A new device for microsurgical sperm aspiration.
A new device for microsurgical aspiration of seminal fluid is presented. It is simple to use, flexible, and watertight, allowing maximum retrieval of seminal fluid.
Epididymis↗
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Use of frozen-thawed testicular sperm for intracytoplasmic sperm injection.
OBJECTIVE: To determine the feasibility of using frozen-thawed testicular spermatozoa for intracytoplasmic sperm injection. DESIGN: Prospective clinical study. SETTING: A university hospital. PATIENT(S): One hundred seventy-five azoospermic men participating in a routine intracytoplasmic sperm injection program. INTERVENTION(S): The men underwent testicular biopsy for cryopreservation of tissue to be used in consecutive intracytoplasmic sperm injection treatment cycles. Their female partners underwent controlled ovarian hyperstimulation for conventional IVF treatment. MAIN OUTCOME MEASURE(S): Fertilization and pregnancy rates. RESULT(S): In 77% of the patients, spermatozoa could be harvested from the testis by an open testicular biopsy technique and used for intracytoplasmic sperm injection after freezing and thawing of testicular tissue. Histopathologic evaluation revealed a Sertoli cell-only pattern in 21%, maturation arrest in 60%, and hypospermatogenesis in 19% of the patients. In 2. 9% of the patients, carcinoma in situ or a germ cell tumor was detected. In all patients, viable spermatozoa could be visualized after the tissue samples were thawed. One hundred thirty-five intracytoplasmic sperm injection treatment cycles were performed, with a fertilization rate of 45% and a clinical pregnancy rate of 30% per oocyte retrieved. CONCLUSION(S): The use of frozen-thawed testicular tissue allows ovarian stimulation of the female partner to be timed and avoids cancellation of ovum pick-up when spermatozoa cannot be retrieved.
Chromosome Aberrations↗
Oocyte quality in patients with severe ovarian hyperstimulation syndrome.
OBJECTIVE: To study the oocyte quality in patients with ovarian hyperstimulation syndrome (OHSS). DESIGN: Retrospective study. SETTING: The Egyptian IVF-ET Center. PATIENT(S): Forty-two patients who developed severe OHSS (group A) were studied for the mean number of oocytes retrieved, percentage of high-quality oocytes, embryo quality, and fertilization, implantation, and pregnancy rates; these patients were compared with an age-matched control group who did not develop OHSS (group B; n = 183) after superstimulation for IVF or intracytoplasmic sperm injection. INTERVENTION(S): In vitro fertilization and ICSI. MAIN OUTCOME MEASURE(S): Fertilization and pregnancy rates. RESULT(S): In group A, the mean number of oocytes retrieved was significantly higher, whereas the percentage of high-quality oocytes and the fertilization rate were significantly lower than that in group B. There were no statistically significant differences in the quality of embryos transferred or the implantation or pregnancy rate between the groups. The percentage of high-quality oocytes and the fertilization rate were significantly lower in patients with polycystic ovaries (PCO) in both groups. CONCLUSION(S): The inferior quality and maturity of oocytes in OHSS reduced the fertilization rate but did not affect the quality or the number of embryos transferred or the pregnancy rate. The effect on oocyte quality could be due to the prevalence of PCO in this group of patients.
Adult↗
Comparison of short and long ovulation induction protocols used in ART applications according to the ovarian response and outcome of pregnancy.
