1 Reproductive Medicine Service, Department of Obstetrics and Gynaecology, Institut Universitari Dexeus, Universitat Autònoma de Barcelona, Passeig Bonanova 67, E-08017 Barcelona and 2 Department of Cellular Biology and Physiology, Universitat Autònoma de Barcelona, E-08193 Bellaterra, Spain
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Abstract |
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Key words: intracytoplasmic sperm injection /meiosis/meiotic chromosome abnormalities/oligoasthenozoospermia/sperm parameters
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Introduction |
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On the other hand, chromosome abnormalities may be responsible for male infertility. Indeed, the prevalence of somatic chromosome abnormalities detectable in the karyotype is 10 times higher in infertile men (5.3%) than in the general population (0.6%) (Egozcue, 1989). Numerical and structural anomalies of the sex chromosomes occur with a high frequency, mainly in azoospermic and severely oligoasthenozoospermic men (Retief, 1986
). In addition, the incidence of synaptic chromosome anomalies restricted to the germ cell line and only detectable by meiotic studies is 47.7% in cases of male infertility (Egozcue et al., 1983
; De Braekeleer and Dao, 1991
).
The aim of this study was to evaluate the relationship between different spermatogenic parameters [testicular size, sperm concentration and motility, baseline serum follicle stimulating hormone (FSH) and count of mature spermatids per tubule] and the presence of meiotic abnormalities (severe arrest and synaptic anomalies) in patients with severe oligoasthenozoospermia and therefore likely to undergo ICSI.
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Materials and methods |
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In all patients, salient features of clinical history and physical examination were recorded as well as results of the following investigations: testicular volume, semen evaluation, baseline serum FSH concentration, and a testicular biopsy.
Testicular dimensions were measured with calipers. Testicular volume was estimated by the following formula (Ley and Leonard, 1985): V = 4/3 p (a/2)2 (b/2), where `a' equals the short testicular axis, and `b' equals the long testicular axis (in cm). Volumes are expressed in ml as the average of the two testicles.
Seminal fluid was collected by masturbation after 35 days of abstinence and in the absence of fever for 3 months before the study. All samples were analysed for volume (ml). Sperm concentration (x106/ml), motility (%) and motile sperm concentration (x106/ml) were assessed by means of the Makler chamber (Makler, 1978; Makler et al., 1980
). Baseline serum FSH was measured by radioimmunoassay with a commercial kit (Abbott Laboratories, S.A.). A single blood sample for each patient was analysed.
A testicular biopsy was taken under local anaesthesia. The biopsy was performed unilaterally for the study of meiosis according to the method described by Egozcue et al. (1983) when a histological diagnosis was already available, and bilaterally for meiotic studies, for histological evaluation (Levin, 1979) and for counting of the number of mature spermatids per 20 tubules (Silber and Rodriguez-Rigau, 1981) when a previous histological diagnosis was not available. Meiosis on testicular biopsy material was independently evaluated by two observers. Three meiotic patterns were defined: normal meiosis and two meiotic abnormalities, i.e. severe arrest (presence of pachytenes and occasional spermatozoa, but no metaphase I figures found) and synaptic anomalies (chromosome pairing anomalies). Peripheral blood karyotypes were also evaluated.
Statistical analysis
The four quantitative parameters were dichotomized as follows: sperm concentration, 1x106/ml versus >1x106/ml; motile sperm concentration,
0.5x106/ml versus >0.5x106/ml; testicular volume, <15 ml versus
15 ml (normal range); and serum FSH concentration, >10 IU/l versus
10 IU/l (normal range). Meiotic abnormalities (arrest and synaptic anomalies) were analysed together and separately. The Student's t-test and the analysis of variance (ANOVA) were used for the comparison of quantitative variables and the chi-square test (
2) for categorical variables. The independent predictive value of significant variables in the univariate analysis was assessed by means of a logistical regression model. All statistical tests were performed at the 5% level of significance. The Statistical Package for the Social Sciences (SPSS) for Windows was used for the analysis of data.
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Results |
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Meiotic studies in testicular samples were performed in all 103 patients. A normal pattern was found in 64 (62.1%), severe arrest in 21 (20.4%), and synaptic anomalies in 18 (17.5%) (Figure 1). The overall rate of meiotic abnormalities was 37.9%. There were no statistically significant differences among patients with normal or abnormal meiotic patterns in relation to mean age, length of time of infertility, seminal volume, percentage of motile spermatozoa, and number of mature spermatids per tubule (Table II
). However, in patients with sperm concentration
1x106/ml, motile sperm concentration
0.5x106/ml and serum FSH concentrations >10 IU/l, meiotic abnormalities were significantly more frequent than normal meiotic patterns (Table III
). Total meiotic abnormalities and synaptic anomalies accounted respectively for 57.8 and 26.7% of patients with sperm counts
1x106/ml and for 54.8 and 22.6% of patients with FSH concentrations >10 UI/l. After multivariate analysis, sperm concentration and serum FSH concentration appeared to be the only independent predictive factors of normal or abnormal meiotic patterns (Table IV
). Motile sperm concentration was not a predictive factor because there was a statistically significant correlation with sperm concentration (r = 0.653; P < 0.0001).
