bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gene Frequencies of the HNA-4a and -5a Neutrophil Antigens in Brazilians and a New Polymerase Chain Reaction-Restriction Fragment Length Polymorfism Method for HNA-5a Genotyping.
Authors: Cardone José D.B. , Chiba Akemi K. , Norcia Angela M.M.I. , Vieira-Filho João P.B. , Bordin José O. 
Published: October 2019

Abstract
The HNA-4 and HNA-5 systems are located on the b2-integrin, that share a common b subunit (b 2 or CD18) noncovalently associated with four different a subunits. The HNA-4a (Mart) and HNA-5a (Ond) antigens are polymorfic variants of a M (CD11b) and a L (CD11a) subunits, respectively, and both are associated with single nucleotide polymorphisms (SNP) leading to amino-acid dimorphisms. HNA-4a has been recently linked to alloimmune neonatal neutropenia while the HNA-5a clinical significance is not clear yet. Using a PCR with sequence-specific primers (SSP) the frequency of allelic polymorphism of HNA-4a among 121 Brazilian blood donors and 114 Amazon Indians was determined. A novel PCR-restriction fragment length polymorphism (RFLP) method digesting the PCR product with the enzyme Bsp1286 I for HNA-5a genotyping was developed, and the gene frequencies of this antigen were determined among 123 Brazilian blood donors and 114 Amazon Indians. In order to validate this proposed new genotyping method, the amplified genomic DNA from 6 previously genotyped samples (2 of each genotype) was purified using a commercial kit (GFX PCR DNA and Gel Band Purification Kit, Amersham Bioscience, Piscataway, NJ, USA), sequenced using a cycle-sequencing kit (BigDye Terminator, PE Applied Biosystems, Foster City, CA, USA) and analyzed on a genetic analyzer (ABI Prism 3100, PE Applied Biosystems). The calculated HNA-4a (+/+), HNA-4a (+/−) and HNA-4a (−/−) genotype frequencies found in blood donors (0.686, 0.273 and 0.041) and Amazon Indians (1.000, 0.000 and 0.000) presented statistically significant differences (p<0.01). The frequencies of HNA-5a (+/+), HNA-5a (+/−) and HNA-5a (−/−) genotypes among blood donors (0.512, 0.399 and 0.089) and Amazon Indians (0.746, 0.219 and 0.035) also differed significantly (p<0.01). Sequencing studies demonstrated absolute concordance with PCR-RFLP genotyping results in all 6 analyzed samples. Overall, the present data indicate that comparing to other population studies, the Brazilian population presents a higher frequency of the HNA-4a-negative allele, suggesting that Brazilians would be more susceptible to HNA-4a alloimmunization by pregnancy or blood transfusion. Moreover, the distribution of the HNA-4 system alleles observed in Amazon Indians is quite similar to that already reported among Korean individuals. Besides that, a new effective and efficient HNA-5a genotyping technique is now available for population studies.
Platelet-Derived Microparticles Promote Clot Stability.
Authors: Loncar Robert , Dzepina Daniel , Stoldt Volker , Zotz Reiner B. , Scharf Rudiger E. 
