Successful cryopreservation of human ovarian cortex tissues using supercooling.
Ontology highlight
ABSTRACT:
SUBMITTER: Moriguchi H
PROVIDER: S-EPMC3406338 | biostudies-literature | 2012
REPOSITORIES: biostudies-literature
Publications
Successful cryopreservation of human ovarian cortex tissues using supercooling.
Moriguchi Hisashi H Zhang Yue Y Mihara Makoto M Sato Chifumi C
Scientific reports 20120727
The development of new method to cryopreserve human ovarian cortex tissues without damage is needed for the improvement of quality of life (QOL) of female cancer patients. Here we show novel cryopreservation method of human ovarian cortex tissues by using supercooling (S.C.) procedure. Our method will be helpful in order to preserve fertility of female cancer patients. ...[more]
Similar Datasets
Project description:BackgroundCryopreservation of leukocytes isolated from the cervicovaginal and colorectal mucosa is useful for the study of cellular immunity (see Hughes SM et al. PLOS ONE 2016). However, some questions about mucosal biology and sexually transmitted infections are better addressed with intact mucosal tissue, for which there is no standard cryopreservation protocol.Methods and findingsTo find an optimal preservation protocol for mucosal tissues, we tested slow cooling (1°C/min) with 10% dimethylsulfoxide (designated "cryopreservation") and fast cooling (plunge in liquid nitrogen) with 20% dimethylsulfoxide and 20% ethylene glycol ("vitrification"). We compared fresh and preserved human cervicovaginal and colorectal tissues in a range of assays, including metabolic activity, human immunodeficiency virus infection, cell phenotype, tissue structure by hematoxylin-and-eosin staining, cell number and viability, production of cytokines, and microbicide drug concentrations. Metabolic activity, HIV infectability, and tissue structure were similar in cryopreserved and vitrified vaginal tissues. However, vitrification led to poor cell recovery from the colorectal mucosa, with 90% fewer cells recovered after isolation from vitrified colorectal tissues than from cryopreserved. HIV infection rates were similar for fresh and cryopreserved ectocervical tissues, whereas cryopreserved colorectal tissues were less easily infected than fresh tissues (hazard ratio 0.7 [95% confidence interval 0.4, 1.2]). Finally, we compared isolation of cells before and after cryopreservation. Cell recoveries were higher when cells were isolated after freezing and thawing (71% [59-84%]) than before (50% [38-62%]). Cellular function was similar to fresh tissue in both cases. Microbicide drug concentrations were lower in cryopreserved explants compared to fresh ones.ConclusionsCryopreservation of intact cervicovaginal and colorectal tissues with dimethylsulfoxide works well in a range of assays, while the utility of vitrification is more limited. Cell yields are higher from cryopreserved intact tissue pieces than from thawed cryopreserved single cell suspensions isolated before freezing, but T cell functions are similar.
| S-EPMC6066204 | biostudies-literature
Project description:Climate change has increased the incidence of coral bleaching events, resulting in the loss of ecosystem function and biodiversity on reefs around the world. As reef degradation accelerates, the need for innovative restoration tools has become acute. Despite past successes with ultra-low temperature storage of coral sperm to conserve genetic diversity, cryopreservation of larvae has remained elusive due to their large volume, membrane complexity, and sensitivity to chilling injury. Here we show for the first time that coral larvae can survive cryopreservation and resume swimming after warming. Vitrification in a 3.5?M cryoprotectant solution (10% v/v propylene glycol, 5% v/v dimethyl sulfoxide, and 1?M trehalose in phosphate buffered saline) followed by warming at a rate of approximately 4,500,000?°C/min with an infrared laser resulted in up to 43% survival of Fungia scutaria larvae on day 2 post-fertilization. Surviving larvae swam and continued to develop for at least 12?hours after laser-warming. This technology will enable biobanking of coral larvae to secure biodiversity, and, if managed in a high-throughput manner where millions of larvae in a species are frozen at one time, could become an invaluable research and conservation tool to help restore and diversify wild reef habitats.
| S-EPMC6200823 | biostudies-other
Project description:Osteoarthritis (OA) is a degenerative joint disease affecting cartilage and is the most common form of arthritis worldwide. One third of OA patients have severe synovitis and less than 10% have no evidence of synovitis. Moreover, synovitis is predictive for more severe disease progression. This offers a target for therapy but more research on the pathophysiological processes in the synovial tissue of these patients is needed. Functional studies performed with synovial tissue will be more approachable when this material, that becomes available by joint replacement surgery, can be stored for later use. We set out to determine the consequences of slow-freezing of human OA synovial tissue. Therefore, we validated a method that can be applied in every routine laboratory and performed a comparative study of five cryoprotective agent (CPA) solutions. To determine possible deleterious cryopreservation-thaw effects on viability, the synovial tissue architecture, metabolic activity, RNA quality, expression of cryopreservation associated stress genes, and expression of OA characteristic disease genes was studied. Furthermore, the biological activity of the cryopreserved tissue was determined by measuring cytokine secretion induced by the TLR ligands lipopolysaccharides and Pam3Cys. Compared to non frozen synovium, no difference in cell and tissue morphology could be identified in the conditions using the CS10, standard and CryoSFM CPA solution for cryopreservation. However, we observed significantly lower preservation of tissue morphology with the Biofreeze and CS2 media. The other viability assays showed trends in the same direction but were not sensitive enough to detect significant differences between conditions. In all assays tested a clearly lower viability was detected in the condition in which synovium was frozen without CPA solution. This detailed analysis showed that OA synovial tissue explants can be cryopreserved while maintaining the morphology, viability and phenotypical response after thawing, offering enhanced opportunities for human in vitro studies.
