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Biomedical subjects

L Gandini

Publications and source records attributed to L Gandini.

At least 37 records · Page 2Linked to original sources
PubMed1999

Immature germ cell separation using a modified discontinuous Percoll gradient technique in human semen.

The difficulty of identifying immature germ cells in unstained, fresh semen has led most laboratories to use the broad definition 'round cells' to indicate cells other than spermatozoa, thus grouping together both leukocytes and immature germ cells. This is also the case in research andrology, where very little attention has been given to immature germ cells in the semen apart from some rare exceptions, such as the attempts to study meiosis. Here we report on the use of a discontinuous Percoll gradient method modified to enable the best separation possible of immature germ cells from the other cells found in the ejaculate, in order to obtain a cellular suspension free of spermatozoa. Our technique (intra-assay variation in duplicates < 10%) demonstrated a high immature germ cell concentration in gradient fractions with 30% to 45% Percoll with a small contamination (1.5-6%) of leukocytes, confirmed by May-Grünwald-Giemsa staining, immunofluorescence and cytofluorimetry. The concentrations of immature germ cells ranged from zero in obstructive azoospermia to 2.0 x 10(6)/ml in oligozoospermia and genital tract infection. The purified immature germ cell suspensions obtained can be useful for diagnostic and research purposes.

Cell Separation↗
PubMed1999

Nuclear chromatin variations in human spermatozoa undergoing swim-up and cryopreservation evaluated by the flow cytometric sperm chromatin structure assay.

The sperm chromatin structure assay (SCSA) is a flow cytometric (FCM) technique which exploits the metachromatic properties of Acridine Orange to monitor the susceptibility of sperm chromatin DNA to in-situ acid denaturation. SCSA was used to study the chromatin structure variations of human spermatozoa in semen, both before and after swim-up and after cryopreservation. Semen samples were provided by 19 healthy normozoospermic subjects attending pre-marriage checks. Each sample was divided into three aliquots: the first aliquot was evaluated without further treatment, the second underwent swim-up, and the third was stored according to standard cryopreservation techniques in liquid nitrogen at -196 degrees C. Samples were also analysed by light and fluorescence microscopy (after Acridine Orange staining to evaluate the number of green fluorescent sperm heads), and by computer-assisted semen analysis. The results showed that post-rise spermatozoa represent a subpopulation characterized by a general improvement of the morphological (reduction of the percentage of abnormal forms and heads, increase of the green head sperm percentage) and kinetic parameters. This subpopulation also exhibited improved chromatin structure properties, confirming that these cells have the best structural and functional characteristics, indicative of optimal fertilizing ability. On the other hand, overall sperm quality deteriorates after cryopreservation. When thawed spermatozoa underwent an additional swim-up round, a general improvement of nuclear maturity was seen in the post-rise spermatozoa.

Adult↗
PubMed1999

Is hyperdiploidy of immature ejaculated germ cells predictive of testis malignancy? A comparative study in healthy normozoospermic, infertile, and testis tumor suffering subjects.

The possibility of diagnosing neoplastic testis pathologies by studying immature germ cells released from the seminiferous epithelium and present in the semen has been reported. It has been suggested that carcinoma in situ (CIS) of the testis and testis tumor may be identified by studying specific surface antigenic determinants or ploidy of chromosome 1 of malignant germ cells recovered from the semen. A noninvasive diagnostic approach of this type would be of great interest if we consider that CIS is supposed to precede the development of testicular germ cell tumors and that the frequency of that preneoplastic condition is increased in specific andrologic pathologies. To evaluate the reliability of this diagnostic approach, we have quantified the presence of immature hyperdiploid germ cells in the seminal fluid of normal healthy subjects, of infertile oligozoospermic patients affected by maldescended testis or left vancocele, and of patients suffering from CIS or testis tumor. Cell ploidy was evaluated on seminal cell fractions highly enriched in immature germ cells, by means of in situ hybridization with a DNA-probe specific for chromosome 1. Our observations indicate that chromosome 1 hyperdiploidy is not necessarily a predictive parameter of testis tumoral pathologies. The percentage of hyperdiploid immature seminal germ cells is in fact increased in nontumoral pathologies associated with infertility, such as cryptorchidism and left varicocele.

