Natural history of autologous sperm cryostorage.

STUDY QUESTION: What is the natural history of elective autologous sperm cryostorage prior to gonadotoxic treatment? SUMMARY ANSWER: We estimate large sample median times to transfer for use, to the man's death or to discard of sperm, and their determinants, as the key operational outcomes of sperm cryostorage. WHAT IS KNOWN ALREADY: No large sample studies of the natural history of sperm cryostorage prior to gonadotoxic treatment are reported. STUDY DESIGN, SIZE, DURATION: This observational single-centre study covered 45 years of outcomes with a survival analysis for sperm cryostorage prior to scheduled gonadotoxic treatment, and its determinants. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study included 3923 men (mean age 30 years) who sought sperm cryostorage for a wide range of cancers and other diseases requiring gonadotoxic treatments. MAIN RESULTS AND THE ROLE OF CHANCE: The median time to transfer for use (n = 371 men 9%) was 2.4 years (quartiles 1.0, 6.0), the median time to death (n = 553 men, 14%) was 1.7 (0.9, 3.3) years, and the median time to discard (n = 1807 men, 46%) was 7.7 (1.7, 11.1) years. In multivariate Cox model regression, the underlying disease, number of storage visits and follow-up visits, and whether sperm were seen at follow-up visits were consistent predictors of times to outcomes. LIMITATIONS, REASONS FOR CAUTION: This study did not investigate sperm cryostorage for reasons other than gonadotoxic treatment, nor the fertilization outcomes of the cryostored sperm. WIDER IMPLICATIONS OF THE FINDINGS: These data provide estimates of the key operational factors for sperm cryostorage programs, prior to potentially sterilizing gonadotoxic treatments, and free from financial or insurance restrictions. STUDY FUNDING/COMPETING INTEREST(S): There was no specific funding for this study. D.J.H. has provided expert witness testimony to antidoping and professional standards tribunals and is supported by an NHMRC Investigator Grant. The other authors have no disclosures. TRIAL REGISTRATION NUMBER: N/A.

Effort in manual operation of reproductive health cryostorage facilities.

RESEARCH QUESTION: What areas of manual IVF cryostorage operations are common to the safe operation of IVF cryostorage facilities and require effort from embryologists? DESIGN: Observational time and motion data were collected by two observers equipped with the digital cameras over 2 weeks at four well-characterized US IVF centres (sites α, ß, γ and δ) from 12 participants performing cryostorage tasks. To understand the work processes of the different sites and assist in the data analysis, informal interviews were conducted with the study participants and laboratory directors. Data were analysed to identify work processes that might be eliminated or diminished by automation and software improvements. RESULTS: On average, it took 3.4 data record queries per retrieval from cryostorage to identify a cane, while the canister was lifted an average of 1.5 times per retrieval, with a mean 11.8 ± 9.2 s per lift. Of the total time spent working with cryostorage equipment, 47.25% was of a fatiguing nature. Sites α, ß and γ utilized one person to fill the liquid nitrogen storage Dewars, while site δ had two technicians working in tandem to move and fill the Dewars, with different frequencies and determination factors for refills and efficiencies. CONCLUSIONS: This time and motion study demonstrated significant time investment, task redundancy and fatiguing working conditions among embryologists using manual cryostorage processes. There was a disparity of processes and space capacity across different laboratories. Some of these issues may be addressed by the integration of automation and technology solutions.

Embryo long-term storage does not affect assisted reproductive technologies outcome: analysis of 58,001 vitrified blastocysts over 11 years.

