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M Margalit,
L Yogev, H Yavetz,
O Lehavi,
R Hauser,
A Botchan,
S Barda,
F Levitin,
M Weiss,
I Pastan,
D H Wreschner,
G Paz,
S E Kleiman
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ABSTRACT: The prostate and testis expression (PATE)-like family of proteins are expressed mainly in the male genital tract. They are localized in the sperm head and are homologous to SP-10, the acrosomal vesicle protein also named ACRV1. Our aim was to characterize the expression and functional role of three PATE-like proteins in the testis and ejaculated sperm.
The expression and localization of PATE-like proteins in human testis biopsies (n= 95) and sperm cells were assessed by RT-PCR, immunohistochemistry and immunofluorescence staining (at least 600 sperm cells per specimen). The function of the PATE protein was tested by the hemizona assay and hamster egg penetration test (HEPT).
PATE and PATE-M genes and proteins were present almost exclusively in germ cells in the testis: immunoflourescence showed that the percentage of germ cells positive for PATE, PATE-M and PATE-B was 85, 50 and 2%, respectively. PATE and PATE-M proteins were localized in the equatorial segment of the sperm head, while PATE-B protein was localized in the post-acrosomal region. A polyclonal antibody (Ab, at 1:50 and 1:200 dilutions) against the PATE protein did not inhibit sperm-zona binding in the hemizona assay (hemizona index of 89.6 ± 10 and 87 ± 36%, respectively). However, there was inhibition of sperm-oolemma fusion and penetration in the HEPT (penetration index: without Ab 7 ± 3.9; Ab dilution of 1:100, 4 ± 3.5; Ab dilution of 1:20, 0.6 ± 1.2, P < 0.001).
Our data suggest that PATE protein is involved in sperm-oolemma fusion and penetration but not sperm-zona binding.
Human Reproduction 03/2012; 27(5):1238-48. · 4.47 Impact Factor
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ABSTRACT: Forty-three subfertile women were treated by intrauterine insemination with washed sperm. Among the couples, infertility was, in 7 cases due to cervical factor infertility, in 17 cases due to subfertile sperm quality, and 19 cases, unexplained. All women ovulated: 9 spontaneously, 26 where treated with clomiphene citrate and 8 with hMG-hCG. One hundred and sixty seven inseminations were performed in 90 ovulatory cycles. Six pregnancies were recorded (14%): three pregnancies among the cervical factor infertility group (42.8%), two among the couples with low sperm quality (11.7%) and one pregnancy in the group with unexplained infertility (5.2%). Thus, it can be concluded that IUI is effective for achieving pregnancy when unfavourable cervical factor is detected. The method is of doubtful value in cases of low sperm quality or unexplained infertility.Zusammenfassung: Mittels einer intrauterinen Insemination wurden 43 subfertile Frauen behandelt, wozu vorher die Spermatozoen gewaschen wurden. Unter diesen Paaren ergab sich in 7 Fllen ein Cervix-Faktor, in 17 Fllen wurde die Subfertilitt durch die Qualitt des Spermas verursacht und in 19 Fllen bestand eine sog. “unerklrte Infertilitt”. Alle Frauen ovulierten: 9 spontan, 26, wenn sie mit Clomiphen behandelt wurden und 8, wenn eine hMG/hCG-Therapie durchgeführt wurde. In 90 Cyclen wurden 167 Inseminationen vorgenommen. Es resultierte eine Schwangerschaftsrate von 14% (6 Graviditten): hiervon entfielen 3 auf die Frauen mit Cervix-Faktor, 2 auf die schlechte Spermaqualittsgruppe und 1 auf die Gruppe der unerklrten Infertilitt. Es wird hieraus die Schlußfolgerung gezogen, daß die intrauterine Insemination geeignet ist, um eine Schwangerschaft herbeizuführen, wenn es sich um einen ungünstigen Cervix-Faktor handelt; dagegen ist der Wert dieser Methode zweifelhaft bei unerklrter Infertilitt und bei schlechter Spermaqualitt.