Adult↗
PubMed1998

Follicular fluid and human granulosa cell cultures: influence on sperm kinetic parameters, hyperactivation, and acrosome reaction.

OBJECTIVE: To evaluate the influence of human granulosa cell (GC) cultures and follicular fluid (FF) on sperm kinetic parameters, hyperactivation, and the acrosome reaction compared with the influence of human tubal fluid (HTF) and Ham's F-10 medium. DESIGN: Sperm kinetic parameters, hyperactivation, and the acrosome reaction were evaluated after 6 hours of incubation in HTF, Ham's F-10 medium, FF, and GC cultures. SETTING: Infertility and In Vitro Fertilization Centre, Reproductive Endocrinology Unit, Institute of Obstetrics and Gynaecology. PATIENT(S): Sixteen normal semen samples. INTERVENTION(S): Sperm kinetic parameters and hyperactivation were analyzed using an automated videomicrography system, the acrosome reaction was performed using a triple-stain technique, and progesterone and 17OH-progesterone levels were measured with the use of commercially available kits. MAIN OUTCOME MEASURE(S): Sperm kinetic parameters, hyperactivation, acrosome reaction. RESULT(S): The percentage of motile sperm, the mean curvilinear velocity, and the mean of the maximum amplitude of lateral head movement were increased significantly after 6 hours of incubation in FF or GC cultures compared with incubation in HTF or Ham's F-10 medium, whereas the mean linearity was decreased significantly. Follicular fluid and GC cultures significantly increased hyperactivation and the acrosome reaction compared with the values obtained using HTF and Ham's F-10 medium. Progesterone and 17OH-progesterone levels were increased significantly after incubation in FF and GC cultures compared with HTF and Ham's F-10 medium. CONCLUSION(S): Follicular fluid and GC cultures increase sperm motility parameters, hyperactivation, and the acrosome reaction. This effect may be related to GC detoxification of the microenvironement or GC secretion of peptides, glycoproteins, growth factors (insulin-like growth factors 1 and 2), or steroids (progesterone and 17OH-progesterone).

17-alpha-Hydroxyprogesterone↗
PubMed1998

Sperm parameters after early left varicocele treatment.

OBJECTIVE: We evaluated whether early treatment for left varicocele can improve sperm parameters. DESIGN: Patient follow-up and comparison with control groups. SETTING: Surgical department of a Children's Hospital and a university infertility-care center. PATIENT(S): Nineteen patients who underwent surgery for left varicocele as adolescents (mean +/- SD, 14.11 +/- 1.59 years) were followed up as adults (mean +/- SD, 19.37 +/- 1.16 years). MAIN OUTCOME MEASURE(S): A testicular examination was performed and varicocele staging was done at the time of surgery. Andrologic examination and semen analysis were done 2-8 years later. Semen results were compared with those of two age-matched control groups of 19 healthy, randomly selected subjects and 19 unoperated patients suffering from left varicocele of the same degree. RESULT(S): The mean values of sperm parameters in the patient study group were similar to those of the healthy controls except that the sperm concentration was significantly lower. However, sperm motility and morphology mean values in the patient study group were significantly higher than those of the unoperated patients. CONCLUSION(S): This study indicates that surgery in adolescence can be useful, possibly because the testis is developing at that age. Large-scale, double-blind controlled trials on early treatment of left varicocele are needed to identify possible markers of disease severity.

Adolescent↗
PubMed1997

DNA integrity in human sperm.