| S-EPMC5117761 | biostudies-literature
Project description:Chemotherapy and radiation therapy can cause gonadal dysfunction in women of reproductive age. Ovarian tissue cryopreservation is performed to restore fertility by allowing transplantation of the patient's frozen-thawed ovarian tissue or through future in vitro maturation and in vitro fertilization of frozen-thawed oocytes. Herein, we describe our initial experience with vaginal natural orifice transluminal endoscopic surgery for ovarian tissue preservation in a young woman with malignant tumor. A 23-year-old woman with anaplastic lymphoma kinase-positive malignant lymphoma was scheduled for hematopoietic stem cell transplantation after experiencing relapse following R-cyclophosphamide, doxorubicin, vincristine, and prednisolone therapy. Ovarian tissue cryopreservation was selected as only MII2 oocytes were collected. Vaginal natural orifice transluminal endoscopic surgery was performed to excise the left ovary. Ovarian tissues were frozen using the vitrification method. The operative time was 37 min, and blood loss was minimal. Pathological examination revealed no metastatic findings of malignant lymphoma and no thermal damage to the ovarian tissue due to bipolar disorder. The patient was discharged on the first day postoperatively, and her postoperative course was uneventful. The vaginal natural orifice transluminal endoscopic surgery technique can provide a safe and effective alternative to laparoscopy or laparotomy for the cryopreservation of ovarian tissue in young patients with cancer. We believe this method has potential application in sexually mature female cancer survivors.
| S-EPMC11003332 | biostudies-literature
Project description:PURPOSE: The aim of this experiment is to detect effects of varying levels of sucrose on vitrified ovarian tissues. METHODS: Ovarian tissues of mice were vitrified-thawed. Mice were randomly assigned to the fresh control group and experimental groups. According to different concentration of sucrose in vitrification solution, the experimental groups were randomly divided into Group I (0.2 M sucrose), Group II (0.4 M sucrose), Group III (0.8 M sucrose) and Group IV (1.6 M sucrose). Cytology was followed throughout the oophorectomy and transplantation period. Hormone levels and density of follicle were measured 1 month after transplantation. RESULTS: The number of days before the resumption of estrous cycles in control group was significantly smaller than those in all of experimental groups. The serum estradiol levels of mice and the follicular density of ovarian grafts in control group were significantly higher than those in all of experimental groups. In addition, the number of days before the resumption of estrous cycles in Group II and Group III were smaller than those in Group I and Group IV. The serum estradiol levels of mice and the follicular density of ovarian grafts in Group II and Group III were significantly higher than those in Group I and Group IV. However, no difference was observed in the number of days before the resumption of estrous cycles and the serum estradiol levels between Group II and Group III. A similar follicular density was also observed in Group II and Group III. CONCLUSION: Sucrose concentration of 0.4 M or 0.8 M in cryoprotective media is suitable for vitrifying mouse ovarian tissues.
| S-EPMC2654937 | biostudies-other
Project description:Physiological adjustments accompanying insect cold acclimation prior to cold stress have been relatively well explored. In contrast, recovery from cold stress received much less attention. Here we report on recovery of drosophilid fly larvae (Chymomyza costata) from three different levels of cold stress: supercooling to -10?°C, freezing at -30?°C, and cryopreservation at -196?°C. Analysis of larval CO2 production suggested that recovery from all three cold stresses requires access to additional energy reserves to support cold-injury repair processes. Metabolomic profiling (targeting 41 metabolites using mass spectrometry) and custom microarray analysis (targeting 1,124 candidate mRNA sequences) indicated that additional energy was needed to: clear by-products of anaerobic metabolism, deal with oxidative stress, re-fold partially denatured proteins, and remove damaged proteins, complexes and/or organelles. Metabolomic and transcriptomic recovery profiles were closely similar in supercooled and frozen larvae, most of which successfully repaired the cold injury and metamorphosed into adults. In contrast, the majority of cryopreseved larvae failed to proceed in ontogenesis, showed specific metabolic perturbations suggesting impaired mitochondrial function, and failed to up-regulate a set of 116 specific genes potentially linked to repair of cold injury.