Andrologia 04/2009; 22(1):29 - 33. · 1.55 Impact Factor
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ABSTRACT: The human X chromosome is enriched with testis-specific genes that may be crucial for male fertility. One is the ubiquitin-specific protease 26 (USP26). Five frequent mutations have been identified: 1737G>A, 1090C > T, 370-371insACA, 494T > C and 1423C>T (with the latter three usually detected in a cluster). Their role in infertility is still controversial. This study assesses the association of the most frequent USP26 mutations with male infertility and male infertility etiology factors.
The study included 300 infertile and 287 fertile men. Data were collected on ethnicity (according to maternal origin) and family history of reproduction. Clinical records from 235 infertile and 62 fertile (sperm bank donors) men were available and summarized. The five mutations were investigated by bioinformatic tools and their frequencies were assessed by restriction analysis. The results were correlated with clinical findings. Segregation of the mutations in four families was analyzed.
The five analyzed mutations were detected in 44 men from both fertile and infertile groups. The cluster and the 1090C>T mutations showed the highest frequency among Arabs and Sephardic Jews of the infertile group, respectively. Inheritance studies showed that mutations were not always associated with the infertility trait. Mutations 1090C>T and 1737G>A were significantly associated with a history of inguinal hernia (P = 0.007 and P = 0.043, respectively). The prevalence of inguinal hernia among men with the 1090C > T mutation was 33.3% (5/15 men), higher than that reported in infertile men (6.7%).
Mutation 1090C > T may be a new genetic risk factor for developing inguinal hernia which may be associated with impaired male fertility.
Human Reproduction 10/2008; 24(2):477-84. · 4.47 Impact Factor
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ABSTRACT: The Y-chromosome AZF regions include genes whose functions and specific roles in spermatogenesis have not been fully clarified. This study investigated the expression of several AZF (USP9Y, DDX3Y/DDX3Yt1, EIF1AY and PRY) and USP9X transcripts in testicular biopsies of 89 azoospermic men who had been classified by histology and cytology assessments.
Expression was analysed by RT-PCR, and some biopsies were evaluated by multiplex RT-PCR. Quantitative PCR was performed in some biopsies to determine the ratio of the testis-specific transcript DDX3Yt1 to the total DDX3Y transcription.
The expression of USP9Y, USP9X and DDX3Y was found in all the specimens tested, whereas DDX3Yt1 expression was diminished or undetectable in several biopsies with impaired spermatogenesis. EIF1AY was detected in all except two of the specimens. Noteworthy, PRY expression was detected mainly in biopsies with germ cells, and this association was significant (P < 0.001). An identical expression profile was obtained by either single or multiplex RT-PCR.
These findings suggest that PRY is usually expressed in germ cells, whereas the other transcripts are also expressed in testicular somatic cells. The absence of EIF1AY expression might sporadically contribute to azoospermia. The decreased ratio of DDX3Yt1/DDX3Y transcript in impaired spermatogenesis suggests that the DDX3Yt1 transcript is under-expressed in impaired spermatogenesis. The findings contribute to the search and selection of the most valuable gene markers potentially useful as additional tools for predicting complete spermatogenesis by multiplex expression analysis.
Human Reproduction 02/2007; 22(1):151-8. · 4.47 Impact Factor
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ABSTRACT: Among azoospermic and severely oligozoospermic men, 7-15% present microdeletions of a region on the long arm of the Y chromosome that has been called AZF (azoospermia factor). Because these deletions present varying relative frequencies in different populations, we decided to ascertain whether their presence was correlated with specific Y-chromosome haplotypes. For that, we evaluated 51 infertile Israeli men, 9 of whom had microdeletions in AZF. Haplotypes were identified using a hierarchical system with eight biallelic DNA markers. We also checked for the presence of the deletion marker 50f2/C, which was absent in all seven patients with isolated AZFc deletion and also in the one patient with isolated AZFb deletion, suggesting that these microdeletions overlap. As expected, haplogroup J was the most common (47%), followed by equal frequencies of haplogroups Y* (xDE, J, K), P* (xR1a, R1b8), K* (xP), and E. In six patients with AZFc deficiencies of comparable size, three belonged to haplogroup J, two belonged to haplogroup P* (xR1a, R1b8), and one belonged to haplogroup R1a. Also, there were no significant differences in the haplotype frequencies between the groups with and without microdeletions. Thus we did not identify any association of a specific haplogroup with predisposition to de novo deletion of the AZF region in the Israeli population.
