Individual Donors Research Articles

Biopsy-derived oral keratinocytes – A model to potentially test for oral mucosa radiation sensitivity

PurposeTo establish stable in vitro growth of keratinocytes from very small biopsy specimens and successfully apply new test systems to determine their radiosensitivity. Materials and MethodsOral mucosa biopsies (diameter: 1.7 mm) from 15 subjects were immobilized with custom-made cups onto culture plates. Outgrowing cells were tested for cytokeratin 5/14 and Ki67, expanded, radiated at different doses, and seeded onto circumscribed areas before being allowed to spread centrifugally. In this newly developed spreading assay, cell-covered areas were measured by image analysis. For statistical analysis, a linear mixed regression model was used; additionally, results were correlated to the radiation dose applied. Colony forming efficiency (CFE) was used to validate the results. DNA damage repair was analysed by gammaH2AX and 53BP1 foci quantification using immunofluorescence microscopy 24 h and 96 h after irradiation. ResultsStable keratinocyte growth continued for up to 7 weeks in 14 biopsies. Cells spread reliably from an initial 16.6 mm2 up to a median of 119.2 mm2 (range: 54.4–290). Radiated cells spread to only 100.7 mm2 (2 Gy; range: 55.3–266.7); 73.2 mm2 (4 Gy; 15–240.4); 47 mm2 (6 Gy; 2–111.9), and 22.7 mm2 (8 Gy; 0–80). Similarly, CFE decreased from 0.223 (0 Gy) to 0.0028 (8 Gy). Using an individual donor as a random factor, cell spread correlated with CFE, where radiation dose was the main driver (decrease by 0.50, adjusted for area). Upon irradiation with 6 Gy, radiation-induced DNA damage was increased after 24 h in all samples, and even after 96 h in 5 out of 7 samples, as detected by a higher number of gammaH2AX/53BP1 foci in irradiated cells (mean 3.7 for 24 h; mean 0.6 for 96 h). ConclusionIn vitro propagation of keratinocytes derived from a small biopsy is feasible. Radiation impairs cellular migration and proliferation, and the newly described spreading assay allows ranking for cellular radioresistance. The keratinocyte model also supports classical functional assays such as clonogenic survival and DNA double strand repair. The clinical relevance awaits upcoming investigations.

Turnover intention and job tenure of US fundraisers

AbstractFundraisers secure financial resources that organizations need to achieve their missions. Raising money, particularly large gifts, can follow years of relationship building with individual donors. When fundraisers leave these efforts can be set back substantially, making fundraiser turnover particularly worrisome and worthy of exploration. This analysis addressed the issue with US survey data (n = 1663) and examinination of three research questions. What are the job tenure and intent to leave of fundraisers? How is fundraiser job tenure affected by intent to leave? What relationships do job tenure and intent to leave have with fundraisers' individual demographics, position attributes, and organizational characteristics? We found that the study participants had current mean job tenures of 3.6 years (median = 2 years) and mean tenures across their fundraising jobs of 3.9 years (median = 3 years). Twenty percent intended to leave their organization and 7% intended to leave fundraising within the next year. Of the tested variables, salary consistently had the largest effects and was the most significant. Older and more experienced fundraisers had longer tenures. The study provides nuanced information about fundraisers' job‐related behaviors, includes careful attention to theory and related research, and presents specific ideas for organizational interventions for increasing fundraiser tenure.

Gating Orbital Memory with an Atomic Donor.

Orbital memory is defined by two stable valencies that can be electrically switched and read out. To explore the influence of an electric field on orbital memory, we studied the distance-dependent influence of an atomic Cu donor on the state favorability of an individual Co atom on black phosphorus. Using low temperature scanning tunneling microscopy and spectroscopy, we characterized the electronic properties of individual Cu donors, corroborating this behavior with abinitio calculations based on density functional theory. We studied the influence of an individual donor on the charging energy and stochastic behavior of an individual Co atom. We found a strong impact on the state favorability in the stochastic limit. These findings provide quantitative information about the influence of local electric fields on atomic orbital memory.

