motile spermatozoa of the same sample were used as a control. The proportion of living spermatozoa among the immotile was also assessed in each sample and was related to the results of the mouse oocyte injection test. The oocyte activation rate after injection of immotile human spermatozoa into mouse oocytes was the same or only slightly lower than after injection with initially motile spermatozoa (87-100% versus 100% respectively). The rate of normal sperm chromosome spreads correlated significantly (r = 0.90, P < 0.05) with the proportion of living immotile spermatozoa in a given sample. It varied from 4 to 48% for samples containing respectively 8 and 40% of living spermatozoa. Most of the mouse oocytes injected and activated with immotile human spermatozoa were arrested during a prolonged period of time at the interphase of the first cell cycle (from 22 to 60%). Others underwent a delayed nuclear envelope breakdown but showed signs of abnormal structure of both male and female or only the male pronuclear chromosomes. Our data demonstrate an irreversible damage of chromosomes in dead ejaculated human spermatozoa and provide an experimental basis for recommending the use of testicular or epididymal spermatozoa for treatment of male infertility due to necrozoospermia.

Animals↗
PubMed1996

Transport intracytoplasmic sperm injection (ICSI): a cost-effective alternative.

PURPOSE: Transport in vitro fertilization (IVF) programs are operational in a lot of countries and especially popular in The Netherlands, where IVF activities are strictly regulated. Since the introduction of intracytoplasmic sperm injection (ICSI) in the IVF laboratory, many laboratories are now setting up this new technique, which necessitates major investments in terms of infrastructure and specialized personnel. METHODS: We present a cost effective alternative, consisting of patient selection, preparation, and oocyte retrieval at one center and transport of oocytes to a second center, where the ICSI procedure and embryo transfer are performed. Since early 1994 several Dutch centers have a transport ICSI program running with the Gent University Infertility Center, and we wish to present the results of our cooperation with two major centers, comparing them to our local results, for the first 10 months of 1994. Patient selection was similar at all three centers: only couples with previously failed in vitro fertilization or having been refused for routine IVF were enrolled in the program. Stimulation schemes and follow-up of the stimulation were different at all three centers. Transport of oocytes was carried out in a transport box or by attaching the closed tubes containing the follicular aspirates to the chest of the husband. Transport times varied between 1.5 and 3 hr, depending on traffic conditions. RESULTS: Up to November 1, 1994, a total of 77 transport ICSI cycles and 294 own ICSI cycles were carried out. Although locally significantly more oocytes were retrieved and thus available for ICSI than in transport cycles, fertilization and pregnancy rates were not different between the two groups. CONCLUSIONS: These results suggest that long-distance transport of human oocytes seems not to be harmful to their capacity to be successfully injected and to further embryonic development and their implantation potential. Transport ICSI seems to be a valuable and cost-effective approach to treat high numbers of patients at a restricted number of highly specialized IVF laboratories, especially in countries where ICSI is not commonly available.

Academic Medical Centers↗
PubMed1996

Does assisted hatching improve implantation rates after in vitro fertilization or intracytoplasmic sperm injection in all patients? A prospective randomized study.

PURPOSE: Preliminary data from some research centers indicate that assisted hatching might be of value to increase embryo implantation rate in the human, at least in selected cases. It is not clear, however, whether this technique would be of benefit for all patients undergoing an embryo transfer. We therefore performed a prospective randomized study to evaluate the effect of assisted hatching on the implantation rate in our in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) program. METHODS: In total, 120 couples undergoing an embryo transfer were randomized between two groups: in one group no assisted hatching was performed (AH-), whereas in the other group the embryos selected for transfer were subjected to partial zona dissection (PZD) immediately prior to the transfer (AH+). Using a computer-generated minimization procedure, patients were allocated to one of the two groups according to four pre-selected criteria: the number of embryos transferred, the cumulative score of transferred embryos, the age of the patient, and the use of ICSI. RESULTS: Pregnancy and implantation rates in the AH+ and AH- groups were, respectively, 42.1 versus 38.1% and 17.9 versus 17.1%. CONCLUSIONS: From our data we conclude that assisted hatching through partial zona dissection prior to embryo transfer does not improve pregnancy and embryo implantation rates in unselected patients undergoing IVF or ICSI.

Adult↗
PubMed1996

Unprotected freezing of human spermatozoa exerts a detrimental effect on their oocyte activating capacity and chromosome integrity.