Human Biology 07/2004; 76(3):469-78. · 1.31 Impact Factor
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ABSTRACT: The present study was conducted to evaluate seasonal variability in the quality of pre- and post-thaw semen parameters among sperm bank donors.
The first two consecutive ejaculates during the months March (spring, 92 males), June (summer, 97 males), September (autumn, 81 males) and December (winter, 97 males) were analysed. A comparison was made between sperm parameters from the same sperm donor at different seasons. Only males who donated semen samples during at least two seasons were enrolled in the study group (n = 103). Sperm specimens were cryopreserved in aliquots with fixed range of 8-12 x 10(6)/ml of progressive motile sperm concentration after thawing.
Differences between months were found in sperm concentration (P = 0.030) and normal morphology (P = 0.038); highest values were found in March and December, and the lowest in September. Mean specimen volume and percent of motile sperm cells did not vary throughout the seasons. The freezability of the donors' sperm dropped dramatically from March to September, as determined by the number of straws (fixed aliquots of 0.5 ml) and total thawed progressive motile sperm that were cryopreserved for each male (P = 0.017 and P = 0.002, respectively).
Cryopreservation of donor sperm is more effective during winter and spring than during the rest of the year.
Human Reproduction 05/2004; 19(4):880-5. · 4.47 Impact Factor
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ABSTRACT: Background
There are no clinical parameters for identification of spermatogenic tissue within the testis of azoospermic subjects. Consequently, multiple biopsies are performed until spermatozoa are found. The aim of this study was to evaluate whether testicular blood flow can be a predictor of spermatozoa retrieval on TESE.Method
Color Doppler ultrasound was performed in 13 azoospermic subjects affected by primary testicular pathology and in 7 fertile subjects. The PI, RI, and S/D ratio reflecting resistance to flow were measured in the mid-portion of the testis in the longitudinal view. All azoospermic subjects underwent TESE in which a biopsy was taken from the head, mid, and tale portion of the testis. The specimens were evaluated for structure and for the presence of spermatozoa.ResultsBlood flow indices were similar in azoospermic subjects as compared to fertile subjects. (mean ± SD; PI = 0.99 ± 0.64, RI = 0.52 ± 0.16, S/D = 2.16 ± 0.74; PI = 0.99 ± 0.19, RI = 0.59 ± 0.07, S/D = 2.51 ± 0.48, respectively). In seven patients, spermatozoa was found in one or more of the testicular specimens. Blood flow indices in this group were similar to the indices in azoospermic patients in whom spermatozoa was not found.Conclusion
At present testicular blood flow as evaluated by color Doppler can not serve as a predictor of spermatozoa retrieval on TESE.
Ultrasound in Obstetrics and Gynecology 01/2002; 16(s1):98 - 98. · 3.01 Impact Factor
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ABSTRACT: Artificial insemination by donor spermatozoa (AID) can prove a valuable treatment for a number of male factor disorders, although its success rate is variable.
Retrospective analysis of the results of 6139 cycles performed in 1001 women during an 18 year period is presented. Pregnancy rates per cycle are presented as a function of: female fertility history, treatment modalities, medication used for induction of ovulation, female age, year of treatment, consecutive cycle effect and the use of fresh versus frozen-thawed spermatozoa.
