bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a Novel Alternatively Spliced Form of AML1-ETO and Analysis of Its Role in Leukemogenesis.
Authors: Kanbe Eiki , Wang Yang , Boyapati Anita , Yan Ming , Peterson Luke F. , Willman Cheryl L. , Rowley Janet D. , Zhang Don-Er 
Published: October 2019

Abstract
The 8;21(q22;q22) translocation is found in 12% of all AML cases, including 40–50% of AML M2 subtype and a small portion of M0, M1, and M4 subtypes. The translocation brings together the DNA sequence encoding the N-terminus of the AML1 (RUNX1) with nearly all of ETO (MTG8) to generate the AML1-ETO fusion protein. The most commonly known AML1-ETO fusion protein (full length AML1-ETO) is encoded by AML1 exons 1 to 4 and by ETO exons 2 to 11 and has 752 a.a. However, a recently discovered ETO exon 9a can provide a protein translation stop codon after the last amino acid encoded by exon 8. The predicted AML1-ETO protein using exon 9a (AML1-ETO9a) has 575 a.a. and does not include the NHR3 and NHR4 domains at the C-terminus of full length AML1-ETO. To demonstrate the presence of the AML1-ETO9a transcript, we performed RT-PCR using RNA prepared from t(8;21) patient samples. Exon 9a transcript was detected in all patient samples when PCR generated a 595 bp fragment with ETO exon 6 and exon 9 primers and was detected in seven of the 14 samples when PCR generated a 1381 bp fragment with AML1 exon 4 and ETO exon 9a primers, indicating the presence of AML1-ETO9a mRNA in patient samples. The predicted molecular weight of AML1-ETO9a protein with 575 a.a. is 63.8 kDa and a 64 kDa AML1-ETO fusion protein has been reported in both Kasumi-1 cells and t(8;21) leukemic blasts. We compared the effects of AML1-ETO and AML1-ETO9a on proliferation and differentiation of hematopoietic cells using FDCPmix cell line, which is an IL-3 dependent multipotent hematopoietic stem cell line. Both forms of fusion proteins inhibited the myeloid lineage differentiation when cells were cultured in the presence of GM-CSF and G-CSF. However, these fusion proteins showed opposite effects on cell cycle progression. AML1-ETO suppressed cell cycle with prolonged G0/G1; AML1-ETO9a enhanced cell cycle with shortened G0/G1. Expression of these fusion proteins in primary bone marrow cells showed a similar result. Most importantly, only AML1-ETO9a efficiently induced acute myeloid leukemia in mice. Thus, the observation that a shorter form of AML1-ETO is a dominant factor in t(8;21) related leukemogenesis exhibits a new paradigm in the understanding of t(8;21) tumorigenesis.
A FVII Deficiency Induces Cardiac Fibrosis in Mice Resulting in Systolic and Diastolic Dysfunction.
Authors: Xu Haifeng , Sandoval-Cooper Mayra , Donahue Deborah L. , Ploplis Victoria A. , Castellino Francis J. 
