Search Results (29)

The advancement of biomedical research depends on efficient data sharing, integration, and annotation to ensure reproducibility, accessibility, and cross-disciplinary collaboration. International collaborative research is crucial for advancing biomedical science and innovation but often faces significant barriers, such as data sharing limitations, inefficient sample management, and scalability challenges. Existing infrastructures for biosample and data repositories face challenges limiting large-scale research efforts. This study presents a novel platform designed to address these issues, enabling researchers to conduct high-quality research more efficiently and at reduced costs. The platform employs a modular, distributed architecture that ensures high availability, redundancy, and interoperability among diverse stakeholders, as well as integrates advanced features, including secure access management, comprehensive query functionalities, real-time availability reporting, and robust data mining capabilities. In addition, this platform supports dynamic, multi-criteria searches tailored to disease-specific patient profiles and biosample-related data across pre-analytical, post-analytical, and cryo-storage processes. By evaluating the platform’s modular architecture and pilot testing outcomes, this study demonstrates its potential to enhance interdisciplinary collaboration, streamline research workflows, and foster transformative advancements in biomedical research. The key is the innovation of a real-time dynamic e-consent (DRT e-consent) system, which allows donors to update their consent status in real time, ensuring compliance with ethical and regulatory frameworks such as GDPR and HIPAA. The system also supports multi-modal data integration, including genomic sequences, electronic health records (EHRs), and imaging data, enabling researchers to perform complex queries and generate comprehensive insights. Full article

Cryopreservation is a widely used method for the long-term preservation of reproductive or somatic cells. It is known that this storage method may negatively affect cell viability, proliferation, differentiation, etc. However, there is a lack of information about whether cryostorage can alter the content of intracellular minerals. Therefore, we focused this study on the analysis of the mineral composition of living cells before and after long-term cold storage. Briefly, three different primary cell lines were established from rabbits as follows: endothelial progenitor cells from peripheral blood (EPCs), endothelial progenitor cells from bone marrow (BEPCs), and mesenchymal stem cells from adipose tissue (AT-MSCs), which were cultured until passage 3 prior to cryopreservation in liquid nitrogen. Samples from freshly cultured and frozen–thawed cells were mineralized and analyzed using inductively coupled plasma-optical emission spectroscopy (ICP-OES) for the content of minerals (macro: Ca, Na, K, and Mg, and micro: Zn, Fe, Cu, Al, Co, Mn, Sr, and Ni). After cryopreservation, we found significantly decreased content of K in frozen–thawed EPCs (p < 0.01) and BEPCs (p < 0.0001) and Ca in AT-MSCs (p < 0.05), while Na was increased in frozen–thawed BEPCs (p < 0.05). Concentrations of Fe and Al were reduced significantly in frozen–thawed EPCs (both p < 0.0001) and AT-MSCs (p < 0.001 and p < 0.0001, respectively). On the contrary, Fe and Al were elevated in frozen–thawed BEPCs (p < 0.0001 and p < 0.01, respectively) together with Ni (p < 0.0001). In addition, decreased Zn (p < 0.05) was observed in cryopreserved AT-MSCs. In conclusion, the ICP-OES technique might be used to analyze the basic elemental composition of animal cells in fresh or frozen–thawed conditions. Nevertheless, additional studies are needed to reveal the possible impact of cryopreservation on cell fate by changing the content of intracellular minerals. Full article

Background/Objectives: Flow cytometry is a convenient tool in immunophenotyping for monitoring the status of immunological conditions and diseases. The aim of this study was to investigate the effect of isolation and cryopreservation by flow cytometric analysis on subpopulations of CD4⁺ T helper (Th), T regulatory (Treg), CD8⁺ T cytotoxic (Tc), CD56⁺ NK, CD19⁺ B and monocytes. Freshly isolated and cryopreserved peripheral blood mononuclear cells (PBMCs) were compared to fresh whole blood. Methods: Peripheral blood was collected from healthy donors and prepared for flow cytometric analysis using the same panels of antibodies throughout the study. Results: Comparisons between fresh (F)- and cryopreserved (C)-PBMCs showed no major differences in percentages of CD4⁺, Th1, Th2 and CD4⁺CD25⁺CD127^low Treg cells. No differences in percentage of CD8⁺ or subpopulations of naive/stem, central or effector memory cells were observed between F- and C-PBMCs. The percentage of CD56⁺ NK cells, CD19⁺ B cells or classical and nonclassical monocytes did not differ between F-and C-PBMCs either. On the contrary, whole blood had lower percentages of Th and NK cells but higher percentages of Th1, Th17, Th1Th17, Tregs, Tc and B cells compared to C-PBMCs, while it had a higher proportion of Tc compared to F-PBMCs. Conclusions: Flow cytometric immunophenotyping minimally differs between freshly isolated and cryopreserved PBMCs. This implies the possibility of cryostorage of cohorts for later analysis. Importantly, care must be taken when comparing results from whole blood with isolated and cryopreserved PBMCs. Collectively, these results can contribute to the standardization of flow cytometric protocols in both clinical and research settings. Full article