BACKGROUND: Recently, the potential detrimental effect that the duration of storage time may have on vitrified samples has raised some concerns, especially when some studies found an association between cryostorage length and decreased clinical results. OBJECTIVE: This study aimed to evaluate the effects of the storage time length of day-5 vitrified blastocysts in 2 study groups: freeze-all cycles and nonelective frozen embryo transfers. STUDY DESIGN: This was a retrospective study that included 58,001 vitrified/warmed day-5 blastocysts from 2 different populations, according to the reason for frozen embryo transfer. Elective frozen embryo transfer comprised freeze-all cycles (N=16,615 blastocysts and 16,615 patients) in which only single embryo transfers and only the first frozen embryo transfer were included. The nonelective frozen embryo transfer group included 41,386 embryos from 25,571 patients where frozen embryo transfer took place using supernumerary embryos after fresh embryo transfer. All the possible frozen embryo transfers were included. Both single embryo transfer and double embryo transfers were included. Donor and autologous oocytes were used. The period covered by this study was 11 years. The blastocyst sample was clustered into deciles, which provided specific storage duration categories. The main outcome was the live birth rate, and secondary outcomes were embryo survival, miscarriage, and clinical and ongoing pregnancy rates according to storage duration. The impact of storage time was assessed by univariable analyses in both groups. The comparison was made between each decile and the last one. A multivariable logistic regression analysis was conducted, including the variables with significant association found in the univariate analysis. Student t test and chi-square tests, or an analysis of variance, were used wherever appropriate. P<.05 was considered statistically significant. RESULTS: There were statistical differences in baseline characteristics of patients included in the study groups. Storage durations ranged from ≤0.67 to ≥4.34 and from ≤1.8 to ≥34.81 months in freeze-all and nonelective frozen embryo transfer, respectively. Embryo survival did not show statistical differences across the categories of storage time in freeze-all and nonelective frozen embryo transfer groups. Statistical differences were found for the live birth rate across some, but not all, the subgroups of storage duration. The multivariable analysis showed no association between storage time and the live birth rate in both groups (nonsignificant). Blastocyst quality, body mass index, number of retrieved oocytes, endometrial preparation, male factor, and uterine factor were related to the drop in the live birth rate in the freeze-all group (P<.05). In the nonelective frozen embryo transfer group, the variables that showed significant association with the live birth rate were age at retrieval and frozen embryo transfer, type of frozen embryo transfer (single embryo transfer or double embryo transfers), number of retrieved oocytes, body mass index, endometrial preparation, origin of sperm sample, and female factor. CONCLUSION: This large study demonstrated no association between storage time and clinical outcome. Other variables, such as the patient's age, embryo quality, body mass index, and etiology, are somewhat responsible for impacting the outcome. This provides evidence for the safety of embryo vitrification, even after long storage periods. This is reassuring for both in vitro fertilization practitioners and patients undergoing frozen embryo transfer of either elective or nonelective embryos.

Influence of Storage Methods on the Vitality and Growth Rate of Macrofungi.

The work contains a comparative analysis of methods for storing pure cultures of macrofungi. The study used 20 species of macrofungi from various taxonomic and ecological-trophic groups. Storage was carried out using five methods: serial subculturing, storage under a layer of distilled water and three cryopreservation protocols: a protocol using blocks of agar medium, a "perlite protocol," and a "grain protocol." For the selected cryostorage methods, various cryoprotective compounds (glycerol, trehalose) were used. Radial growth rate was used as a criterion for the state of crops. The values of the radial growth rate obtained immediately after isolation of the pure culture were chosen as the control. It has been shown that the most favorable for preserving the physiological activity of cultures are the storage method under a layer of distilled water, "perlite" and "grain" cryopreservation protocols.

Cryostorage management of reproductive cells and tissues in ART: status, needs, opportunities and potential new challenges.

Among the wide range of procedures performed by clinical embryologists, the cryopreservation of reproductive cells and tissues represents a fundamental task in the daily routine. Indeed, cryopreservation procedures can be considered a subspecialty of medically assisted reproductive technology (ART), having the same relevance as sperm injection or embryo biopsy for preimplantation genetic testing. However, although a great deal of care has been devoted to optimizing cryopreservation protocols, the same energy has only recently been spent on developing and implementing strategies for the safe and reliable storage and transport of reproductive specimens. Herein, we have summarized the content of the available guidelines, the risks, the needs and the future perspectives regarding the management of cryopreservation biorepositories used in ART.