Variations in the Composition of Human Milk Oligosaccharides Correlates with Effects on Both the Intestinal Epithelial Barrier and Host Inflammation: A Pilot Study.

Background: Human milk oligosaccharides are complex, non-digestible carbohydrates that directly interact with intestinal epithelial cells to alter barrier function and host inflammation. Oligosaccharide composition varies widely between individual mothers, but it is unclear if this inter-individual variation has any impact on intestinal epithelial barrier function and gut inflammation. Methods: Human milk oligosaccharides were extracted from the mature human milk of four individual donors. Using an in vitro model of intestinal injury, the effects of the oligosaccharides on the intestinal epithelial barrier and select innate and adaptive immune functions were assessed. Results: Individual oligosaccharide compositions shared comparable effects on increasing transepithelial electrical resistance and reducing the macromolecular permeability of polarized (Caco-2Bbe1) monolayers but exerted distinct effects on the localization of the intercellular tight junction protein zona occludins-1 in response to injury induced by a human enteric bacterial pathogen Escherichia coli, serotype O157:H7. Immunoblots showed the differential effects of oligosaccharide compositions in reducing host chemokine interleukin 8 expression and inhibiting of p38 MAP kinase activation. Conclusions: These results provide evidence of both shared and distinct effects on the host intestinal epithelial function that are attributable to inter-individual differences in the composition of human milk oligosaccharides.

Biocompatible Gas Plasma Treatment Affects Secretion Profiles but Not Osteogenic Differentiation in Patient-Derived Mesenchymal Stromal Cells.

Cold physical plasma (CPP), a partially ionized gas that simultaneously generates reactive oxygen and nitrogen species, is suggested to provide advantages in regenerative medicine. Intraoperative CPP therapy targeting pathologies related to diminished bone quality could be promising in orthopedic surgery. Assessment of a clinically approved plasma jet regarding cellular effects on primary bone marrow mesenchymal stromal cells (hBM-MSCs) from relevant arthroplasty patient cohorts is needed to establish CPP-based therapeutic approaches for bone regeneration. Thus, the aim of this study was to derive biocompatible doses of CPP and subsequent evaluation of human primary hBM-MSCs’ osteogenic and immunomodulatory potential. Metabolic activity and cell proliferation were affected in a treatment-time-dependent manner. Morphometric high content imaging analyses revealed a decline in mitochondria and nuclei content and increased cytoskeletal compactness following CPP exposure. Employing a nontoxic exposure regime, investigation on osteogenic differentiation did not enhance osteogenic capacity of hBM-MSCs. Multiplex analysis of major hBM-MSC cytokines, chemokines and growth factors revealed an anti-inflammatory, promatrix-assembling and osteoclast-regulating secretion profile following CPP treatment and osteogenic stimulus. This study can be noted as the first in vitro study addressing the influence of CPP on hBM-MSCs from individual donors of an arthroplasty clientele.

Characteristics of Pooled Wharton's Jelly Mesenchymal Stromal Cells (WJ-MSCs) and their Potential Role in Rheumatoid Arthritis Treatment.

Mesenchymal stromal cells (MSC) from Wharton's jelly of umbilical cord is primitive and serve as an inexhaustible source of stem cells with greater potential in clinics. The existence of heterogeneity among the donor MSCs makes it difficult to predict the properties and clinical outcome of WJ-MSCs. We developed a strategy to minimize the donor to donor heterogeneity and produce consistency in biological properties by pooling three individual donors WJ-MSCs. Further, evaluated the effectiveness of the pooled MSCs in regulating the disease severity of Rheumatoid arthritis (RA) in animal models. WJ-MSCs were isolated from umbilical cord obtained from different donors, characterised and pooled based on the gender of baby. The biological properties of the pooled WJ-MSCs were compared to the individual WJ-MSCs. Further, the pooled WJ-MSCs were analysed for their safety profile in both in vitro and in vivo settings. The efficiency of pooled WJ-MSCs in regulating RA pathogenesis was also analysed in mice models of Collagen induced arthritis (CIA). We identified differences in proliferation capacity, pro inflammatory gene expression levels among individual WJ-MSCs isolated from different donors and the variation is also attributed to gender difference. WJ-MSCs pooled and cultured from different donor's exhibit all the MSC characteristics and exhibited superior immunosuppressive capabilities. In the in vivo toxicity study, pooled MSCs are found to be safe, and further in the RA preclinical studies, they were found to decrease the disease severity in these animals. Pooled WJ-MSCs reduces heterogeneity of individual donors and have superior immunosuppressive property. It is also effective in reducing the disease severity in the experimental animal models of RA.

