Antioxidant enzyme profile and lipid peroxidation products in semen samples of testicular germ cell tumor patients submitted to orchiectomy

ABSTRACT Purpose To determine enzymatic antioxidant and lipid peroxidation levels in seminal plasma of patients orchiectomized for testicular tumors. Materials and Methods The study included 52 patients: 26 control men and 26 orchiectomized patients for testicular tumor, of which 12 men had seminoma tumor and 14 men non-seminoma tumor. After semen analysis performed according to the WHO guidelines, an aliquot of semen was centrifuged and the seminal plasma was collected. Lipid peroxidation was performed by thiobarbituric acid reactive substances (TBARS) assay and antioxidant profile was assessed by analyzing catalase, glutathione peroxidase (GPx) and superoxide anion (SOD) activities using colorimetric assays with a standard spectrophotometer. Data were tested for normality and compared using one-way ANOVA (p<0.05). Results Seminoma and non-seminoma groups presented lower sperm concentration and morphology when compared to control group (p=0.0001). Both study groups (seminoma and non-seminoma) presented higher TBARS levels when compared to control group (p=0.0000013). No differences were observed for SOD (p=0.646) andGPx (p=0.328). It was not possible to access the enzymatic activity of catalase in any group. Conclusion Patients with testicular tumor present increased semen oxidative stress, but no differences were observed in antioxidant levels, even after orchiectomy. This indicates that most likely an increased generation of oxidative products takes place in these patients.

Antioxidant enzyme profile and lipid peroxidation products in semen samples of testicular germ cell tumor patients submitted to orchiectomy.

PURPOSE: To determine enzymatic antioxidant and lipid peroxidation levels in seminal plasma of patients orchiectomized for testicular tumors. MATERIALS AND METHODS: The study included 52 patients: 26 control men and 26 orchiectomized patients for testicular tumor, of which 12 men had seminoma tumor and 14 men non-seminoma tumor. After semen analysis performed according to the WHO guidelines, an aliquot of semen was centrifuged and the seminal plasma was collected. Lipid peroxidation was performed by thiobarbituric acid reactive substances(TBARS) assay and antioxidant profile was assessed by analyzing catalase, glutathione per-oxidase (GPx) and superoxide anion (SOD) activities using colorimetric assays with a standard spectrophotometer. Data were tested for normality and compared using one-way ANOVA (p<0.05). RESULTS: Seminoma and non-seminoma groups presented lower sperm concentration and morphology when compared to control group (p=0.0001). Both study groups (seminoma and non-seminoma) presented higher TBARS levels when compared to control group (p=0.0000013). No differences were observed for SOD (p=0.646) and GPx (p=0.328). It was not possible to access the enzymatic activity of catalase in any group. CONCLUSION: Patients with testicular tumor present increased semen oxidative stress, but no differences were observed in antioxidant levels, even after orchiectomy. This indicates that most likely an increased generation of oxidative products takes place in these patients.

Follicular fluid lipid peroxidation levels in women with endometriosis during controlled ovarian hyperstimulation.

This observational study aimed to establishing a relationship between lipid peroxidation and endometriosis in women undergoing controlled ovarian hyperstimulation. A total of 79 women were divided into two groups: (i) controls (tubal or male factor); and (ii) endometriosis (stages III/IV). The endometriosis diagnosis was confirmed by videolaparoscopy and the controlled ovarian stimulation protocol was similar to all patients. Follicular fluid (FF) lipid peroxidation levels were determined through the quantification of malondialdehyde. Statistical analysis was performed using parametric and non-parametric tests, logistic regression was performed to estimate the chance of achieving a pregnancy in each group and a moving average was calculated for the endometriosis group. Peroxidation levels in the endometriosis group were significantly higher when compared to controls. The moving average showed a decrease of MDA levels in the endometriosis group with increasing female age. Moreover, women with endometriosis who were under 33 years of age were 4.3 times more likely to achieve a pregnancy than women above that age. In conclusion, endometriosis is associated with increased FF oxidative stress (OS) in patients undergoing in vitro fertilization (IVF). Also, increasing age is associated with a decrease in severity of the oxidative status, but a decreased chance of pregnancy.

Alterations in the proliferative/apoptotic equilibrium in semen of adolescents with varicocele.