Since the 1970s there have been conflicting reports of decreasing sperm counts in man and increasing testicular cancer. There is a hypothetical link between apparent adverse trends in several measures of human reproductive health and exposure to endocrine disrupters. Rodent bioassays are not suited for the large-scale screening of such chemicals because of their costs, complexity, and ethical concerns. Various in vitro assays have been used to examine the effects of these chemicals, but none has directly used semen as one of the target tissues in man. The present study has examined in the alkaline Comet assay in human sperm the effect of two estrogens--beta-estradiol and the phytoestrogen daidzein--and 1,2-epoxybutene, a metabolite of 1,3-butadiene, and compared them with the effects of the known reprotoxin, dibromochloropropane, in two fertile and two infertile frozen sperm samples and two fresh fertile samples. While differences were detected in the frozen fertile and infertile samples with flow cytometry, in the Comet assay both frozen and fresh samples exposed to the chemicals in vitro from fertile and infertile men produced similar altered responses by comparison with untreated samples. The integrity of DNA is necessary not only for the noncancerous state, but also for the accurate transmission of genetic material to the next generation. Thus this assay may be useful for examination of chemicals in fresh and frozen sperm samples.

DNA↗
PubMed1997

Unilateral cryptorchidism corrected in prepubertal age: evaluation of sperm parameters, hormones, and antisperm antibodies in adult age.

OBJECTIVE: To evaluate whether prepubertal orchidopexy for unilateral cryptorchidism can reduce adult dysspermia. DESIGN: Patient follow-up comparison with control groups. SETTING: Surgical and endocrinologic sections of a children's hospital and a university infertility-care center. PATIENT(S): Seventy-one patients with unilateral cryptorchidism who underwent orchidopexy in prepubertal age (6.4 +/- 2.8 years) were followed up as adults (20.0 +/- 2.8 years). MAIN OUTCOME MEASURE(S): Patients underwent testicular examination and hormonal evaluation, 49 of these had semen analysis and antisperm antibody tests. Semen results were compared with those of two age-matched control groups: a group of 20 healthy, randomly selected subjects and a group of 20 patients operated on in postpubertal age for cryptorchidism. RESULT(S): Unilateral reduced testis size was found in 30.1% of patients, eight patients had a low LH level, eight had a low T level, and none had abnormal FSH values. Antisperm antibodies were found in 1 of 49 cases. Cluster analysis of sperm parameters showed that the mean values of patients were worse than those of the healthy controls but better than those of the subjects operated on in postpubertal age. CONCLUSION(S): This study indicates that prepubertal orchidopexy can given better results than postpubertal correction.

Adult↗
PubMed1997

The in-vitro effects of nicotine and cotinine on sperm motility.

Swim-up spermatozoa from the seminal samples of non-smokers, usually not exposed to passive smoking, were treated in vitro with nicotine (NIC) and cotinine (COT) at the average levels found in smokers' seminal plasma and at levels 500 times higher than this average. This was done to evaluate the action of these drugs on sperm motility. Each sample was allowed to swim up in Tyrode's solution with or without the drug; the study was carried out at time 0 and +1, +2, +4, +8 and +24 h of incubation, using a light microscope and a CASA system (experiment 1). In addition, the direct action of smoke on spermatozoa was studied using aspirated cigarette smoke (experiment 2). Kinetic parameters were then measured at 30 min, 45 min and 60 min starting from the last smoke injection. The first experiment showed that NIC and COT at average levels did not produce statistically significant variations of the kinetic parameters studied up to 24 h. However, the much higher concentration significantly altered all the kinetic variables in relation to the time of incubation. The second experiment with smoke in toto demonstrated a sharp reduction in all the sperm kinetic parameters. This reduction was seen after 30 min exposure to smoke and increased progressively until almost complete immotility at 1 h of exposure. These results suggest that NIC and COT are not responsible for the harmful effects of cigarette smoke on sperm kinetic parameters reported in the literature.

Adult↗
PubMed1997

Antisperm antibody detection: 1. Methods and standard protocol.