This retrospective study was designed to determine whether there is any difference between short and long protocol ovulation induction with Gonadotropin Releasing Hormone agonist (GnRHa) and gonadotropins used in Assisted Reproductive Technology (ART) applications according to the number of retrieved oocytes, oocyte maturity, fertilization rates, embryo quality and the outcome of pregnancies. 240 cycles consisting of in vitro fertilization (IVF) cycles without andrologic factor and intracytoplasmic sperm injection (ICSI) cycles were evaluated. 112 cycles which were induced by short protocol GnRHa and Follicle Stimulating Hormone (FSH) + Human Menopausal Gonadotropin (HMG) combinations and 128 cycles which were induced by long protocol GnRHa and FSH + HMG combinations were compared according to the number of retrieved oocytes, cancellation rate of cycles, oocyte maturity, fertilization rates, embryo quality and pregnancy rates. The cancellation rate for short protocol cycles were found to be significantly higher than those with long protocol. The number of retrieved oocytes, mature oocytes and fertilized oocytes were also found significantly lower. The quality of embryos did not show any significant difference between these groups. The clinical pregnancy rates were evidently found to be high in the long protocol cycles. As a conclusion we have found that while the number of retrieved oocytes, mature oocytes, fertilized oocytes and clinical pregnancy rates were increasing, the cancellation rate of cycles were decreasing significantly in ART cycles induced by long protocol.
Cell Count↗
In vivo maturation of oocytes by extending the interval between human chorionic gonadotropin administration and oocyte retrieval.
OBJECTIVE: To find whether the percentage of mature oocytes can be increased in the following ICSI cycle, in a selected group of patients with >or=47% immature oocytes, with the prolongation of hCG-to-oocyte retrieval interval. DESIGN: Randomized study. SETTING: In vitro fertilization unit in a medical center in Israel. PATIENT(S): From January 2003 to June 2005, in 72 of 2,650 intracytoplasmic sperm injection cycles (2.8%), >or=47% of the total aspirated oocytes were immature (group A). In the following treatment cycle, with similar controlled ovarian hyperstimulation, hCG was planned to be injected in the same interval (group B) as well as 3-4 hours earlier (group C). In fact, the hCG-to-oocyte retrieval interval in group C was statistically significantly longer (38.6 +/- 1.2 hours) compared with that in group B (35.3 +/- 0.7 hours). INTERVENTION(S): The clinical characteristics of the patients, ovarian stimulation, hormonal profile, and the intracytoplasmic sperm injection outcome of the various groups were assessed. MAIN OUTCOME MEASURE(S): The number and maturity of the aspirated oocytes. RESULT(S): The mean number of ampules needed for stimulation, treatment duration, serum E(2), and P level on the hCG day were similar in the various groups. Escaped ovulation did not occur in any of the patients. A larger number of aspirated oocytes, a statistically significantly larger percentage of mature oocytes (72%), and a statistically significantly lower percentage of metaphase I (12%), germinal vesicle (18%), and degenerated oocytes (8%) were found in group C compared with group A (50%; 16%, 21%, and 13%, respectively). Similar morphological distribution of the aspirated oocytes was observed among groups B and C. Clinical implantation and pregnancy rates were higher in group C compared with group B. CONCLUSION(S): In a selected group of patients with >or=47% immature oocytes per retrieval, a larger cohort of mature oocytes can be recruited by extending the hCG- to-oocyte retrieval interval in a subsequent cycle. This improved outcome directly affects the quality of embryos available for transfer and thus increases pregnancy rate.
Adult↗
Matrix metalloproteinases 2 and 9 in follicular fluids of patients undergoing controlled ovarian stimulation for ICSI/ET.
The objective of this study was to detect the level of matrix metalloproteinases 2 and 9 in the follicular fluid of patients undergoing controlled ovarian stimulation and to correlate these levels with hormonal and clinical parameters. Follicular fluids were collected from thirty patients participating in an intracytoplasmic sperm injection/embryo transfer (ICSI/ET) program in "Otmar Bauer" Assisted Reproduction Center, Alexandroupolis, Greece. Follicular fluid was collected at oocyte retrieval and MMP-2 and MMP-9 were identified by gelatin zymography. Gelatin zymography of the follicular fluids revealed bands consistent with proMMP-2 and proMMP-9. ProMMP-2 was the predominant band in the majority of samples. The levels of MMPs varied considerably among the samples. A significant positive correlation between MMP-2 and the peak estradiol levels was found.
