OBJECTIVETo investigate the influence of different methods of semen preservation and processing on sperm DNA integrity.

METHODSWe collected semen samples from 100 normozoospermic male volunteers and, following homogeneous mixing, preserved them by means of snap freezing, slow freezing, or at the room temperature for 4 and 24 hours. Meanwhile we processed the semen by washing, swim-up, and density gradient centrifugation (DGC). Then we obtained the sperm DNA fragmentation index (DFI) by sperm chromatin dispersion test and measured total sperm motility and DFI after cultured for 24 hours following processing.

RESULTSThe sperm DFIs after 4 hours of preservation by snap freezing, slow freezing, and at the room temperature were (27.3 ± 6.4)%, (26.9 ± 6.1)%, and (24.7 ± 6.8)%, respectively, and that after preserved at the room temperature for 24 hours was (35.6 ± 9.0)%, with statistically significant differences between the first three and the 24-hour room temperature preservation groups (P < 0.05) but not among the former three groups (P > 0.05). The sperm DFI was significantly higher in the samples processed by washing ([13.7 ± 2.0]%) than in those processed by swim-up ([9.1 ± 1.3]%) and DGC ([8.0 ± 2.5]%) (P < 0.05), and it was the lowest in the DGC group after 24-hour culture ([11.5 ± 4.2]%) as compared with the other groups (P < 0.05).

CONCLUSIONSperm DNA integrity is influenced by different semen preservation conditions and processing methods.

Centrifugation, Density Gradient ; DNA Fragmentation ; Humans ; Male ; Semen ; Semen Analysis ; Semen Preservation ; methods ; Sperm Motility ; Spermatozoa ; cytology

OBJECTIVETo investigate the effects of smoking on sperm apoptosis and semen quality of healthy adult males in the main urban area of Chongqing.

METHODSAccording to the smoking habit, we divided 235 healthy adult males into a non-smoking group (n = 89) and a smoking group (n = 146). Then we detected the routine semen parameters by the computer-assisted semen analysis system and obtained the parameters of sperm apoptosis (the ratios of AN-/PI-, AN+/PI-, AN+/PI+ and AN-/PI+ sperm) by flow cytometry combined with Annexin V-FITC/PI fluorescence staining.

RESULTSThe rate of early apoptotic sperm (AN+/PI-) was higher in the smoking than in the non-smoking group ([8.1 +/- 5.1]% vs [6.8 +/- 3.8]%; P = 0.039), but there were no significant differences between the two groups in the rate of late apoptotic sperm (AN+/PI+) ([5.6 +/- 5.2]% vs [5.5 +/- 5.1]%; P = 0.87), as well as in such routine semen indexes as semen volume, sperm density, sperm motility, sperm vitality and normal sperm morphology (P = 0.30, 0.82, 0.37, 0.81 and 0.84, respectively).

CONCLUSIONThe rate of early apoptotic sperm is higher in smokers than in non-smokers, suggesting that smoking may induce early damage to sperm cells. Compared with routine semen parameters, sperm apoptosis is a more sensitive biomarker to reflect smoking-induced damage to sperm.

Apoptosis ; China ; Humans ; Male ; Semen ; Semen Analysis ; Smoking ; Sperm Count ; Sperm Motility ; Spermatozoa ; cytology

Leukocytospermia is a most common cause of male infertility, but the distribution, origin and role of leukocytes in semen are still controversial. Some reports on leukocytospermia have indicated its negative effects on semen parameters and even in vitro fertilization (IVF). Recent literature has made it clear that the most deleterious effect of leukocytospermia is that the increased reactive oxygen species (ROS) may cause sperm damage, leading to significantly increased male infertility. The treatment and prevention of leukocytospermia have been proven of help for improving semen parameters.
Humans ; Infertility, Male ; etiology ; Leukocyte Count ; Leukocytes ; Male ; Reactive Oxygen Species ; metabolism ; Semen ; cytology ; Spermatozoa ; cytology

OBJECTIVETo discuss the clinical significance of determining immature germ cells (IGC) in human semen.

METHODSDiscontinuous Percoll gradients technique was employed to separate different cells and May-Grunwald-Giemsa staining and fluorescein isothiocyanate (FTTC)-Mab-CD45 was adopted to identify IGCs and leukcocytes in semen. The IGCs in 30 semen samples were determined including 10 fertile and 20 infertile cases.