PURPOSE: To verify if the presence of varicocele (grades II and III) with and without seminal alterations, using the 5th centile cutoff values in table A1.1 of the World Health Organization (WHO, 2010) manual, alters the seminal plasma levels of proteins DNASE1 (deoxyribonuclease-1) and IGFBP7 (Insulin-like growth factor-binding protein 7), which are related to apoptosis regulation and cell proliferation, respectively, demonstrating that these proteins are important for correct spermatogenesis. METHODS: This cross sectional study was performed at the Sao Paulo Federal University Paulo between May 2014 and April 2016. A total of 61 male adolescents were included in this study, of which 20 controls without varicocele (C), 22 with varicocele and normal semen analysis (VNS) and 19 with varicocele and altered semen analysis (VAS). Seminal plasma from each patient was used for Western blotting analysis of individual protein levels. Values of each protein were normalized to a testicular housekeeping protein (PARK7-protein deglycase DJ-1). RESULTS: Levels of IGFBP7 protein are increased in varicocele. Levels of DNASE1 are progressively decreased in varicocele (lower in varicocele and normal semen analysis, lowest in varicocele and altered semen analysis) when compared to adolescents without varicocele. DNASE1 levels are positively correlated with sperm concentration and morphology (correlation values of 0.400 and 0.404, respectively; p values of 0.001 and 0.001, respectively). CONCLUSION: In conclusion, in adolescents, seminal plasma levels of IGFBP7, responsible for proliferative activity, are increased in varicocele grades II and III, and DNASE1, responsible for apoptosis regulation, are lower in varicocele, lowest in varicocele and low semen quality. These proteins demonstrate molecular alterations brought upon by varicocele. Moreover, DNASE1 is capable of discriminating a varicocele that causes alterations to semen quality from one that does not. We propose that the initial response of varicocele is to increase proliferative activity which, if followed by regulation of apoptosis, may lead to the ejaculation of a population of sperm that are in accordance with WHO cutoff values but, in the presence of dysregulated apoptosis, leads to lower sperm concentration and morphology.

Association between the seminal plasma proteome and sperm functional traits.

OBJECTIVE: To analyze the seminal plasma proteome and biological functions associated with sperm functional alterations. DESIGN: Cross-sectional study. SETTING: University andrology and research laboratories. PATIENT(S): A total of 156 normozoospermic men. INTERVENTION(S): Sperm mitochondrial activity, acrosome integrity, and DNA fragmentation were evaluated in a semen aliquot. Remaining semen was centrifuged, and seminal plasma was utilized for proteomic analysis (liquid chromatography-tandem mass spectrometry). Patients were divided into percentiles (15%) to form the following groups: substudy 1, high (control, n = 26) and low (study, n = 23) sperm mitochondrial activity; substudy 2, high (control, n = 23) and low (study, n = 22) sperm acrosome integrity; and substudy 3, low (control, n = 22) and high (study, n = 22) sperm DNA fragmentation. Groups were compared using univariate and multivariate analyses. Differentially expressed proteins were used for functional enrichment analysis. MAIN OUTCOME MEASURE(S): Seminal plasma proteome and postgenomic pathways are associated with several sperm functional traits. RESULT(S): In total, 506, 493, and 464 proteins were observed in substudies 1, 2, and 3, respectively. Enriched functions in substudy 1 were intramolecular oxidoreductase activity, aminoglycans catabolism, endopeptidases inhibition, lysosomes, and acute-phase response (study group). In substudy 2, main enriched functions were phospholipase inhibition, arachidonic acid metabolism, exocytosis, regulation of acute inflammation, response to hydrogen peroxide, and lysosomal transport (study group). In substudy 3, enriched functions were prostaglandin biosynthesis and fatty acid binding (study group). We proposed eight, six, and eight seminal biomarkers for substudies 1, 2, and 3, respectively. CONCLUSION(S): Seminal plasma proteome reflects sperm mitochondrial activity reduction, acrosome damage, and DNA fragmentation, with several postgenomic functions related to these alterations.

Lipidomics analysis of follicular fluid by ESI-MS reveals potential biomarkers for ovarian endometriosis.