Adult↗
Comparative effects of autologous and homologous seminal plasma on the viability of largely extended boar spermatozoa.
Sperm handling, associated to artificial reproduction technologies (ART) such as in vitro fertilization (IVF) or the use of flow cytometry for cell analysis or sorting imposes volumetric extension of the sperm suspension and decreases sperm viability, presumably because of the removal of seminal plasma (SP) components. This study evaluated whether a 10% v/v of autologous SP (retrieved from the same donor boar) or homologous SP (e.g. from any of the four fertile boars included, other than the one providing the spermatozoa) would differently affect the viability of boar spermatozoa subjected to large extension in a simple saline medium [phosphate-buffered saline and 0.1% ethylenediaminetetraacetic acid (EDTA), PBSm] to a concentration of 0.3 x 10(6) spermatozoa/ml and incubated for 2 h at 30 degrees C. Sperm viability was monitored as membrane integrity [using the fluorophore carboxyfluorescein diacetate (C-FDA) and propidium iodide (PI)], mitochondrial function (using the fluorophore R-123) and motility characteristics [using Computer Assisted Sperm Analysis (CASA)]. Substraction of the SP and extension followed by incubation in PBSm significantly (p < 0.05) decreased sperm viability, which could be restored by addition of autologous SP. Furthermore, exposure of the extended spermatozoa to homologous SP (from any other individual boar) significantly (p < 0.05) varied with the source of the sire; some boars exerting beneficial effects (even surpassing the effects of the autologous SP; p < 0.05) while at least one boar negatively (p < 0.05) influencing the viability of the incubated spermatozoa. It is concluded that SP should be present when incubating highly extended spermatozoa. As a result of the obvious differences among boars, it would be advantageous to examine the ability of SP to maintain sperm viability prior to the use of SP pools during sperm handling in vitro.
Animals↗
A prospective, randomized comparison of two starting doses of recombinant FSH in combination with cetrorelix in women undergoing ovarian stimulation for IVF/ICSI.
BACKGROUND: A prospective randomized study was carried out in two centres to compare the number of oocytes retrieved after two different starting doses of recombinant human FSH (rhFSH) (Gonal-F) in women undergoing ovarian stimulation for IVF/intracytoplasmic sperm injection (ICSI) cycles using the multiple dose regimen of the gonadotrophin-releasing hormone (GnRH) antagonist cetrorelix (Cetrotide) to prevent induction of the premature LH surge. METHODS: Sixty women were randomized to receive rhFSH 150 IU ('low'), and 60 women to receive rhFSH 225 IU ('high') as the starting dose for the first 5 days of stimulation. From stimulation day 6 and onwards, including the day of human chorionic gonadotrophin (HCG) administration, the women received 0.25 mg of cetrorelix as a daily dose. The primary endpoint was the number of oocytes retrieved. RESULTS: The mean number (+/- SD) of oocytes was 9.1 +/- 4.4 and 11.0 +/- 4.6 in the 'low' and 'high' groups respectively (P = 0.024). The mean number of 75 IU ampoules of rhFSH was significantly lower in the 'low' group (23.0 +/- 6.3 versus 30.5 +/- 5.6, P < 0.0001). The ongoing pregnancy rate per started cycle and per embryo transfer were 25.9 and 28.8% versus 25.4 and 26.8% respectively in the 'low' and 'high' rhFSH groups (P = NS). CONCLUSIONS: When using a starting dose of 225 IU rhFSH combined with the multiple dose of 0.25 mg cetrorelix from stimulation day 6, significantly more oocytes were obtained than with a starting dose of 150 IU rhFSH.
Cell Count↗
Comparison of G1.2/G2.2 and Sydney IVF cleavage/blastocyst sequential media for the culture of human embryos: a prospective, randomized, comparative study.