Overall pregnancy rate of 12.6% and cumulative pregnancy rate after 12 months of treatment of 75% were achieved. Age was found to be the most important determinant for success rate.
Since the establishment of AID treatments, the mean age of the population of women receiving treatment has increased each year. Consequently, success rate did not improve, even with the use of more sophisticated medical modalities.
Human Reproduction 12/2001; 16(11):2298-304. · 4.47 Impact Factor
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ABSTRACT: The objective of this study was to determine the seminal concentrations of four different catecholamines and their association with semen quality. Seminal concentrations of adrenaline, noradrenaline, 3,4-dihydroxy-phenylalanine (DOPA), and 3,4-dihydroxy-phenyl acetic acid (DOPAC) were determined in 13 healthy volunteers, using high-performance liquid chromatography with an electrochemical detector. In addition, semen analysis was performed. Noradrenaline and DOPA were present in all specimens with a concentration of 15 181+/- 2951 pg ml(-1) and 4023 +/- 429 pg ml(-1) (mean +/- SE), respectively. These concentrations are respectively 19 times (range: 3-44) and twice (range: 1-3) as high as the maximal normal concentration in plasma. Adrenaline was present in 10 and DOPAC in seven of 13 specimens. No correlation was found between the concentration of any of the catecholamines evaluated and semen characteristics. In conclusion, noradrenaline and DOPA are present in human semen at concentrations that are much higher than maximal normal values in plasma. Adrenaline and DOPAC were also found in some of the samples. The concentrations of catecholamines in semen are not associated with semen quality.
Andrologia 12/2001; 33(6):347-50. · 1.55 Impact Factor
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ABSTRACT: The aim of this study was to evaluate the change in the expression of mannose-ligand receptors following a freezing-thawing procedure, in order to assess its impact on sperm capacitation and acrosome reaction. Twenty semen samples were obtained from fertile donors. Sperm samples were divided into two equal volumes. One aliquot was cryopreserved and the other aliquot was incubated at 32 degrees C. After 2 h the frozen sample was thawed and both samples were further incubated at 32 degrees C to allow capacitation. Mannose receptors were examined following 4 and 22 h of incubation using a mannosylated-BSA-FITC probe. The expression of mannose-ligand receptors on the sperm plasma membrane was determined according to the fluorescence pattern: pattern I represents pre-capacitation, pattern II represents capacitated spermatozoa and pattern III represents acrosome-reacted spermatozoa. After 4 h incubation in capacitating medium, the percentages of patterns I, II and III were 90, 7 and 3% for fresh spermatozoa and 89, 8 and 3% for frozen-thawed spermatozoa, respectively (P > 0.05). Following 22 h of incubation, the percentages of patterns I, II and III were 84, 11 and 5 for fresh spermatozoa and 83, 11 and 6% for frozen-thawed spermatozoa, respectively (not significant at P > 0.05). The percentages of patterns II and III in fresh and frozen-thawed spermatozoa were increased by the same magnitude with longer incubation in the capacitating conditions. It was concluded that the freezing-thawing procedure for human spermatozoa does not affect the expression of mannose-ligand receptors and the dynamics of sperm pre-fertilization processes.
Andrologia 10/2001; 33(5):272-6. · 1.55 Impact Factor
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ABSTRACT: Azoospermia is defined as the absence of spermatozoa in the ejaculate, although some foci of spermatogenesis may exist in the testes of these men. Currently, there are no clinical, seminal or hormonal parameters for identifying spermatogenesis within the testis sufficient for achieving genetic offspring. As a result, multiple biopsies are performed at several arbitrary sites of both testes in search of spermatozoa. We developed a power Doppler (PD) ultrasound (US) image-based technique that predicts sites with the greatest potential for spermatogenesis. PDUS images of the testes of azoospermic men were acquired at seven cross-sections to reconstruct a 3-D matrix for constructing a spatial map of preferential regions where spermatozoa are most likely to exist. This technique may obviate the need for arbitrary multiple biopsies that inflict some degree of damage upon testicular tissue, and may increase the success rate of identifying viable spermatozoa in testicular biopsies.