Deep technology for the optimization of cryostorage.

Cryopreservation, for many reasons, has assumed a central role in IVF treatment cycles, which has resulted in rapidly expanding cryopreserved oocyte and embryo inventory of IVF clinics. We aspire to consider how and with what resources and tools "deep" technology can offer solutions to these cryobiology programs. "Deep tech" has been applied as a global term to encompass the most advanced application of big data analysis for the most informed construction of algorithms and most sophisticated instrument design, utilizing, when appropriate and possible, models of automation and robotics to realize all opportunities for highest efficacy, efficiency, and consistency in a process.

The impact of Covid-19 on gamete shipping by Australian and New Zealand patients.

BACKGROUND: Cross-border surrogacy and egg donor arrangements are an increasingly common means to family building. Establishing patterns of use has always been difficult in relation to Australian patients. Accurate data is stymied by lack of documentation of international third-party reproductive care available to Australian authorities. When international travel bans came into effect, it is hypothesised that those planning to use cross-border reproductive care had to rely significantly more on local in vitro fertilisation (IVF) clinics for services such as sperm freezing, embryo creation and gamete release procedures. AIM: To quantify and characterise the impact of the Covid-19-related travel ban on international and interstate gamete shipping by Australian IVF clinics. MATERIALS AND METHODS: Thirty-one Australian and New Zealand IVF clinics were invited to provide de-identified data on interstate and international gamete export applications from two 12 month time periods pre- and during Covid-19-related international travel lockdowns. Seven IVF organisations provided data on: patient age; type of gametes exported; destination country/state; and date gamete release approved. RESULTS: Most gametes (78%) were shipped to another Australian IVF clinic and 22% internationally. Patient-initiated shipping domestically and internationally both showed significant increases when comparing pre- and post-Covid datasets. Of the 21 destination countries reported for international shipments, the US was the commonest (39%), followed by Ukraine (21%) and Canada (9%). CONCLUSIONS: The inability of involuntarily infertile patients to travel internationally, rather than halt cross-border reproductive care, has led to a significant increase in the uptake of gamete shipping. The high proportion of internationally shipped gametes going to the US and Ukraine is likely a reflection of the availability of surrogates and donors and more amenable legal frameworks.

Viable human brain microvessels for the study of aging and neurodegenerative diseases.

The brain microvasculature is altered in normal aging and in the presence of disease processes, such as neurodegeneration or ischemia, but there are few methods for studying living tissues. We now report that viable microvessels (MV) are readily isolated from brain tissue of subjects enrolled in studies of neurodegenerative diseases who undergo rapid autopsy (performed with <12 h postmortem interval - PMI). We find that these MV retain their morphology and cellular components and are fairly uniform in size. Sufficient MV (~3-5000) are obtained from 3 to 4 g of tissue to allow for studies of cellular composition as well as extracellular matrix (ECM). Using live/dead assays, these MV are viable for up to 5 days in tissue culture media (2D) designed to support endothelial cells and up to 11 days post-isolation in a 3-dimensional (3D) matrix (Low Growth Factor Matrigel™). Assays that measure the reducing potential of live cells \demonstrated that the majority of the MV maintain high levels of metabolic activity for a similar number of days as the live/dead assays. Functional cellular components (such as tight junctions and transporter proteins) and ECM of MV in tissue culture media, and to a lesser extent in 3D matrices, were readily visualized using immunofluorescence techniques. MV in tissue culture media are lysed and protein content analyzed, but MV in 3D matrix first require removal of the supporting matrix, which can confound the analysis of MV ECM. Finally, MV can be preserved in cryoprotective media, whereby over 50% retain their baseline viability upon thawing. In summary, we find that MV isolated from human brains undergoing rapid autopsy are viable in standard tissue culture for up to 5 days and the timeframe for experiments can be extended up to 11 days by use of a supportive 3D matrix. Viable human MV allow for temporal and spatial analysis of relevant cellular and ECM components that have implications for microvascular function in neurodegenerative diseases, vascular brain injury, and neurotrauma.