OBJECTIVE: To compare two commercially available sequential media, G1.2/G2.2 and Sydney IVF cleavage/blastocyst media, as supports for human embryo culture. DESIGN: Prospective randomized study. SETTING: University-based IVF clinic. PATIENT(S): Two hundred forty-nine patients undergoing IVF treatment for the first or second time, randomly allocated at the time of oocyte retrieval, to either culture in G1.2/G2.2 or Sydney IVF media. INTERVENTION(S): Oocyte recovery, IVF or intracytoplasmic sperm injection, embryo culture, transfer on day 3 or day 5/6. MAIN OUTCOME MEASURE(S): Developmental stage on day 3, blastocyst rate, pregnancy outcome as assessed by beta hCG positive test, implantation rates, and ongoing pregnancies. RESULT(S): Embryos cultured in G1.2/G2.2 media displayed a faster kinetics of cleavage, compaction, blastulation, and hatching, but a lower day 3 embryo quality than those grown in Sydney IVF media. For patients with at least five embryos, G1.2/G2.2 media yielded higher implantation rates (26.2%) in our day 3 embryo transfer program when compared to Sydney IVF medium (15.5%), whereas similar implantation rates were obtained for day 5/6 embryo transfer for both media (43.1% and 36.1%, respectively). CONCLUSION(S): In our day 3 embryo transfer program, G1.2/G2.2 media were superior to Sydney IVF media, whereas both media yielded similar outcomes in our blastocyst transfer program.
Adult↗
Azoospermia in two Labrador retrievers.
Azoospermia is described in two sibling Labrador Retriever dogs. Clinical investigations following failure to sire pups after normal matings revealed testicular hypoplasia and degeneration. Sperm were absent on repeated ejaculate examination in both dogs. Histopathological examination of testicular needle aspirate biopsy and whole testicle of the first dog displayed an absence of spermatids and spermatocytes. Seminiferous tubules containing Sertoli cells with or without primary spermatogonia were present in the second dog. Peritubular lymphocyte accumulation was also present in both dogs. The dogs had been conceived using frozen-thawed semen.
Alkaline Phosphatase↗
Conservative and surgical semen retrieval in patients with spinal cord injury.
Assisted semen retrieval was applied in 219 men with spinal cord injury (212 supranuclear, seven infranuclear lesions). Vibrostimulation in supranuclear lesions was successful in 133, and in five more after physostigmine injection. Electroejaculation was successful in all seven infranuclear lesions and in four supranuclear patients failing with vibrostimulation. Eight more supranuclear patients responded to electroejaculation and physostigmine. Surgical retrieval was applied in 27 patients. Nine Wagenknecht spermatoceles showed only one success, from all 18 implanted Brindley reservoirs semen could be collected. Three patients, in whom no pregnancy could be induced after Brindley reservoir implantation, had testicular sperm aspirated. In 109 patients who wanted to have offspring, 73 pregnancies could be induced in 46 couples, leading to 54 births (four twins), 16 abortions and three pregnancies. Conservative semen retrieval was possible in 82 of these men (63 pregnancies in 37 couples) and surgical methods were used in 27 (10 pregnancies in nine couples).
Adult↗
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↗
Clinical experience with recombinant follicle-stimulating hormone (FSH) and urinary FSH: a retrospective case- controlled analysis.
