Cell and Tissue Banking最新文献

The purpose of this study was to compare two methods for harvesting human dental pulp stem cells (hDPSCs) and investigate their impact on the DPSC viability and differentiation capacity. Healthy premolar teeth were collected from children aged 9 - 17 years old undergoing orthodontic treatment with two or four premolar extractions. The included premolars were randomly allocated to the apical group (removal of the dental pulp through the apical foramen after tooth extraction) or the coronal group (isolating the pulp tissue through the crown prior to tooth extraction). A total of 148 healthy premolar teeth from 46 patients were extracted, including 74 pulp tissues for the apical group and 74 for the coronal group. Due to bacterial and fungal infections, 37 (18 apical and 19 coronal) DPSC lines were obtained. Flow cytometric analysis and MTT assays showed no statistical differences in cell viability between the two methods. Odontogenic differentiation, assessed through immunocytochemical staining and Alizarin Red staining, also showed no visual or statistical differences between the apical and coronal approaches. In conclusion, the biological characteristics of hDPSCs harvested through the crown were equal to apically prelevated hDPSCs.

Cryobags play a critical role in freezing, storing, and transporting cellular therapy products but are prone to fractures, which can disrupt patient outcomes and facility workflows. This study evaluated the incidence, risk factors, and impact of cryobag fractures in a large inventory of cellular therapy products in Brazil. A retrospective cohort study included 4514 cryobags from 2262 peripheral blood stem cell collections processed between 2015 and 2024 at a single center supporting nine transplant facilities. Cryobags were frozen at - 80 °C and stored in nitrogen tanks. Fractures and leaks were identified through routine visual inspections. Among the cryobags, 15 (0.3%) fractured, with 12 detected at processing facility and 3 after release. The fracture rate was 0.37 per 100 bag-years, with a cumulative incidence of 1% at 3.62 years. Of these, 8 were discarded, and 7 were salvaged and infused into six patients. Two salvaged cryobags underwent bedside recovery, while five were recovered aseptically in the processing facility. Positive bacterial cultures were commonly found in salvaged products. In multivariate analysis, a higher total nucleated cell count per cryobag remained an independent risk factor for fracture (OR = 1.005; 95% CI: 1.0002-1.0099; p = 0.046). Following implementation of quality improvement initiatives based on root cause analysis, no further fractures were observed. These findings highlight the importance of monitoring cell concentration and adjusting cryopreservation protocols to mitigate risks. Adding overwraps may provide additional protection for cryobags at higher risk, reducing the likelihood of microbial contamination and improving the safety and reliability of cellular therapy products.

The purpose of this study was to investigate the role of Calcium-Sensing Receptor in the chondrogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and to provide a new target for cartilage defect repair. BMSCs were cultured in vitro, and cultured in the complete culture medium with gradient concentration of calcium sensitive receptor inhibitor and activator, and the optimum dose was selected by CCK-8 experiment. The experiment was divided into four groups. After 7, 14 and 21 days of intervention, the intracellular calcium concentration was detected by laser confocal microscope, the differentiation of cartilage was detected by toluidine blue staining, and the expression of cartilage marker proteins (Col- II, Agg and Sox9) was detected by immunocytochemical staining and Western Blot. The CCK-8 assay results showed that the optimal concentrations of Gd and NPS were 300 μM and 10 μM, respectively. After 7, 14, and 21 days of culture, intracellular calcium fluorescence decreased, with notably higher cartilage differentiation in the NPS inhibitor group. Col-II, Agg and Sox9 chondrocyte marker proteins increased with culture time in all groups, with significantly higher levels in the inhibitor group compared to others, followed by the cartilage induction solution group, and then the activator group. Inhibition of calcium sensitive receptors can promote chondrogenic differentiation of rat BMSCs by regulating Sox9, affecting Col- II and Agg.

The transmissibility rate of the SARS-CoV-2 virus, which causes COVID-19, in organ and tissue donation is not known. Considering all issues related to the risk of SARS-CoV-2 transmission and the risk of tissue shortages due to donor exclusion, this in vitro study evaluated the detection of SARS-CoV-2 RNA in bone tissue models after bone processing for tissue bank. Bone processing has activity against SARS-CoV-2 and not RNA was detected in the end of the process. The use of bone tissue can be considered in case of shortage.