Method bias in calcium measurement

The RCPAQAP offers a Liquid (Serum) Chemistry Program sourced from pooled serum from an individual male and female haemochromatosis donors which by definition is fully commutable and should behave the same as patient samples. RCPA/AACB Harmonised Calcium reference intervals are widely adopted in Australia, so the assumption is that there are no significant method biases between analytical platforms. We report on method differences in the 2020 and 2021 Liquid Chemistry Program which could potentially impact patient care.

Empires of Antiquities: Modernity and the Rediscovery of the Ancient Near East, 1914–1950

Empires of Antiquities: Modernity and the Rediscovery of the Ancient Near East, 1914–1950

HlaR: A rapid and reproducible tool to identify eplet mismatches between transplant donors and recipients

Eplet mismatch load, both overall and at the single molecule level, correlates with transplant recipient outcomes. However, precise eplet assessment requires high-resolution HLA typing of both the donor and recipient. Anything less than high-resolution typing requires imputation of HLA types. The currently available methods to identify eplet mismatch are both tedious and demanding. Therefore, we developed a software package and user-friendly web application (hlaR), that simplifies the workflow of eplet analysis, provides functions to impute high-resolution from low-resolution data and calculates both overall and single molecule eplet mismatch for single or multiple donor recipient pairs. Compared to manual assessments using currently available tools (namely, HLAMatchMaker), hlaR resulted in only minimal discrepancy in eplet mismatches (mean absolute difference of 0.56 for class I and 0.86 for class II for unique sum across loci). Additionally, output of the single molecule eplet function compared well to manual calculation, with an average single antigen count increase of 0.19. Importantly, the hlaR tool permits rapid and reproducible imputation and eplet mismatch including comparison between eplet reference tables (e.g. HLAMatchMaker version 2 or 3). Users can import data from a spreadsheet rather than relying on keystroke entry of individual donor and recipient data, thus reducing the risk of data entry errors. The resulting improved scalability of the hlaR tool is highlighted by plotting analysis time against the size of the input dataset. The new hlaR tool can provide eplet mismatch data with a streamlined workflow. With decreased effort from the end user, eplet matching and mismatch load data can be further incorporated into both research and clinical use.

Competitively disrupting the neutrophil-specific receptor–autoantigen CD177:proteinase 3 membrane complex reduces anti-PR3 antibody-induced neutrophil activation

CD177 is a neutrophil-specific receptor presenting the proteinase 3 (PR3) autoantigen on the neutrophil surface. CD177 expression is restricted to a neutrophil subset, resulting in CD177pos/mPR3high and CD177neg/mPR3low populations. The CD177pos/mPR3high subset has implications for antineutrophil cytoplasmic autoantibody (ANCA)–associated autoimmune vasculitis, wherein patients harbor PR3-specific ANCAs that activate neutrophils for degranulation. Here, we generated high-affinity anti-CD177 monoclonal antibodies, some of which interfered with PR3 binding to CD177 (PR3 “blockers”) as determined by surface plasmon resonance spectroscopy and used them to test the effect of competing PR3 from the surface of CD177pos neutrophils. Because intact anti-CD177 antibodies also caused neutrophil activation, we prepared nonactivating Fab fragments of a PR3 blocker and nonblocker that bound specifically to CD177pos neutrophils. We observed that Fab blocker clone 40, but not nonblocker clone 80, dose-dependently reduced anti-PR3 antibody binding to CD177pos neutrophils. Importantly, preincubation with clone 40 significantly reduced respiratory burst in primed neutrophils challenged with either monoclonal antibodies to PR3 or PR3–ANCA immunoglobulin G from ANCA-associated autoimmune vasculitis patients. After separating the two CD177/mPR3 neutrophil subsets from individual donors by magnetic sorting, we found that PR3–ANCAs provoked significantly more superoxide production in CD177pos/mPR3high than in CD177neg/mPR3low neutrophils, and that anti-CD177 Fab clone 40 reduced the superoxide production of CD177pos cells to the level of the CD177neg cells. Our data demonstrate the importance of the CD177:PR3 membrane complex in maintaining a high ANCA epitope density and thereby underscore the contribution of CD177 to the severity of PR3–ANCA diseases.