OBJECTIVE: To compare the efficacy and efficiency of recombinant FSH (rFSH) and urinary FSH (uFSH). DESIGN: Retrospective case controlled analysis. SETTING: An assisted reproduction unit at a university center. PATIENT(S): 1388 patients undergoing long protocol in vitro fertilization/embryo transfer (IVF-ET) using buserelin acetate from day 2 of the cycle and either rFSH (follitropin beta) (n = 694) or uFSH (n = 694) with equal number of ampules started (rFSH: 50 IU, uFSH: 75 IU). INTERVENTION(S): Patients were included in the two groups of treatment after matching for similarity in age and type of treatment (IVF or intracytoplasmic sperm injection). MAIN OUTCOME MEASURE(S): Total dose of FSH, ovarian response, and IVF outcome. RESULT(S): Patients who received uFSH experienced a shorter period of stimulation, and a higher number of oocytes were collected. The total FSH used was lower in the rFSH group, and they required a lower FSH dose per oocyte retrieved. The implantation and pregnancy rates were similar between the uFSH and rFSH groups. In both groups implantation and pregnancy rates were higher when intracytoplasmic sperm injection was performed as compared with IVF. CONCLUSION(S): The implantation and pregnancy rates are similar when either rFSH or uFSH is used (when compared on an ampule-to-ampule basis, rFSH: 50 IU, and uFSH: 75 IU). However, a significantly lower total FSH dose was used in the rFSH group with a lower FSH dose per oocyte collected.
Adult↗
Consequences of high ovarian response to gonadotropins: a cytogenetic analysis of unfertilized human oocytes.
Several reports have shown decreased fertilization and implantation rates in high-responder patients to gonadotropins. In this cytogenetic study of unfertilized human oocytes, we have investigated possible reasons for this clinical evidence. Three groups of patients were established according to the number of oocytes retrieved: group 1 (n = 21), 1 to 5 oocytes; group 2 (n = 32), 6 to 10 oocytes; and group 3 (n = 35), greater than or equal to 11 oocytes. Patients in group 3 had lower estradiol levels per follicle developed, lower follicular volume, higher incidence of diploid oocytes, and higher rate of oocytes with an additional set of prematurely condensed sperm chromosomes of the presynthetic gap than groups 1 and 2. These results suggest that women with greater than or equal to 11 oocytes retrieved have a relative higher cytoplasmic immaturity that could explain the lower fertilization and implantation rates found in other studies.
Female↗
Glass wool column filtration, an advantageous way of preparing semen samples for intracytoplasmic sperm injection: an auto-controlled randomized study.
The characteristics of the glass wool filtration technique for the preparation of semen samples in an intracytoplasmic sperm injection programme (ICSI) were compared with a two-layer Percoll density gradient procedure. Half of each of 25 semen samples were prepared by each technique. The oocytes from each patient were randomly injected, half with spermatozoa prepared by glass wool filtration and half with Percoll-separated spermatozoa from the same semen sample. The percentage of recovered motile spermatozoa, the total motile sperm count, the percentage of morphologically normal forms, the fertilization rate, the cleavage rate and embryo quality obtained with both preparations were analysed. The 95% confidence intervals obtained through the Altman-Bland analysis showed that the percentage of motile spermatozoa recovered was 3.5-9.9% higher with the glass wool filtration technique, and this was highly correlated (r = 0.92, P = 0.0001) to the method. Similarly, the total number of spermatozoa available for ICSI was 0.18 x 10(6)-2.44 x 10(6) higher with the glass wool column and was highly correlated to the method (r = 0.956, P = 0.0001). Also, the percentage of normal forms was 1.25-3.31% higher after glass wool filtration but was poorly correlated to the method (r = 0.47, P = 0.017). Out of 100 metaphase II oocytes injected with glass wool-extracted spermatozoa, 77 fertilized and 72 cleaved. Out of 97 metaphase II oocytes injected with Percoll-selected spermatozoa, 71 fertilized and 69 cleaved. These results were not statistically different. The mean +/- SEM embryo quality score for the glass wool group (2.90 +/- 0.27) was the same as that for the Percoll group (2.80 +/- 0.24). No fertilization failures occurred and 11 patients (44% per oocyte retrieval) became pregnant. It was concluded that glass wool filtration for semen preparation in an ICSI programme offers higher sperm recovery and sperm morphology of superior quality than with the classic two-layer Percoll gradient method, without affecting the fertilization rate and embryo quality.
Cell Separation↗
The outcome of intracytoplasmic sperm injection in patients with retrograde ejaculation.