The influence of unprotected freezing of mammalian spermatozoa on their oocyte activating capacity and chromosome integrity is unknown. However, this type of sperm treatment has been used in assisted reproduction by intracytoplasmic sperm injection in cattle and humans. The mouse oocyte injection test was used to analyse the influence of unprotected freezing of human spermatozoa on their reproductive characteristics. Mouse oocytes were microinjected with intact human spermatozoa or spermatozoa treated with two cycles of unprotected freeze-thawing. Oocytes surviving the injection were either cultured without further treatment or exposed to ethanol solution to induce parthenogenetic activation. Both injected and activated oocytes were used for sperm chromosome analysis. The results revealed a significant reduction in oocyte activating capacity and a tenfold increase in the incidence of structural chromosomal abnormalities in human spermatozoa treated by unprotected freezing. We conclude that unprotected freezing of human spermatozoa has a detrimental effect on their reproductive characteristics. Our data also provide a new perspective on the stability of mammalian spermatozoa to physical factors and demonstrate the importance of detailed analysis of the stability of sperm structures for successful development of new approaches in assisted reproduction.

Animals↗
PubMed1996

Oocyte morphology does not correlate with fertilization rate and embryo quality after intracytoplasmic sperm injection.

The fertilization rates and further development of 528 human metaphase II oocytes directly injected by a single spermatozoon were analysed with respect to their morphological features at the light microscopy level at the time of retrieval. The deviations of oocyte morphology which were most frequently observed, after removal of cumulus cells, were dark incorporations, dark zona pellucida, large perivitelline space, spots, vacuoles, refractile bodies and irregular shape. These deviations correlated neither with the fertilization rate nor with the embryo quality score, as compared to 'ideal' oocytes. Since the majority of oocytes displayed deviations from the 'ideal' morphotype but were still fertilized and developed in culture at a normal rate, they were probably as normal as 'ideal' oocytes. Since some of these morphotypes, such as refractile bodies, have been shown to be associated with failure of fertilization, it seems that intracytoplasmic sperm injection may be an appropriate method of treatment for couples in whom repeated failure of in-vitro fertilization is associated with the retrieval of dysmorphic oocytes in the presence of normal semen characteristics.

Cytoplasm↗
PubMed1996

Analysis of the oocyte activating capacity and chromosomal complement of round-headed human spermatozoa by their injection into mouse oocytes.

Intracytoplasmic sperm injection (ICSI) in the human is a very effective procedure which allows the fertilization of the majority of oocytes even in cases of extreme oligoasthenoteratozoospermia. Round-headed acrosomeless human spermatozoa, however, form an exception to this rule, because in about half of the couples with globozoospermia all oocytes remain unfertilized after injection. The incapacity of the spermatozoon to activate the oocyte following injection of round-headed spermatozoa could be the underlying mechanism. To investigate this hypothesis, activation rates of mouse oocytes injected with spermatozoa from a patient with globozoospermia were compared with those obtained after injection with normal spermatozoa. Of mouse oocytes surviving the injection with donor spermatozoa, 95% underwent activation, compared to none of the 88 mouse oocytes surviving the injection with round-headed spermatozoa. After fixation, prematurely condensed sperm chromosomes were found in these oocytes. Parthenogenetic activation of mouse oocytes (8% ethanol at 40 min after injection) injected with round-headed spermatozoa led to the activation of 96% of oocytes. These oocytes developed normally to the first mitosis and were fixed for analysis of the sperm karyotypes. The incidence of chromosomal abnormalities of round-headed spermatozoa (6%) was similar to that in spermatozoa from a fertile donor (9%). These data provide further information on the basic defect in cases of globozoospermia and demonstrate that globozoospermia is not associated with sperm karyotype abnormalities.

Adult↗
PubMed1995

Frequent deletion of chromosome 19 and a rare rearrangement of 19p13.3 involving the insulin receptor gene in human ovarian cancer.

Human ovarian cancer cells usually have multiple specific chromosomal deletions which can be detected by cytogenetic analysis or molecular techniques. Tumour suppressor genes might be located in these deleted chromosomal segments. The importance of these different loci is usually estimated from the frequency with which they are deleted. Here we report a 59% loss of heterozygosity for chromosome 19 in the DNA of human invasive epithelial ovarian cancer from a series of 37 patients. In all cases informative on both chromosomal arms a subchromosomal loss is observed. Analysis of the same tumours for chromosome 17p and 11p loss suggests that loss of chromosome 19p/q is less important than 17p loss, but more important than 11p loss. The deletion of chromosome 19q seems to be associated with distant, hematogeneous metastasis (stage IV). In two patients with high grade tumours, the deletion involves a rearrangement of the insulin receptor locus (19p13.3). This suggests that some of the previously described frequent cytogenetic 19p+ markers and 19p13.3 breaks observed in high grade ovarian cancers, might actually occur in the insulin receptor gene.

Adolescent↗
PubMed1995

Study of numerical aberrations of chromosome 1 by fluorescent in situ hybridization and DNA content by densitometric analysis on (pre)-malignant cervical lesions.