RESULTSIGCs concentrated in gradient fractions with 30% to 45% Percoll and leukocytes concentrated in 50%-55% Percoll fractions. The concentration of IGCs was (0.70 +/- 0.40) x 10(6) ml in the fertile group and (1.28 +/- 0.70) x 10(6)/ml in the infertile group(P < 0.05). There was no statistical correlation between the IGC concentration and the sperm density, vitality and normal morphology(P > 0.05).

CONCLUSIONThe use of the discontinuous Percoll gradient method can reach the best separation of IGCs in the ejaculate and it is possible to be used as a clinical index to reflect semen quality.

Cell Separation ; Centrifugation, Density Gradient ; Humans ; Male ; Semen ; cytology ; Spermatozoa ; cytology

OBJECTIVETo assess the spermatogenic function of the infertile patients with Y-chromosomal microdeletion.

METHODSThirty-five 23-44 years old patients with microdeletions of Y chromosome were included in this study. Three semen analyses confirmed that 26 cases were non-obstructive azoospermia and 9 oligospermia with sperm count < 1 x 10(6)/ml. They were divided into 3 groups by the locus of deletion, 5 cases of AZFa + b + c deletion in group 1, 4 cases of AZFb + c and 3 cases of AZFb deletion in group 2, and 23 cases of AZFc deletion in group 3. Semen was collected and centrifuged, the supernatant removed and the centrifugate applied on the clean slides after dilution. Following Wright's-Giemsa staining, the slides were viewed under the microscope. Testis histopathological biopsy was performed for 6 of the cases.

RESULTSIn group 1, no spermatogenic cells were observed but only Sertoli cells in 1 case, with a consistency between the result of spermatogenic cell test and that of testis biopsy. In group 2, spermatogenic cell tests revealed spermatocytes in 6 cases, 2 were proved by testis biopsy with sperm maturation arrest in the primary spermatocyte stage, and spermatogenic cells of all developmental stages were seen in 1 AZFb deletion patient with the same sperm maturation arrest as the above two. In group 3, only primary spermatocytes were detected by spermatogenic cell test in 5 oligospermia patients but no spermatogenic cells in the 15 azoospermia cases, and biopsy revealed 2 cases of Sertoli cell-only syndrome.

CONCLUSIONThe spermatogenic cell test can effectively assess the spermatogenic function of AZF deletion patients. Non-invasive and easily accepted by patients, it is highly recommendable for the evaluation of spermatogenesis of patients with Y-chromosomal microdeletion.

Adult ; Chromosome Deletion ; Chromosomes, Human, Y ; Humans ; Infertility, Male ; etiology ; genetics ; pathology ; Male ; Semen ; cytology ; Semen Analysis ; Testis ; pathology

Fertility preservation is a hotspot of research in reproductive medicine, and that of male adolescent cancer patients is drawing even more attention from reproductive and oncologic clinicians. Both cancer and its treatment can decrease semen quality and even induce irreversible damage to fertility. Sperm cryopreservation is an effective method for fertility preservation. In the past few years, marked advances have been made in the cryopreservation, transplantation, and in vitro culture of testis tissue and stem spermatogonial cells. Although still experimental, these approaches may offer some options to those with no mature sperm in the testis. Unfortunately, very few people know and participate in the studies of fertility preservation and the utilization rate of cryopreserved sperm remains low. Therefor reproductive physicians and oncologists are required to make more efforts to search for effective fertility preservation methods for male adolescent cancer patients.
Adolescent ; Cryopreservation ; Fertility Preservation ; methods ; Humans ; Male ; Neoplasms ; therapy ; Semen Analysis ; Semen Preservation ; methods ; Spermatogonia ; Testis ; cytology

Sperm cryopreservation has revolutionized the field of assisted reproduction. However, it causes cryodamage to the structure and function of spermatozoa during the freezing-thawing process. Optimization of sperm cryopreservation is necessary for the preservation of male fertility. Cryodamage can be reduced effectively by such methods as improvement of semen quality before freezing, spermatozoal selection, addition of optimal cryoprotectants and application of appropriate thawing techniques. Recent studies focus on cryobiology, improvement of freezing-thawing methods, especially for poor quality semen, and evaluation criteria for post-thaw spermatozoa.
Cryopreservation ; methods ; Humans ; Male ; Semen Preservation ; methods ; Sperm Count ; Sperm Motility ; Spermatozoa ; cytology ; physiology

AIMTo establish a method for cynomolgus monkey sperm cryopreservation in a chemically defined extender.

