Human sperm cryopreservation is an increasingly mature technique in assisted reproduction. However, conventional sperm cryopreservation is not suitable for the cryopreservation of small numbers of sperm. The solution to the cryopreservation of small numbers of sperm may contribute a lot to the clinical treatment of asthenospermia, oligospermia and azoospermatism. Recently, many researchers focus on searching for appropriate carriers for the cryopreservation of small numbers of sperm. This article outlines the effects of current cryopreservation methods including empty zona pellucida, microdrops, other mocrocarriers, testicular tissue cryopreservation and testicular sperm and epididymal sperm refrigeration.
Cryopreservation ; methods ; Humans ; Male ; Semen Preservation ; methods ; Testis

Human sperm cryopreservation has been widely applied in human assisted reproduction technologies, but ultra-low temperature may damage the structure and function of sperm, which in turn may affect the success rate of human reproductive technology. However, this view is not yet universally accepted. Researchers around the world are endeavoring for the mechanisms of cyrodamage and discoveries of cryo-protection. This article gives an overview on the types of human sperm cryodamage, mechanisms of sperm functional changes, and latest discoveries of cryo-protection.
Cryopreservation ; Humans ; Male ; Semen Preservation ; adverse effects ; methods

Despite the worldwide application of sperm-freezing technology as a means of preserving male fertility functions, few reports are seen about the transmission of microorganisms to cryopreserved sperm via contaminated liquid nitrogen (LN). Although the risk of cross-contamination between samples stored in liquid nitrogen is "vanishingly small", it must be accepted as a finite risk and all reasonable measures taken to reduce the likelihood of its occurrence. Moreover, all methods employed, including collection, cryopreservation, storage and thawing of human sperm as well as the clinical use of cryobanked human sperm, must reduce the risk of contamination from any source throughout the entire process.
Cryopreservation ; methods ; Equipment Contamination ; Humans ; Male ; Semen Preservation ; methods

OBJECTIVETo provide a more effective cryoprotective medium (CPM), effect of union of albumin and egg yolk on human sperm cryopreservation was studied.

METHODSEgg yolk-glycerol-sodium citrate was regarded as CPM of the control group and egg yolk-glycerol-sodium citrate with different concentrations of albumin (1, 2, 3, 4, 5 g/L) were regarded as CPMs of experiment groups. Before and after cryopreservation, sperm movement parameters were assessed by using computer-aided sperm analyzer (CASA) system, and then egg yolk-glycerol-sodium citrate group added 1 g/L albumin was selected, whose movement parameters were the best among the experimental groups, and egg yolk-glycerol-sodium citrateto group as the control to compare sperm survival rate, membrane integrity, function of mitochondrion and ultrastruction.

RESULTSSperm in egg yolk-glycerol-sodium citrate added I g/L albumin showed significantly higher motility, viability than those in the control group and other experimental groups (P < 0.05). Sperm in egg yolk-glycerol-sodium citrate group added 1 g/L albumin had significantly higher survival rate, head unpigmenting rate than those in control group (P < 0.05). Sperm in egg yolk-glycerol-sodium citrate group added I g/L albumin manifested significantly higher succinate dehydrogenase (SDH) activity than that in control group (P < 0.05) and had better ultrastructure than that in control group.

CONCLUSIONUnion of two kinds of albumin and egg yolk has better effects on human sperm cryopreservation than those of solitary use of egg yolk. The action of albumin is related to its concentration, and albumin combining with egg yolk may have plus and complementary effects on human sperm cryopreservation.

Adult ; Albumins ; Cryopreservation ; Egg Yolk ; Humans ; Male ; Semen Preservation ; methods

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

OBJECTIVETo study the effect of direct fumigation on the post-thaw recovery rate of cryopreserved spermatozoa, and to search for a best method for human sperm cryopreservation.

METHODSWe collected semen samples from 100 donors conforming to the normal reference values in WHO Laboratory Manual for the Examination and Processing of Human Semen (5th Ed), divided them into two groups, and subjected them to cryopreservation by programmable freezing (Group A) and direct fumigation (Group B), respectively. We detected the progressive motility of pre-freezing and post-thaw sperm with a computer-assisted semen analyzer, and compared the effects of the two methods on the functional integrity of sperm membrane and the rate of abnormal sperm using the percentage of hypo-osmotic swelling sperm and modified Papanicolaou staining.