Clinical, obstetric and perinatal outcomes after vitrified-warmed euploid blastocyst transfer are independent of cryo-storage duration.

RESEARCH QUESTION: The study aimed to retrospectively evaluate the impact of cryo-storage duration on clinical, obstetric and perinatal outcomes after vitrified-warmed euploid blastocyst transfer. DESIGN: This was an observational study including 2688 vitrified-warmed euploid single blastocyst transfers that was conducted at a private IVF centre between May 2013 and March 2020. It included a total of 1884 women (age 38 ± 3 years) undergoing at least one transfer after preimplantation genetic testing for aneuploidies. The euploid blastocysts transferred were clustered into seven groups according to the cryo-storage duration between vitrification and warming: ≤60 days (n = 646; control group), 61-90 days (n = 599), 91-180 days (n = 679), 181-360 days (n = 405), 361-720 days (n = 144), 721-1080 days (n = 118) and >1080 days (n = 97). The primary outcome was the live birth rate (LBR) per transfer. The secondary outcomes were miscarriage rate, obstetric and perinatal issues. The data were adjusted for confounders through logistic or linear regressions. RESULTS: A significantly lower LBR was reported for transfers performed within 91-180 days (n = 291/679, 42.9%; P = 0.017), 181-360 days (n = 169/405, 41.7%; P = 0.016) and 361-720 days (n = 57/144, 39.6%; P = 0.034) versus ≤60 days (n = 319/646, 49.4%). However, this was mainly due to top-quality embryos being transferred first when more euploid blastocysts were available, thereby leaving lower quality ones for subsequent procedures. Indeed, the multivariate odds ratios adjusted for confounders showed similar results across all cryo-storage duration clusters. No difference was reported also for all secondary outcomes. CONCLUSIONS: Cryo-storage duration even beyond 3 years from blastocyst vitrification does not affect clinical, obstetric and perinatal outcomes.

The effect of extended cryo-storage following vitrification on embryo competence: a systematic review and meta-analysis.

PURPOSE: Few studies explored whether prolonged cryo-storage after vitrification affects embryo competence and perinatal outcomes. This systematic review and meta-analysis aims at highlighting any putative impact of cryo-storage duration on cryo-survival, miscarriage, live birth and major malformations. METHODS: A systematic review was performed using MEDLINE (PubMed), ISI Web of Knowledge, Scopus and Embase databases up to June 2021. Data were combined to obtain a pooled OR, and meta-analysis was conducted using a random effects model. Out of 1,389 screened abstracts, 22 papers were assessed for eligibility, and 5 studies were included (N = 18,047 embryos). Prolonged cryo-storage was defined as > 12 months (N = 3389 embryos). Subgroup analysis was performed for untested vitrified cleavage stage embryos (N = 1739 embryos) and for untested and euploid vitrified blastocysts (N = 13,596 and 2712 embryos, respectively). RESULTS: Survival rate, miscarriage, live birth and major malformation rates were all similar in the two groups. CONCLUSION: These data further support the safety of long-term cryo-storage of human embryos beyond 12 months. This is reassuring for good prognosis patients with surplus embryos, couples seeking a second child from supernumerary embryos and women postponing the transfer for clinical or personal reasons.

Simple method of thawing cryo-stored samples preserves ultrastructural features in electron microscopy.