To compare the differences in biological and biomechanical characteristics of cadaver tendon treated by deep freezing, 95wt.% ethanol, tributyl phosphate (TBP), and sodium dodecyl sulfate (SDS) respectively. A total of 50 fresh tendon specimens were randomly divided into five groups, and four groups were treated with deep freezing (- 80 °C), 95wt.% ethanol, 1% TBP, or 1% SDS, respectively. Hematoxylin-eosin (HE), toluidine blue (TB) and 4',6-diamidino-2-phenylindole (DAPI) staining, and transmission/scanning electron microscope observations were performed. Then, cyclic creep and tensile tests were conducted to investigate the biomechanical properties. The content of residual DNA was tested. The HE, TB and DAPI staining showed a significant decrease in tendon cells following treatments, compared with fresh specimens. SDS, TBP, alcohol can almost completely decellularize the tendon, and deep-frozen group remained a few number of tendon cells. The residual DNA content was significantly lowered, with an average percentage of 50.97%, 79.16%, 88.91% and 72.56%, for groups of deep freezing, alcohol, TBP and SDS, respectively. The arrangement of collagen fibers was significantly disrupted, and the gap between fibers was widened, following treatments by alcohol, TBP and SDS. However, the biomechanical properties were generally similar among the five groups, with significantly lowered cyclic creep for ethanol group and lowered Young's modulus for SDS group exclusively. The four treatments can effectively reduce the number of residual cells and DNA content. Among them, cryogenic treatment has almost no damage to tendon histology and biomechanics, while ethanol, SDS and TBP decellularization methods cause different degrees of damage.

To qualify for bone donation many criteria must be met. For procurement, two time intervals-a postmortem interval (PMI) of 6 h (hours) until cooling of the deceased and a maximum procurement time of tissues within 24/36 h postmortem are mandatory. Expanding the procurement to donors with a longer PMI would be helpful e.g. to have more time to contact relatives for consent for donation. A loss of biomechanical quality of bone tissues should not be expected in the PMI until procurement, but the question of microbiological contamination remains. Therefore, samples from the iliac crest were procured under standard procurement conditions between 48 and 54 h (n = 14, group 1) and 72-78 h (n = 7, group 2) postmortem (pm). Samples were immediately frozen after procurement at < - 18 °C and sent to the German Institute for Tissue and Cell Replacement (DIZG) for mechanical preparation. Additionally, chloroform treatment was performed at the tissue bank of the Charité Berlin. After each step the samples were refrozen and sent to a contracted microbiological lab for bioburden testing according to the European Pharmacopoeia. Samples were tested after procurement, preparation, and chloroform treatment respectively. As acceptance criterion a maximum amount of 9 × 10⁴ colony forming units per gram (CFU/g) tissue was defined. All samples were within the defined limit with a maximum value of 2.5 × 10² CFU/g. These values are in the range of bone tissues procured under non-strict-aseptic procurement conditions within 24 or 36 h and are within the capacity of the peracetic acid sterilization process used by several tissue banks.

Despite significant progress in cartilage regeneration therapeutics, several challenges remain in achieving optimal results under in vivo conditions. The present research evaluated the chondrogenic potential of poly(glycerol sebacate) copolymer nanofibrous scaffold (PGS NF) loaded with growth differentiation factor-5 incorporated sugar glass nanoparticles (SGnP-GDF5), in combination with allogenic bone marrow-derived mesenchymal stem cells (BM-MSC) in a rabbit model. A full-thickness chondral defect of 4 mm diameter was created in the trochlear facet of the left femur of rabbits using a Brad point drill bit. PGS NF was used in group B, BM-MSC laden PGS NF in group C, SGnP-GDF5 loaded PGS NF in group D, and BM-MSC laden SGnP-GDF5 loaded PGS NF in group E. Five animals from each group were sacrificed on days 60 and 90 post-treatment. The samples were assessed based on gross morphology, histopathology, scanning electron microscopy (SEM), and micro-computed tomography (micro-CT) analysis to evaluate regeneration. The SGnP-GDF5 PGS NF group and the BM-MSC laden SGnP-GDF5 PGS NF group exhibited superior cartilage regeneration, closely resembling hyaline cartilage. Histopathological evaluation revealed a columnar pattern of chondrocytes, along with an optimal concentration of proteoglycans and collagen in the extracellular matrix of the newly formed cartilage, indicating robust regeneration in both groups. Furthermore, the SEM and micro-CT analysis findings highlighted the exceptional quality of the repaired tissue in these groups. The release of GDF5 from SGnP and the expedient microenvironment provided by the NF scaffold augmented chondrogenic differentiation, resulting in superior cartilage tissue regeneration.

