bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Role of Rho/Rac Pathways in Proplatelet Formation of Megakaryocytes.
Authors: Murai Kazunori , Kowata Shugo , Ito Shigeki , Sato Mamiko , Ishida Yoji 
Published: October 2019

Abstract
We reported that Rho and/or Rac signal pathways were involved in proplatelet formation (PPF) of megakaryocytes in the last ASH Meeting. Recently Nagata et al. reported that 3β-hydroxysteroid dehydrogenase (3β-HSD) was a target of p45 NF-E2 and that estradiole, synthesized by 3β-HSD in megakaryocytes, stimulated PPF (GENES & DEVELOPMENT 2003). In this study we focused on the signal pathways, activated by estradiole, and the predominant pathway in the process of PPF using murine megakaryocytes. Murine megakaryocytes were isolated from bone marrow cells by Percoll® density centrifugation and BSA gradient method. The purity of the isolated megakaryocytes was more than 40% in all experiments.
Isolated megakaryocytes (300/well) were cultured in serum free condition with 17β-estradiol (E2) or/and Y27632 (Rho kinase inhibitor) or PP1(Src family kinase inhibitor) or LY294002 (PI3K inhibitor). Y27632 inhibits Rho pathway and the 2 latters inhibits Rac activation. Y27632 alone stimulated PPF of murne megakaryocytes (0 μM:100%, 1μM:116.2±0.2%,10μM:129.2±4.25% (p<0.01),100μM: 159.8±13.3% (p<0.01))(n=4). E2 alone stimulated PPF of murne megakaryocytes in the dose-dependent manner (0μM:100%,1μM:108.1±4.2%,10μM:118.7±9.5%(p<0.05),100μM: 123.7±3.7% (p<0.01))(n=4). While, PP1 or LY294002 alone inhibited PPF of murine megakaryocytes (PP1; 0μM: 100%, 2μM: 82.1±7.3%(p<0.05), 20μM: 36.2±4.5%(p<0.01), LY294002; 0μM: 100%, 1μM: 94.8±2.3%, 10μM:66.5±4.7%(p<0.01),100μM:43.9±4.2%(p<0.01))(n=4). Based on these results, Rho pathway inhibition or activated Rac stimulated PPF. The addition of E2 to Y27632 resulted in no inhibition/stimulation of PPF (E2/Y27632; 0 nM/10 μM: 100%, 2.3 nM/10 μM: 113±3.5%, 23 nM/10 μM 116.9±9.0%, 230 nM/10 μM: 115.8±4.6 %, 0 nM/100 μM:100%, 2.3 nM/100 μM: 97.2±6.9%, 23 nM/100 μM: 93.1±5.1%, 230 nM/100 μM: 99.4±5.7%)(n=4).The addition of E2 to PP1 or LY294002 resulted in the dose-dependent manner reduction of PPF (E2/PP1; 230 nM/0 μM: 100%, 230 nM/2 μM: 61.9±8.5%, 230 nM/20 μM: 51.0±4.1%, E2/LY294002; 23 nM/0μM: 100%, 23 nM/10μM: 66.8±2.0%, 23 nM/100μM: 68.4±3.3%)(n=4). The degree in reduction with E2/PP1 or E2/LY294002 was similar to that with PP1 or LY294002 alone.These results strongly suggested that that PPF of megakaryocytes, stimulated by E2, might not be involved in Rho but Rac pathway.
Next, we investigated the predominance between Rho and Rac pathways in the process of PPF.The addition of PP1 or LY294002 to Y27632 resulted in no inhibition of PPF (PP1/Y27632; 0μM/100 μM:100%, 2μM /100 μM:105.6±6.0%, 20 μM/100 μM: 107.5±4.7, LY294002/Y27632; 0μM/100 μM:100%, 1μM/100 mμM:92.5±13.8%, 10μM /100 μM:92.6±6.6%, 100μM/100 μM: 97.8±8.0%, ( n=4)).