Kidney Transplantation Outcome Predictions (KTOP): A Risk Prediction Tool for Kidney Transplants from Brain-dead Deceased Donors Based on a Large European Cohort

BackgroundEuropean kidney donation shortages mandate efficient organ allocation by optimizing the prediction of success for individual recipients. ObjectiveTo develop the first European online risk tool for kidney transplant outcomes on the basis of recipient-only and recipient plus donor characteristics. Design, setting, and participantsWe used individual recipient and donor risk factors and three outcomes (death, death with functioning graft [DWFG], and graft loss) for 32 958 transplants within the Eurotransplant kidney allocation system and the Eurotransplant senior program between January 2006 and May 2018 in eight European countries to develop and validate a risk tool. Outcome measurements and statistical analysisCox proportional-hazards models were used to analyze the association of risk factors with overall patient mortality, and proportional subdistribution hazard regression models for their association with graft loss and DWFG. Prediction models were developed with recipient-only and recipient-donor risk factors. Sensitivity analyses based on time-specific area under the receiver operating characteristic curve (AUC) with leave-one-country-out validation were performed and calibration plots were generated. Results and limitationsThe 10-yr cumulative incidence rate was 37% for mortality, 12% for DWFG, and 41% for graft loss. In recipient-donor models the leading risk factors for mortality were recipient diabetes (hazard ratio [HR] 10.73), retransplantation (HR 3.08 per transplant), and recipient age (HR 1.08). Effects were similar for DWFG. For graft loss, diabetes (subdistributional HR [SHR] 1.32), increased donor age (SHR 1.02), and prolonged cold ischemia time (SHR 1.02) had increased SHRs. All p values were <0.001. ConclusionsPreviously identified risk factors for outcomes following kidney transplants allow for outcome prediction with 10-yr AUC values of up to 0.81. Patient summaryUsing European data, we estimated individual risks to predict the success of kidney transplants and support physicians in decision-making. An online tool is now available (https://riskcalc.org/ktop/) for predicting kidney transplant outcomes both before and after a donor has been identified.

IgG3 and IgM Identified as Key to SARS-CoV-2 Neutralization in Convalescent Plasma Pools.

Analysis of convalescent plasma derived from individuals has shown that IgG3 has the most important role in binding to SARS-CoV-2 antigens; however, this has not yet been confirmed in large studies, and the link between binding and neutralization has not been confirmed. By analyzing plasma pools consisting of 247-567 individual convalescent donors, we demonstrated the binding of IgG3 and IgM to Spike-1 protein and the receptor-binding domain correlates strongly with viral neutralization in vitro. Furthermore, despite accounting for only approximately 12% of total immunoglobulin mass, collectively IgG3 and IgM account for approximately 80% of the total neutralization. This may have important implications for the development of potent therapies for COVID-19, as it indicates that hyperimmune globulins or convalescent plasma donations with high IgG3 concentrations may be a highly efficacious therapy.