Published: October 2019

Abstract
Microparticles (MP) are the plasma membrane fragments. They are formed along with membrane remodelling processes of most stimulated eukaryotic cells or generated upon cellular stimulation including platelets. After having been considered “inert cell debris” previously, recent findings suggested that MP can modulate distinct cellular responses in the related microenvironment. For example, the concentration of circulating platelet-derived MP is increased in acute myocardial infarction and stroke. It is hypothesized that platelet-derived MP promote hemostasis and thrombosis. However, the precise role of MP is still unknown. In this study, we evaluated the influence of platelet-derived MP on clot stability. Anticoagulated blood (3.8% sodium citrate) was obtained from healthy blood donors. Platelet-rich plasma was centrifuged at 1500g (10 min, 22°C). The pelleted platelets were washed three times in PBS (pH 7.4), resuspended in 1 ml of the same buffer and activated with human collagen of type I (10 min, 35°C) at a final concentration of 10 μg/ml. The supernatant (1500g 10 min, 22°C) containing activated platelet MP was centrifuged at 13,000g (30 min, 4°C). The pellet of MP was resuspended in 450 μl of PBS. MP were identified by scanning electron microscopy and flow cytometry following immunolabelling with an anti-GPIbα FITC-monoclonal antibody. The influence of platelet MP onto clot stability, determined as platelet contractile force (PCF) and clot elastic modulus (CEM), was evaluated with a Hemodyne haemostasis analyzer (Hemodyne, Richmond, USA). Mean PCF and CEM in blood of healthy donors (n=7) were 6.5±3 Kdynes and 9.6±6 Kdynes/cm2, respectively. Addition of 100 μl of platelet-derived MP increased PCF (forces generated by platelets within a clot) without reaching statistical significance (mean increase of 11% as compared to controls without MP). By contrast, addition of platelet MP significantly enhanced CEM as measure of clot stability from 9.6±6 Kdynes/cm2 to 94 ± 62 Kdynes/cm2, (p<0.05). In experiments conducted with platelet-rich plasma or platelet-poor plasma instead of anticoagulated whole blood, no influence of added platelet-derived MP on clot stability was observed. In a patient with thrombocytopenia (70,000/μl) supplementation of whole blood with platelet MP increased CEM by 70%. Our ex vivo experiments demonstrate that collagen-induced platelet-derived MP can modulate clot stability. However, this effect is restricted to anticoagulated whole blood and not observed in platelet-rich plasma or plasma alone. Therefore, interaction of platelet-derived MP with other cellular elements than platelets, e.g. monocytes, may be relevant to promote clot stability. In general, microparticles may be a pharmacological target in the management of hemostatic disorders.
Impact of Joint Bleeding on Joint Function in a Murine Model of Hemophilia A.
Authors: Mejia-Carvajal Carmen R. , Valentino Leonard A. , Hakobyan Narine , Kazarian Tamara 
Published: October 2019

Abstract
Hemophilia is a congenital bleeding disorder that commonly results in musculoskeletal complications due to recurrent bleeding into joints. After an acute joint bleed, there is pain, swelling and limited motion due to the increase in intra-articular pressure and inflammation. Blood in the joint space provokes a proliferative and inflammatory disorder known as hemophilic synovitis (HS), which causes mechanical dysfunction and eventually destruction of the articular surface and bone. Among persons in the USA with severe hemophilia, nearly half have some restriction in activity (Evatt 2004) and it is estimated that 73% of boys enter adolescence with arthropathy and between 13–16 years of age, two or more major joints are affected (Brackmann,1992). Detail examination of the earliest changes in joint function following 1–3 joint hemorrhages is difficult to perform in patients. In this project, a murine model of HS (Valentino,2004) was used to examine the impact of a minimum number of hemarthrosis on joint function. The hypothesis tested was that there is a relationship between hemarthrosis and joint dysfunction. It was expected that joint dysfunction would be observed following few bleeds. Also, it was expected that a single hemarthrosis will result in permanent joint dysfunction. To test these hypotheses, 25 mice deficient in factor VIII coagulant activity were injured, times three, at weekly intervals to induce hemarthrosis then their ability to ambulate monitored using a Rotorod: Economex apparatus, which is intended for testing coordination and impairment of locomotor agility of laboratory animals. Mice were trained to ambulate on the rotating rod then injured and retested 2, 4 and 7 days after each hemarthrosis. A 2nd group of 25 mice was not injured and served as controls. At the conclusion of the experiment, mice were sacrificed and the knee joints examined by gross and histological methods. Correlations were drawn between the ambulatory dysfunction and the pathological changes observed. Preliminary data showed:
mice can be trained to ambulate on the Rotarod, acute hemarthrosis temporarily impairs their ability to ambulate, and after mice recover from acute injury they regain their lost ability to ambulate to the point that their performance is better than previous.