Published: October 2019

Abstract
Mice severely deficient in coagulation Factor (F)VII that survive to adulthood have been developed, thus allowing the study of spontaneous bleeding and inflammatory phenotypes under conditions of a deficiency of extrinsic coagulation, especially in organs e.g., the heart, that continually undergo normal mechanical challenge. Mice genetically modified to produce very low levels (∼1% of wild-type) of coagulation FVII (FVIItTA/tTA) spontaneously developed heart abnormalities that were revealed by echocardiography. There were no significant differences between groups at baseline for the hemodynamic and respiratory parameters: temperature and heart rate (HR). The end-systolic functions of FVIItTA/tTA mice heart were comparable to wild-type (WT) controls. However, there were significant changes in the end-diastolic parameters of FVIItTA/tTA hearts. The diameter at end-diastole in FVIItTA/tTA mice was smaller compared to WT mice, which was due to the stiffness of the interventricular wall. When viewed from the parasternal short-axis, these mice had evidence of hypokinesis, indicating alteration of the elastic properties of the left ventricular (LV) wall. Correspondingly, the stroke volume, ejection fraction, fractional shortening, and cardiac output were lower in FVIItTA/tTA mice, compared to WT mice. Mitral valve inflow was compromised in FVIItTA/tTA mice, as demonstrated by the reduced early/late (E/A) filling ratio. We found that the spontaneous impaired cardiac function in FVIItTA/tTA mice was associated with an increased ratio of heart weight to body weight. Decreases in ventricular size, accompanied by reductions in systolic and diastolic functions, suggest a restrictive cardiomyopathy, consistent with an infiltrative myopathic process. Microscopic analysis of mouse hearts showed severe patchy fibrosis in FVIItTA/tTA mice. Hemosiderin deposition in hearts of these mice, along with large increases in inflammatory cells, ultimately resulting in widespread collagen deposition, were found. Significant increases in mRNA levels of TGF-beta, TNF-alpha, and a variety of matrix metalloproteinases, beginning at early ages in FVIItTA/tTA mice, supported the fibrotic pathology. The results of this study demonstrated that hemorrhagic and inflammatory responses to a severe FVII deficiency resulted in the development of cardiac fibrosis, manifest echocardiographically as a restrictive cardiomyopathy, with significantly compromised ventricular diastolic and systolic functions. These data also suggest that a FVII deficiency differentially regulates the expression level of inflammation and extracellular matrix proteins, which result in compromised cardiac function.
Apoptosis Stimulating Protein of p53 (ASPP2) Haploinsufficiency Promotes Irradiation-Induced High Grade Lymphomas in a Mouse Model.
Authors: Kampa Kerstin M. , Akoba Jared D. , Chen Dexi , Beemer Kelly , Gay Joel , Zhu Zhiyi , Padiernos Emerson , Corless Christopher , Baily Alexis , Flemming William H. , Lopez Charles D. 
Published: October 2019

Abstract
Apoptosis Stimulating Protein of p53 (ASPP2) binds the tumor suppressor protein p53 and promotes damage-induced apoptosis in part through stimulation of p53 apoptotic function. We have previously demonstrated that low ASPP2 levels correlate with poor clinical outcome in patients with diffuse large B-cell lymphoma treated with anthracycline-based chemotherapy, which is consistent with its role as a tumor suppressor. We targeted the ASPP2 allele in a mouse model by homologous recombination using a knockout vector that replaced exons 10–17 with a neoR gene. Although ASPP2−/− is an early embryonic lethal event, ASPP2+/– mice appeared developmentally normal and reproduce. Importantly, ASPP2+/– mice developed tumors spontaneously at a higher frequency than sibling ASPP2+/+ controls. We now show that after ionizing irradiation, ASPP2+/– mice develop high grade lymphatic malignancies compared to ASPP2+/+ mice. ASPP2+/+ and +/– mice were irradiated, with either 6 Gy or 10,5 Gy, 8 weeks after birth. Following irradiation, mice were monitored weekly by blood smears and flow cytometry if suspicious cells were observed. Tumor formation was detected as early as 17 weeks post-irradiation. In the 10,5 Gy group, ASPP2+/– mice developed thymic lymphomas (7/31), while no tumors were detected in ASPP2+/+ mice (0/14). Tumor-free survival was statistically significant (p= 0,045, log-rank test). In the 6 Gy dose group 9 out of 28 of the ASPP2+/– mice developed high-grade thymic lymphomas compared to 0/20 ASPP2+/+ mice. Tumor-free survival was statistically significant (p=0,024, log-rank test). Immunophenotyping and flow cytometry on mice with circulating tumor cells from whole unfractionated gated viable bone marrow (Bm) cells revealed that the Bm was typically almost completely replaced by CD4neg, CD5+, CD8+ leukemic/lymphoma cells that are negative for B-cell (B220) and myeloid markers (Mac-1/Gr-1). Preliminary analysis demonstrated that MEFs (mouse embryonic fibroblasts) from ASPP2+/– mice treated with doxorubicin, serum starvation or irradiation had an attenuated G1/G0 checkpoint arrest. Ongoing experiments are being conducted in thymocytes prepared from mice of different ASPP2 and p53 genotypes. Our data demonstrates that ASPP2 plays an important role in suppressing the development of lymphomas after irradiation in a mouse model. Consequently, disruption of the p53-ASPP2 axis may be important in lymphomagenesis and response to therapy. Furthermore, our data provides a rationale for the future study of ASPP2 expression in patients during the formation of secondary hematologic malignancies after therapies with DNA damaging agents.