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Discussion |
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The population studied suffered from severe non-obstructive oligoasthenozoospermia, i.e. low motile sperm concentration (mean 0.49x106/ml), moderately hypoplastic testicular volume (mean 11.74 ml); slightly elevated baseline serum FSH level (mean 10.7 IU/l), and low count of mature spermatids per tubule (mean 5.33) and met the criteria for inclusion in an ICSI programme. In this population, we found a high incidence of meiotic abnormalities (37.9%), 17.5% of which corresponded to synaptic anomalies. This incidence is ~2.5 times higher than in heterogeneous male infertility (Egozcue et al., 1983), suggesting that the higher frequency of meiotic chromosome abnormalities may often be responsible for infertility in patients with severe oligoasthenozoospermia.
The present results also demonstrate that most total meiotic abnormalities (66.7%) and synaptic anomalies (66.7%) were found in patients with sperm concentrations 1x106/ml, affecting 57.8 and 26.7% of the patients respectively. When the sperm concentration was
0.5x106/ml, 32% of patients had a normal meiotic pattern, 28% had a severe arrest and 40% had synaptic anomalies (P < 0.001). The incidence of synaptic anomalies is about five to six times higher than for the general infertile male population. These data suggest a relationship between the incidence of meiotic chromosome abnormalities and the degree of impairment of spermatogenesis. In our opinion, this high incidence of meiotic abnormalities in severe oligoasthenozoospermia makes meiotic studies in male infertility advisable before ICSI if they are available. Meiotic studies can be performed on testicular tissue samples (Egozcue et al., 1983
; Lange et al., 1997
) or in semen (Sperling and Kaden, 1971
; Templado et al., 1980
), although in the ejaculate sufficient material for a consistent diagnosis is only obtained in 2530% of cases.
Meiotic abnormalities include severe meiotic arrest and synaptic anomalies. Severe meiotic arrest due to synaptic anomalies or to unknown causes results in an arrest of spermatogenesis, usually at the stage of primary spermatocyte, resulting in oligozoospermia or azoospermia. Synaptic anomalies limited to the germ cell line in patients with normal karyotype and, therefore, only detectable by meiotic studies, are usually associated with an incomplete meiotic arrest and oligoasthenozoospermia (or azoospermia in case of complete meiotic arrest) and/or subsequent production of chromosomically abnormal spermatozoa (sperm aneuploidies or diploidies), which may be responsible for male infertility, spontaneous abortions during the first trimester of pregnancy or fetal chromosome abnormalities (Egozcue et al., 1983). In the present series, a former history of spontaneous abortions was recorded in three cases (2.9%). All three patients had a normal karyotype but synaptic anomalies were found in two of them. The single patient with an abnormal karyotype (46,XY/47,XXY) had a normal meiotic pattern.
In our group of 31 patients with increased FSH levels (>10 IU/l), total meiotic abnormalities occurred in 54.8% and synaptic anomalies in 22.6%. In contrast with others (Novero et al., 1997), this high incidence of meiotic abnormalities leads us to recommend the inclusion of FSH measurement in the routine evaluation of infertile male patients with oligoasthenozoospermia for ICSI.
We conclude that in male infertility due to oligoasthenozoospermia, sperm concentrations 1x106/ml and/or baseline serum FSH levels >10 IU/l are significant predictors of meiotic abnormalities. In these cases, meiotic chromosome studies can be performed to identify and characterize cytogenetic errors. This would allow the characterization of a particular `high-risk' group that needs genetic counselling and prenatal diagnosis in case of establishment of a pregnancy in ICSI cycles.
Meiotic studies are predictive, but they not provide information on the final status of the gametes. On the other hand, fluorescent in-situ hybridization (FISH) on decondensed sperm heads only provides information on the status of the chromosome pairs analysed with the probes used, may be difficult to perform on a significant number of spermatozoa in cases with very low sperm counts. If possible, we would recommend sperm chromosome studies, although they take a long time and are expensive, which makes them unpractical in clinical work.
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Acknowledgments |
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Notes |
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References |
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Submitted on July 28, 1998; accepted on November 17, 1998.