Cardiovascular diseases have become the leading cause of death worldwide in recent years. Therefore, blood vessel transplantation is an alternative to reduce complications and deaths resulting from cardiovascular diseases. Therefore, this study aims to analyze specific family refusal to donate blood vessels for transplantation in an Organ Procurement Organization. This is a cross-sectional study carried out in accordance with Family Authorization for Organ and Tissue Donation Terms of patients diagnosed with brain death by an Organ Procurement Organization in the state of São Paulo, Brazil. The variables analyzed were sex, age, cause of death, hospital institution and refusal to donate blood vessels. Descriptive and inferential data analysis was performed. Of 2,447 donors, 37.71% refused to donate vessels, influenced by sex, age group of 41-59 years, traumatic brain injury as the cause of death and public hospitals. Refusal varied with age, as it decreased up to 11 years and increased after 60 years. From 2001 to 2020, refusal decreased by 57%, 64%, 68% and 69% for the age groups of 12-19 years, 20-40 years, 41-59 years and 60 years and over, respectively. Donors from public institutions were 18% less likely to refuse donation. Refusal to donate blood vessels for transplantation was associated with all variables analyzed. More research is needed to support strategies that reduce the rejection of these tissues.

Mesenchymal Stem cells (MSCs) have a wide range of therapeutic applications due to their self-renewal and multi-lineage differentiation ability; large-scale production of MSCs is possible only with a highly efficient medium, which facilitates increased proliferation of MSCs within a short period. Recently, Human Platelet Lysate (hPL) has emerged as a promising substitute for fetal bovine serum (FBS) for cell expansion. The goal of this study is to optimize a stable gel formulation for the 3D expansion of MSCs using hPL as a matrix material for the improved proliferation of Human Umbilical Cord Tissue derived MSCs (hUCT-MSCs) in comparison to FBS and hPL-supplemented media in 2D culture. To assess the potential benefits of the hPL gel system, in promoting cell proliferation capacity, hUCT-MSCs were cultured on hPL gel coated-dish supplemented with hPL CM, and in FBS CM. Among the varying concentrations, 20% hPL gel was optimized to have more functional stability and shorter gelation time. SEM analysis and gel degradation study at different concentrations revealed the structural integrity and morphology of the gel. Microscopic images and histological staining by H&E were conducted to understand the multi-layered proliferation of hUCT-MSCs in hPL Gel. Flow cytometry analysis reported the expression of positive markers for human umbilical cord MSCs, namely CD 90⁺ and CD 105⁺, in hPL Gel and hPL Complete Medium (CM) similar to that in FBS. The CCK8 Assay carried out for each culture system, generated OD values respective to cell viability and proliferation. OD values of 1.65 nm, 1.27 nm, and 0.92 nm on average were observed for hPL Gel, hPL CM, and FBS control, respectively. Cells in hPL gel showed a 50% higher proliferation rate of viable cells compared to other culture media. AO/EtBr staining with FBS CM, hPL CM, and hPL gel revealed an increase in viable cells and a decrease in early apoptotic and necrotic cells in hPL Gel. In conclusion, the results of this study highlight the potential of hPL-based gels as superior matrices for multi-layered and enhanced proliferation of hUCT-MSCs.

Tibial reconstruction for major bone loss is a challenging surgical situation. For more than 30 years, massive bone allografts entered our therapeutic arsenal as a major tool to reconstruct large bone defect. Despite allograft is valued for its convenience, it is still burdened by a high complication rate. This retrospective monocentric study focuses on the clinical outcomes of massive tibial allografts and their outcome's prediction. Between 1987 and 2022, the files of 148 massive tibial allografts were retrospectively reviewed (registration number B403201523492). Survival curves were calculated based on the allograft success or failure. Survival curves without revision surgery were calculated following the same design. Finally, multiple logistic regression models were set up to point out variables that influence the allograft survival. After 30 years, 87.2% of the patient retained limb function. However, 55% of the allograft failed and had to be removed (mean survival time is 20.06 ± 2.07 years (CI 16.0-24.1)). The estimate mean survival time is 10.26 ± 1.60 years (CI 7.1-13.4) with less than 20% survival for the allografts without revision surgery after 30 years. Tumor and septic indications worsen the prognosis as well as the number of revision surgeries and the osteochondral allograft type. In contrast, PSI use or traumatological indications improve the allograft survival. Despite remaining an excellent surgical reconstructive option, massive tibial allografts show a high revision surgery rate. Thanks to our multiple logistic regression models, we can start to predict and improve the final outcomes of these complex allograft reconstruction.