Varicocelectomy improves sperm parameters, sperm DNA integrity as well as the other critical semen features.

Varicocele is a major entity defined within male infertility. In this report we have studied the influence of laparoscopic varicocelectomy on semen quality, biochemical parameters of seminal plasma and sperm DNA fragmentation. In this study, the semen samples from patients with left-side varicocele of grade II-III before and after laparoscopic varicocelectomy were compared to healthy individuals and separated into three groups. The volume of semen, sperm concentration (106/ml), motility (%), viability (%) and normal morphology (%) were assessed. Total antioxidant capacity (TAC), catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA) together with other biochemical substances in seminal plasma as alpha-glucosidase (α-Glu), fructose (Fr) and citric acid (CA) were determined by ELISA method. The spermatozoa activity including ion-transports through sodium, potassium ATPase (Na+, K+-ATPase) and calcium, magnesium ATPase (Ca2+, Mg2+-ATPase) were determined by using spectrophotometry. In addition, flow cytometry method for detection of sperm DNA fragmentation was used. The results showed, that three months after varicocelectomy such intervention led to significant postoperative improvement in volume of semen (p<0.001), total sperm count (p<0.001), sperm motility (p<0.001) and spermatozoa with normal morphology (p<0.001). We found decreased α-Glu levels due to varicocelectomy (p<0.05). There has been shown a high positive correlation between Na+, K+-ATPase and Ca2+, Mg2+-ATPase activity with total number of spermatozoa (p<0.05). The TAC levels and DNA fragmentation values after varicocelectomy can be considered as significant indicators of good prognosis after surgical intervention. It has to be emphasized that α-Glu levels and total sperm count expressed statistically significant both positive and negative predictive values for semen assessment. Varicocelectomy may lead to significant improvement of semen quality although the observations must be correlated with clinical pregnancies observed thereafter.

Sperm mitochondrial dysfunction and oxidative stress as possible reasons for isolated asthenozoospermia.

Reduced sperm motility, defined as asthenozoospermia, is a frequent cause of male infertility, and is mainly connected with the dysfunction of sperm mitochondria. The aim of this study was to identify the proteins, and thereby the metabolic pathways, responsible for asthenozoospermia, using 2-DE and MALDI-TOF MS, and correlate the results obtained with those of two mitochondrial tests: JC-1 and MitoSox Red. The JC-1 test was performed to test sperm mitochondrial activity, and the MitoSox Red test was performed to check whether the observed sperm poor motility is associated with mitochondrial reactive oxygen species (ROS) production. To identify proteins strictly connected with reduced sperm motility, men with isolated asthenozoospermia (n = 4 versus 10 normozoospermic controls) alone were included in the study. The proteomic analyses resulted in the identification of 25 sperm proteins that are differentially expressed in asthenozoospermic individuals. Most of the identified proteins were downregulated and were involved in energy production; however, we have also identified structural sperm proteins and proteins secreted by the epididymis. The latter, together with the results from MitoSox Red assay, may provide insights into the pathophysiological basis of asthenozoospermia.

Proteomic identification of sperm antigens using serum samples from individuals with and without antisperm antibodies.

The aim of the study was to identify human sperm antigens reacting with polyclonal antisperm antibodies. Protein sperm extracts were subjected to electrofocusing, and next immune reactions (immunoblotting) were carried out with positive for antisperm antibodies and control (not containing antisperm antibodies) serum samples. Proteomic analysis of human sperm proteins resulted in identification of 80 sperm antigens that could be divided into three groups: antigens specific for patients with antisperm antibodies (32), antigens recognised by both infertile patients and control sera (35) and antigens detected by control serum samples only (13). Among antigens specific for infertile patients, there were 12 sperm entities known to be involved in fertilisation process. We have also characterised three protein entities identified only by sera of infertile women. Altogether, the proteomic analysis resulted in identification of 27 sperm entities not reported previously in human sperm proteome. Identified proteins are sperm antigens that could be potentially responsible for immunological infertility. The study also sheds new light on the sperm antigens in aspect of gender specificity. The investigation of human sperm proteome by the use of antisperm antibodies-containing sera of infertile individuals not only may indicate new proteins but also can draft their immunological nature.