Preservation of ultrastructural features in biological samples for electron microscopy (EM) is a challenging task that is routinely accomplished through chemical fixation or high-pressure freezing coupled to automated freeze substitution (AFS) using specialized devices. However, samples from clinical (e.g. "biobanking" of bulk biopsies) and preclinical (e.g. whole mouse tissues) specimens are often not specifically prepared for ultrastructural analyses but simply immersed in liquid nitrogen before long-term cryo-storage. We demonstrate that ultrastructural features of such samples are insufficiently conserved using AFS and developed a simple, rapid, and effective method for thawing that does not require specific instrumentation. This procedure consists of dry ice-cooled pre-trimming of frozen tissue and aldehyde fixation for 3 h at 37 °C followed by standard embedding steps. Herein investigated tissues comprised human term placentae, clinical lung samples, as well as mouse tissues of different composition (brown adipose tissue, white adipose tissue, cardiac muscle, skeletal muscle, liver). For all these tissues, we compared electron micrographs prepared from cryo-stored material with our method to images derived from directly prepared fresh tissues with standard chemical fixation. Our protocol yielded highly conserved ultrastructural features and tissue-specific details, largely matching the quality of fresh tissue samples. Furthermore, morphometric analysis of lipid droplets and mitochondria in livers of fasted mice demonstrated that statistically valid quantifications can be derived from samples prepared with our method. Overall, we provide a simple and effective protocol for accurate ultrastructural and morphometric analyses of cryo-stored bulk tissue samples.

Absence of SARS-CoV-2 (COVID-19 virus) within the IVF laboratory using strict patient screening and safety criteria.

RESEARCH QUESTION: Is there a risk of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral exposure and potential cross-contamination from follicular fluid, culture media and vitrification solution within the IVF laboratory using strict patient screening and safety measures? DESIGN: This was a prospective clinical study. All women undergoing transvaginal oocyte retrieval were required to have a negative SARS-CoV-2 RNA test 3-5 days prior to the procedure. Male partners were not tested. All cases used intracytoplasmic sperm injection (ICSI). The first tube of follicular fluid aspirated during oocyte retrieval, drops of media following removal of the embryos on day 5, and vitrification solution after blastocyst cryopreservation were analysed for SARS-CoV-2 RNA. RESULTS: In total, medium from 61 patients, vitrification solution from 200 patients and follicular fluid from 300 patients was analysed. All samples were negative for SARS-CoV-2 viral RNA. CONCLUSIONS: With stringent safety protocols in place, including testing of women and symptom-based screening of men, the presence of SARS-CoV-2 RNA was not detected in follicular fluid, medium or vitrification solution. This work demonstrates the possibility of implementing a rapid laboratory screening assay for SARS-CoV-2 and has implications for safe laboratory operations, including cryostorage recommendations.

Fertility treatment during the COVID-19 pandemic: A systematic review.

The COVID-19 pandemic has led to challenges in fertility preservation practices and has led to ethical issues, especially in developing countries. This paper provides a systematic review on this topic. At the beginning of the pandemic, several countries issued directions to suspend fertility treatments except among cancer patients. However, fertility preservation practices resumed gradually. The pandemic has evoked three major issues. First, many voices call for treating infertility as an essential medical condition in individual cases. There is no or negligible risk of transmission of COVID-19 through fertility treatment procedures or pregnancy. Second, there are weaknesses in health systems, especially in African countries. Third, there is enhanced discrimination and, in particular, a need to seriously consider inequality and social stratification in Africa. Oncofertility practices may be unevenly provided. The use of telemedicine to reduce nonessential contacts and the role of the Oncofertility Consortium in developing countries are highlighted.

The Impact of SARS-CoV-2 on Sperm Cryostorage, Theoretical or Real Risk?