PURPOSE: The aim of the present study was to analyze the lipid profile of follicular fluid from patients with endometriosis and endometrioma who underwent in vitro fertilization treatment (IVF). METHODS: The control group (n = 10) was composed of women with tubal factor or minimal male factor infertility who had positive pregnancy outcomes after IVF. The endometriosis group consisted of women with endometriosis diagnosed by videolaparoscopy (n = 10), and from the same patients, the endometriomas fluids were collected, which composed the endometrioma group (n = 10). From the follicular fluid and endometriomas, lipids were extracted by the Bligh and Dyer method, and the samples were analyzed by tandem mass spectrometry. RESULTS: We observed phosphatidylglycerol phosphate, phosphatidylcholine, phosphatidylserine, and phosphatidylnositol bisphosphate in the control group. In the endometriosis group, sphingolipids and phosphatidylcholines were more abundant, while in the endometrioma group, sphingolipids and phosphatidylcholines with different m/z from the endometriosis group were found in high abundance. CONCLUSION: This analysis demonstrated that there is a differential representation of these lipids according to their respective groups. In addition, the lipids found are involved in important mechanisms related to endometriosis progress in the ovary. Thus, the metabolomic approach for the study of lipids may be helpful in potential biomarker discovery.

Follicular fluid alterations in endometriosis: label-free proteomics by MS(E) as a functional tool for endometriosis.

Endometriosis is a chronic gynecological condition that affects 10-32% of women of reproductive age and may lead to infertility. The study of protein profiles in follicular fluid may assist in elucidating possible biomarkers related to this disease. For this, follicular fluid samples were obtained from women with tubal factor or minimal male factor infertility who had pregnancy outcomes after in vitro fertilization (IVF) treatment (control group, n = 10), women with endometriosis (endometriosis group, n = 10), along with the endometrioma from these same patients were included (endometrioma group, n = 10). For proteomic analysis, samples were pooled according to their respective groups and normalized to protein content. Proteins were analyzed by in tandem mass spectrometry (MS(E)) Spectra processing and the ProteinLynx Global Server v.2.5. was used for database searching. Data was submitted to the biological network analysis using Cytoscape 2.8.2 with ClueGO plugin. As a result, 535 proteins were identified among all groups. The control group differentially or uniquely expressed 33 (6%) proteins and equal expression of 98 (18%) proteins was observed in the control and endometriosis groups of which 41 (7%) proteins were further identified and/or quantified. Six (1%) proteins were observed in both the endometriosis and endometrioma groups, but 212 (39%) proteins were exclusively identified and/or quantified in the endometrioma group. There were 9 (1%) proteins observed in both the control and endometrioma groups and there were 139 (25%) proteins common among all three groups. Distinct differences among the protein profiles in the follicular fluid of patients included in this study were found, identifying proteins related to the disease progression and IVF success. Thus, some pathways related to endometriosis are associated with the presence of specific proteins, as well as the absence of others. This study provides a first step to the development of more sensitive diagnostic tests and treatment.

Follicular fluid lipid fingerprinting from women with PCOS and hyper response during IVF treatment.

PURPOSE: Polycystic ovary syndrome (PCOS) is an endocrine-metabolic disorder that leads to lower natural reproductive potential and presents a challenge for assisted reproductive medicine because patients may exhibit immature oocyte retrieval and a higher risk of ovarian hyper stimulation syndrome during in vitro fertilization (IVF) treatment. This study aimed to identify potential lipid biomarkers for women with PCOS and a hyper response to controlled ovarian stimulation. METHODS: Follicular fluid samples were collected from patients who underwent IVF, including normal responder women who became pregnant (control group, n = 11), women with PCOS and a hyper response to gonadotropins (PCOS group, n = 7) and women with only hyper response to gonadotropins (HR group, n = 7). A lipidomic analysis was performed by electrospray ionization mass spectrometry, and candidate biomarkers were analyzed by tandem mass spectrometry experiment. RESULTS: The lipid profiles indicated particularities related to differences in phosphatidylcholine (PCOS and HR), phosphatidylserine, phosphatydilinositol and phosphatidylglycerol (control), sphingolipids (PCOS) and phosphatidylethanolamine (control and HR). CONCLUSIONS: These findings contribute to the understanding of the molecular mechanisms associated with lipid metabolism in the PCOS-related hyper response, and strongly suggest that these lipids may be useful as biomarkers, leading to the development of more individualized treatment for pregnancy outcome.