The conservation of genetic resources in pig breeds, notably the Iberian pig, is crucial for genetic improvement and sustainable production. Prolonged storage in liquid nitrogen (LN₂) is recognized for preserving genetic diversity, but potential adverse effects on seminal quality remain debated. This study aims to assess the impact of ten years of storage at different LN₂ levels and to optimize thawing protocols for Iberian pig sperm. Sperm samples from 53 boars were cryopreserved and stored at varying LN₂ levels and, a decade later, the samples were thawed at 37 °C for 20 s or at 70 °C for 8 s. Sperm motility, membrane integrity, acrosome status, and DNA fragmentation were evaluated in year 0 and year 10. Overall, no significant differences were observed in post-thaw sperm quality between storage levels in year 0 or year 10. But thawing at 70 °C 8 s showed significant improvements, particularly in samples that were always stored in LN₂, in all analyzed parameters except fragmentation, which was not affected by cryostorage. This study suggests that the long-term preservation of Iberian pig sperm does not affect quality over time, regardless of whether the samples were fully submerged in LN₂. Furthermore, it is determined that thawing at 70 °C for 8 s maximizes post-thaw sperm quality, especially in those samples stored constantly submerged in LN₂. Full article

Green ash (Fraxinus pennsylvanica) and white ash (F. americana) populations are currently experiencing major declines across their native ranges in North America due to infestation by the exotic insect pest emerald ash borer (Agrilus planipennis). The development of a reliable method for the long-term storage of green and white ash germplasm in the form of embryogenic cultures using cryopreservation would be a considerable aid to ash conservation efforts. We compared recovery percentages of cryopreserved green and white ash embryogenic cultures using vitrification versus slow cooling methods. Three Plant Vitrification Solution 2 (PVS2) exposure durations (40, 60, and 80 min) for vitrification and three DMSO concentrations (5%, 10%, and 15%) for slow cooling were tested for their effects on the percentage of cultures that regrew following cryostorage. Vitrification resulted in a higher overall culture recovery percentage (91%) compared to cultures that were cryostored using the slow cooling approach (39%), and a more rapid initiation of regrowth (5 days versus 2–3 weeks) resulted. Recovery from cryostorage by cultures using the slow cooling approach varied significantly (p < 0.05) between experiments and with genotype (p < 0.05). The recovery of vitrified tissue from cryostorage did not vary with genotype, species, or PVS2 exposure duration (p > 0.05). The vitrification cryopreservation protocol provides a reliable and versatile alternative to the traditional slow cooling method, strengthening our ability to preserve valuable ash germplasm for conservation and restoration. Full article

Recent advances in anticancer treatment have significantly improved the survival rate of young females; unfortunately, in about one third of cancer survivors the risk of ovarian insufficiency and infertility is still quite relevant. As the possibility of becoming a mother after recovery from a juvenile cancer is an important part of the quality of life, several procedures to preserve fertility have been developed: ovarian surgical transposition, induction of ovarian quiescence by gonadotropin-releasing hormone agonists (GnRH-a) treatment, and oocyte and/or ovarian cortical tissue cryopreservation. Ovarian tissue cryostorage and allografting is a valuable technique that applies even to prepubertal girls; however, some patients cannot benefit from it due to the high risk of reintroducing cancer cells during allograft in cases of ovary-metastasizing neoplasias, such as leukemias or NH lymphomas. Innovative techniques are now under investigation, as in the construction of an artificial ovary made of isolated follicles inserted into an artificial matrix scaffold, and the use of stem cells, including ovarian stem cells (OSCs), to obtain neo-folliculogenesis and the development of fertilizable oocytes from the exhausted ovarian tissue. This review synthesizes and discusses these innovative techniques, which potentially represent interesting strategies in oncofertility programs and a new hope for young female cancer survivors. Full article