Ultrasound in Medicine & Biology 10/2001; 27(9):1171-6. · 2.29 Impact Factor
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Fertility and Sterility 05/2001; 75(4):821-2. · 3.56 Impact Factor
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Fertility and Sterility 05/2001; 75(4):828-9. · 3.56 Impact Factor
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ABSTRACT: Substantial involvement of the Y chromosome in sexual development and spermatogenesis has been demonstrated. Over the last decade, varying extent of Y chromosome microdeletions have been identified among infertile patients with azoospermia or oligozoospermia. These microdeletions were clustered in three main regions named AZFa, AZFb, and AZFc. Analysis of the Y chromosome microdeletion was found to be of prognostic value in cases of infertility, both in terms of clinical management as well as for understanding the aetiology of the spermatogenesis impairment. However, the accumulated data are difficult to analyse, due to the variable extent of these deletions, the different sequence-tagged sites (STS) used to detect the microdeletions, and the non-uniformity of the histological terminology used by different investigators. This debate discusses the chances of finding testicular spermatozoa in men with a varying extent of Y chromosome microdeletions. The genotype and germ cell findings in men with AZFa microdeletions as well as those that include more than a single AZF region are reviewed, as is the effect of Y chromosome AZF microdeletions on the maturity of the Sertoli cells.
Human Reproduction 04/2001; 16(3):399-402. · 4.47 Impact Factor
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ABSTRACT: To investigate the expression of deleted in azoospermia (DAZ), RNA-binding motif (RBM), and chromodomain y1 (CDY1) genes in the testes of men with azoospermia with variable histopathologies.
Prospective study.
Andrology laboratory of a university-affiliated maternity hospital.
Sixty-six men with azoospermia.
Testicular sperm extraction.
The results of gene expression in testicular tissue tested by RT-PCR were correlated with those of histopathologically and microscopically examined minced testicular tissue. Y chromosome microdeletion testing and karyotyping were performed, as was direct sequencing of CDY1-PCR products.
CDY1-minor expression was detected in all biopsies in which mature spermatids/spermatozoa were observed by histological analysis and/or in the minced tissue. CDY1-minor expression was also detected in two biopsies with arrest at the spermatocyte stage during which no mature spermatids/spermatozoa were observed. A previously unreported CDY1-minor alternative splicing transcript was identified. DAZ and RBM gene expressions were detected in all biopsies in which at least a few germinal cells in early stages were found and in one biopsy histologically determined as Sertoli cell only.
Our preliminary results suggest that CDY1-minor expression might increase the prospect for complete spermatogenesis, while RBM and DAZ expression can only be indicative of the presence of germinal cells.
Fertility and Sterility 02/2001; 75(1):166-73. · 3.56 Impact Factor
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ABSTRACT: Testicular biopsies of infertile men are often characterized by mixed histologic patterns, with different types of spermatogenic impairments being found in adjacent seminiferous tubules. RNA-binding motif (RBM) is a nuclear protein expressed exclusively in the male germ cell line. We reasoned that RBM might be a useful marker to identify germ cells in testicular sections, particularly in biopsies with mixed histologic phenotype and small focal concentrations of spermatogenesis. Testicular biopsies from azoospermic men were immunohistochemically evaluated for RBM expression. RBM expression was detectable in spermatogonia, spermatocytes, and round spermatids in biopsies of men with obstructive azoospermia and normal spermatogenesis. No specific cell staining was shown in cases of Sertoli-cell-only (SCO) syndrome. In biopsies of patients with spermatogenic disorders, all the germ cells were stained up to and including the stage level of the arrest in spermatogenesis. This approach enabled identification of small focal concentrations of spermatogenesis in a biopsy previously classified as being SCO by hematoxylin and eosin staining. Thus, RBM can be a useful immunohistochemical marker for the specific identification of germ cells and provide greater accuracy in the histopathologic evaluation of testicular biopsies.