Cryopreservation of human gametes and embryos as well as human reproductive tissues has been characterized as an essential process and aspect of assisted reproductive technology (ART). Notably, sperm cryopreservation is a fundamental aspect of cryopreservation in oncological patients or patients undergoing gonadotoxic treatment. Given that there is a risk of contamination or cross-contamination, either theoretical or real, during the procedures of cryopreservation and cryostorage, both the European Society for Human Reproduction and Embryology (ESHRE) and the American Society for Reproductive Medicine (ASRM) have provided updated guidelines for preventing or reducing the contamination risk of sexually transmitted viruses. Given the ongoing and worldwide COVID-19 pandemic, there is considerable interest in what measures should be taken to mitigate SARS-CoV-2 contamination during cryopreservation and cryostorage of semen samples. The SARS-CoV-2 virus is the virus that causes COVID-19, and whose transmission and infection is mainly aerosol-mediated. Several ART professional societies, including ESHRE and ASRM have proposed measures to mitigate the spread of the SARS-CoV-2 virus. Whether the proposed safety directives are enough to mitigate the possible SARS-CoV-2-contamination of sperm samples during cryopreservation or whether the policies should be re-evaluated will be discussed in this review. Additionally, insights regarding the possible impact of COVID-19 vaccination on the safety of sperm cryopreservation will be discussed.

Storage, Utilization, and Disposal of Hematopoietic Stem Cell Products in Patients with Multiple Myeloma.

High-dose chemotherapy (HD-CHT) and autologous blood stem cell transplantation (ABSCT) represent the standard of care in multiple myeloma (MM) for transplantation-eligible patients. Up to 3 HD-CHT/ABSCT treatments may be administered during the course of disease, including during late-onset relapse. Transplantation centers routinely collect more than 1 peripheral blood stem cell (PBSC) graft; however, subsequent HD-CHT/ABSCT treatments are often not performed, for various reasons. Currently, little is known about the actual utilization rate of stored PBSCs. The collection, storage, and disposal of PBSC products was analyzed in a large cohort of patients with MM (n = 1114) over a 12-year period with a minimum follow-up of 6 years. The final dataset analysis was performed in March 2019, which was set as the reference date. Based on institution-specific charges, the costs for PBSC collection, processing, and storage were estimated. The median number of sufficient PBSC transplantations per patient was 3 (range, 0 to 6), which were stored in a median of 3 (range, 1 to 11) cryopreserved bags (overall, n = 3644). A total of 95% of all patients (n = 1059) underwent at least 1 HD-CHT/ABSCT treatment. However, multiple ABSCTs were performed in 51% of the patients (n2/3 ABSCTs = 538), and only 14% of the patients underwent ABSCT 3 times (n3 ABSCTs = 149). Only a small proportion of collected PBSC bags (5%; n = 109) were used after being stored for longer than 5 years. Overall, 23% of the products (n = 830) were discarded, and 16% (n = 566) were kept in storage until the reference date. From a retrospective standpoint, the collected and discarded (definitively not used) or stored (potentially not used) cryostored PBSCs were associated with considerable costs for long-term cryostorage of approximately €1,600,000. We identified considerable discrepancies between the collection/storage and utilization of PBSCs. This is associated with significant efforts and costs on the one hand; on the other hand, disposal may raise legal and ethical questions. Therefore, we implemented comprehensive guidelines for the systematic reevaluation of stored PBSC grafts at our institution.

Fertility preservation during the COVID-19 pandemic: mitigating the viral contamination risk to reproductive cells in cryostorage.

Reopening fertility care services across the world in the midst of a pandemic brings with it numerous concerns that need immediate addressing, such as the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on the male and female reproductive cells and the plausible risk of cross-contamination and transmission. Due to the novelty of the disease the literature contains few reports confirming an association of SARS-CoV-2 with reproductive tissues, gametes and embryos. Cryobanking, an essential service in fertility preservation, carries the risk of cross-contamination through cryogenic medium and thus calls for risk-mitigation strategies. This review aims to address the available literature on the presence of SARS-CoV-2 on tissues, gametes and embryos, with special reference to the possible sources of cross-contamination through liquid nitrogen. Strategies for risk mitigation have been extrapolated from reports dealing with other viruses to the current global crisis, for safety in fertility treatment services in general, and specifically for oncofertility.