Isolated Alpha-2 Antiplasmin Deficiency Presenting as a Spontaneous Bleeding Disorder in a 63 Year Old Man.
Authors: Harish Vallathucherry C. , Lawson Heather L. , Zhang Lin , Huff Jason D. , Owen John 
Published: October 2019

Abstract
Spontaneous disruption of hemostasis in adults is a common clinical presentation. However, in a few patients, no cause is found. We were faced with such a situation when a 63 year old Caucasian male presented with a large spontaneous hematoma of his left thigh. There was no prior history of bleeding disorders in our patient or amongst his family members. Initial investigations did not show any conclusive coagulation factor abnormalities. Fibrinogen, D-dimer, liver function tests and platelet aggregation studies were normal. Subsequent tests showed that while his activatable plasminogen and euglobin clot lysis were normal, the function of his alpha-2 antiplasmin was at 35% of normal. Laurell immunoassay demonstrated a type 1 deficiency with antigen levels proportionately decreased at 30% of normal. Treatment with low dose epsilon amino caproic acid resulted in resolution of the hematoma and control of bleeding.
We therefore sought to determine the cause of his isolated alpha-2 antiplasmin deficiency. Alpha-2 antiplasmin is a serpin which targets plasmin. It is synthesized in the liver and deficiency results in a bleeding disorder.Congenital deficiencies of alpha-2 antiplasmin are uncommon bleeding disorders. However, since heterozygotic mutations of alpha-2 antiplasmin can present with initial bleeds late in life, we sequenced his alpha-2 antiplasmin gene. This gene is located on chromosome 17, spanning 16 Kb. It has 10 exons which we amplified by PCR in 13 segments and sequenced by the di-deoxy method. All splice junctions and the 70 base pairs upstream of exon 1 were normal. Three heterozygous polymorphisms were found, ruling out major gene deletions. The polymorphisms are as follows: Exon 2, C to T causing substitution of Alanine by Valine in the signal peptide, Exon 3, C to T causing substitution of Arginine by Tryptophan, Exon 10, G to A causing substitution of Arginine by Lysine. Each of these 3 polymorphisms have been found in blood donors.
This raises important issues about his deficiency. First, lack of production could be due to defective translational factors or epigenetic factors regulating gene transcription. It could also be due to the Alanine to Valine change in the signal peptide which we are currently investigating. Secondly, non-inhibitory antibodies to alpha-2 antiplasmin could explain his type 1 deficiency. Finally, age-related vascular and connective tissue defects may unmask a dormant inherited bleeding disorder. Unfortunately, his demise due to the development of nephrotic syndrome and sepsis may leave us without answers to these questions. There is no case linking nephrotic syndrome with the use of epsilon amino caproic acid.
In conclusion, since response to anti- fibrinolytics can be dramatic, deficiencies of alpha-2 antiplasmin must be considered in patients presenting at any age with a spontaneous bleeding disorder.
Hyperconcentrated (Dry) Versus Standard Platelet Apheresis: An In Vitro Quality Study.
Authors: Perotti Cesare , Bergamaschi Paola , Viarengo Gianluca , Del Fante Claudia , Bellotti Laura , Bressan Daniela , Balduini Carlo , Salvaneschi Laura 
Published: October 2019

Abstract
Introduction: Platelet concentrates (PC) collected by apheresis are effective in supporting deeply thrombocytopenic patients. The reduced risk of multiple allogeneic exposure and transmissible infectious diseases together with the high WBC depletion and diminished transfusion reactions are the main advantages offered by PC transfusion. At moment, the availability of several synthetic solutions for platelets storage permits to prepare hyperconcentrate(dry) apheresis platelets with the advantage of reducing febrile non-hemolytic transfusion reactions and, in low body weight patients the citrate toxicity, without the necessity of further manipulations. The aim of this study was to test the quality of 20 dry platelets (DP) in comparison to 20 standard plateletpheresis (SP) concentrates.
