RESUMO
We evaluated sperm quality after a 3-month smoking cessation programme by sperm analysis, objective sperm motility analysis, protein tyrosine phosphorylation in capacitating conditions and DNA fragmentation (TUNEL). Sperm analysis after smoking cessation revealed a distinctive improvement in sperm concentration, fast spermatozoa (≥35 µm/s), sperm vitality, percentage of spermatozoa recuperated after an enrichment technique and protein tyrosine phosphorylation. However, no changes were observed in the number of germinal cells in the ejaculate, sperm morphology and sperm DNA fragmentation. It is concluded that physicians should strongly advise their patients to quit smoking before undergoing medical treatment or assisted reproduction techniques to achieve pregnancy.
Assuntos
Abandono do Hábito de Fumar , Espermatozoides/fisiologia , Feminino , Fertilização , Humanos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Masculino , Pessoa de Meia-Idade , Fosforilação , Gravidez , Taxa de Gravidez , Tirosina/metabolismoRESUMO
OBJECTIVES: To establish the validity of the TUNEL assay in determining sperm DNA fragmentation, the relationship between the degree of fragmentation and the seminal parameters and the sample needed to conduct the test. MATERIAL AND METHODS: We used semen samples from healthy fertile men (n=33), patients who consulted for infertility with a prescription for the TUNEL assay (n=77) and patients with intracytoplasmic sperm injection failure (n=20), analyzed according to the 2010 WHO. The TUNEL/propidium iodide test was performed by flow cytometry, on baseline and post-swim-up samples. RESULTS: The cutoff value for the TUNEL assay (ROC curves) was 26%, with a sensitivity and specificity of 85% and 89%, respectively. The pre-swim-up and post-swim-up medians of the results from the TUNEL assay showed no significant differences (17.0% vs. 12.9%, respectively). However, 39.1% of the samples showed a difference greater than 15 in absolute value between the results of the baseline and post-swim-up TUNEL assays. The linear correlation study of the morphology, mobility and vitality using the post-swim-up TUNEL assay showed a greater correlation than preselection, with significant results (r: -0.394, P<.0001; r: -0.461, P<.0001; r: -0.526, P<.0001). CONCLUSIONS: The TUNEL assay is a valid test for clinical use. DNA fragmentation is a factor independent from traditional semen tests. We found a greater susceptibility to damage generated in the laboratory procedures in the samples with lower quality. The sample of choice for evaluating DNA fragmentation will depend on whether the clinician is treating a natural or assisted fertilization.
Assuntos
DNA , Marcação In Situ das Extremidades Cortadas , Infertilidade Masculina/diagnóstico , Infertilidade Masculina/genética , Espermatozoides , Adulto , Humanos , Masculino , Reprodutibilidade dos TestesRESUMO
The first approach to assessing male fertility is to study a spermiogram, where special attention is given to sperm count, motility and morphology, while less attention is given to other cells in the ejaculate. Normal spermatogenesis requires a balance between cell death and proliferation; therefore, the number of germ cells (GC) in the ejaculate is less than the number of sperm. We propose a new index for altered spermatogenesis, i.e., the rate GC/sperm. We investigated a patient with oligozoospermia and a GC/sperm ratio greater than one, which indicated that spermatogenesis had been damaged. Complementary cytological tests were employed to characterized GC status: Papanicolaou stain, transmission electronic microscopy (TEM), vitality test, AgNOR and TUNEL assay. We also correlated cell morphology with ultrastructure studies that showed apoptosis. Nuclear apoptosis is characterized by vacuolization, misshapen nuclei, and "half moon," dispersed, uncondensed, disrupted and smudged chromatin. Cytoplasmic apoptosis is characterized by vacuolization, cytoplasmic protrusions, lamellar bodies, and swollen endoplasmic reticulum and mitochondria. To date, only testicular biopsy has been used to diagnose complete or incomplete testicular arrest. Our investigation is the first to determine a cytological feature in semen samples that could be used as a biological marker for abnormal spermatogenesis and for predicting the transition from oligospermia to azoospermia.