Rivaroxaban — An Oral, Direct Factor Xa Inhibitor — Prevents Arterial Thrombotic Occlusion in Electrolytically Injured Rat Carotid Arteries.
Authors: Haertlein Barbara , Parry Tom J. , Chen Cailin , Perzborn Elizabeth , Andrade-Gordon Patricia , Damiano Bruce P. 
Published: October 2019

Introduction: Rivaroxaban is an oral, direct Factor Xa (FXa) inhibitor in advanced clinical development for the prevention and treatment of thromboembolic disorders, including arterial indications such as acute coronary syndromes (ACS). The aim of this study was to determine whether rivaroxaban can prevent arterial thrombotic occlusion in electrolytically injured rat carotid arteries.
Methods: Anesthetized, male Sprague-Dawley rats were equipped with an intravenous (i.v.) catheter to administer drug or vehicle and an intra-arterial catheter for arterial pressure monitoring and blood sampling. The left carotid artery was exposed and a bipolar electrode placed proximally around it. A pulsed Doppler flow probe was then placed distally around the vessel. Blood was obtained before drug or vehicle administration to determine activated clotting time (ACT), prothrombin time (PT), Russell’s viper venom time (RVVT), activated partial thromboplastin time (aPTT) and thrombin-antithrombin (TAT) complex concentration; baseline carotid blood flow was recorded for 20 minutes. Vehicle (polyethylene glycol:ethanol:water or saline, 0.5 mL/kg/min), rivaroxaban (0.3, 1 and 3 mg/kg; dosing volume 0.5 mL/kg/min), or the low molecular weight heparin enoxaparin (10 mg/kg) were infused i.v. over 30 seconds. After infusion, direct current (3 mA) was applied to the left carotid artery for 5 minutes. Blood flow was monitored for 30 minutes after injury. Blood was sampled again 14.5 and 30 minutes after injury for ACT determination, and after 30 minutes for PT, RVVT, TAT and aPTT determination.
Results: Rivaroxaban dose-dependently increased the median time to arterial thrombotic occlusion (TTO), and was significantly more effective than vehicle at doses of 1 and 3 mg/kg (vehicle, 13.2 minutes; rivaroxaban 1 and 3 mg/kg, >30 minutes; P<0.05 by Kaplan-Meier analysis). Compared with enoxaparin (10 mg/kg), rivaroxaban inhibited TTO at doses of 1 and 3 mg/kg more effectively. Rivaroxaban (0.3–3 mg/kg i.v.) dose-dependently prolonged ACT, PT and RVVT, and substantially decreased the formation of TAT complexes, but did not substantially affect aPTT. Enoxaparin prolonged ACT, aPTT and RVVT, and decreased TAT complex formation, but had no effect on PT.
Conclusions: Rivaroxaban potently inhibited electrolytically induced arterial thrombotic occlusion in rat carotid arteries, resulting in substantial changes in coagulation parameters. The coagulation parameter changes were consistent with inhibition of FXa activity as indexed by the RVVT. These findings suggest that rivaroxaban may be an effective anticoagulant in arterial thrombotic disorders, such as ACS.
Studies of the Affinity and Stoichiometry of Convulxin Binding to Glycoprotein VI: Clues into Its Potent Agonism.
