bluuebunny/crossref_metadata_2025_split huggingface dataset:
Membrane Protein and Lipid Properties of Freeze-Dried Human Red Blood Cells.
Authors: Banerjee Mitali , Bali Rachna , Torok Zsolt , Satpathy Gayana R. , Little Erika , Tablin Fern , Crowe John H. , Tsvetkova Nelly M. 
Published: April 2020

Abstract
Post-rehydration membrane properties are an important indicator of effective freeze-drying of mammalian blood cells. In general, cellular membranes are extremely susceptible to damage during freezing and drying. The disaccharide trehalose, naturally occurring in anhydrobiotic organisms, has been shown to prevent such damage through direct interactions with the phospholipids and proteins, and by forming an amorphous glassy matrix around the membranes. Thermal properties and lipid composition of plasma membranes from freeze-dried trehalose loaded RBCs were studied using Fourier Transform Infrared Spectroscopy (FTIR) and analytical thin layer chromatography (TLC). FTIR results showed that plasma membranes from freeze-dried and rehydrated RBCs have major phase transitions between 10 and 20°C, and between 30 and 40°C, similarly to membranes isolated from fresh RBCs. Analysis of the lipid composition of freeze-dried RBCs showed profiles of the freeze-dried RBCs that are very similar to the ones of membranes from freshly isolated RBCs, suggesting that the freeze-drying procedure did not affect the lipid composition. SDS-PAGE and immunoblotting analysis showed that the cytoskeletal components of the membrane such as spectrin, ankyrin, band 3 and glycophorin A which form a flexible meshwork of proteins underlying the lipid bilayer are also well preserved in the freeze-dried RBCs. The study demonstrates that freeze-drying of trehalose loaded human erythrocytes does not result in significant changes in the lipid and protein structure. These findings have immediate applications in clinical medicine for long-term storage of RBCs. Supported by DARPA grant N66001-03-1-9827
Severe Acute Hemolytic Transfusion Reaction Following ABO-Mismatched Platelet Transfusion: Should ABO-Mismatched Platelet Transfusion Policy Be Evaluated.
Authors: Zubair Abba , Grant Rhonda , Black Neta , Bertholf Marsha , Roy Archana , Roy Vivek 
Published: April 2020

Abstract
Transfusion of ABO-mismatched platelets is common practice. Although a unit of single donor apheresis platelet (SDP) contains as much plasma and passive anti-A or/and anti-B antibodies (allo-agglutinins) as a unit of fresh frozen plasma, ABO- mismatched platelet transfusions are considered safe and hemolytic transfusion reactions have only rarely been reported. We report a case of a 67-year old male, blood group A, with mantle cell lymphoma who received 1 SDP for chemotherapy induced thrombocytopenia. Several prior platelet transfusions had been uneventful. After 150 ml of irradiated, leukoreduced, SDP from blood group O donor had been infused, he developed nausea, light-headiness, chills, and back pain. He became bradycardic and hypotensive. Platelet transfusion was discontinued and intravenous dexamethasone and rapid saline infusion was administered. Later, his hemoglobin dropped from 10.6 to 7.4 g/dl and total serum bilirubin increased by 9 fold. A direct antiglobulin test (DAT) on post-transfusion sample demonstrated 1+ reactivity at room temperature. An elution test performed on the patient’s circulating red blood cells contained anti-A antibodies. These results are consistent with a severe acute hemolytic transfusion reaction. The donor anti-A and anti-B antibodies titers at room temperature and at 37°C were 512 and 2048 with A1 cells and 128 and 256 with B cells, respectively.This case illustrates the rare possibility of hemolytic transfusion reaction occurring as a result of ABO-mismatched platelets from group O donor with high-titer anti-A antibodies. SDP (in contrast to pooled platelets) offers the advantages of reduced risk of infection transmission and alloimmunization by limiting exposure to one donor only. However, it increases the opportunity for hemolytic transfusion reaction to occur if the donor has high-titer antibodies which would have been diluted if pooled platelets were used. As SDP use becomes more widespread, the risk of clinically significant hemolytic reactions is likely to increase, especially in seriously ill patients who tolerate these reactions particularly poorly. Moreover, increasingly, apheresis platelets are being donated by a small group of highly motivated donors, raising the chances of repeated transfusions from the same donors. We propose that the safety of ABO-mismatched platelet transfusions be re-evaluated. Effectiveness (and costs) of potential preventive strategies such as screening all SDP units for high-titer antibodies and/or plasma volume reduction of all mismatched SDP units should be studied.
Sustained Suppression of Hepatitis C by Pegylated Interferon and Ribavirin in Hemophilic Patients.
Authors: Santagostino Elena , Mancuso Maria E. , De Filippi Francesca , Rumi Maria G. , Colombo Massimo , Mannucci Pier M. 
Published: April 2020