PROBLEM: Today, antisperm antibody (ASA) detection is one of the most important steps in the evaluation of male infertility. This practice is generally accepted even though there is still some disagreement about the meaning of antisperm immunity and a good deal of controversy about the test regarded as the most suitable for the detection of antibodies directed against sperm antigens. International workshops have tried to standardize universally accepted protocols. A panel of three or four methods is generally advised to provide a correct and complete screening of patients with antisperm immunity. METHOD OF STUDY: This paper reports on the results of a serum exchange workshop for the standardization of the ASA detection in the sera carried out under the auspices of the Italian Society of Endocrinology, which was used as reference laboratory. A careful description of the most widely used methods is reported and proposed as a standard protocol also on the basis of the results of the correlation studies carried out by our group is also reported. CONCLUSIONS: If the proposed methods and procedures will be accepted by the members of the Alps-Adria Society for Immunology of Reproduction, a Study Group for Sperm Antibody Testing, could start its activity having the goal of reaching a consensus on methodology and carrying out a European serum exchange workshop.

Agglutination Tests↗
PubMed1997

Antisperm antibody detection: 2. Clinical, biological, and statistical correlation between methods.

PROBLEM: In clinical andrology, the detection of antisperm antibodies (ASA) is regarded as one of the most important steps in the study of male infertility. This practice is generally accepted even though there is still some disagreement about the true meaning of antisperm immunity, and there remains a good deal of controversy about the test regarded as the most suitable for the detection of antibodies directed against sperm antigens. International Workshops have tried to standardize universally accepted protocols. A panel of three or four methods is generally advised to provide a correct and complete screening of patients with antisperm immunity. The aim of the present paper is to report our studies on the correlation between direct methods (IBT, MAR test) and indirect methods (gelatin agglutination test [GAT], and tray agglutination test [TAT]) and to establish whether biological models can explain the antibody test results. An attempt was also made to establish a "predictive threshold" to explain even apparently discordant direct and indirect results.

Agglutination Tests↗
PubMed1996

Effects of prolonged autovehicle driving on male reproduction function: a study among taxi drivers.

This study had the purpose of exploring the possible association between the work exposures of professional drivers and their reproductive health, by studying a group of 201 taxi drivers in the city of Rome. Data on work and reproductive history were collected by interviews. Biological markers examined in 72 subjects included salivary testosterone levels, sperm quality (i.e., sperm concentration, sperm morphology, and motility), and fertility experience, including time to pregnancy. Their spermatologic profile was compared with that of a control group of 50 healthy subjects of similar age and smoking habits. The results showed that taxi drivers, compared to the controls, had a significantly lower prevalence of normal sperm forms (45.8% vs. 64.0%); this was particularly true for those with a longer time on this job. This result was confirmed by a multivariate analysis in which confounders such as age, smoking, and alcohol consumption were controlled. The other sperm parameters did not differ in the study and the control groups. Among the life-style factors, we found smoking to be associated with poorer sperm morphology. Moderate alcohol consumption was associated with a better seminologic profile, while the pattern in respect to coffee intake was inconclusive. Subjects with poor semen quality also more frequently exhibited longer time to pregnancy of their partner. The results suggest that prolonged urban automobile driving might be a risk factors for sperm quality, and particularly for sperm morphology, but the finding needs further confirmation.

Adult↗
PubMed1995

Environmental tobacco smoke: nicotine and cotinine concentration in semen.

Forty-three nonsmokers attending an infertility clinic reported their degree of passive exposure to environmental tobacco smoke (ETS) and provided samples of blood and seminal plasma. Levels of nicotine and cotinine in biological fluid were evaluated by a radioimmunoassay. The data indicate that exposure to ETS results in measurable nicotine and cotinine levels in seminal plasma. Seminal plasma cotinine concentration also shows a significant positive correlation with degree of reported exposure.

Adult↗
PubMed1995

Study of antisperm antibodies bound to the sperm cell surface and their relationship to circulating ASA.