HLA-DQA1*0505 sharing and killer immunoglobulin-like receptors in sub fertile couples: report from the 15th International Histocompatibility Workshop.

This aim of the study was to investigate whether human leukocyte antigen (HLA)-DQA1*0505 sharing or the maternal killer immunoglobulin-like receptor (KIR) repertoire is associated with recurrent spontaneous abortion (RSA) or repeated implantation failure (RIF). The study included 224 couples with RSA, 61 couples with RIF, 182 fertile couples, and 10 couples with successful in vitro fertilization and embryo transfer (IVF)/ET at first cycle. HLA-DQA1*0505 typing using polymerase chain reaction-sequence-specific oligonucleotide (PCR-SSO) was performed in 185 RSA (117 with alloimmune abnormalities and 68 of autoimmune etiology), 61 RIF and 182 control couples, and KIR genotyping using polymerase chain reaction-sequence-specific primer (PCR-SSP) in 167 RSA and 55 RIF cases as well as 46 RSA and 10 IVF controls. No differences in DQA1*0505 sharing were found between patients and controls. In RSA and RIF women, the ratio of inhibitory to activating KIRs was slightly lower (1.53 and 1.85 vs 2.03 in controls). The analysis of maternal inhKIR and fetal HLA-C molecule pairs showed that the 'less inhibiting' combination KIR2DL3-C1 was found in higher percentage in subfertile (mainly RIF) than in fertile couples. In contrast, the percentage of cases possessing the 'strong inhibiting' combination KIR2DL1-C2 was lower in the RSA and RIF groups in comparison with that in the control groups (17.36% vs 23.91 and 16.36% vs 40%, respectively). In women with >or= 6 implantation failures, the KIR2DL1-C2 combination was not found in any of them (P = 0.0014), and the KIR2DL3-C1 combination was not found in the control IVF group. The results oppose the suggestion that increased HLA-DQA1*0505 sharing predispose to RSA or RIF. The KIR2DL3-C1 combination (or lack of the KIR2DL1-C2 one) is associated with implantation failure.

Idiopathic infertility in married couples in the light of cytogenetic analysis and sperm penetration assay.

We have selected 47 couples with unexplained infertility in order to analyse a possible link between sperm dysfunction studied in males in in vitro conditions and karyotype analysis of somatic cells. In order to identify so called "idiopathically infertile" couples we had to exclude any change in reproductive organs in both partners or in spermiogram which would qualify any of spouses into known category of infertility. We have revealed chromosome aberrations (translocations and marker chromosomes) in 19% of infertile males and in 6% of infertile females. Idiopathically infertile males had an overall decreased ability of sperm function (measured by proportion of penetrated hamster oocytes by human sperm) in comparison to fertile controls, however, still well placed within physiological range of values. Only sperm from a patient with identified translocation was clearly below the normal level of penetration (20% of penetrated oocytes), however, also the patients with revealed chromosome variant polymorphisms presented statistically lower values of penetration in comparison to fertile controls (39% vs 57%, p<0.05). On the contrary, patients with marker chromosomes did not exhibit affected sperm function. It can be speculated that only particular chromosome aberration in group of idiopathically infertile males may affect sperm functional capability (measured in vitro), however, the intragonadal genetic analysis has to be recommended in order to confirm such a causative link.

Antizona and antisperm antibodies in women with endometriosis and/or infertility.