Vegetatively propagating aquatic angiosperms, the Lemnaceae family (duckweeds) represents valuable genetic resources for circular bioeconomics and other sustainable applications. Due to extremely fast growth and laborious cultivation of in vitro collections, duckweeds are an urgent subject for cryopreservation. We developed a robust and fast DMSO-free protocol for duckweed cryopreservation by vitrification. A single-use device was designed for sampling of duckweed fronds from donor culture, further spin-drying, and subsequent transferring to cryo-tubes with plant vitrification solution 3 (PVS3). Following cultivation in darkness and applying elevated temperatures during early regrowth stage, a specific pulsed illumination instead of a diurnal regime enabled successful regrowth after the cryopreservation of 21 accessions of Spirodela, Landoltia, Lemna, and Wolffia genera, including interspecific hybrids, auto- and allopolyploids. Genome size measurements revealed no quantitative genomic changes potentially caused by cryopreservation. The expression of CBF/DREB1 genes, considered as key factors in the development of freezing tolerance, was studied prior to cooling but was not linked with duckweed regrowth after rewarming. Despite preserving chlorophyll fluorescence after rewarming, the rewarmed fronds demonstrated nearly zero photosynthetic activity, which did not recover. The novel protocol provides the basis for future routine application of cryostorage to duckweed germplasm collections, saving labor for in vitro cultivation and maintaining characterized reference and mutant samples. Full article

Vitrification and ultrarapid laser warming are crucial for the cryopreservation of animal embryos, oocytes, and other cells of medicinal, genetic, and agricultural value. In the present study, we focused on alignment and bonding techniques for a special cryojig that combines a jig tool and jig holder into one piece. This novel cryojig was used to obtain a high laser accuracy of 95% and a successful rewarming rate of 62%. The experimental results indicated that our refined device improved laser accuracy in the warming process after long-term cryo-storage through vitrification. We anticipate that our findings will lead to cryobanking applications that use vitrification and laser nanowarming to preserve cells and tissues from a wide range of species. Full article

The conservation of genetic resources in cryocollections requires reliable protocols for the cryopreservation and the regeneration of the preserved material. With Ulmus glabra, the regeneration of thawed buds by in vitro organogenesis has suffered from low shoot growth and high contamination rates. The dehydration of the buds before cryopreservation improved the shoot growth rate and ameliorated the contamination rate of in vitro cultures initiated from thawed buds, although the degree of success varied depending on the donor tree. Full article

The different pre- and post-treatments are critical in cryopreservation procedures and affect the shoot tip regrowth after freezing. In the present study, the long-term storage of four citrus cultivars [Bodrum Mandarin (Citrus deliciosa Ten.); Klin Mandarin (Citrus nobilis Lauriro); White grapefruit and Red grapefruit (Citrus paradisi L.)] were carried out by droplet vitrification methods, and the critical points for effective cryopreservation of these species were determined. In this study, we investigated the effect of explant size, cold hardening treatments, sucrose concentrations, and media combinations on shoot regrowth after cryopreservation. The highest shoot tip regrowth, ranging from 13.3 to 33.3%, was achieved when they were obtained from 0.3 to 0.7 mm explants excised from cold hardened seedlings at 4 °C for three days that were then precultured in a medium containing 0.25 M of sucrose and treated with PVS2 at 0 °C for 45 min. In addition, it has been determined that a regeneration medium containing boric acid (H₃BO₃) or ferric ethylenediaminetetraacetate (FeEDDHA) increased the regeneration up to 33.3% after cryopreservation. Full article

Cryopreservation is the final step of stem cell production before the cryostorage of the product. Conventional methods of adding cryoprotecting agents (CPA) into the cells can be manual or automated with robotic arms. However, challenging issues with these methods at industrial-scale production are the insufficient mixing of cells and CPA, leading to damage of cells, discontinuous feeding, the batch-to-batch difference in products, and, occasionally, cross-contamination. Therefore, the current study proposes an alternative way to overcome the abovementioned challenges; a highly efficient micromixer for low-cost, continuous, labour-free, and automated mixing of stem cells with CPA solutions. Our results show that our micromixer provides a more homogenous mixing of cells and CPA compared to the manual mixing method, while the cell properties, including surface markers, differentiation potential, proliferation, morphology, and therapeutic potential, are well preserved. Full article

Mesenchymal stem cells (MSCs) manifest vast opportunities for clinical use due both to their ability for self-renewal and for effecting paracrine therapeutic benefits. At the same time, difficulties with non-recurrent generation of large numbers of cells due to the necessity for long-term MSC expansion ex vivo, or the requirement for repeated sampling of biological material from a patient significantly limits the current use of MSCs in clinical practice. One solution to these problems entails the creation of a biobank using cell cryopreservation technology. This review is aimed at analyzing and classifying literature data related to the development of protocols for the cryopreservation of various types of MSCs and tissue-engineered structures. The materials in the review show that the existing techniques and protocols for MSC cryopreservation are very diverse, which significantly complicates standardization of the entire process. Here, the selection of cryoprotectors and of cryoprotective media shows the greatest variability. Currently, it is the cryopreservation of cell suspensions that has been studied most extensively, whereas there are very few studies in the literature on the freezing of intact tissues or of tissue-engineered structures. However, even now it is possible to develop general recommendations to optimize the cryopreservation process, making it less traumatic for cells. Full article