Human Pathlogy 02/2001; 32(1):36-41. · 2.88 Impact Factor
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Human Reproduction 01/2001; 15(12):2685-6. · 4.47 Impact Factor
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ABSTRACT: The study was conducted to evaluate the results of IUI treatment in a homogenous group with male factor infertility, and to assess the correlation of sperm variables, including sperm morphology by strict criteria, with pregnancy achievement after IUI. A total of 108 couples with no apparent female aetiology for infertility underwent 264 intrauterine insemination treatment cycles. A comparison was made between the sperm variables in two groups in which the achievement of pregnancy differed. The percentage of motile spermatozoa, degree of motility and normal morphology (by strict criteria) were significantly higher in the pregnant group compared with that of the nonpregnant group. A significant difference in pregnancy rates per couple after intrauterine insemination was demonstrated among three groups according to the percentage of sperm morphology, i.e. poor (< 4%), fair (4-14%) or good (> 14%) (11.1%; 36.1% and 50.0%, respectively). Intrauterine insemination is a valid mode of treatment in cases with male infertility, provided that normal morphology by strict criteria is higher than 4%.
Andrologia 01/2001; 33(1):13-7. · 1.55 Impact Factor
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ABSTRACT: Lymphomas are a group of diseases, prevalent at reproductive age. Fertility is notoriously reduced among lymphoma patients. This study evaluates pre- and post-treatment semen concentration and motility, and factors associated with semen quality deterioration. We followed-up 33 patients with non-Hodgkin's lymphoma or with Hodgkin's disease during the years 1987-1997 who were referred for semen cryopreservation. Pretreatment semen analysis, and hormonal profile were recorded at diagnosis and at least 1 year after completion of the treatment, and compared. Medical records for disease type, disease stage and treatment protocols were related to long-term sperm outcome. Hormonal concentrations were not predictive of post-treatment sperm concentration. In patients with localized disease, initial sperm concentration and motility tended to be preserved, compared with patients with widespread disease (P: = 0. 016). In Hodgkin's disease patients, treatment with the adriamycin, bleomycin, vinblastine and dacarbazine (ABVD) protocol was superior to the mechloretamine, vincristine, procarbazine and prednisone with ABV protocol regarding germinal toxicity (P: = 0.0008). The post-treatment sperm outcome was better in patients treated with local irradiation than in those who did not undergo irradiation (P: = 0.0027). No predictive tools for post-treatment fertility were found and, therefore, every patient with a lymphoma should have his semen cryopreserved at diagnosis.
Human Reproduction 10/2000; 15(9):1985-8. · 4.47 Impact Factor
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ABSTRACT: The use of testicular spermatozoa for intracytoplasmic sperm injection introduced a new treatment modality for management of male infertility. Since testicular biopsies of non-obstructive azoospermic men are not homogenous in their histological patterns, identification with certainty of focal spermatogenesis might be difficult, particularly in those with small foci of spermatogenesis. We used an immunohistochemical marker of the male germ line, an antibody generated against RBM (RNA-binding-motif), to recognize with high precision the presence of germ cells in the biopsy. Biopsies of 30 men with azoospermia, most with non-obstructive azoospermia and a few with obstruction of the vas deferens, were evaluated. Immunohistochemical staining for RBM protein contributed to the detection and accuracy of the identification of germ cells. Furthermore, this immunohistochemical technique aided the histopathologist to focus on even small foci of spermatogenesis. Absence of the protein expression confirmed the diagnosis of Sertoli-cell-only syndrome. The results indicate that expression of RBM can be a diagnostic marker for identifying the germ cells of small concentrations of spermatogenesis. This method can enhance the accuracy of histopathological evaluation of testicular biopsies that had formerly relied mainly on hematoxylin-and-eosin staining.
Harefuah 10/2000; 139(5-6):179-82, 247.