Assuntos
Apoptose/fisiologia , Azoospermia/patologia , Células Germinativas/patologia , Contagem de Espermatozoides/métodos , Motilidade dos Espermatozoides , Espermatozoides/patologia , Tomografia com Microscopia Eletrônica , Células Germinativas/metabolismo , Células Germinativas/ultraestrutura , Humanos , Infertilidade Masculina/patologia , Masculino , Oligospermia/patologia , Espermatogênese , Espermatozoides/metabolismo , Espermatozoides/ultraestruturaRESUMO
We propose that evaluation of protein tyrosine phosphorylation (TP) status in ejaculated spermatozoa under capacitating conditions in an experiment that mimics "in vitro" the physiology of sperm from ejaculation through the female genital tract could potentially be used as a prognostic test for functional competence of sperm in fertilization. Our purpose was to elucidate whether there is a relation between conventional sperm parameters, occurrence of TP and pregnancy outcome obtained from intrauterine insemination (IUI). Semen samples were analyzed according to WHO criteria. TP levels were determined by immunocytochemistry under four different conditions: 1) ejaculated sperm, 2) postselection sperm, 3) postselection sperm incubated 5 h at 37 degrees C and 5% CO(2), and 4) postselection sperm incubated overnight at 37 degrees C and 5% CO(2). Data on sperm tyrosine phosphorylated proteins did not correlate with sperm concentration, progressive motility or normal sperm morphology. TP increased under capacitating conditions and showed a time dependent pattern except for five outlier cases. IUI was performed in 12 selected couples who had neither female nor male infertility factors. The three pregnancies had a time dependent pattern for TP. Of the unsuccessful cases, one had an outlier TP pattern. It appears that a TP time dependent pattern is necessary for fertilization.
Assuntos
Andrologia , Imuno-Histoquímica/métodos , Laboratórios , Espermatozoides/metabolismo , Tirosina/metabolismo , Ejaculação , Feminino , Fertilização in vitro , Humanos , Infertilidade Masculina/metabolismo , Masculino , Fosforilação , Gravidez , Resultado da Gravidez , Prognóstico , Contagem de Espermatozoides , Espermatozoides/citologia , Espermatozoides/fisiologiaRESUMO
This is a retrospective study of clinical experience collected at the University Clinical Hospital over a 19-year period. Semen samples were analyzed according to WHO criteria. In the postmasturbatory urine, sperm count was performed. Data were expressed as total sperm number in urine (TSNU) and using a retroejaculation index. Patients were categorized into four groups according to the presence of sperm in the studied samples: a) in semen and urine; b) only in urine; c) only in semen; d) neither in semen nor in urine. A control group included nonretroejaculator patients. Retroejaculator patients are those whose TSNU is superior to 3.8 x 10(6) and the RI superior to 2.16%. While diagnosing retroejaculation, the only presence of sperm in the postmasturbatory urine is not adequate. The proposed index added to total sperm number in urine and semen volume may identify true retroejaculator patients.
Assuntos
Ejaculação/fisiologia , Sêmen/citologia , Espermatozoides/citologia , Urina/citologia , Humanos , Infertilidade Masculina/diagnóstico , Infertilidade Masculina/etiologia , Masculino , Estudos Retrospectivos , Contagem de EspermatozoidesRESUMO
Spermatozoa travel a long distance to meet and fertilize the oocyte, so sperm motility is a requisite for normal fertilization. Asthenozoospermia, or low sperm motility, is a common cause of human male infertility. This is a retrospective study (1992-1999) to document the prevalence of this pathology in infertile men and to clarify the probable factors associated to its etiology. The prevalence was 18.71% for asthenozoospermia and 63.13% for asthenozoospermia associated with oligo- and/or teratozoo-spermia; thus, 81.84% of the studied samples had altered motility. Leukocytospermia, the ratio of germ cells/sperm, anti-sperm antibodies, consistency, biochemical markers of accessory sex glands, and sperm response after swim-up were studied in normospermic (N), asthenozoospermic (A), and combined asthenozoospermic (C) samples. No significant difference was found in the frequency of leukocytospermia among groups. The rate of germ cells/(spermatozoa + germ cells) between C and N (p < .01) and C and A (p < .01) was statistically different, while no difference was found on comparing N and A. MAR-test over 40% was found in 6% of the A samples and 7.6% of the C, while no positive values were observed in the N group. The percentage of hyperviscous samples was higher in the low sperm motility samples than in the normal group. Data on concentration of the biochemical markers seem to be decreased in asthenozoospermia. Pure and combined asthenozoo-spermia showed different behavior in sperm recovery after swim-up. Two different asthenozoospermias could be defined: the pure one where sperm environment is involved (immunological factor, hyperviscosity, and secretory gland function) and the combined, where the testis is comprised.