Cryostorage of silver barb (Barbodes gonionotus) semen in dry shipper: Efficacy, risk of bacterial cross-contamination and effects of Aeromonas hydrophila on post-thaw sperm quality.

This study was conducted to investigate the efficacy of dry shipper for the cryostorage of silver barb (Barbodes gonionotus) sperm, the subsequent risk of bacterial cross-contamination, and the effects of Aeromonas hydrophila on post-thaw sperm. Semen was diluted with calcium-free Hank's balanced salt solution containing 10% ME2SO, frozen at -8 °C/min and stored for 14 d in a dry shipper. A significant decline (P < 0.05) in the post-thaw sperm motility and viability of samples kept in the dry shipper for 14 d showed a reverse correlation (P < 0.05) with a slight increase in temperature within the dry shipper. The levels of contaminated bacteria in the compartments of the dry shipper were significantly (P < 0.05) lower than those detected in the liquid nitrogen tank. Bacteria from the atmosphere could recontaminate the chambers of the dry shipper and liquid nitrogen tank after 14 d. Bacillus was the most common bacteria isolated from the dry shipper, liquid nitrogen tank, circulating air, bench surface and outer surface of straws. There was no cross-contamination of A. hydrophila from contaminated straws to pathogen-free straws kept in either cryogenic tank. Post-thaw sperm motility and sperm viability significantly (P < 0.05) declined during cryostorage in the dry shipper and liquid nitrogen tank due to the introduction of A. hydrophila and the interaction effect of A. hydrophila and freezing. This study reports, for the first time, the efficacy of a dry shipper for the cryostorage of fish sperm for at least 14 d without a risk of bacterial cross-contamination.

Sperm cryostorage in a dry tank: An accurate alternative.

This prospective study aimed to determine the effects of dry nitrogen cryostorage on human sperm characteristics in comparison with liquid nitrogen cryostorage. For this purpose, 42 men undergoing routine semen analysis (21 normozoospermia and 21 with altered semen parameters) were analyzed. After slow freezing, half of the straws of each sample were randomly stored in liquid and dry tanks, at the top and bottom levels of the latter. After 6 months storage, thawed samples were treated by density gradient centrifugation and sperm characteristics were compared. There was no difference in sperm progressive motility (15.1% ± 14.2% vs. 15.1% ± 12.7%; p = 0.76), sperm vitality (25.5% ± 17.7% vs. 26.2% ± 19%; p = 0.71), percentages of acrosome-reacted spermatozoa (38% ± 8.5% vs. 38.5% ± 7.4%; p = 0.53) and DNA fragmentation spermatozoa (27.3% ± 12.4% vs. 28.5% ± 12.9%, p = 0.47) after cryostorage in the dry or the liquid nitrogen tank. Moreover, we did not observe differences between either cryostorage system for normal and altered sperm samples. This lack of difference was also observed whatever the floor level of cryostorage in the dry tank. The temperature measurement of the dry tank showed a stable temperature at -194 °C throughout storage whatever the storage floor level, guaranteeing the stability of the low temperatures suitable for human sperm storage. Because of its greater safety, dry storage without contact with the liquid phase should be preferred and can be a useful alternative for the cryostorage of human sperm samples.

Collection, Cryostorage, Transplantation, and Disposal of Hematopoietic Stem Cell Products.