Materials and methods: A total of 40 apheresis procedures were performed by the single-needle Cobe Trima separation device (Gambro BCT, Lakewood, CO, USA) collecting either DP or SP concentrates. Within 1h after collection, the bag containing DP was added with the appropriate amount (70% of DP final volume) of synthetic solution for platelets storage (SSP, MacoPharma). Both DP and SP concentrates were stored at room temperature with gentle agitation for 4 days. For both concentrates, platelet yield was calculated and in vitro studies of membrane glycoproteins expression and aggregation at day +1 and day +4 were carried out.
Results: The comparison between 20 DP and 20 SP concentrates in terms of ability to aggregate in vitro and membrane glycoproteins expression at day +1 and day +4 of storage is reported in table A and B respectively.
Conclusions: The in vitro tests documented a major activation of dry platelets. In particular, the ability to aggregate was reduced in the 20 DP concentrates analised and this phenomenon was more evident at day +4 of storage. The alteration of membrane glycoproteins expression (markers of storage lesion) confirms the lower in vitro quality of DP concentrates. The effectiveness of this new blood component in vivo should be evaluated in a controlled clinical trial.
Table A. At collection Day +1 Day +4 SP DP SP DP SP DP Collagen μg/ml 4 93 88 97 82 80 53 ADP 10 μM 32 11 24 16 16 5 Ristocetin 1.5 mg/ml 91 97 77 81 71 52 Collagen 10 μg/ml + Adrenaline 10 μM 98 95 93 94 93 80 ADP 10 μM + Adrenaline 10 μM 87 72 76 61 57 30 Table B. At collection Day +1 Day +4 SP DP SP DP SP DP GPIb alfa (MFI) 5.06 5.75 6.31 6.13 5.26 4.54 GPIIb-IIIa (MFI) 35.47 36.71 34.61 37.7 31.76 40.9 GP IV (MFI) 11.49 11.4 11.67 10.28 11.65 11.38 GP 53 24.23 27.11 21.74 25.91 21.46 31.84 GMP 140 21.79 29.29 22.65 30.38 20.58 34.89
Safety of Central Venous Catheter Removal in the Setting of Thrombocytopenia and Peripheral Blood Stem Cell Collection.
Authors: Mantha Simon , Wu Yanyun 
Published: October 2019

Abstract
Thrombocytopenia is not uncommonly seen in patients undergoing peripheral blood stem cell collection. It is most likely secondary to the administration of chemotherapy and G-CSF, as well as the apheresis process itself. Peripheral blood stem cell collection causes a variable loss of platelets, which are collected along with the mononuclear cells. The ensuing thrombocytopenia can be at times severe, but the bleeding risk in this setting is unclear, as is the need to administer platelets prophylactically when the central catheter used for collection is removed. In this study we retrospectively analyzed data on 40 patients who had a central venous line removed immediately after their last autologous stem cell collection and had a platelet count of less than 50000/μL before the apheresis procedure. Patients were divided into two groups, according to whether or not they had received a platelet transfusion. The expected pre-catheter removal platelet count was calculated, according to the patient’s blood volume, the number of platelets in the stem cell product and the amount of platelets transfused in random donor platelet units (RDP) equivalents. Each chart was reviewed for the occurrence of catheter removal-associated bleeding. The results are shown in table 1. Post-apheresis platelet measurement is not routinely performed, due to its inconvenience to the patient and delay in catheter removal. Most clinicians use the pre-apheresis value as a guide to transfuse platelets before taking out the central line. As shown in table 1, in spite of significantly lower measured platelet counts before the harvest procedure in the transfused group, the calculated final counts were similar, due to the transfusion of allogeneic platelets. The patients’ weight was significantly lower in the transfused group, and this might actually have contributed to their lower platelet counts after repeated peripheral stem cell collections. There was no bleeding associated with catheter removal in either group. One patient had persistent oozing at the catheter insertion site, likely secondary to an unusually large incision. Removal was uneventful however. In this study, removing the central venous catheter was performed safely without platelet transfusion in patients who had a pre-apheresis platelet count above 30000/μL. It might be safer to administer prophylactic platelet transfusions to patients with lower counts.