In an attempt to determine whether the fluorescent in situ hybridization (FISH) can be used as a rapid approach for the identification of aneuploidy in premalignant cervical smears, a centromeric probe for chromosome 1 was used. The results from the FISH experiments were compared with measurements of the overall DNA content obtained by means of an image analysis system. With progression to neoplasia, a decrease of the frequency of cells with two spots was observed, due to an increasing polysomy of chromosome 1. As far as the DNA content was concerned, an increasing DNA index and 5C-exceeding ratio (fraction of cells with a DNA content higher than 5C) was observed. Classification of the FISH results by a linear discriminant analysis revealed that 67.6% of the cases were classified in agreement with the CIN classification. These data suggest that chromosome 1 may be considered as a marker chromosome for pre-malignant cervical lesions and that the DNA content measurements are complementary to the FISH results.

Adult↗
PubMed1995

Sperm plasma membrane damage prior to intracytoplasmic sperm injection: a necessary condition for sperm nucleus decondensation.

In the present study we investigated the relevance of sperm immobilization prior to intracytoplasmic sperm injection (ICSI) in the fertilization process. Using supravital staining of the spermatozoa with eosin and studying sperm decondensation with 2 mM dithiothreitol (DTT) in conditions imitating sperm handling during ICSI, we demonstrated that immobilization of the spermatozoon by squeezing its tail between the glass pipette and the bottom of the dish damages the sperm plasma membrane. Polyvinylpyrrolidone (PVP), which is usually present in the drop with the spermatozoon to facilitate its handling, was found to impede the access of both eosin and DTT to the sperm nucleus. We conclude that (i) sperm immobilization prior to ICSI damages the sperm plasma membrane, that (ii) this damage is sufficient for thiol-reducing agents to gain access to the sperm nucleus, and finally that (iii) PVP possibly interferes with sperm nucleus decondensation.

Cell Membrane↗
PubMed1995

Timing of sperm and oocyte nuclear progression after intracytoplasmic sperm injection.

We investigated the time course of human oocyte activation after intracytoplasmic sperm injection (ICSI) by observing the oocyte chromosome configuration at different times after injection. One day old human oocytes were injected with spermatozoa and subjected to cytogenetic analysis at 2, 3, 4 and 5 h after injection. We found that anaphase is initiated in the vast majority of the oocytes between 2 and 3 h after injection, and that by 4-5 h after injection most of the oocytes have reached the chromatin mass stage. Two distinguishable stages of sperm nucleus transformation were observed. The first phase-swelling-was reached within 2 h after the injection and was independent of oocyte activation. The second phase-the "brush'-like stage or decondensed chromatin stage-was found only in activated oocytes. Moreover, this stage was not reached before the chromatin mass stage (late telophase) of the oocyte. The same proportion of metaphase II oocyte chromosome configurations and unchanged sperm nuclei was found at any given time after injection. We conclude that: (i) ICSI allows users to obtain an almost synchronized population of activated oocytes; (ii) anaphase II is initiated in the majority of oocytes not later than 2-3 h after injection and telophase II is reached approximately 5 h after injection; and (iii) there are two distinguishable phases of sperm nucleus transformation after ICSI: oocyte activation-independent swelling of the sperm head and oocyte activation-dependent chromatin decondensation which is coupled to the beginning of oocyte chromosome decondensation.

Cell Nucleus↗
PubMed1995

Human oocyte activation following intracytoplasmic injection: the role of the sperm cell.

The aim of this study was to investigate whether the human spermatozoon participates in the activation of human oocytes following intracytoplasmic sperm injection (ICSI) and if so, by what mechanism. In the first series of experiments, we randomized human oocytes which had remained unfertilized after in-vitro fertilization (IVF) or ICSI, for intracytoplasmic injection with live spermatozoa, spermatozoa presumed to be dead and no spermatozoa. Secondly, unfertilized human oocytes and freshly ovulated mouse oocytes were randomized for intracytoplasmic and sub-zonal injection with human sperm cytosolic fraction (CF) before and after heat treatment. We found that oocyte injection with initially motile spermatozoa induces human oocyte activation at a significantly higher rate than injection with dead spermatozoa (61 versus 0%; P < 0.001) or injection without a spermatozoon (61 versus 14%; P < 0.001). Intracytoplasmic injection of CF activated both human and mouse oocytes at the same rate as sperm injection of human oocytes (activation rates of 70 and 65% respectively). This effect was greatly reduced by heat treatment of the CF. From these experiments we conclude firstly that the human spermatozoon injected intracytoplasmically contributes to human oocyte activation and secondly that the spermatozoon releases into the oocyte a heat-sensitive, intracellularly active factor, which is not species-specific.