Raman-based spectrophenotyping of the most important cells of the immune system

IntroductionHuman peripheral blood mononuclear cells (PBMCs) are a heterogeneous population of cells that includes T and B lymphocytes. The total number of lymphocytes and their percentage in the blood can be a marker for the diagnosis of several human diseases. Currently, cytometric methods are widely used to distinguish subtypes of leukocytes and quantify their number. These techniques use cell immunophenotyping, which is limited by the number of fluorochrome-labeled antibodies that can be applied simultaneously. ObjectiveB and T lymphocytes were isolated from peripheral blood obtained from healthy human donors. MethodsThe immunomagnetic negative selection was used for the enrichment of B and T cells fractions, and their purity was assessed by flow cytometry. Isolated cells were fixed with 0.5% glutaraldehyde and measured using confocal Raman imaging. K-means cluster analysis, principal component analysis and partial least squares discriminant methods were applied for the identification of spectroscopic markers to distinguish B and T cells. HPLC was the reference method for identifying carotene in T cells. ResultsReliable discrimination between T and B lymphocytes based on their spectral profile has been demonstrated using label-free Raman imaging and chemometric analysis. The presence of carotene in T lymphocytes (in addition to the previously reported in plasma) was confirmed and for the first time unequivocally identified as β-carotene. In addition, the molecular features of the lymphocytes nuclei were found to support the discriminant analysis. It has been shown that although the presence of carotenoids in T cells depends on individual donor variability, the reliable differentiation between lymphocytes is possible based on Raman spectra collected from individual cells. ConclusionsThis proves the potential of Raman spectroscopy in clinical diagnostics to automatically differentiate between cells that are an important component of our immune system.

Myths and Misconceptions Preventing College Going Students from Voluntary Blood Donation

Background and Objectives: Blood services are facing a shortage of blood all over the world. Demand for blood is rising day by day and current blood donation is insufficient to meet the demand. This study was planned to assess the myths of college-going students regarding blood donation, address their misconceptions, and raise awareness among college students on the importance of blood donation. Methods: The study population comprised 1000 students above the age of 18 years who qualify for blood donation after assessment of suitability to donate blood as per the blood donor questionnaire and consent form of the department. Results: Myths and misconceptions were more prevalent among nonblood donor students. The most common reason for not donating blood, as stated by nonblood donor students, was fear of needle pain in 137 (27.4%), followed by being not fit to donate in 134 (26.8%) and the fear of becoming weak in 95 (19%). Conclusion: The most common reason comes out to be related to health of the individual donor; almost one in two college-going students (45.8%) fear that either they are not fit enough to donate blood (26.8%) or that they will become weak (19%) after blood donation.

Experience of individual donor nucleic acid testing on screening of blood donors for human immunodeficiency virus, hepatitis c virus, and hepatitis b virus at an Apex blood bank of Northern India

Experience of individual donor nucleic acid testing on screening of blood donors for human immunodeficiency virus, hepatitis c virus, and hepatitis b virus at an Apex blood bank of Northern India

Shotgun Immunoproteomics for Identification of Nonhuman Leukocyte Antigens Associated With Cellular Dysfunction in Heart Transplant Rejection.

The International Society for Heart and Lung Transplant consensus panel notes that too little data exist regarding the role of non-HLA in allograft rejection. We developed a novel shotgun immunoproteomic approach to determine the identities and potential roles non-HLA play in antibody-mediated rejection (AMR) in heart transplant recipients. Serum was collected longitudinally from heart transplant recipients experiencing AMR in the absence of donor-specific anti-HLA antibodies (n = 6) and matched no rejection controls (n = 7). Antidonor heart affinity chromatography columns were formed by recipient immunoglobulin G immobilization at transplantation, acute rejection, and chronic postrejection time points. Affinity chromatography columns were used to capture antigens from individual patient's donor heart biopsies collected at transplantation. Captured proteins were subjected to quantitative proteomic analysis and the longitudinal response was calculated. Overlap in antigen-specific response between AMR and non-AMR patients was only 8.3%. In AMR patients, a total of 155 non-HLAs were identified, with responses toward 43 high prevalence antigens found in ≥50% of patients. Immunofluorescence staining for representative high prevalence antigens demonstrated that their abundance increased at acute rejection, correlating with their respective non-HLA antibody response. Physiological changes in cardiomyocyte and endothelial cell function, following in vitro culture with patient immunoglobulin G, correlated with response toward several high prevalence antigens. This work demonstrates a novel high-throughput strategy to identify clinically relevant non-HLA from donor endomyocardial biopsy. Such a technique has the potential to improve understanding of longitudinal timing of antigen-specific responses and their cause and effect relationship in graft rejection.