| S-EPMC5849770 | biostudies-literature
Project description:The inability to preserve vascular organs beyond several hours contributes to the scarcity of organs for transplantation1,2. Standard hypothermic preservation at +4 °C (refs. 1,3) limits liver preservation to less than 12 h. Our group previously showed that supercooled ice-free storage at -6 °C can extend viable preservation of rat livers4,5 However, scaling supercooling preservation to human organs is intrinsically limited because of volume-dependent stochastic ice formation. Here, we describe an improved supercooling protocol that averts freezing of human livers by minimizing favorable sites of ice nucleation and homogeneous preconditioning with protective agents during machine perfusion. We show that human livers can be stored at -4 °C with supercooling followed by subnormothermic machine perfusion, effectively extending the ex vivo life of the organ by 27 h. We show that viability of livers before and after supercooling is unchanged, and that after supercooling livers can withstand the stress of simulated transplantation by ex vivo normothermic reperfusion with blood.
| S-EPMC6776681 | biostudies-literature
Project description:Ovarian tissue cryopreservation is considered as a useful option to preserve fertility for cancer patients. This study purposed to evaluate the change of telomere length and senescence markers during human ovarian tissue cryopreservation. Ovarian tissues were obtained from women who underwent benign ovarian surgery in the gynecology research unit of a university hospital with prior consent and IRB approval. DNA was extracted from the ovarian tissues using a DNeasy tissue kit and telomere lengths in the DNA samples were determined by real time PCR before and after cryopreservation. All tissues were stained with hematoxylin-eosin and subjected to immunohistochemistry and TUNEL assays. Other senescence markers, including p53, p16, p21, and phospho-pRb proteins, were evaluated using western blot analysis. Ovarian tissues were collected from ten patients and prepared for slow freezing with the same size of diameter 4 mm and 1 mm thickness. Mean age of patients was 26.7?±?6.2 years (range, 16-34 years), and ovarian tissues were cryopreserved and thawed 4 weeks after cryopreservation. The mean telomere length was significantly decreased after cryopreservation (9.57?±?1.47 bp vs. 8.34?±?1.83 bp, p?=?0.001). Western blot analysis revealed that p53, p16, and p21 proteins increased and phospho-pRb protein expression decreased after ovarian tissue cryopreservation. Ovarian tissue cryopreservation and transplantation is regarded as one of promising options for fertility preservation. However, clinicians and researchers should be aware of possible irreversible DNA changes such as shortening of telomere length and alterations of other senescence markers in human ovarian tissues.
| S-EPMC7838193 | biostudies-literature
Project description:BackgroundCryopreservation of ovarian tissue has the potential to preserve female germ cells of endangered mammals. In the present study, a freezing protocol successfully used for human tissue, was adapted for preserving ovarian tissue of domestic and non-domestic felids. Ovaries from non-domestic felid species were obtained from seven freshly euthanized and two recently deceased wild felids kept in different European Zoos. In addition, ovaries from domestic cats were obtained after ovariectomy from local veterinary clinics for methological adaptations.Ovarian cortex was dissected and uniform sized pieces of 2 mm diameter were obtained. Using a slow freezing protocol (-0.3°C per min) in 1.5 mol/L ethylene glycol, 0.1 mol/L sucrose, the pieces were cultured for up to 14 days both before and after cryopreservation. The integrity of primordial follicles was assessed by histology, and the impact of different protein sources (FCS or BSA) and Vitamin C was determined during two weeks of culture.Results and conclusionDuring culture the number of primordial follicles decreased within the ovarian pieces (p < 0.05). This effect was less pronounced when FCS was used as the protein source instead of BSA. Supplementation with Vitamin C had a detrimental effect on follicle survival. Since the procedure of cryopreservation had no effect on the follicle survival after one week of culture we conclude that the freezing protocol was suitable for felids. This is the first report of preserving a huge amount of follicles within ovarian tissue by slow freezing performed in several wild feline species.
| S-EPMC3614466 | biostudies-literature
Project description:Primordial germ cells (PGCs) are the precursors of germline cells that generate sperm and ova in adults. Thus, they are promising tools for gene editing and genetic preservation, especially in avian species. In this study, we established stable male and female PGC lines from 6Hungarian indigenous chicken breeds with derivation rates ranging from 37.5 to 50 percent. We characterized the PGCs for expression of the germ cell-specific markers during prolonged culture in vitro. An in vivo colonization test was performed on PGCs from four Hungarian chicken breeds and the colonization rates were between 76 and 100%. Cryopreserved PGCs of the donor breed (Partridge color Hungarian) were injected into Black Transylvanian Naked Neck host embryos to form chimeric progeny that, after backcrossing, would permit reconstitution of the donor breed. For 24 presumptive chimeras 13 were male and 11 were female. In the course of backcrossing, 340 chicks were hatched and 17 of them (5%) were pure Partridge colored. Based on the backcrossing 1 hen and 3 roosters of the 24 presumptive chimeras (16.6%) have proven to be germline chimeras. Therefore, it was proven that the original breed can be recovered from primordial germ cells which are stored in the gene bank. To our knowledge, our study is a first that applied feeder free culturing conditions for both male and female cell lines successfully and used multiple indigenous chicken breeds to create a gene bank representing a region (Carpathian Basin).
| S-EPMC8271167 | biostudies-literature