Lipid profiling of follicular fluid from women undergoing IVF: young poor ovarian responders versus normal responders.

This study identified possible lipid biomarkers in follicular fluid from women with poor ovarian response. These biomarkers indicate pathophysiological pathways and have potential diagnostic applications. An observational case-control study of young women undergoing ovarian stimulation for in-vitro fertilization was conducted. The participants were categorized into a poor ovarian response group and a normal ovarian response to stimulation group. All of the women underwent the same ovarian stimulation protocol, and follicular fluid was collected after ovarian aspiration. Analyses were performed using matrix-assisted laser desorption/ionization mass spectrometry. Principal component analysis and Volcano plots were used to describe follicular fluid classification models based on the lipid profiles. A total of 10 lipids were differentially expressed between the study and control groups. Of these lipid ions, three belonged to the phosphatidylcholine subclass and were present in higher concentrations in the control group. The other seven differential lipids were present in the study group and classified into four lipid subclasses: phosphatidylethanolamines, phosphatidylglycerols, phosphatidylinositols, and diacylglycerols. These distinctive lipids may be involved in hormonal responses and oocyte development processes and may be useful as biomarkers for therapeutic intervention in women with poor ovarian response.

Changes in the seminal plasma proteome of adolescents before and after varicocelectomy.

OBJECTIVE: To compare seminal plasma protein profiles before and after varicocele correction to assess if surgical intervention alters the protein profile. DESIGN: Prospective study. SETTING: Academic research environment. PATIENT(S): Nineteen adolescent boys with varicocele grades II or III. INTERVENTION(S): Two semen samples were collected before bilateral subinguinal microsurgical varicocelectomy, and two semen samples were collected 3 months after surgery. Seminal plasma protein profiles were determined with the use of two-dimensional gel electrophoresis. Proteins were separated in 18-cm 3-10 pH strips and 10%-17.5% gradient gels. Gels were stained, scanned, and compared with the use of Imagemaster 2D platinum 7.0. Spots of interest were removed from gels, and protein digestion was performed with the use of trypsin. Digests were identified with the use of electrospray ionization-quadrupole/time-of-flight tandem mass spectrometry (ESI-QTOF MS/MS), and spectra were analyzed with the use of the Mascot software. MAIN OUTCOME MEASURE(S): Proteins uniquely or overexpressed in each period (before or after varicocelectomy). RESULT(S): Nineteen spots were differentially expressed between pre- and postsurgery samples. Identified proteins were albumin, proteasome subunit alpha type 6, alpha-1-antitrypsin, fibronectin, CD177, prostatic acid phosphatase, specific prostatic antigen, alpha-2-antiplasmin, vitamin D-binding protein, gastricsin, clusterin, semenogelin-1, semenogelin-2, superoxide dismutase, protein-glutamine gamma glutamyltransferase-4, and prolactin-inducing protein. CONCLUSION(S): Varicocelectomy is associated with changes in the seminal plasma protein profile. Understanding specific pathways leading to male infertility may further assist physicians in demonstrating deviation from homeostasis in male infertility. In addition, it may be possible to observe if surgical intervention does indeed revert altered pathways toward a homeostatic state.

Proteomic analysis of follicular fluid from women with and without endometriosis: new therapeutic targets and biomarkers.

Endometriosis is a gynecological disease that affects women of reproductive age. The protein profiles of women with endometriosis who were able or unable to achieve pregnancy and women without endometriosis who did achieve pregnancy were compared in this study. The follicular fluid was collected from 21 patients undergoing in vitro-fertilization treatment, according to the following groups: nine women in the control group (Group C), four women with endometriosis who achieved pregnancy (Group E.P), and eight women with endometriosis who did not achieve pregnancy (Group E.NP). Follicular fluid proteins were separated using 2D-electrophoresis, and their spots were compared, excised, and submitted to LC-ESI-MS/MS for proteins identification. The analysis showed 29 differentially expressed spots among the groups, and from these, 21 proteins were identified. Analysis showed some functional enrichment in the E.P group, including response to oxidative stress and apoptosis, while the E.NP group showed functions related to response to reactive oxygen species and positive regulation of apoptosis. These data suggest that endometriosis leads to differential protein expression in the follicular fluid, which can influences the outcome of pregnancy. These proteins may be potential targets for better diagnostics and new therapeutic intervention in affected women, as well as assisting in comprehending the physiopathologic mechanisms underlying endometriosis.