These results indicated that Rho pathway inhibition stimulated PPF of megakaryocytes, even if activated Rac decreased. Therfore, Rho pathway is more predominant than Rac pathway in PPF of megakaryocytes.
Angiogenic Cytokines: Vascular Endothelial Growth Factor and Basic Fibroblast Growth Factor Levels in Serum of Children with Hematopoietic Malignancies and Solid Tumors.
Authors: Palgan Izabela , Wysocki Mariusz , Palgan Krzysztof , Debski Robert , Styczynski Jan 
Published: October 2019

Abstract
Angiogenesis is a process of blood vessel formation from the preexisting capillaries. It plays an essential role of growth and development of cancer. The most important angiogenic stimulators are Vascular Endothelial Growth Factor (VEGF) and basic Fibroblast Growth Factor (bFGF). The aim of the study was to evaluate the serum levels of VEGF and bFGF in children with solid tumors, hematological malignancies and in healthy controls. Patients: 110 children (33 with solid tumors and 77 with hematological malignancies, including 46 with acute lymphoblastic leukemia and 17 with acute myeloblastic leukemia) (56 male and 54 female) aged 2–18 years (median 8.5 years) and 70 healthy controls (31 male and 39 female) aged 3–18 years (median 13 years). For the quantitative determination the Quantikine VEGF and bFGF immunoassay, a solid-phase enzyme-linked immunosorbent assay method was used. Elevated VEGF and bFGF were defined as being higher than the 95 percentile value in control group. Results: At the time of diagnosis the range of VEGF serum levels in all cancer patiens was 0–2691.3 pg/ml, and in remission 6.08–967.67 pg/ml; while the range of bFGF at diagnosis was 0–64.48 pg/ml, and in remission 0–32.52 pg/ml. In all children with cancer, serum levels of VEGF and bFGF were significantly more often elevated than in healthy controls (p<0.004; p<0.0005). At diagnosis, almost 12% patients had simultaneously elevated serum levels of VEGF and bFGF. Among patients with solid tumors at the time of diagnosis, elevated levels of VEGF and bFGF was observed in 42% and 60%, respectively. There were no differences between serum levels of these factors in patients in remission and in control group. In patients with progression, serum levels of VEGF and bFGF were higher, when compared to levels of these factors at diagnosis. The range of serum levels of VEGF in children with solid tumors at the time of diagnosis was 85.96–2691.3 pg/ml (median 365.48 pg/ml), and in remission 6.08–787.16 pg/ml (median 204.1 pg/ml). The median level of VEGF at diagnosis in children with hematopoietic malignancies (range 0–1869.6 pg/ml, median 111.6 pg/ml) was lower than in children with solid tumors (p<0.0005). In children with acute leukemias at diagnosis, serum levels of VEGF were significantly lower than in children with lymphomas (p<0.0003) and in controls (p<0.05). These results suggest that evaluation of serum levels of VEGF and bFGF can be helpful for monitoring activity of cancer, particularly in solid tumors and in the future can be a target of anti-angiogenic therapy.
Microgranular Variant (M3V) of Acute Promyelocytic Leukemia (APL) Does Not Have a Worse Prognosis than Classical APL in the Atra Era: A Report of 153 Patients Treated on Intergroup 0129 and Pethema LPA96 and LPA99.
Authors: Tallman Martin S. , Kim Haesook T. , Schiffer Charles A. , Appelbaum Frederick R. , Feusner James H. , Ogden Angela K. , Shepherd Lois , Willman Cheryl L. , Bloomfield Clara D. , Rowe Jacob M. , Larson Richard A. , Wiernik Peter H. , Martin Guillermo , Rayon Chelo , de la Serna Javier , Rivas Concha , Gonzalez-San Miguel Jose D. , Deben Guillermo , Sanz Miguel A. 