Abstract
Pegylated interferon (PegIFN) and ribavirin therapy is the standard treatment option for patients with chronic hepatitis C. In a multicenter open trial, we assessed the efficacy and tolerability of PegIFN and ribavirin in 72 HIV negative hemophiliacs (age: 21–64 years) with persistently high ALT values. HCV genotype was 1 in 72%, 3 in 14%, 2 in 12% and 4 in 3%. Treatment: PegIFN alpha2b 1.5 mcg/Kg/week for 48 weeks in genotypes 1/4 or 24 weeks in genotypes 2/3 plus ribavirin 800–1200 mg/day based on viral type. Treatment was stopped in patients lacking virological response at month 6. All patients were assessed for 6 months post-treatment. Results: 11 patients withdrew for side-effects (6) or non-compliance (5). Sixteen patients (22%) did not achieve a primary virological response and 4 (6%) had a virological breakthrough. Therefore, 41 patients (57%) had had an end-of-treatment response. Post-treatment follow-up is ongoing in 7 patients. In 65 patients who completed the study, 2 were relapsers and 32 (49%) had a sustained virological response (SVR, 82% of genotypes 2/3 and 38% of genotypes 1/4). Neutropenia (&lt;800 cells), high ALT and diabetes were reasons for treatment interruption. The median fall in hemoglobin levels was 3.1 g/dL. Leucopenia (40%), weight loss (39%), and fatigue (35%) were frequent side-effects. Thirty patients (42%) required dose reduction of ribavirin (21, 29%) and PegIFN (22, 30%). SVR rates were similar in patients who adhered to protocol and those requiring dose reduction (56% vs 45%). Conclusions: combination therapy with PegIFN and ribavirin improves SVR in HIV negative hemophiliacs with chronic hepatitis C.
Extracellular UTP Affects In Vitro Migration and In Vivo Homing and Engraftment of CD34+ Hematopoietic Stem Cells.
Authors: Rossi Lara , Fogli Miriam , Ferrari Davide , Pizzirani Cinzia , Vaselli Diletta , Bertolini Francesco , Di Virgilio Francesco , Baccarani Michele , Lemoli Roberto M. 
Published: April 2020

Abstract
Homing and engraftment of hematopoietic stem cells (HSCs) depend on the complex interplay between chemokines, cytokines, growth factors and adhesion molecules in the intricate architecture of bone marrow (BM) microenvironment. We have recently reported that CD34+ HSCs express P2Y and P2X receptors (P2Rs) for extracellular nucleotides. P2Rs activation by ATP and UTP produced stimulatory effects, in synergy with other hematopoietic cytokines, on clonogenic CD34+ and lineage-negative CD34− progenitor cells. ATP and UTP also expanded the more primitive CD34+- derived long-term culture-initiating cells. Extracellular nucleotides are also emerging as chemotactic factors for different cell types, including vascular endothelial cells, arterial smooth muscle cells and neutrophils, and regulate the trafficking of specific dendritic cell populations. In this study we asked if the extracellular nucleotide UTP modulates the migration of HSCs in response to the chemotactic peptide CXCL12/SDF-1α. Very low concentrations of UTP (10 μM) significantly improved stem cell migration, assayed with the dual-chamber migration system and evaluated by cell counting and clonogenic assays. Phenotypical analysis of CXCR4 antigen showed that SDF-1α gradients mainly attract HSCs expressing high levels of this receptor. Conversely, UTP induced the migration of HSCs expressing low levels of CXCR4. Furthermore, we evaluated actin polymerization as index of HSCs capacity to respond to chemotactic stimulation and to home to the BM microenvironment. The combined stimulation of CD34+ cells with UTP and SDF-1α produced a synergistic increase in actin polymerization, as indicated by the enhanced fluorescence intensity of FITC-phalloidin. Adhesion assays, performed in fibronectin-coated wells, also showed that UTP can stimulate the adhesion of HSCs to fibronectin, a key component of the extracelluar bone marrow microenvironment, without any substantial change in VLA-4 and VLA-5 expression levels. Moreover, xenogenic transplant studies showed that short-term pre-incubation with UTP significantly increased the engraftment efficiency of CD34+ HSC in nonobese diabetic / severe combined immunodeficiency mice. Taken together, our data suggest that UTP may affect HSC migration and homing to the BM, in synergy with the chemotactic peptide SDF-1α, perhaps activating a CXCR4-indipendent pathway.
Hematopoietic Stem Cells “Stemness” Genes Contain Many Genes Mutated or Abnormally Expressed in Hematopoietic Malignancies.
Authors: Toren Amos , Bielorai Bella , Jacob-Hirsch Jasmine , Fisher Tamar , Zelikovsky Sharon , Givol David , Domany Eitan , Itskovitz-Eldor Joseph , Kventsel Iris , Rosenthal Ester , Amariglio Ninette , Rechavi Gideon 
Published: April 2020