OBJECTIVE: To measure the levels of antigamete antibodies in serum and peritoneal fluid of women with endometriosis and/or infertility. DESIGN: Antibody activity against human sperm and porcine oocytes was analyzed in selected subgroups of women. SETTING: Clinic of reproduction. PATIENT(S): Women with endometriosis and/or infertility. INTERVENTION(S): No treatment was implemented before peritoneal fluid and blood sample collection. MAIN OUTCOME MEASURE(S): Quantitative ELISA. RESULT(S): Four groups of women (n = 98) were analyzed for the presence of antizona and antisperm antibodies: infertile with endometriosis (n = 30), idiopathic infertility (n = 28), fertile with endometriosis (n = 20), and healthy fertile controls (n = 20). Antibodies were analyzed simultaneously in serum and peritoneal fluid. No statistically significant differences in antibody levels were detected in serum samples among the analyzed groups. The median values for antizona and antisperm antibodies in peritoneal fluid were significantly higher in women with idiopathic infertility than in the control group. In women with unexplained infertility, a high degree of correlation (Spearman) was found between the presence of antizona antibodies in peritoneal fluid and serum (r = 0.579). A positive predictive value of 80% was calculated for the presence of antizona antibodies (>5 ng/oocyte) in the peritoneal fluid of patients with infertility. CONCLUSION(S): Antizona antibodies locally produced in the peritoneal fluid have diagnostic value for infertility status; however, they cannot be treated as a marker or prognostic factor for minimal endometriosis and/or its treatment.

Sperm antigens recognized by antisperm antibodies present in sera of infertile adults and prepubertal boys with testicular failure.

Immunoblotting of a repertoire of sperm antigens reacting with antisperm antibodies present in sera of infertile adults and prepubertal boys with testicular failure was performed. In the subgroups selected for this study, 55% of examined infertile women, 65% of infertile men and 64% of prepubertal boys with gonadal failure gave positive results by Western blotting with extracted sperm antigens. Sperm antigens with molecular weights of 57, 58, 62, 63 and 66 kDa were the most immunodominant entities recognized by antisperm antibodies from prepubertal boys. No positive reactions were detected by Western blotting in a control population of fertile adults, whereas in a group of prepubertal healthy boys only one sample revealed reactivity against sperm antigens of 58 and 70 kDa.

Antisperm antibodies in prepubertal boys and their reactivity with antigenic determinants on differentiated spermatozoa.

PROBLEM: Antisperm antibodies induced in prepubertal boys with testicular failures were characterized by using four techniques of antibody detection. The reactivity of circulating antisperm antibodies in prepubertal boys and the reactivity of antibodies in sera samples of adult fertile and infertile males were compared against the same sperm antigenic pools (live or fixed spermatozoa, or sperm antigenic extracts). METHOD OF STUDY: The incidence of antisperm antibodies in sera samples of 69 prepubertal boys with testicular failures and 21 samples obtained from adult, male individuals was assessed by indirect immunobead binding test (IDIBT), flow cytometry measurement, enzyme-linked immunosorbent assay, and Western blotting. Immunoblot analysis was performed by using sperm extracts of glycosylated and deglycosylated solubilized membrane antigens. RESULTS: Sera samples were studied in a group composed of healthy prepubertal boys (n = 7) and prepubertal boys with testicular failures (n = 69). Applied tests of antibody detection revealed striking differences in a group of boys with testicular pathology. With IDIBT, 7% of the sera samples were found positive, whereas with flow cytometry measurement, 48% of the sera samples were positive. Immunosorbent assay (fixed sperm) indicated 32% positive cases in the same group. The sera samples were found to be positive in 65% of immunoblotting reactions with glycosylated antigens and in 70% of immunoblotting reactions with deglycosylated antigens. All applied detection assays were clearly negative on sera samples from fertile, adult males. Western immunoblotting indicated an immunodominant antigenic determinant of 58 kDa. CONCLUSIONS: Tests of antibody detection with the use of live sperm (IDIBT and flow cytometry measurements) presented low sensitivity (8% and 48%, respectively) in a group of prepubertal boys. This observation underlines the difficulties in assigning the prospective prognosis of future fertility status in prepubertal boys with antisperm antibodies.