Table 1 Untransfused group Transfused group P N 34 6 Patient weight in kg (median, range) 93 (52–125) 76 (63–89) 0.03 Measured platelet count pre-apheresis X 1000/μL (median, range) 40 (28–49) 24 (19–30) <0.01 Number of platelets removed X 1011 (median, range) 0.51 (0.05–1.44) 0.48 (0.14–2.34) 0.58 Number of transfused RDP equivalents per patient (median, range) 0 5 (4–6) - Calculated platelet count post-apheresis (median, range) 32 (19–44) 39 (8–45) 0.56 Number of bleeding episodes 0 0 -
Role of TPO in the Differentiation of Embryonic Stem Cells into Hematopoetic Cells.
Authors: Wang Zheng , Andri Pramono , Julia Skokowa , Karl Welte 
Published: October 2019

Abstract
Thrombopoetin (TPO) is a primary regulator of megakaryocyte and platelet production. However, studies in c-mpl-deficient mice and in congenital amegakaryocytic thrombocytopenia-patients with non-sense c-mpl mutation who develop pancytopenia during the first years of life suggest that TPO also play an important role on early hematopoesis. We demonstrated that TPO enhances FLK-1 (VEGF-receptor) expression on hemangioblasts during murine embryonic stem cell differentiation in embryoid body-liquid cultures (up to 73%). To extend our studies, we investigated the TPO signaling in FLK-1 positive cells. ES cells at different time point of differentiation showed that TPO enhances c-mpl-, BMP4-, Notch-, HOXB4-, HOXB9-, HOXA10-, Runx1-and CD133- mRNA expression. To investigate mesoderm formation, we also analyzed GATA-4 and T-brachyury mRNA level expression. Interestingly, we found that TPO alone did not increase GATA-4- and T-brachyury- mRNA expression, suggesting that TPO requires other cytokines to form the mesoderm. We also found that TPO could maintain VEGF-A mRNA expression level during differentiation of ES-cells. We hypothesize that VEGF expression together with c-mpl expression is required in hematopoetic differentiation of ES cell. This activity of Tpo was also observed during Rhesus monkey embryonic stem cell differentiation into hematopoetic cell. Only combinations of TPO and VEGF were capable of increasing CD34 positive hematopoietic progenitor cells (up to 8%), but TPO alone failed to induce high levels of CD34+ cell. In addition, analysis of gene expression during hemangioblast development demonstrated that TPO was capable of increasing the expression of VEGF receptors (FLK-1) and TPO receptors (c-mpl) in mice and primates. The in-vitro differentiation of mouse and rhesus monkey ES cells provides an opportunity to better understand the role of TPO in the early stage of hematopoietic development from ES cells to mature hematopoietic cells.
Pharmacokinetics and Pharmacodynamics of CNTO 528, a Novel Erythropoietin Receptor Agonist in Normal and Anemic Rats.
Authors: Bugelski Peter , Nesspor T. , Spinka-Doms T. , Makropoulos D. , Eirikis E. , Fisher J. , Volk A. , Shamberger K. , James I. , Fisher P. , Pool C. , Jang H. , Miller B. , Huang C. , Heavner G. , Knight D. , Ghrayeb J. , Scallon B. 
Published: October 2019

Abstract
CNTO 528 is a member of a group of novel stimulants of erythropoiesis that has recently entered clinical trials. Although CNTO 528 bears no sequence homology to erythropoietin (EPO), it is an EPO receptor agonist, capable of rescuing EPO-dependent cells from apoptosis in vitro and stimulating erythropoiesis in vivo. The purpose of these experiments was to explore the pharmacodynamics and pharmacokinetics of CNTO 528 in normal rats and to evaluate its efficacy in rat models of anemia. In vitro, CNTO 528 was approximately 10-fold less potent on a molar basis than recombinant human EPO (rhEPO) in stimulating the growth of UT-7EPO cells. Despite the lower in vitro potency, when compared to rhEPO and darbepoetin in normal rats, a single subcutaneous dose of CNTO 528 caused a longer-lived reticulocytosis and a longer-lived increase in hemoglobin. Also, CNTO 528 caused only minor changes in red cell distribution width (RDW) or mean cell volume (MCV) led to the release of mature reticulocytes and had no effect on mean platelet volume (MPV). We have also shown that CNTO 528 was efficacious in rat models of anemia and in a rat model of pure red cell aplasia. Taken together, our data show that CNTO 528 is a novel stimulant of erythropoiesis in rats.
