bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequential Analysis of the α- and β-Globin Gene Expressions during Erythropoietic Differentiation from Primate ES Cells.
Authors: Katsutsugu Umeda , Heike Toshio , Shinoda Gen , Arai Masato , Ma Feng , Suemori Hirofumi , Luo Hong Yuan , Chui Devid H.K. , Torii Ryuzo , Shibuya Masabumi , Nakatsuji Norio , Nakahata Tatsutoshi 
Published: April 2020

Abstract
Globin switching of erythrocytes during primate (human and monkey) hematopoietic development has been most thoroughly investigated not only as a model of tissue- and temporally specific transcriptional control but also as a tool for drug discovery against hemoglobinopathies. However, the regulatory mechanisms of globin switching in primates remains to be unresolved in primates mainly due to lack of available model system to date which reproduces the process of hematopoiesis to reflect accurately in vivo development. Recently primate ES cell lines were established, which are expected to serve as an experimental model for tissue growth and development, along with an efficacy and toxicity screening system for new drugs and a cell source for regeneration therapy. Among them, we previously demonstrated that the transition from primitive into definitive erythropoiesis was induced from primate ES cells by coculture with OP9 stromal cells. We also demonstrated that the VEGFR-2high CD34+ cells, emerging onto OP9 stromal layer after initial 6-day differentiation, contain the hemogenic progenitors.
In this study, we quantitatively analyzed temporal pattern of embryonic, fetal, and adult globin expression in the α (ζ to α), and β (ε to γ and γ to β) cluster both at the transcriptional and translational level in erythrocytes induced from primate ES cells by the coculture system. When VEGFR-2high CD34+ cells were harvested and reseeded onto OP9 stromal cells in the presence of EPO, IL-3, SCF, and TPO, two waves of erythropoiesis occurred sequentially thereafter. Immunostaining and real-time reverse transcription-polymerase chain reaction analyses of floating mature erythrocytes revealed that the globin switches occurred in parallel with the erythropoietic transition. However, the temporal patterns of switching were asynchronous in the α- and β-globin genes; sudden fall of ζ-globin expression coincides increase of β-globin expression, while ε-globin expression gradually decreased and were still detected in many erythrocytes. Colony-forming assays demonstrated that the replacement of primitive clonogenic progenitor cells by definitive ones occurred during culture. The decline in embryonic ζ- and ε-globin expressions at the translational level occurred in a single definitive erythroid progenitor. The β-globin expression in individual definitive erythroid progenitors is up-regulated in the presence of OP9 stromal cells. Thus, this system reproduces early hematopoietic development in vitro and can serve as a model for analyzing the mechanisms of globin switch in human.
GATA-1, but Not GATA-2, Antagonizes PU.1 Mediated Transcriptional Activity at the CBFA2T3 (ETO2, MTG16) Promoter through a Mechanism Dependent on GATA DNA Binding.
Authors: Saunthararajah Yogenthiran , Yang SiJun , Kadkol ShriHari , Baraoidan Marie , Ibanez Vinzon , Mortazavi Fariborz 
Published: April 2020