Remarks on the Anonymous Collective Sponsorships in Post-Byzantine Epirus (Greece): The Case of an Eighteenth-Century Painting Workshop

Co-operative patronage is based on the joint effort of individuals, lay or clerical, couples, families, colleagues, ecclesiastical and military authorities, or fellow citizens. Through the donor inscriptions are revealed the different categories of such co-operative patronage in Byzantine and Post-Byzantine society. In the Greek-speaking Post-Byzantine world, such types of anonymous groups of donors and benefactors most often came from a community as a whole, or certain inhabitants of a region, while collective donations by groups of monks were also widespread. The present paper examines the practice of anonymous collective sponsorships in Post-Byzantine Epirus, presenting the surviving monuments from the sixteenth to the seventeenth century and, in detail, the cases of anonymous collective sponsorships in a specific painting workshop of the eighteenth century, that of the so-called Kapesovite painters. In Post-Byzantine period the special privileges from the Ottomans and the development of trade, contributed to the Epirus’s cultural development. The tectonic transformations in the residential network of Epirus began in the late sixteenth century and increased after the seventeenth century. During the eighteenth century, the flourishing of Post-Byzantine art is a fact, indicating the gradual rise to prevalence of the parishes and the communities over the monastic establishments and individual donors. The financial and commercial privileges, especially after the treaty of Kucuk Kaynarca (1774), contributed decisively to religious monuments’ construction or renovation. The financial circumstances and the social cohesion of the Orthodox Christians in Epirus favored the increase of anonymous collective sponsorship in the eighteenth century. The monuments of that period evidence a significant amount of co-operative patronage, in which “anonymity” starred among the donors. The anonymous collective sponsorships indicates the community’s cohesion and the benefactor’s desire to create a legacy for future generations.

From the Editor’s Desk...

From the Editor’s Desk...

What convinces donors? An analysis of donation‐based crowdfunding projects from nonprofit charities: The case of South Korea

AbstractDespite the potential usefulness of donation‐based crowdfunding as a fundraising tool for nonprofit social welfare organizations, academic understanding of what leads to a successful campaign is limited. Drawing on an information processing model, this research analyzes 889 donation‐based crowdfunding campaigns undertaken by nonprofit social welfare organizations using HappyBean, the most representative crowdfunding platform in South Korea. This study examines whether different types of project information influence the percentage of individual donors giving to the campaign's funding goal. Results show that the emotional impression of a campaign title, visual information, message quality, and message contents are significantly related to the extent of crowdfunding success. This study provides recommendations for nonprofit social welfare organizations to improve the way in which they operate crowdfunding campaigns and communicate with potential funders.

A Direct MS-Based Approach to Profile Human Milk Secretory Immunoglobulin A (IgA1) Reveals Donor-Specific Clonal Repertoires With High Longitudinal Stability

Recently, a mass spectrometry-based approach was introduced to directly assess the IgG1 immunoglobulin clonal repertoires in plasma. Here we expanded upon this approach by describing a mass spectrometry-based technique to assess specifically the clonal repertoire of another important class of immunoglobulin molecules, IgA1, and show it is efficiently and robustly applicable to either milk or plasma samples. Focusing on two individual healthy donors, whose milk was sampled longitudinally during the first 16 weeks of lactation, we demonstrate that the total repertoire of milk sIgA1 is dominated by only 50-500 clones, even though the human body theoretically can generate several orders of magnitude more clones. We show that in each donor the sIgA1 repertoire only changes marginally and quite gradually over the monitored 16-week period of lactation. Furthermore, the observed overlap in clonal repertoires between the two individual donors is close to non-existent. Mothers provide protection to their newborn infants directly by the transfer of antibodies via breastfeeding. The approach introduced here, can be used to visualize the clonal repertoire transferred from mother to infant and to detect changes in-time in that repertoire adapting to changes in maternal physiology.