Effect of smoking on the functional aspects of sperm and seminal plasma protein profiles in patients with varicocele.

STUDY QUESTION: What are the effects of smoking on the functional aspects of the sperm, the levels of lipid peroxidation and the protein profile of seminal plasma in patients with varicocele? SUMMARY ANSWER: In men with varicocele, smoking is associated with altered semen quality, decreased sperm functional integrity and seminal oxidative stress. Alterations in seminal plasma protein profiles are also present and may explain the altered semen phenotype. WHAT IS KNOWN ALREADY: Varicocele is a major cause of male infertility. It reduces testicular blood renewal with a consequent accumulation of toxic substances. Thus, it can potentiate the toxic effects of environmental exposure to genotoxic substances such as those found in cigarette smoke. STUDY DESIGN, SIZE AND DURATION: A cross-sectional study was performed in 110 patients presenting with variococele to the Human Reproduction Section of the Sao Paulo Federal University (2006-2010). The patients were divided into a control group of non-smokers, a moderate smokers group and a heavy smokers group. PARTICIPANTS/MATERIALS, SETTING AND METHODS: Semen parameters were analysed by standard methods. Sperm DNA integrity and mitochondrial activity were assessed by Comet assays and by 3,3'-diaminobenzidine deposition, respectively. The level of lipid peroxidation in semen was determined by malondialdehyde quantification. Proteomic studies were performed by 2D-electrophoresis and mass spectrometry. MAIN RESULTS AND THE ROLE OF CHANCE: Both groups of smokers showed reduced semen quality in comparison with the control group. In the groups of smokers, sperm DNA integrity and mitochondrial activity were also decreased and lipid peroxidation levels were increased. Proteomic analyses revealed 20 proteins differentially expressed between the study groups. LIMITATIONS AND REASONS FOR CAUTION: A study including smokers without varicocele is still warranted as these results apply only to smokers who present varicocele. WIDER IMPLICATIONS OF THE FINDINGS: Patients with varicocele who are exposed to tobacco smoking present more important alterations to semen quality and sperm functional integrity and show changes in the seminal plasma proteome. This suggests testicular, and possibly systemic, adverse effects of smoking. STUDY FUNDING/COMPETING INTEREST(S): Funding for the study was provided by Fundação de Amparo à Pesquisa do Estado de São Paulo (Fapesp) (2007/59423-7) and by the Division of Urology, Human Reproduction Section at the São Paulo Federal University.

Ovodoação: a questão do sigilo / Egg donation and the issue of disclosure

O objetivo deste estudo foi identificar a intenção de revelar ou não para o filho, familiares e amigos a ovodoação e os motivos subjacentes. Participaram do estudo dez casais receptores de oócitos do Programa de Ovodoação da Universidade Federal de São Paulo. Foram realizadas entrevistas individuais, e os dados foram analisados segundo o método qualitativo. A maioria dos participantes relatou que não havia contado sobre a ovodoação para outras pessoas, assim como não tinha a intenção de contar para o filho. Foram alegados diversos motivos, acompanhados de temores relacionados às possíveis reações das crianças. O sigilo sobre a doação de gametas é uma questão complexa, que não deve ser respondida com regras simples e gerais, devendo-se considerar as especificidades de cada família e o contexto em que vivem.(AU)

The aim of this study was to identify the intention among parents to disclose conception via egg donation to their child, relatives and friends and the underlying reasons. The study considered ten couples to have received oocytes via the Egg Donation Programme at the Federal University of São Paulo. Most couples preferred not to disclose their conception via egg donation to friends or relatives and did not intend to inform their child. Various reasons were reported, as well as fears over the future child's potential reaction. Disclosure regarding gamete donation is a complex issue and approaches should not apply simple and general rules, but rather take into consideration each family's specific circumstances and context.(AU)