Abstract
Umbilical cord blood (CB) and mobilized peripheral blood (PB) are relatively new sources of hematopoietic stem cells (HSC) that have been increasingly used in clinical transplantations. The genetic basis of the underlying mechanism of HSC self- renewal and differentiation has not been elucidated yet.
Several groups have recently used microarray technology to study the common characteristics of stem cells from different tissues (“stemness”) and the typical features of each source of stem cell (“specificity”). Most groups focused their study on mice, used relatively small cDNA microarrays, and used CD34 as the cell surface marker for stem cells isolation. We studied human stem cells characterized by expression of the more primitive CD133 antigen, and used the Affymetrix Human Hu133A oligonucleotide arrays containing 22,215 probe sets to study the expression profile of these cells. An unsupervised hierarchical clustering of 14,025 “valid” probe sets showed a clear distinction between the CD133 + cells representing the stem cell population, and CD133 − cells that represent various stages of cell differentiation. CD133+ cells isolated from CB were compared to CD133− cells identifying 304 genes that were up regulated by at least two folds and the comparison between PB CD133+ cells and CD133− cells identified 218 genes which were up regulated by at least two-folds. These genes were considered as source specific and maybe relevant to the unique properties of CB and PB derived HSC.
We were concentrated in the 244 genes that were found to be up regulated by at least two folds in the CD133 positive cells as compared to the CD133 negative cells and were common to both CB and PB. Comparison of these “stemness” genes, to the lists of “stemness” genes that were identified by 2 recent studies that analyzed mainly murine HSC identified 39 (Ramalho et al.) and 83 (Ivanova et al.) common genes.
Twenty-four genes were common to another study that analyzed human HSC (Georgantas et al.).
Among these common “stemness” genes we identified 4 groups of genes that have an important role in hematopoiesis: Growth factor receptors (Receptor thyrosine kinases (RTK) and c-mpl), a group of transcription factors which includes several homeobox genes and TGF-β targeted genes, genes that have an important role in the process of development and genes involved in cell growth.
Among these 4 groups we identified 16 “stemness” genes (MPL, FLT3, HOXA9, MEIS 1, MLLT3, KIT, TIE, GATA-2, HOXA5, BAALC, HLF, MYCN, EVI1, MYB, FHL1, and HMGA2) that are known to be mutated or abnormally regulated in acute leukemias. It can be suggested that perturbation of expression of key hematopoietic stemness machinery genes may lead to abnormal proliferation and leukemia.]
The Higher CD34+ Cell Counts in Cord Blood Than Those in Neonatal Peripheral Blood Correlate with Increased Expression of Cytokine Receptors and Cell Adhesion Molecules on the CD34+ Cells in Cord Blood.
Authors: Lee Young-Ho , Kim Joong-Pyo , Sung Myung-Hwa , Ko Ri-Young , Han Jin-Yeong 
Published: April 2020