Authors: Herr Andrew B. , Horii Katsunori 
Published: October 2019

Abstract
The primary cell-surface receptor that mediates platelet activation upon exposure to collagen is glycoprotein VI (GPVI). GPVI can be activated by fibrous collagen, soluble collagen mimics, or several snake venom proteins. Upon binding of these agonists, GPVI signaling activates key integrins and triggers secretion of platelet granule contents, leading in turn to activation of neighboring circulating platelets. One of the most potent GPVI agonists from snake venom is convulxin (CVX), an α4β4 C-type lectin-like protein from the South American rattlesnake, Crotalus durissus terrificus. Using analytical ultracentrifugation, we have discovered that CVX exists in solution as a stable dimer of α4β4 rings, creating eight potential binding sites for GPVI. Using a combination of kinetic and equilibrium surface plasmon resonance binding experiments, we show that CVX binds GPVI with high picomolar to low nanomolar affinity at each site, and that all eight sites are likely to be available for binding. Analysis of the previously published crystal structures of CVX suggests that its potency could be due to both high-affinity binding and clustering of GPVI on a single platelet as well as potential bridging of adjacent platelets via the dimeric CVX architecture.
Essential Roles for Mef2c in Lymphoid Commitment and B-Cell Function.
Authors: Stehling-Sun Sandra , Jimenez Rebecca , Hu Andrew , Camargo Fernando D. 
Published: October 2019

Abstract
MEF2 transcription factors are well-established regulators of muscle development. Recently, work in murine models has identified one of these factors, Mef2c, as an important regulator in the pathogenesis and the development of acute myeloid leukemia (AML). However, little is know about the molecular mechanism and physiological role of Mef2c in hematopoiesis. Using conditional gene ablation, we have discovered an unexpected role for MEF2c in hematopoietic stem cells (HSCs), where it is required for pan-lymphoid commitment. Competitive repopulation experiments using Mef2c-null HSCs deleted by means of the Mx1-Cre/poly(IC) approach, revealed completely normal monocytic, granulocytic and erythroid differentiation capacities by mutant cells. Generation and renewal of myeloid progenitors and HSCs was also normal. However, contribution to lymphoid lineages (T-cells, B-cells and natural killer cells) was dramatically reduced. Mef2c-deleted HSCs were able to generate lymphoid primed multipotent progenitors (LMPPs) and expressed normal levels of Flt-3 and the master lymphoid regulator ikaros. However, expression of the interleukin-7 receptor (IL-7R) and the number of phenotypically defined common lymphoid progenitors (CLPs) were substantially reduced. We have found two conserved Mef2c-binding sites in the promoter of the Il-7R gene, indicating that Mef2c could directly regulate Il-7R transcription. This and other potential molecular mechanisms of Mef2c-mediated lymphoid commitment will be discussed. We have also studied the effects of lineage-specific deletion of Mef2c in both myeloid and lymphoid populations. Whereas deletion in myelomonocytic cells using the LysM-Cre strain resulted in no anomalies, B-cell specific ablation with the CD19-Cre line revealed major phenotypical and functional abnormalities. CD19-Cre:Mef2cf/f mice show impaired germinal center formation and reduced antibody production in response to T-cell dependent antigens. In addition Mef2c-null mature B-cells fail to express the mature marker CD23, the low affinity receptor for IgE, which we show is a direct transcriptional target. As a consequence of CD23 reduction, CD19-Cre:Mef2cf/f mice have increased IgE production, thus indicating a potential role of Mef2c in allergic disease. Our work here sheds new light on the molecular mechanisms of lymphopoiesis and identifies MEF2 factors as critical hematopoietic transcriptional regulators.
Effect of Deferasirox on Midazolam Pharmacokinetics.
Authors: Arrowsmith-Bensasson Caroline , Rojkjaer Lisa , Sethuraman Venkat , Skerjanec Andrej 
Published: October 2019

Abstract
Introduction: Deferasirox (Exjade®, ICL670) is an oral iron chelator approved for the treatment of iron overload due to chronic transfusions in more than 80 countries. Deferasirox is mainly eliminated via biliary excretion following glucuronidation. Oxidative metabolism, presumably by CYP1A and CYP2D6 enzymes, accounts for approximately 6% and 2%, respectively. Deferasirox has been shown to be a weak inhibitor of CYP3A4/5 in vitro. It is possible that deferasirox could potentially alter the metabolism of other drugs eliminated via this pathway. Midazolam, a rapid-onset benzodiazepine, is almost exclusively metabolized via CYP3A4/5 making it an ideal probe to investigate the effect of an inhibitor/inducer of CYP3A4/5 in vivo. This drug-drug interaction study was undertaken to investigate the effect of deferasirox on midazolam pharmacokinetics (PK).