PROBLEM: Direct and indirect tests for antisperm antibody (ASA) detection are commonly used in laboratories for the diagnosis of male immunological infertility. Even though the two kinds of tests study the same immunological phenomena, frequently no perfect correlation is found even on large series of patients and with precise statistical models. The aim of the present investigation was to try to establish whether biological models can explain the antibody test results and if a predictive threshold can be established for expected positivity/negativity. METHOD: The data relate to 667 patients, who were screened using the Direct Immunobead test (d-IBT) for ASA bound to the sperm surface and with the gelatin and tray agglutination test (GAT and TAT) in sera for circulating ASA. Correlation were studied and cases of no correlation were further analyzed. RESULTS: The number of patients with a clinically significant positivity to d-IBT (binding percentage > or = 20%) was 134 (20.1% of total patients). The analysis of the results of direct and indirect tests shows that the discordances are due to differences in the Ig class of immunization or to the site of epitopes involved. In fact, as far the Ig class is concerned, if d-IBT shows a prevalent or exclusive IgG or IgA positivity, this results in a poor correlation between binding percentage to d-IBT and GAT-TAT titers. If both the Ig classes are involved, the level of positivity of the two kinds of test are strictly related. As far as the site of Ag-Ab reaction on the sperm surface is concerned, the wide immunization involving all the surface sperm antigens (mixed), shown by d-IBT, is related to higher indirect test titers. CONCLUSION: Local and systemic antisperm immunizations are strictly related and a predictive threshold of expectation can be established to explain even apparently discordant direct and indirect results.

Agglutination Tests↗
PubMed1994

Glutathione treatment of dyspermia: effect on the lipoperoxidation process.

We recently introduced reduced glutathione into the therapeutic protocols in some selected cases of dyspermia. This therapy improved semen quality both in a pilot follow-up study and in a double-blind cross-over trial. This improvement was seen in patients with varicocele and germ-free genital tract inflammation, two pathologies in which production of reactive oxygen species or other toxic compounds could have a pathogenic role. Polyunsaturated fatty acids of phospholipids play a major role in membrane constitution and function and are one of the main targets of the lipoperoxidative process. Therefore, to understand the therapeutic action of reduced glutathione, we selected infertile patients and studied the modifications produced by the therapy in seminal parameters, biochemical sperm membrane parameters, and the pattern of fatty acids of phospholipids from blood serum and red blood cell membranes (a model widely accepted as representative of general cell membrane status). The results showed an improvement in both sperm parameters and cell membrane characteristics. This study suggests that biochemical modifications in membrane constitution could explain the seminal results of glutathione therapy. On the other hand, it seems likely that only subjects with systemic membrane disturbances associated with andrological pathologies express this membrane damage in spermatozoa, resulting in dyspermia. This sperm alteration can be partially reversed by glutathione therapy if the structural cell membrane damage is not too severe.

Adult↗
PubMed1994

Non-human primate models for testing the efficacy and safety of periodontal regeneration procedures.

Experimentally-produced defects in the non-human primate are preferred for testing the efficacy and safety of periodontal regenerative therapies. These animals closely resemble the human in dental anatomy and periodontal wound healing physiology. Defects can be produced which do not display spontaneous regeneration and which are contralaterally identical. Thus, within animals, untreated controls can be obtained to indicate the level of supporting tissues prior to therapy. The use of non-human primate models is recommended for science transfer experiments involving invasive surgery and the testing of potentially harmful new devices and pharmaceuticals.

Acute Disease↗
PubMed1993

Computer assisted sperm motility analysis at the moment of induced pregnancy during gonadotropin treatment for hypogonadotropic hypogonadism.

Two azoospermic patients with isolated hypogonadotropic hypogonadism (IHH) no. 1,24-year-old, and no. 2,31-year-old), never treated before, were given gonadotropin therapy (GT), (FSH, 225 IU/week, and HCG, 4000 IU/week) and followed up monthly for sperm characteristics. After 6 (no. 1) and 9 (no. 2) months of treatment, immotile spermatozoa appeared in the semen. Two months later motile spermatozoa also appeared. A motile sperm concentration sufficiently high for computer motion analysis (CASA) was achieved 1 month later 9th and 12th month, respectively. At the 13th (no. 1) and 17th (no. 2) month of GT, CASA showed a small population of spermatozoa with a very good motility pattern. In the next ovulatory cycle the patients' partners became pregnant.

Adult↗