Many transplantation centers routinely collect 1 or more autologous peripheral blood stem cell (PBSC) grafts in patients with hemato-oncologic and autoimmune disorders. However, subsequent high-dose chemotherapy and autologous blood stem cell transplantation (ABSCT) are often not performed, for various reasons. Currently, little is known about the actual utilization rate of stored PBSCs. We retrospectively analyzed the collection, storage, and disposal practices of PBSC products from a large cohort of patients (n = 1020) with hematologic, oncologic, and autoimmune disorders at our institution over a 12-year period. Patients with multiple myeloma were excluded. Based on our institution-specific charges, we estimated the costs for PBSC collection/processing and storage. The median number of sufficient PBSC collections per patient in the whole cohort was 2 (range, 1 to 6). We could demonstrate that only 67% of all patients who had collected sufficient PBSCs for transplantation actually underwent ABSCT, and only a small minority of all patients (4%) underwent multiple ABSCTs. The actual use of the stored PBSC grafts varied among disease entities from >80% to 0%. From a retrospective standpoint, the collected and discarded (definitively not used) or stored (potentially not used) cryostored PBSCs were associated with considerable costs of collection, cryopreservation, and long-term cryostorage. Although keeping open the therapeutic option for future transplantations may be important, there is currently a huge discrepancy between collection/storage practices and actual utilization of the cryopreserved PBSCs, at a considerable cost and strain on patients. Our study provides a rationale for reevaluating the present standards.

Sperm cryopreservation prior to gonadotoxic treatment: experience of a single academic centre over 4 decades.

STUDY QUESTION: What is the natural history of outcomes of sperm cryostorage at an Australian tertiary academic centre? SUMMARY ANSWER: Cryostorage is feasible in virtually all men facing gonadotoxic therapy but the timing of sperm disposal varies according to the reason for it. WHAT IS KNOWN ALREADY: Gonadotoxic treatment for cancer or non-cancer diseases damages spermatogenesis and impairs male fertility. Sperm cryopreservation is an established technique to preserve male fertility prior to gonadotoxic treatment. STUDY DESIGN, SIZE, DURATION: A retrospective review of clinical, anthropometric, semen analysis and hormonal data from 1978 to 2017 involving 2717 men comprising 2085 men with cancer, 234 non-cancer disease and 398 healthy controls, in a single tertiary academic centre with the same clinic and laboratory staff. PARTICIPANTS/MATERIALS, SETTING AND METHODS: Sperm output was analysed according to diseases, the feasibility of sperm cryostorage notably for adolescents, regional access to an urban cryostorage facility, the determinants of sperm output and time-dependent disposal of cryostored sperm. Semen samples were assessed by contemporaneous WHO methods. MAIN RESULTS AND THE ROLE OF CHANCE: Of 2085 men with cancer, 904 (43%) had haematological malignancies, 680 (33%) testicular cancers and 136 (6.5%) were adolescents. Most men (89%) and adolescents (80%) could collect sperm. Sperm output for all cancers and non-cancer diseases was lower than controls. Sperm output correlated positively with total testicular volume (r = 0.44, P < 0.0001) and negatively with serum FSH and LH (r = -0.24, -0.12, respectively, both P < 0.0001) but not testosterone. For all stored samples, the median time in cryostorage was 8.5 years, 7% were transferred for use to induce pregnancy (median time 2.5 years) and 62.2% were discarded as no longer needed (return of fertility, 35.9% median 3.5 years; death, 26.3%, median 6.5 years), the high disposal rate reflecting regular annual follow-up to establish ongoing need for continued cryostorage. Cryostorage facilities are not available in remote and rural areas of the State and the proportion of outer regional and remote area residents cryostoring sperm was only about half that compared with urban residents. LIMITATIONS, REASONS FOR CAUTION: This study does not report the pregnancy outcomes of the patients who used the cryostored sperm, due to recent limitations on health data privacy. WIDER IMPLICATIONS OF THE FINDINGS: Sperm cryostorage is feasible for virtually all men, including sufficiently mature adolescents, who can collect semen to insure future paternity as well as making positive psychological preparation for the patient's survival. Disposal of cryostored material when no longer required is efficient with regular follow-up. Sperm cryopreservation should be an integral part of comprehensive treatment plan in men receiving gonadotoxic treatment but remains underutilized. STUDY FUNDING/COMPETING INTEREST(S): There was no external funding for this study and there were no relevant conflicts of interest.