Cell Death↗
PubMed1995

Prospective randomized study of clomiphene citrate and gonadotrophins versus goserelin and gonadotrophins for follicular stimulation in assisted reproduction.

We performed a prospective randomized study of goserelin, a long-acting gonadotrophin-releasing hormone agonist (GnRHa) and human menopausal gonadotrophin (HMG) versus clomiphene citrate and HMG for follicular stimulation in assisted reproduction to investigate whether the use of this GnRHa provides a clear advantage in terms of pregnancy per treatment cycle in unselected patients, who entered a first trial of assisted reproduction. From a retrospective analysis comparing the two stimulation protocols, a relative increase of the pregnancy rate per cycle of 50% was anticipated. To detect this difference with a power of 90%, 300 patients had to be included. The main prognostic factors affecting the outcome of assisted reproduction were equally divided among the two groups by a minimization procedure. The pregnancy rates per cycle were significantly better in the goserelin/HMG group than in the clomiphene citrate/HMG group, both for all procedures of assisted reproduction combined (36.8 versus 24.5%; P < 0.02) and for the main procedure of in-vitro fertilization (IVF) (37.0 versus 23.5%; P < 0.02). Differences in pregnancy rates per oocyte retrieval and per embryo transfer were less pronounced (37.8 versus 30.8%; P = 0.40 and 44.4 versus 36.8%; not significant). On the other hand, stimulation with goserelin/HMG was associated with a higher number of ampoules of HMG (44.9 versus 9.9; P < 0.0001), a longer duration of stimulation (11.2 versus 8.7 days; P < 0.0001) and an incidence of ovarian stimulation of 4.5% (7/154) versus 0% in the clomiphene citrate/HMG group. Goserelin was well tolerated and proved to be very reliable as an adjunct of follicular stimulation in assisted reproduction.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Detection of premalignant stages in cervical smears with a biotinylated probe for chromosome 1.

Fluorescence in situ hybridization with (peri-)centromeric probes is an easy method to detect numerical aberrations in nonmitotic and mitotic cells. In this study, cervical smears of premalignant and malignant stages (26 controls, 15 CIN I, 12 CIN II, and 15 CIN III cervical smears) were analyzed for the presence of numerical aberrations of chromosome 1 with a centromeric DNA probe (1q12). With more severe stages a decrease of disomy was observed, merely due to a gain of extra copies of chromosome 1; in some cases, however, monosomy was detected. The frequencies of disomy for chromosome 1 ranged from 65.3% to 95.0% in the controls, from 71.3% to 94.3% in CIN I, from 59.2% to 91.5% in CIN II, and from 23% to 96.2% in CIN III. Polysomy ranged from 0% to 5.7% in the controls, from 0% to 14.4% in CIN I, from 0.9% to 30.8% in CIN II, and from 0.8% to 69.6% in CIN III. Monosomy ranged from 2.6% to 34.1% in the controls, from 0% to 17.5% in CIN I, from 3.6% to 27.5% in CIN II, and from 0.9% to 31.4% in CIN III. The results show that screening for aneuploidy of chromosome 1 allows a good discrimination between control samples and dysplasia. These data suggest that chromosome 1 may be a marker chromosome. They are in accordance with previous cytodensitometric analyses, where already in the preneoplastic stages an increased DNA content (polyploidization with subsequent aneuploidization) is observed.

Adult↗
PubMed1994

Cytogenetic analysis of human oocytes parthenogenetically activated by puromycin.

PURPOSE: Treatment of aged human oocytes by puromycin allows a high rate of parthenogenetic activation and development until the first cleavage division. This technique was used for the study of the chromosome complement of oocytes which remained unfertilized after in vitro fertilization. Three hundred four unfertilized oocytes were treated with 10 micrograms/ml puromycin for 6-8 hr and further cultured for 12-15 hr. RESULTS: Activation occurred in 90.5% of the oocytes. Heterozygous diploids with two pronuclei predominated (61%), which is in contrast to the mouse, where the majority of oocytes activated by puromycin are uniform haploids (89%). CONCLUSIONS: Therefore we conclude that puromycin treatment induces retention of the second polar body in human oocytes, unlike in mouse oocytes treated in the same way. Chromosome analysis performed on 182 oocytes suggested a nondisjunction (ND) rate for the second meiotic division of 12.7%. This is a low figure considering the fact that puromycin itself has been reported to induce nondisjunction. For the first meiotic division a ND rate of only 5.6% was found. This rate is lower than the one found in metaphase II arrested oocytes and we believe that this difference is due to the technical differences between the study of meiotic and that of mitotic chromosomes.

Cell Nucleus↗