Abstract
The hematologic values of neonatal peripheral blood show a characteristic series of changes during neonatal period. The hematopoietic stem cells in cord blood (CB) are more primitive and abundant than those in neonatal peripheral blood (NB). However, there have been no studies regarding the reasons for those differences. Thus, We investigated the kinetics of nucleated cells and CD34+ cells in CB and neonatal peripheral blood. We obtained the CB and NB samples from 14 normal full-term babies for this study, which were collected immediately after delivery as well as at 24 hours and 48 hours after delivery. We analysed the expressions of CD34, CD34/CXCR4, CD34/CD49d and CD34/CD44 as well as CD3, CD19 and CD16/56 on the isolated mononuclear cells by using FACSort. We also performed CFU-GM counts and apoptotic analysis with isolated mononuclear cells. The total white blood cell counts as well as absolute neutrophil counts of NB at delivery were higher than those of CB (p=0.04) and increased until 24 hours, when decreased until 48 hours after delivery. There were no differences of absolute lymphocyte counts, CD3+ cell and CD19+ cell counts among CB and NB at 0, 24 and 48 hours postnatally, however, the CD16/56+ cell counts were lower in CB than in NB and gradually decreased until 48 hours after delivery (p=0.011). The number of CFU-GM counts (p=0.02) and CD34+ cells (p=0.049) as well as CD34+CXCR4+ cells (p=0.002), CD34+CD49d+ (p=0.001) and CD34+CD44+ cells (p=0.001) in CB were more significantly higher than those of NB at delivery, when decreased gradually until 48 hours after delivery. The incidence of apoptosis were statistically not significant among CB and NB at 0, 24 and 48 hours postnatally. In conclusion, the higher leukocyte counts for several days after delivery were correlated with neutrophilic increment which may be influenced by stressful events during delivery. The more higher CFU-GM counts and CD34+ cell counts in CB than those in NB might be correlated with increased expression of cytokine receptors and cell adhesion molecules on the CD34+ cells in CB.
Differential Gene Expression Profiles of Cord Blood Lineage-Negative Cells Prior to and Following Selective Amplification.
Authors: Shi Meng-jiao , Hermann Jason , Laning Joseph , Hoppe Henry , Kraus Morey , Visser Jan W.M. 
Published: April 2020

Abstract
Using a selective-amplification system, we have ex vivo expanded umbilical cord blood (UCB) derived lineage-negative− (Lin−: CD2−, CD3−, CD14−, CD16−, CD19−, CD24−, CD56−, CD66b−, and Glycophorin A−) cells. To assess at the molecular level the effect of the ex vivo expansion on properties of UCB Lin−-cells, we compared gene expression profiles between cells at day 0 and cells cultured in serum-free medium supplemented with rhFlt-3L, rhSCF and rhTPO (100ng/ml each) for 14 days with a reselection using the same lineage depletion antibody cocktail at the 7th day of culture. In this study, we utilized a human stem cell gene cDNA array (SuperArray Bioscience Corp.), which include genes encoding surface markers, growth factor/cytokines, extracellular matrix molecules and cell cycle regulators for embryonic, neural, mesodermal and hematopoietic stem/progenitor cells. Of the 266 total genes in the array, we detected 85 genes expressed above background in either of the two populations. Among these expressed genes, 19 were found only in the day_0 (pre-culture) population and 11 were found only in the day_14 cultured cells. Ex vivo expansion under these conditions had no significant effect on the majority of the genes, which include highly expressed homing-related (CXCR4, CD44) and cell adhesion (CD31) molecules. Further analysis revealed that, compared to the levels in the original day_0 Lin− population, 7 genes showed more than 3-fold increases and 13 genes showed more than 3-fold decreases in their levels of expression in the cultured day_14 cell population. The up-regulated genes included those involved in cell cycling, proliferation and migration or anti-apoptosis (cyclin E1, IGFR-2, alpha(4)beta(1)-integrin, Mdm2, Cystatin C, ALK-5). The down-regulated genes included those involved in embryological or neural development (GCMb, FGF11), a subset of integrin molecules (integrin alpha E, beta 3 and 5) and an inhibitor of BMP signaling (CER1). In conclusion, our results indicate that selective ex vivo expansion conditions promote expression of genes related to the growth and expansion of targeted hematopoietic precursor cells while retaining expression of homing-related genes. Further studies are slated to link, where possible, these transcriptional expression patterns with expression and functions of these genes at the protein and cellular levels.
Figure Figure
Nicotinamide, a Potent SIRT2 Inhibitor, Delays Differentiation of Hematopoietic Progenitor Cells.
Authors: Peled Toni , Adi Sophie , Glukhman Elina , Grynspan Frida , Nagler Arnon , Fibach Eitan , Daniely Yaron 
Published: April 2020