Published: October 2019

Abstract
The microgranular variant (M3V) accounts for 15–20% of patients (pts) with acute promyelocytic leukemia (APL). Historically, it has been associated with a higher white blood cell count (WBC) at diagnosis and an inferior outcome compared to classical morphology. Pts with APL treated with ATRA plus chemotherapy strategies have a 3–5 year DFS of 70–80%. The prognosis of pts with M3V treated with ATRA-containing chemotherapy strategies has not been adequately studied. This analysis included 153 patients with M3V entered on the North American Intergroup protocol 0129 (INT0129), a randomized phase III study of either ATRA (INT, ATRA)or daunorubicin/cytarabine for induction (INT, DA) and ATRA maintenance or observation after intensive consolidation or PETHEMA protocols LPA96 and LPA99 in which all pts received ATRA plus idarubicin for induction, intensive consolidation and ATRA plus low-dose chemotherapy for maintenance (PETHEMA, ATRA). The median age was 39 years (range 3–79), the median WBC was 14.5 K/μL (range 0.4–550), and the median platelet count was 25 K/μL (range 1.8 – 207.0). The median follow-up time for pts on INT0129 is 9.7 years, and for pts on PETHEMA LPA96 and LPA99 is 3.2 years. The 3-year disease-free (DFS) and overall survival (OS) analyses are shown in the table below.
N Median (in mo) (95% CI) DFS p-value N Median (in mo) (95% CI) OS p-value *comparison between ATRA (INT+PETHEMA) and DA; **comparison among WBC groups. NE: not estimable or not reached in this analysis     INT+PETHEMA, ATRA 106 NE 77% <0.0001* 128 NE 75% <0.0001*     INT, ATRA 19 NE 63% 24 NE 71%     PETHEMA, ATRA 87 NE 81% 104 NE 76% INT, DA 20 8.3(6.7, 12.6) 15% 25 19.4(10.4, 41.8) 32% INT+PETHEMA, ATRA     0<WBC<10K 47 NE 89% 0.01** 52 NE 90% 0.07**     10K≤WBC<50K 37 NE 78% 48 NE 69%     WBC≥50K 22 NE 54% 28 NE 58% INT, DA     0<WBC<10K 8 8.4(3.0, 24.6) 13% 0.94** 10 19.5(8.6, 41.8) 30% 0.32**     10K≤WBC<50K 6 9.2(5.2, 12.6) 17% 8 56.6(10.3, NE) 50%     WBC≥50K 6 7.9(7.4, 15.9) 17% 7 13.6(6.8, 22.2) 14%
In a Cox regression model, when age, gender, WBC, platelet, Hgb, ATRA treatment (ATRA vs DA), and source of pts (INT vs. PETHEMA) are included, only ATRA induction treatment (HR=0.16, p-value=0.0005) appears to be a significant prognostic factor for DFS, and age, WBC, and ATRA induction treatment (HR=0.38, p-value=0.03) for OS. These data suggest that the outcome for pts with APL M3V is excellent, influenced by the WBC at diagnosis and is not different than that of pts with classical morphology when treated with ATRA-containing strategies.
Study on the Relations between HLA-Class II Antigens and Idiophatic Thrombocitopenic Purpura in Adults.
Authors: de Paz Raquel , Cano Pedro , de la Concha Emilio Gomez , Montes Jose Antonio Rodriguez , Hernandez-Navarro Fernando 
Published: October 2019

Abstract
Background: The mechanism of autoimmunity of chronic idiopathic autoimmune thrombocytopenia (cAITP) is not fully understood but may involve binding of antigenic peptides to HLA antigens. We studied MHC class II phenotypes in adult patients with cAITP.