Methods: This was a single-center, open-label, one-sequence, cross-over study conducted in healthy volunteers. There were two treatment periods. During Period 1, volunteers received a midazolam 5 mg oral solution and midazolam PK were performed at 0–12 hours. During Period 2, volunteers received doses of deferasirox 30 mg/kg daily for 5 days and on Day 4, a midazolam 5 mg oral solution was given. Midazolam PK were performed at 0–48 hours.
Results: Twenty-two healthy volunteers were enrolled, and 21 completed the study. Mean age was 32 years, 59% of volunteers were male and all were Caucasian. Descriptive statistics for mean plasma concentration of midazolam and its metabolite, 1-hydroxymidazolam, are summarized below:
AUCinf (h.ng/mL) AUClast (h.ng/mL) Cmax (ng/mL) Tmax (h) T½ (h) Midazolam PK Midazolam alone, n=22 Mean (SD) 68.3 (28.4) 63.8 (26.4) 34.4 (17.4) 0.44 (0.17) 3.7 (0.9) Deferasirox + midazolam, n=22 Mean (SD) 61.7 (37.4) 60.3 (37.2) 27.1 (13.6) 0.47 (0.26) 4.4 (1.4) 1-hydroxymidazolam PK Midazolam alone, n=22 Mean (SD) 22.3 (8.8) 20.8 (7.9) 12.8 (6.7) 0.48 (0.15) 3.3 (1.5) Deferasirox + midazolam, n=22 Mean (SD) 25.6 (10.9) 23.8 (10.0) 11.7 (5.3) 0.5 (0.24) 6.1 (3.5)
There was a statistically, but not clinically, significant reduction (17%) in midazolam exposure (AUC) in the presence of deferasirox in comparison with midazolam administered alone. Similarly, Cmax was decreased on average by 23%.
Conclusion: An increase in midazolam exposure, which would have been expected for an inhibitory effect of deferasirox on the CYP3A4-mediated metabolic pathway, was not observed. A decrease in midazolam exposure is suggestive of an induction effect, albeit weak in nature. Drugs that are substrates for CYP3A4 can be co-administered with deferasirox if necessary.
Production of Inflammatory Mediators by Nanoparticle Stimulated Macrophages.
Authors: Hall David J. , Ufkin Melanie , Frear Meghan E. 
Published: October 2019

Abstract
Nanoparticles are receiving attention because of their many potential applications in drug delivery. The ability to treat humans with Polymethylmethacrylate (PMMA) nanospheres and gold nanoparticles is dependent on the biocompatibility of the nanoparticles. One measure of biocompatibility is the interaction between monocyte derived macrophages (MDM) and nanoparticles. Macrophages initiate many immune reactions with a variety of mechanisms, one such reaction is the release of nitric oxide (NO). Our research shows that RAW 264.7 (murine) and THP-1(human) macrophage cell lines, when treated with gold or PMMA nanoparticles, show the production of nitric oxide and degradation of IKB-α, but not reactive oxygen species. In this study, we used primary human MDMs as a more relevant model system. We report that gold nanoparticles and PMMA nanospheres did not cause the production of the same amounts of NO as their cell line counterparts. Western blot analysis showed the degradation of IKB-α during a 60-minute time course in primary human MDM. These studies suggest that the safety concerns over nanoparticle use are not supported for primary human MDM.
Increased Expression of TLR2 and TLR4 in Mononuclear Cells in Patient with Immune Thrombocytopenic Purpura.