RESULTSStatistically significant differences were found in post-thaw sperm progressive motility between the Groups A and B ([34.0 +/- 18.4]% vs [43.0 +/- 19.5]%, P<0.05), both remarkably decreased as compared with pre-freezing ([57.0 +/- 16.7]%, P<0.05). Such differences were also found in the post-thaw recovery rate of progressively motile sperm between the two groups ([52.2 +/- 20.6]% vs [67.1 +/- 20.0]%, P<0.05). The post-thaw percentage of hypo-osmotic swelling sperm was obviously decreased in both Groups A and B ([67.1 +/- 11.1]% and [70.6 +/- 10.0]%) in comparison with pre-freezing ([84.5 +/- 7.5]%, P<0.05), with significant differences between A and B (P<0.05). However, the rate of sperm abnormality was evidently increased in Groups A and B ([85.0 +/- 8.7% and [85.7 +/- 9.1]%), significantly higher than pre-freezing ([77.8 +/- 9.6]%, P<0.05), but with no significant differences between A and B (P>0.05).

CONCLUSIONDirect fumigation is superior to programmable freezing for its easier operation, wider application, and higher sperm recovery rate.

Adult ; Cryopreservation ; methods ; Humans ; Male ; Semen Analysis ; Semen Preservation ; methods ; Sperm Motility ; Spermatozoa ; Young Adult

AIMTo evaluate two extenders and two cryoprotectant agents (CPA) for alpaca semen cryopreservation.

METHODSSemen samples were obtained from four adult alpacas (Lama pacos) and frozen using extender I (TRIS, citrate, egg yolk and glucose) or extender II (skim milk, egg yolk and fructose), each containing either glycerol (G) or ethylene glycol (EG) as CPA. Consequently, four groups were formed: 1) extender I-G; 2) extender I-EG; 3) extender II-G; and 4) extender II-EG. Semen was diluted in a two-step process: for cooling to 5 degrees (extenders without CPA), and for freezing (extenders with CPA). Viability and acrosome integrity were assessed using trypan blue and Giemsa stains.

RESULTSWhen compared, the motility after thawing was higher (P >0.05) in groups II-EG (20.0% +/- 6.7%) and II-G (15.3 % +/- 4.1% ) than that in groups I-G (4.0 % +/- 1.1%) and I-EG (1.0 % +/- 1.4%). Viable spermatozoa with intact acrosomes in groups II-EG (18.7 % +/- 2.9%) and II-G (12.7 % +/- 5.9%) were higher than that in groups I-G (5.7% +/- 1.5%) and I-EG (4.0% +/- 1.0%).

CONCLUSIONThe skim milk- and egg yolk-based extenders containing ethylene glycol or glycerol to freeze alpaca semen seems to promote the survival of more sperm cells with intact acrosomes than the other extenders.

Animals ; Camelids, New World ; Cryoprotective Agents ; administration & dosage ; Freezing ; Male ; Semen ; Semen Preservation ; Sperm Motility

In the 1960s, sperm cryopreservation was developed as a method to preserve fertility. Currently, techniques for the cryopreservation of human spermatozoa have been widely used in assisted reproduction. However, although sperm cryobiology has made notable achievements, the optimal method for the recovery of viable spermatozoa after cryopreservation remains elusive. Postthawing sperm quality can be affected by cryoprotectants, ice formation, storage conditions, and osmotic stress during the freezing process. This review discusses recent advances in different cryopreservation techniques, cryoprotectants, and freezing and thawing methods during cryopreservation and new indications for the use of cryopreserved spermatozoa.
Humans ; Male ; Semen Preservation/methods* ; Sperm Motility ; Semen ; Cryopreservation/methods* ; Spermatozoa ; Cryoprotective Agents/pharmacology*

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

The use of artificial insemination (AI) in buffalo (Bubalus bubalis) is limited by poor ovarian activity during the hot season, seasonal qualitative patterns in semen, low resistance of sperm cells in the female tract, difficulties in estrus detection, and variable estrus duration. Although AI procedures are commonly used in bovine, use of AI has been limited in buffalo. In the zootechnical field, different studies have been conducted to develop techniques for improvement of fertilizing ability of buffalo spermatozoa after AI. In this study, for the first time, the use of alginate encapsulation and cryopreservation of buffalo spermatozoa is described, and the same procedure was performed with Holstein Friesian (Bos taurus) semen. Results obtained from in vitro analyses indicate that the encapsulation process does not have detrimental effects (compared to controls) on quality parameters (membrane integrity, progressive motility, path average velocity) in either species. Similarly, there were no detrimental effects after cryopreservation in either species. The fertilizing potential of encapsulated and cryopreserved semen was evaluated after AI in 25 buffalo and 113 bovine females. Pregnancy rates were not affected in either species. The results of this study show proof of concept for the use of frozen semen controlled-release devices in buffalo.
Buffaloes* ; Cryopreservation* ; Estrus ; Estrus Detection ; Female ; Humans ; In Vitro Techniques ; Insemination, Artificial ; Pregnancy Rate ; Seasons ; Semen ; Semen Preservation ; Spermatozoa* ; Water*