Methods: We have typed 64 adult Caucasian (Spanish) patients with cAITP (17 male and 59 female; median age 47 years range 17–77 years), and compared with the data from to control groups with geographic and ethnic differences. 200 Spanish control samples and 617 American control samples were analyzed. Patients with cAITP were evaluated according to their response to treatment with steroids, splenectomy, dapsona and Anti-CD20. According to the clinical response to the therapy we were able to form two subgroups; patients with good response, normal platelet count during therapy/after splenectomy and poor, less than 50x109/L or no response. Splenectomy was performed in 19 patients and resulted in a bad response in 11 patients of them. 25 of these patients showed a good and 39 a poor response. Class II typing was performed using polymerase chain reaction (PCR) with sequence specific primers.
Results: The comparison of antigen frequencies of the whole group of patients with cAITP and the two groups of healthy controls (Spanish and American samples) revealed significant increase of the HLA-DR12 and HLA-DR10 alleles of the MHC class II especifities (p<0.05). Moreover, the HLA-DR2 and HLA-DR4 phenotype was significantly decreased in the subgroup of patients with good response to treatment (p<0.05), with a high incidence in the subgroup of patients with bad response to all treatment, including progression to splenectomy and refractory to new treatments like Anti-CD20. No other significance was found.
Conclusions: Our data showed that DRB12 and DRB10 were more prevalent in ITP patients than in the two different control groups. Moreover, the difference in the genetic distribution of HLA-DR2 and HLA-DR4 in patients with good and poor response to treatment may indicate that ITP is a more heterogeneous condition than previously suspected, and this genetic marker may improve the early management of patients by better predicting prognosis.
Activation of the Extrinsic Coagulation Pathway in Inflammatory Exudates from Acute Lung Injury: Influence of Hydration on Factor Xa Generation and Inhibition.
Authors: McGee Maria P. , Hite Robert D. , Morris Peter E. 
Published: October 2019

Abstract
The exudative phase of ALI (acute lung injury) is characterized by loss of the endothelial/alveolar barrier, disruption of local water homeostasis, and intra-alveolar activation of blood coagulation. In previous studies, ALI exudates sampled by BAL (bronchoalveolar lavage) demonstrated increased tissue factor and antiproteinase activities compared to BAL from healthy lungs. This study applied osmotic stress techniques to explore the hypothesis that local water activity influences the procoagulant and antiproteinase activities recovered in human BAL fluids. Samples were prepared from 8 injured and 7 normal lungs by sequentially centrifuging BAL fluid at 200 and 40,000g to remove cells and surfactant particles, respectively. Factor Xa generation and inhibition were measured by chromogenic assay under standard (buffer pH 7.4, 0.13 NaCl, 1.3 mM CaCl2, 0.5mg/ml ovalbumin) and osmotic stress (~0.6 atm in standard buffer with polyethylene glycol 8000, ~26 Å radius) conditions by adding either purified factor X (200 nM) or factor Xa (50 nM) to the samples. Factor Xa generation curves had a sigmoidal shape, and the yields in 25 min-long reactions were variable, as expected, and ranged from 4 to 47 nM. Average activity values were 16-fold higher in ALI than in normal lungs, while tissue factor antigen levels measured by ELISA were 6-fold higher than in normal and ranged from 3.5 to 108 ng/ml. Under osmotic stress, the yields and rates increased in samples from healthy lungs, while the responses in ALI samples were mixed. Four showed decreases; 2 increases and 2 showed no change. Antiproteinase capacity, measured as the decrease in factor Xa activity, was variable but higher in ALI samples, and in both ALI and normal lungs, it increased in response to osmotic stress. These results imply that local hydration and dehydration directly influence procoagulant/anticoagulant activities in bronchoalveolar exudates, regardless of other factors, and that impaired alveolar fluid clearance may contribute directly to coagulation imbalances in ALI.
Gene Expression Profiles Distinguish Children Who Develop Therapy-Related Myeloid Leukemia.