Abstract
CD38, originally described as a differentiation marker, has emerged as an important multifunctional transmembrane protein. Its most intriguing and well-characterized function is its ability to catalyze the synthesis of cyclic ADP-ribose (cADPR) from NAD. Of particular interest is its presence on the inner membrane of the nucleus, suggesting that CD38/cADPR may play a direct role in mediating nuclear activation and gene expression. Our studies on ex vivo expansion of Hematopoietic Stem Cells (HSCs) have led us to test whether alteration of CD38 function carries the potential of affecting cell fate decisions of HSCs. Inhibition of CD38 enzymatic activity was achieved by treating CD34+ cell cultures with nicotinamide (NA), a well-known base-exchange inhibitor demonstrated to inhibit the synthesis of cADPR from NAD.
We report here that exogenously added nicotinamide (5–10 mM) to CD34+ cell cultures supplemented with cytokines (SCF, TPO, IL-6, FLt3, +/− IL-3) resulted in significant enrichment of CD34+CD38− (79±9.3%, n=9) and CD34+CD38−Lin− (19±3%, n=8) cells, as compared with control cultures treated only with cytokines (6.3±1.8%, n=9, and 0.7±0.06%, n=8, respectively, p&lt;0.01). The functionality of these early progenitor subsets was demonstrated using the extended LTC-CFC assay, performed in the absence of NA. These results raised the intriguing possibility that cADPR production may have a pivotal role in regulation of CD34+ cell fate. However, inhibition of cADPR downstream signal transduction pathways by its specific antagonist, 8-amino-cADPR did not yield any effect on CD34+ cell cultures, excluding the possibility that nicotinamide modulates CD34+ cell fate solely by inhibition of cADPR synthesis.
Nicotinamide is also a well-known potent inhibitor of SIRT2, a unique NAD(+)-dependent type III histone deacetylase (HDAC) with mono-ADP-ribosyltransferase activity involved in gene silencing, metabolism, apoptosis and aging. NA blocks NAD(+) hydrolysis by binding to an adjacent conserved pocket, and is therefore suggested as the physiologically relevant regulator of SIRT2 enzymes. This additional function of nicotinamide raises the intriguing possibility that HSC enrichment achieved by nicotinamide treatment may be related to specific inhibition of SIRT2 deacetylase activity and modulation of chromatin architecture leading to re-activation of previously silenced genes.
In line with this hypothesis, Milhem et all. recently reported that addition of trichostatin A, a specific HDAC (type I and II) inhibitor, along with a DNA hypomethylating agent, modulated HSC fate ex vivo resulting in the retention of stem cell phenotype, number, and function (Blood, 2004; 103; 4102).
Ongoing work is aimed at elucidating whether inhibition of SIRT2 is specifically involved in NA mechanism of activity leading to modulation of hematopoietic stem cell fate in ex vivo conditions.
Effect of Serum on Long-Term Cultures of Human Umbilical Cord Blood Megakaryocyte Progenitors.
Authors: Smith Stephen L. 
Published: April 2020

Abstract
Platelet recovery following umbilical cord blood (CB) transplants can be delayed for several months. In vitro expansion and transplantation of CB megakaryocyte progenitor cells (MPC) may be one way to increase circulating platelets thereby shortening the period of thrombocytopenia. The in vitro expansion and differentiation of CB MPC derived from enriched CB CD34+ cells was studied after immuno-magnetic bead isolation. Isolated CB CD34+ cells were incubated in serum-free culture medium supplemented with a Tpo peptide agonist (TpoA), Flt-3L, Stem Cell Factor (SCF) and Vascular Endothelial Growth Factor (VEGF). At week 2, the cultures were divided and 1) maintained in serum-free medium, 2) supplemented with 5% human serum and, 3) supplemented with 5% mouse serum. Using flow cytometry and histocytochemistry, the cultured cells were evaluated for MPC using CD41a, CD36, von Willebrand factor (VWF) and also for myeloid progenitors using CD15 and CD33. The mean total percent MPC (CD41a+ and CD36+) at week 2 was 53 ±17% and 49 ±12%, respectively. After 2–4 weeks of additional culture the %MPC declined to less than 30% in cultures with or without serum supplements. However, at weeks 5–8 of additional culture the %MPC increased in the cultures supplemented with human or mouse serum. The mean %MPC was 45 ±19% (serum-free), 55 ±16% (human serum) and, 71 ±5% (mouse serum) in cultures at week 5–8 of additional culture. The remaining cells in the cultures were myeloid progenitors (CD15+ or CD33+). Total cell numbers at week 5–8 were inhibited 3 to 5x-fold in mouse serum supplemented cultures compared to serum free and human serum supplemented cultures. In addition, VWF+ cells could be detected after 5 weeks of additional culture when either human or mouse serum was present. These data help to explain the delay in platelet recovery typically associated with cord blood transplants but, indicate that additional serum factors are present to enhance megakaryocyte differentiation. These results also suggest that the transplant of cultured CB MPC may prove beneficial for the treatment of thrombocytopenic patients and normal platelet recovery.