Nearly four million yearly deaths can be attributed to respiratory diseases, prompting a huge worldwide health emergency. Additionally, the COVID-19 pandemic’s death toll has surpassed six million, significantly increasing respiratory disease morbidity and mortality rates. Despite recent advances, it is still challenging for many drugs to be homogeneously distributed throughout the lungs, and specifically to reach the lower respiratory tract with an accurate sustained dose and minimal systemic side effects. Engineered nanocarriers can provide increased therapeutic efficacy while lessening potential biochemical adverse reactions. Poly(lactic-co-glycolic acid) (PLGA), a biodegradable polymer, has attracted significant interest as an inhalable drug delivery system. However, the influence of the nanocarrier surface charge and its intratracheal instillation has not been addressed so far. In this study, we fabricated red fluorescent PLGA nanocapsules (NCs)—Cy5/PLGA—with either positive (Cy5/PLGA+) or negative surface charge (Cy5/PLGA-). We report here on their excellent colloidal stability in culture and biological media, and after cryo-storage. Their lack of cytotoxicity in two relevant lung cell types, even for concentrations as high as 10 mg/mL, is also reported. More importantly, differences in the NCs’ cell uptake rates and internalization capacity were identified. The uptake of the anionic system was faster and in much higher amounts—10-fold and 2.5-fold in macrophages and epithelial alveolar cells, respectively. The in vivo study demonstrated that anionic PLGA NCs were retained in all lung lobules after 1 h of being intratracheally instilled, and were found to accumulate in lung macrophages after 24 h, making those nanocarriers especially suitable as a pulmonary immunomodulatory delivery system with a marked translational character. Full article

Horticultural crops comprise various economic species extending from fruits, nuts, vegetables, spices and condiments, ornamentals, aromatic, and medicinal plants. Ornamental and fruit plants are produced mainly for their nutritional and aesthetic values, respectively. Unfortunately, many tropical and subtropical species are in danger of extinction because of climate change and (a)biotic stresses. It is imperative to preserve the germplasms of these species for the present and future genetic improvement programs. Cryopreservation, i.e., maintenance of tissues at the ultralow temperature of liquid nitrogen, is a promising long-term preservation technique, alternative to seed or in vitro banks, which can be applied for both vegetatively and generatively (through seeds) propagated crops, including those with recalcitrant seeds. It is a technology of choice not only for the preservation of plant biodiversity but also for virus elimination in the proficient administration of large-scale micropropagation. The main advantages of cryopreservation are the lowering of in vitro culture expenditures, needed space, contamination risk, and operator errors. However, tropical species are temperature delicate and one of the foremost challenging issues is preconditioning treatments that stimulate physiological reactions to sufficiently enhance tolerance to dehydration and cryogenic procedures. In recent years, several cryopreservation methods based on encapsulation-vitrification, droplet-vitrification, the use of aluminum cryo-plates, and cryo-mesh have been established. Combined cryo-techniques, gene/DNA conservation, as well as studies on perceiving bio-molecular events and exploring the multistage process from the beginning to end of cryopreservation are receiving more emphasis. The development of cryobiomics delivers a conceptual framework to assess the significance of cell signaling mechanisms on cellular functions, the influence of cryoinjury factors on sample viability, and the implications for genetic stability following cryo-storage. The aim of this mini-review article is to provide a succinct synthesis of the developed cryogenic procedures and their use for the storage and exchange of genetic resources of tropical and subtropical horticultural crops, particularly fruit crops and ornamental plants under the threat of extinction. Full article

Holm oak populations are severely affected by oak decline syndrome, and reliable methods of conserving the plant material are required. A vitrification-based cryopreservation method was used for the first time for the long-term conservation of holm oak embryogenic cultures. Successful cryopreservation was achieved after determining the best developmental stage of the somatic embryos used and the optimal incubation period in plant vitrification solution 2 (PVS2). Embryos were recovered from individual nodular embryogenic structures (NES) derived from four embryogenic lines after preculture on a medium containing 0.3 M sucrose, incubation in PVS2 vitrification solution for 15 min at 25 °C and direct immersion in liquid nitrogen (LN). Embryo recovery rates of 16.7–63.3% were obtained after cryostorage for four years in LN. In addition to the embryo developmental stage and the PVS2 treatment time, the genotype can also significantly affect embryo recovery after LN storage. There were no significant differences in plant regeneration or polyploid stability between somatic embryos and plants derived from control embryos (not cryopreserved) and cryopreserved embryos. The findings indicate that embryo proliferation, plant conversion and polyploid stability are maintained in material recovered from the vitrification solution and subsequently cryopreserved. Full article