Authors: Bogni Alessia , Cheng Cheng , Liu Wei , Yang Wenjian , French Deborah , Mukatira Suraj , Downing James R. , Pui Ching-Hon , Relling Mary 
Published: October 2019

Abstract
In children with acute lymphoblastic leukemia (ALL), failure due to therapy-related myeloid leukemia (t-ML) is a devastating complication. Using a target gene approach, only a few host genetic risk factors for t-ML have been defined. Microarray analysis of gene expression allows for a more genome-wide approach to identify possible genetic risk factors for t-ML. We assessed gene expression profiles (12625 gene probe sets) using oligonucleotide-based arrays in diagnostic ALL blasts from 228 children treated on St. Jude ALL protocols (Total XIII) that included etoposide; 13 of these children developed t-ML. A group of 83 probe sets were significantly related to the time-dependent risk of t-ML, with principal component analysis plot (right panel) separating patients who developed t-ML from the others. Hierarchical clustering of the 83 probe sets grouped patients into 3 clusters (n=163, n=52, n=13), with the cumulative incidence of t-ML being significantly higher in the last cluster (p < 0.0001, left panel) compared to those of the other gene-expression-defined clusters.
Figure Figure
A permutation test indicated that probe sets selected by chance are unlikely to obtain the observed distinct clusters (p=0.045). Distinguishing genes included transcription-related oncogenes (v-Myb, Pax-5), cyclins (CCNG1, CCNG2 and CCND1) and Histone H4. Common transcription factor recognition elements among similarly up- or down-regulated genes included several involved in hematopoietic differentiation or leukemogenesis (Maz, PU.1, FOXO4). This approach has identified several genes whose expression differentiates patients at risk of t-ML, and provides targets for assessing the germline predisposition to leukemogenesis.
Haemophilia A Genotype and Inhibitor Risk: Preliminary Results of a French Haemophilia Centre.
Authors: Borel-Derlon Annie , Slaoui Mounia , Costa Jean Marc , Lavergne Jean Maurice 
Published: October 2019

Abstract
Caen Haemophilia centre monitored 343 haemophilia A patients, 62 of them with a severe defect. We report here the preliminary results of genotypes and factor VIII (FVIII) inhibitors response for the first 59 and particularly for 15 patients with FVIII inhibitors. For severe haemophilia A, the first step is a long PCR for intron 22 inversion, then PCR for intron 1 inversion. The second step is a direct sequencing of the FVIII gene in severe form without inversion and in moderate or mild disease. 41% (16 / 39) of the patients with severe haemophilia A have an intron 22 inversion, the remainder patients have single point mutations (17), principally on exon 14, large deletion (1), small deletion (3) or splice mutations (3). In the severe form, 10 of these patients have developed inhibitors: 6 with intron 22 inversion, 3 non sense mutations on exons 14 or 18, 1 large deletion on exon 14 and 1 splice mutation on intron 22. In the moderate or mild form, 20 patients have missense mutations, 5 of them have developed inhibitors. The genotype of three patients has been previously reported to be associated with a high incidence of inhibitors in moderate and mild form: Arg 593 Cys by loss of tolerance to exogenous and endogenous FVIII (1), Arg 2150 His by reducing the binding of FVIII to von willebrand factor (2). For the other two patients, we identified two missense mutations: Val 2017 Ala, Tyr 1786 Ser. We are processing genotypes for each haemophilia A patients to evaluate the genotype - phenotype relationship and the FVIII inhibitor risk related to genetic and environmental factors.
Functional Vanilloid Receptor VR1 Is Expressed on Megakaryocytes and Platelets, and Acts as a Sensor for Hypertheimia Generating Endocannabinoids.
Authors: Maruyama Ikuro , Soejima Kentaro , Shimi Toshiaki , Abeyama Kazuhiro , Hashiguchi Teruto 
Published: October 2019

Abstract
Recent investigations have been revealing that platelet can sense shear stress and cold stimulation. These mechanical and physical stimuli alter the platelet functions, including induction of activation and irreversible dysfunction. These data may suggest that platelets, most small-sized cells, can efficiently circulate in the microcirculatory vasculature, and can sense the peripheral physical conditions including temperature.