Authors: Cheng Yunfeng , Zou Shanhua , Li Feng 
Published: October 2019

Abstract
Immune thrombocytopenic purpura (ITP) is an autoimmune disease characterized by platelet destruction resulting from autoantibodies against self-antigens and T-cell mediated cytotoxicity. Toll-like receptors (TLRs) are pattern recognition receptors important in mediating the immune response and their activation can lead to production of cytokines. Recent data suggest that TLR2 and TLR4 are crucial for the production of inflammatory cytokines and play central role in autoimmune diseases, yet little is known about their roles in ITP. Here we examined the gene expressions of TLR2 and TLR4 in ITP patients. We hypothesize that significant differences will exist between pre-treatment and post-treatment in ITP patients with similar changes reflected in the plasma concentration of cytokines. Total RNA was extracted from mononuclear cells obtained from 12 ITP patients and 15 healthy subjects. TLR2 and TLR4 mRNA expression levels were analyzed using a quantitative real-time PCR method and their protein expressions were validated by western blot. Plasma concentrations of cytokines IL-2, IFN-γ and TNF-α were measured by ELISA. Correlation analyses were carried out between the mRNA expression levels of TLR2 or TLR4 and the plasma levels of IL-2, IFN-γ and TNF-α. The gene expression of TLR2 and TLR4 were significantly increased in ITP patients comparing to healthy control group (p < 0.05 and p < 0.01, respectively). In addition their mRNA expression levels were decreased back into normal range after remission in 8 patients (p > 0.05, compared to healthy control group). Significantly positive correlations were found between the TLR2 mRNA expression level and the plasma concentration of IFN-γ or TNF-α (R = 0.75, p < 0.05; R = 0.83, p < 0.05, respectively). Changes in the gene expression of TLR4 and in the plasma concentration of IFN-γ or TNF-α were also significantly correlated (R = 0.82, p < 0.05; R = 0.88, p < 0.05, respectively). Directional changes in TLR2 / TLR4 and IFN-γ /TNF-α expression were concordant. However, there was no correlation found between TLR2 / TLR4 and IL-2. Differences in TLR2 and TLR4 expression strongly correlated with changes in IFN-γ and TNF-α suggest that the increased gene expressions of TLR2 and TLR4 in ITP patients may contribute to the pathophysiological progression of this disease by increasing the secretion of IFN-γ and TNF-α. Additional studies need to be performed to further clarify the role of TLRs -cytokines pathway in ITP.
Membrane Microparticles from Patients with Steroid-Induced Avascular Necrosis of Femoral Head Increase the Expression of Fas in Endothelial Cells In Vitro.
Authors: Weng Xi-sheng , Wu Zhi-hong , Wei Yi , Qiu Gui-xing 
Published: October 2019

Abstract
High-dose or long-term usage of steroid have been suggested to be at great risk of developing avascular necrosis of femoral head (ANFH). A theory proposed to decipher the mechanism behind the development of steroid-induced ANFH involves vascular compromise and cell death. Membrane microparticles (MMPs) are fragments shed from plasma membrane blebs of virtually all cell types when submitted to a number of stress conditions, including apoptosis. It has been reported that high-dose dexamethasone cause dysfunction and apoptosis of endothelial cells; and MMPs isolated from the plasma of patients with myocardial infarction of preclampsia were found to cause damage in isolated arteries in vitro. We hypothesize that MMPs generated after high-dose or long-term administration of steroid facilitates the apoptosis of endothelial cells initiated by steroid, which contributes to the development of ANFH. MMPs were isolated from the blood of 4 healthy individuals, and 5 patients demonstrated to have ANFH by magnetic resonance imaging (MRI) and X-ray. The mean duration and accumulated dose of steroid administration were 8 months and 2000 mg respectively. The 3rd generation of human umbilical vein endothelial cells (HUVEC) was exposed to MMPs (corresponding to 0.2-fold circulating plasma level) prepared above. After 48 hours, part of cells was lysed to make total cell lysate. And the rest were used to prepare mRNA. The expression level of Fas was detected with reverse-transcript PCR and Western Blot. Our results shown that treatment of HUVEC with MMPs from ANFH patient’s blood significantly increases the transcription and expression of Fas, indicating that MMPs derived from patients with steroid-induced ANFH exacerbates high-dose steroid-induced apoptosis of endothelial cells by enhancing the expression of Fas.