Ovodoação: a questão do sigilo / Egg donation and the issue of disclosure

O objetivo deste estudo foi identificar a intenção de revelar ou não para o filho, familiares e amigos a ovodoação e os motivos subjacentes. Participaram do estudo dez casais receptores de oócitos do Programa de Ovodoação da Universidade Federal de São Paulo. Foram realizadas entrevistas individuais, e os dados foram analisados segundo o método qualitativo. A maioria dos participantes relatou que não havia contado sobre a ovodoação para outras pessoas, assim como não tinha a intenção de contar para o filho. Foram alegados diversos motivos, acompanhados de temores relacionados às possíveis reações das crianças. O sigilo sobre a doação de gametas é uma questão complexa, que não deve ser respondida com regras simples e gerais, devendo-se considerar as especificidades de cada família e o contexto em que vivem.

The aim of this study was to identify the intention among parents to disclose conception via egg donation to their child, relatives and friends and the underlying reasons. The study considered ten couples to have received oocytes via the Egg Donation Programme at the Federal University of São Paulo. Most couples preferred not to disclose their conception via egg donation to friends or relatives and did not intend to inform their child. Various reasons were reported, as well as fears over the future child's potential reaction. Disclosure regarding gamete donation is a complex issue and approaches should not apply simple and general rules, but rather take into consideration each family's specific circumstances and context.

Effect of varicocele on sperm function and semen oxidative stress.

OBJECTIVE: To assess the effect of varicocele on sperm DNA integrity, mitochondrial activity, lipid peroxidation and acrosome integrity. PATIENTS AND METHODS: In all, 30 patients with a clinically diagnosed varicocele of grade II or III and 32 men without a varicocele were evaluated for sperm DNA fragmentation (comet assay), mitochondrial activity (3,3'-diaminobenzidine assay), lipid peroxidation (malondialdehyde) and acrosome integrity (fluorescent probe labelled peanut agglutinin). RESULTS: The varicocele group showed fewer spermatozoa with intact DNA (grade II, P= 0.040), more cells with inactive mitochondria (class III, P= 0.001), fewer cells with active mitochondria (class I, P= 0.005) and fewer spermatozoa with intact acrosomes (P < 0.001). Finally, no significant differences were observed in lipid peroxidation levels. CONCLUSION: Men with varicocele showed an increase in sperm DNA fragmentation and a reduction in mitochondrial activity and acrosome integrity. However, lipid peroxidation levels remained unchanged.

Adolescent varicocele: improved sperm function after varicocelectomy.

OBJECTIVE: To assess the effect of varicocelectomy on sperm function (DNA integrity and mitochondrial activity) and levels of lipid peroxidation in seminal plasma of adolescents. DESIGN: Prospective study. SETTING: Patients recruited from a local public school. PATIENT(S): Adolescents (14-19 years old), Tanner stages IV or V with varicocele grades II or III, attending a local public school. INTERVENTION(S): Two semen collections with a one week interval between collections before bilateral varicocele repair using subinguinal microsurgical varicocelectomy, and two semen collections with a one week interval between collections three months after the surgery. MAIN OUTCOME MEASURE(S): Rate of sperm DNA fragmentation as assessed by the Comet assay and categorized as classes I (no DNA fragmentation) to IV (high DNA fragmentation). Rate of mitochondrial activity as assessed by the diaminobenzidine assay and categorized as grades I (all mitochondria active) to IV (all mitochondria inactive). Levels of lipid peroxidation in seminal plasma by a colorimetric method that quantifies a lipid peroxidation subproduct (malondialdehyde). RESULT(S): Concerning DNA integrity, the samples after varicocelectomy showed more spermatozoa with intact nuclear DNA (grade I) and less spermatozoa with Comet grades II, III, and IV. Regarding mitochondrial activity, the samples after varicocelectomy showed less cells with inactive mitochondria (class III). No differences were observed in classes I, II, and IV. Concerning lipid peroxidation, no significant differences were observed. CONCLUSION(S): This study was able to demonstrate that varicocelectomy in adolescents is associated with increased sperm DNA integrity and mitochondrial activity. However, levels of seminal products of lipid degradation (malondialdehyde) are not different.

Expression of the Fas-ligand gene in ejaculated sperm from adolescents with and without varicocele.