Here we show that human megakaryocytic cell line, MEG-01 and platelets express vanilloid receptor VR1, a receptor for capsaicin. It has been identified that VR1 is linked to thermal sensor, and this can sense hyperthermia around 43 °. Therefore we examined the physiological significance of VR1 expressed on MEG-01 and platelets.
First we identified the expression of VR1 on human platelets and MEG-01 by immunoblotting, flowcytometer and RT-PCR, and identified the expression of VR1 on these cells. Hyperthermia stimulation of MEG-01 at 40–43 ° generated a endocannabinoids, 2-arachidonylglycerol(2-AG) and anandamide. Since this effect of hyperthermia stimulation was abolished by the incubation with capsazepine, an antagonistic analogue of capsaicin, it was suggested that the effect of hyperthermia might through VR1. It is well known that 2-AG and anandamide act on their specific receptors, CB1, CB2 and VR1 expressed on various cell-types, and act as an endocannabinoids, and accomplish diverse physiological functions, including vessel dilatation. Hyperthermia stimulation also affected the cell-cycle and induced G1-phase arrest in MEG-01 cells, partially through autocrine manner of generated endocannabinoids by acting of CB1 and CB2 receptors. Moreover prolonged hyperthermia stimulation up to 6 hours induced cellular apoptosis in MEG-01 by releasing cytocrome C and resulting caspase cascade activation.
Based on these results, we hypothesize that VR1 on platelets may act as a hyperthermal sensor and generate endocannabinoid, and this may regulate the diverse systems including vessel functions.
Clinical Outcomes of Recombinant Activated Factor VII (rVIIa) Use in a Tertiary Care Hospital: 70 Treatment Courses in 63 Patients.
Authors: DeLoughery Thomas G. , Boshkov Lynn K. , Schreiber Martin A. , Thorburg Per 
Published: October 2019

Abstract
rVIIa was originally developed and released for use in hemophilia patients with inhibitors. Recently the suggested uses for rVIIa have expanded dramatically. However, most of the data on the novel uses of rVIIa have been published as case reports or limited series. We reviewed our inpatient experience with rVIIa over the past 4 years. Diagnoses included CNS bleeding (12 of 14 on warfarin), inherited coagulation defects (2 patients with Glanzmann’s thrombasthenia (GT), one with factor VII deficiency and one with XI deficiency), inhibitors (1 hemophilia A, 2 hemophilia B, and 8 acquired factor VIII). “Last ditch” included 4 patients bleeding from severe GVHD after stem cell transplant, 3 cases of refractory sepsis, bleeding due to pulmonary hypertension, and CNS bleed due to tumor), surgery (4 of 7 cardiac) and trauma. 6 patients (2 inhibitor, 1 surgery, and 3 inherited) received more than one course of rVIIa. The average dose was 90 ug/kg. Overall 42 patients (60%) were alive at discharge with the congenital coagulation deficient and acquired inhibitor patients having 100% survival. Patients with liver disease or those patients given rVIIa as a “last resort” had the worst survivals despite being effective for controlling bleeding in most patients. rVIIa is effective for patients with GT, factor deficiencies, or inhibitors and its use is associated with good outcomes. In patients with bleeding due to their underlying medical problems, rVIIa was modestly effective at controlling bleeding. However, their outcomes were determined by the survivability of their disease process. For effective use of rVIIa patient selection appears to be crucial.
Diagnosis Number % Alive at D/C Effective% Avg # Doses Given CNS Bleed 14 57% 85% 1.2 Inherited Defects 7 100% 100% 10 Inhibitor 16 100% 93% 19 “Last Ditch” 9 0 66% 1.3 Liver Disease 6 16% 50% 1.6 Surgery 7 85% 100% 1.5 Trauma 13 46% 92% 1.3