METHODSSemen samples were collected by electro-ejaculation from four sexually mature male cynomolgus monkeys. The spermatozoa were frozen in straws by liquid nitrogen vapor using egg-yolk-free Tes-Tris mTTE synthetic extender and glycerol as cryoprotectant. The effects of glycerol concentration (1 %, 3 %, 5 %, 10 % and 15 % [v/v]) and its equilibration time (10 min, 30 min, 60 min and 90 min) on post-thaw spermatozoa were examined by sperm motility and sperm head membrane integrity.

RESULTSThe post-thaw motility and head membrane integrity of spermatozoa were significantly higher (P0.05) for 5 % glycerol (42.95 +/- 2.55 and 50.39+/- 2.42, respectively) than those of the other groups (1%: 19.19 +/- 3.22 and 24.84 +/- 3.64; 3%: 34.23 +/- 3.43 and 41.37 +/- 3.42; 10%:15.68 +/- 2.36 and 21.39 +/- 3.14; 15%: 7.47 +/- 1.44 and 12.90 +/- 2.18). The parameters for 30 min equilibration(42.95 +/- 2.55 and 50.39 +/- 2.42) were better (P0.05) than those of the other groups (10 min: 31.33 +/- 3.06 and 38.98 +/- 3.31; 60 min: 32.49 +/- 3.86 and 40.01 +/- 4.18; 90 min: 31.16 +/- 3.66 and 38.30 +/- 3.78). Five percent glycerol and 30 min equilibration yielded the highest post-thaw sperm motility and head membrane integrity.

CONCLUSIONCynomolgus monkey spermatozoa can be successfully cryopreserved in a chemically defined extender, which is related to the concentration and the equilibration time of glycerol.

Animals ; Cryopreservation ; Cryoprotective Agents ; Glycerol ; chemistry ; Macaca fascicularis ; Male ; Semen Preservation ; Spermatozoa ; cytology

Semen analysis is a basic test to evaluate male reproductive function. In recent years, urgent needs for the standardization of semen analysis have been emphasized among andrologists worldwide. This review discusses the standardization and quality control (QC) for the analysis of sperm quality parameters, including sperm concentration, motility and morphology. The key to sperm concentration analysis is the standardization of sperm-counting chamber, thus Cell-VU chamber may be the first choice. The analysis of sperm motility and morphology is too subjective to be reliable. Therefore, the computer-aided semen analysis (CASA) system may be the final selection. QC of semen analysis mainly lies in the selection of QC materials and the administration of external QC and internal QC. Meanwhile, the charts and arithmetic methods should be established to monitor QC of semen analysis.
Humans ; Male ; Quality Control ; Reference Standards ; Semen ; cytology ; Sperm Count ; instrumentation ; standards ; statistics & numerical data ; Sperm Motility ; Spermatozoa ; cytology ; physiology

OBJECTIVETo investigate the relationship of nitric oxide (NO) and nitric oxide synthase (NOS) with varicocele (VC) infertility.

METHODSFifty-three infertile men, 21 with varicocele and 32 with subvaricocele, were enrolled as Group 1, 29 infertile patients with oligoasthenozoospermia but without varicocele as Group 2 and 28 normal fertile controls as Group 3. The NO content and NOS activity in the seminal plasma and peripheral blood were measured by nitric acid reductase method, and the semen parameters of VC determined by computer-assisted semen analysis (CASA).

RESULTSSignificant differences were noted between Group 1 and the other two in the NO content and NOS activity in the seminal plasma (P < 0.05) but not in the peripheral blood (P > 0.05). In Group 1, the NO content and NOS activity were increased in both the seminal plasma and peripheral blood with the enhanced diameter of the varicose spermatic vein, with a significant difference only in the seminal plasma between the varicocele and subvaricocele patients (P < 0.05), and the same increase was observed with decreased sperm concentration (> or = 20 x 10(6)/ml and < or = 10 x 10(6)/ml) and motility (> or = 50% and < or = 25%), with significant differences (P < 0.05).

CONCLUSIONNO plays an important role in the VC-induced decrease of seminal quality. For the diagnosis of VC, the determination of the NO content and NOS activity in the seminal plasma is of more significance than that in the peripheral blood, and the earlier the determination, the greater its clinical value for both the diagnosis and treatment of VC.

Adult ; Humans ; Infertility, Male ; etiology ; metabolism ; Male ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; metabolism ; Semen ; cytology ; metabolism ; Semen Analysis ; Sperm Count ; Varicocele ; complications ; metabolism ; Young Adult