PURPOSE: To assess FasL mRNA levels in ejaculated sperm from adolescent patients with and without varicocele. METHODS: Semen was obtained by masturbation following 2-4 days of ejaculatory abstinence, from 14 adolescents with varicocele grades II and III (study group), and 20 adolescents without varicocele (control group). Seminal analysis was done according to World Health Organization guidelines and morphology using Kruger's strict criteria. The Fas-ligand (FasL) gene expression was performed using reverse transcription and real-time quantitative polymerase chain reaction (RQ-PCR) analysis, according to the expression level of the housekeeping cyclophilin A gene. A Student's t-test was applied to compare the groups, and Spearman's rank test in order to verify possible correlations (p < 0.05). RESULTS: Quantitative RQ-PCR demonstrated that the expression of FasL mRNA in sperm from the varicocele group was higher than in the control group. Also, sperm concentration was higher in the controls, when compared to the varicocele group. When submitted to correlation analysis, adolescents with varicocele presented a correlation between sperm concentration and FasL gene expression levels (r = -0.470), not observed in controls. CONCLUSION: Our results allow us to conclude that, in adolescents with varicocele presenting lower sperm concentration, FasL mRNA levels are higher than in adolescents without varicocele.

Individual and seminal characteristics of patients with testicular germ cell tumors.

OBJECTIVE: To analyze the characteristics of patients with testicular germ cell cancer and compare patients' sperm quality according to histologic type (seminomatous and nonseminomatous tumors). DESIGN: Prospective study. SETTING: Sperm bank at a university. PATIENT(S): One hundred consecutive patients with testicular tumor who had been referred to our infertility center for cryopreservation, between 2004 and 2006. INTERVENTION(S): A questionnaire, through personal interview, was given to all patients and collection of seminal data before cryopreservation was performed. MAIN OUTCOME MEASURE(S): Patient characteristics, including age, time between diagnosis and orchiectomy, history of cryptorchidism, histologic type, and seminal analysis were taken into consideration. RESULT(S): The mean age of the patients at the time of diagnosis was 26.9 years. The mean time between cancer suspicion and the diagnosis of neoplasm was 58.9 days, and 19.4 more days were necessary until orchiectomy was performed. Eleven patients had a history of cryptorchidism. Thirty-seven patients had seminomatous tumors. Men with a seminoma present a higher number of motile and morphologically normal sperm in the ejaculate than men with a nonseminoma, although individual semen variables are not different. CONCLUSION(S): The majority of the patients with testicular cancer, referred to our infertility center, are very young, single, do not have children, and are unaware of their fertility potential status by the time diagnosis is made. Men with a nonseminoma present semen of lower quality.

Is sperm cryopreservation an option for fertility preservation in patients with spinal cord injury-induced anejaculation?

OBJECTIVE: To evaluate the effects of cryopreservation on sperm mitochondrial activity and nuclear DNA integrity in men with spinal cord injury. DESIGN: Prospective controlled study. SETTING: Patients in an academic research environment. PATIENT(S): Men with and without spinal cord injury-induced anejaculation. INTERVENTION(S): Electroejaculation or penile vibrating stimulation semen cryopreservation using a commercial TEST-yolk-buffer technique. MAIN OUTCOME MEASURE(S): Rate of sperm DNA fragmentation as assessed by the comet assay, graded in Classes I (high DNA integrity) to IV (high DNA fragmentation). Mitochondrial activity as assessed by a method in which active mitochondria precipitate 3,3'-diaminobenzidine. Cells were classified as I (all active) to IV (all inactive). Semen was cryopreserved in a Test-yolk buffer, and motility, DNA fragmentation, and mitochondrial activity were analyzed precryopreservation and postthaw. RESULT(S): Before cryopreservation, when the study (SCI) and control groups were compared, no statistically significant differences were found with respect to concentration or total sperm count; however, the SCI group presented significantly lower ejaculate volume, decreased sperm morphology, and an increase in the round cell and neutrophils counts. In both groups, cryopreservation was associated with an increase in DNA fragmentation, a decrease in mitochondrial activity, and a decrease in motility, of which the latter was of greater importance in the control group. CONCLUSION(S): Cryopreservation causes a decrease in conventional seminal variables as well as in mitochondrial activity and DNA fragmentation. However, these were no more detrimental to sperm from men with SCI than to sperm from the control group.