Pegfilgrastim and Daily Granulocyte Colony-Stimulating Factor (G-CSF) Patterns of Use and Neutropenia-Related Outcomes in Cancer Patients in Spain: Results of the Learn Study.
Authors: Almenar D. , Mayans J. , Juan O. , Bueno J.M. García , Jalón Ji , Frau A. , Guinot M. , Cerezuela P. , Buscallà E. García , Sánchez J. ,   On Behalf of the LEARN Study Group
Published: October 2019

Abstract
Background: Daily G-CSFs(Filgrastim, lenograstim) are widely used to reduce duration of chemotherapy-induced neutropenia(CIN) and incidence of febrile neutropenia(FN) in cancer patients(pts). Data on their patterns of use and effectiveness in routine clinical practice are however limited. The introduction of the once-per-cycle G-CSF pegfilgrastim(Neulasta®) in Spain in 2003 may have changed patterns of use of daily G-CSFs and CIN-related outcomes.
Methods: Multicentre, retrospective, observational study of daily G-CSF and pegfilgrastim patterns of use and outcomes in adult subjects with non-myeloid malignancies receiving myelosuppressive chemotherapy(CT). Consecutive patient medical records with documented use of daily G-CSF or pegfilgrastim were abstracted from 10 Spanish centers during 2003.
Endpoints: percentage of proactive(primary prophylaxis) vs reactive(secondary prophylaxis/treatment) use of G-CSFs, duration of treatment with G-CSF, and CIN-related outcomes(dose delay, dose reduction, incidence of FN, hospitalization and antibiotic consumption).
Results: 248 charts documented pegfilgrastim or daily G-CSF use; 75 pts received pegfilgrastim only; 111 pts received daily G-CSF only(99 Filgrastim, 12 lenograstim); 62 pts received both daily G-CSF and pegfilgrastim during their CT(data not shown). Most common tumor types were lung(25%), breast(20%), malignant lymphomas(20%). Pattern of use (% pts on primary or secondary prophylaxis, or treatment at any time during CT) was:pegfilgrastim(39%,48%,17%, respectively) vs daily G-CSF(40%,48%,30%, respectively). Median number of injections/cycle in the daily G-CSF group was 6 (range 1–13) in primary prophylaxis, and 5 (range 1–11) in secondary prophylaxis and treatment. CIN-related outcomes are shown in the table below.
Conclusions: Patterns of use of daily G-CSFs and pegfilgrastim were similar for primary and secondary prophylaxis, but a potential trend to less frequent treatment use in the pegfilgrastim group was observed. CIN-related complications, including incidence of FN, were observed to be lower in pts receiving pegfilgrastim.
CT-related complications% pts (95% CI) Pegfilgrastim(n=75) Daily G-CSF(n=111) Dose Delay 44% (33; 55) 46% (36; 55) Dose Reduction 14.7% (8.2; 24.6) 20.7% (14.2; 29.2) Dose Reduction due to Neutropenia 6.7% (2.5; 15.0) 20.7% (14.1; 29.2) Febrile Neutropenia (FN) 10.7% (5.3; 19.9) 24.3% (17.2; 33.1) Hospitalization due to FN 9.3% (4.3; 18.3) 19.8% (13.4; 28.3) Antibiotic Consumption due to FN 8.0% (3.4; 16.7) 17.1% (11.2; 25.3)