The objective of this retrospective clinical study was to assess the benefit of assisted fertilization in cases of anejaculatory infertility due to retrograde ejaculation. We report the outcome of intracytoplasmic sperm injection (ICSI) treatment. In 16 couples in which the men suffered from retrograde ejaculation. We performed 35 cycles of ICSI with spermatozoa retrieved from post-ejaculatory urine. The patients had been instructed to alkalinize the urine by ingesting sodium bicarbonate before the procedure. The fertilization rate averaged 51.2%. Seven clinical pregnancies were achieved. Three spontaneous first trimester abortions occurred, but three live offspring were delivered and one pregnancy is ongoing. In conclusion, the use of ICSI may be feasible for patients with retrograde ejaculation who are resistant to medical treatment and whose sperm quality is so low or unpredictable that intrauterine insemination or conventional methods of in-vitro fertilization are not possible.
Adult↗
Pentoxifylline is not useful in enhancing sperm function in cases with previous in vitro fertilization failure.
OBJECTIVE: To assess the efficiency of the addition of pentoxifylline to spermatozoa for in vitro fertilization (IVF) in cases with previous fertilization failure. DESIGN: After retrieval, oocytes were divided at random and inseminated with spermatozoa prepared with or without pentoxifylline. SETTING: In vitro fertilization program of the Centre for Reproductive Medicine of the Dutch-speaking Brussels Free University, Belgium, a tertiary referral institution. PATIENTS: Couples in this study were undergoing a second or third IVF attempt for the treatment of male factor infertility. INTERVENTIONS: Sperm samples were divided into two equal parts. The first part was prepared with 3.6 mM pentoxifylline (treatment group) and the other part without pentoxifylline (control group). After swim-up, oocytes were randomly inseminated with spermatozoa from either treatment. MAIN OUTCOME MEASURES: Fertilization and cleavage rates after IVF. RESULTS: Overall a fertilization rate of 8.8% was achieved with no significant differences between treatment and control groups. Cleavage rate was 81.5% and was not different between groups. CONCLUSIONS: This study demonstrates no therapeutic advantage of using pentoxifylline in IVF for male factor infertility in cases with previous fertilization failure.
Adult↗
The culture of human epididymal epithelium and in vitro maturation of epididymal spermatozoa.
OBJECTIVE: To promote human sperm maturation in vitro. DESIGN: Spermatozoa from the proximal epididymis were coincubated with epididymal epithelial fragments. SETTING: Hospital and research institute. PATIENTS, PARTICIPANTS: Tissue samples were obtained from men undergoing epididymovasostomy procedures or vasectomy. INTERVENTIONS: Fragments of epididymal epithelium formed everted epithelial spheres that in the presence of androgen maintained cell integrity. Coincubation for up to 48 hours of caput epididymal spermatozoa with 3-day-old epithelial cultures from the cauda epididymis was undertaken. MAIN OUTCOME MEASURES: Morphology of epididymal epithelium was assessed by light and electron microscopy. Pulse labeling of tissue in vitro with 35S-methionine was performed with analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis technique and fluorography. Spermatozoa were assessed for progressive motility and their capacity to bind to salt-stored human zona pellucidae. RESULTS: Epididymal fragments formed everted epithelial spheres that maintained cell integrity and functional morphology for 5 to 7 days. Specific proteins were synthesized in culture, in particular, proteins of 20, 22, 40, and 66 kd. Coincubation of caput epididymal spermatozoa with cultures from the cauda epididymis induced a significant increase in progressive sperm motility and sperm binding to salt-stored human zona pellucidae compared with control cultures of epithelium incubated in the absence of androgens or overgrown with fibroblasts. CONCLUSIONS: Aspects of human sperm maturation processes can be mimicked in vitro using coculture techniques with epididymal epithelium. This method may be valuable for improving the fertilizing capacity of human spermatozoa retrieved from the proximal region of the excurrent ducts.
Cells, Cultured↗