Abstract
CBFA2T3 (ETO2, MTG16), a target of chromosomal translocation in acute myeloid leukemia, has its highest expression in hematopoietic cells compared to other tissues. This suggests that its expression is regulated by major hematopoietic transcription factors. The proximal promoter from −171 to −65 bp has greater than 90% identity between mouse and human and contains recognition sites for major hematopoietic transcription factors PU.1, GATA-1 and GATA-2. Using chromatin immuno-precipitation and the MPD hematopoietic cell-line, this segment was pulled down with endogenous PU.1, GATA-1 and GATA-2. In luciferase reporter gene assays, PU.1 and GATA-2, but not GATA-1, activated the promoter. As would be expected from these findings, CBFA2T3 levels declined during terminal erythroid differentiation of primary hematopoietic cells. GATA-1, but not GATA-2, antagonized PU.1 mediated activation but this effect of GATA-1 was abrogated by mutation of the GATA DNA binding sites. Both GATA-1 and GATA-2 have been reported to antagonize PU.1 transcriptional activity by antagonizing PU.1 interactions with c-Jun (Zhang et al, Proc Natl Acad Sci USA1999;96:8705–8710); however, the DNA binding dependent mechanism reported here allows GATA-2 and GATA-1 to have contrasting relationships with PU.1 and may be the basis for the co-operation of GATA proteins with PU.1 in some contexts yet antagonism of PU.1 activity in others.
Transcriptome Analysis of Human Hematopoietic Progenitor Cells during In Vitro Erythroid Differentiation.
Authors: Keller Margaret A. , Addya Sankar , Vadigepalli Rajanikanth , Ponte Christine M. , Banini Bubu , Delgrosso Kathleen , Surrey Saul 
Published: April 2020

Abstract
The hematopoietic progenitor cell (HPC) has the potential to differentiate into multiple cell types. Deciphering transcriptional regulatory networks necessary for HPCs to undergo lineage commitment is critical. Toward this goal, we established a serum-free in vitro differentiation system for human HPCs using CD34−positive cells isolated from peripheral blood of normal donors. Cells were cultured in the presence of Epo, SCF and IL-3 for two-weeks, and assessed for total hemoglobin and percent fetal hemoglobin. Adult-derived HPCs express large amounts of Hb A after approximately one week of culture with little expression of Hb F. Transcriptome analysis, using Affymetrix Human Genome U133 Plus 2.0 Arrays on RNA from days 1, 3, 5, 7, 9, 11 and 13 of culture shows more than 1500 differentially-expressed (DE)(&gt; 2-fold up or down) genes, a subset of which were validated independently by real-time RT-PCR. Up-regulated erythroid-specific genes included those for blood group antigens (Kell, Duffy, Rhesus), membrane proteins (ankryin 1, α- and β-spectrin, Band 4.1, Band 4.2, Band 4.9, glycophorin A), receptors (transferrin and erythropoietin receptors), heme biosynthesis enzymes (ALAS, ALAD, UROS, UROD, CPOX, PPOX, FECH), and transcription factors important in globin-gene expression (EKLF, BKLF, NFE2, GATA1) as well as α-, β- and γ-globin mRNAs. Down-regulated genes included those expressed in hematopoietic stem cells (GATA2, CD34) and non-erythroid hematopoietic lineages (MHC class I and II genes, CD33, CD53). The DE genes were clustered into a limited number of expression patterns, suggesting coordinate regulation of genes within each cluster. The DE gene set was examined in silico for co-regulation using transcriptional regulatory network analysis employing PAINT v3.2 software (Vadigepalli R et al., OMICS.2003; 7(3):235–52). GATA binding sites were represented on 149 of the DE gene promoters. Several transcriptional regulatory elements (TRE) were statistically significantly enriched in the DE gene set when compared to the array gene set. These include TRE for AP-1, CREB, GATA-1 and Oct-1. Co-regulation of genes in the different gene clusters will be analyzed by PAINT and findings validated by examination of TRE:transcription factor interactions. These studies should help define timing of transcription factor occupancy in relation to epigenetic changes occurring prior to transcriptome alteration during differentiation of human erythroid cells.
Pulmonary Mortality after High-Dose Oral Busulfan and Autologous Stem Cell Transplant (ASCT) in Patients with Non-Hodgkin Lymphoma (NHL).
Authors: Kalaycio Matt , Pohlman Brad , Rybicki Lisa , Sobecks Ronald , Kuczkowski Elizabeth , Andresen Steven , Brown Stacey , Bolwell Brian 
Published: April 2020

Abstract
Many high-dose chemotherapy preparative regimens include agents known to cause pulmonary toxicity such as BCNU. Chemotherapy induced pulmonary toxicity may be fatal. From 1/1/93 to 12/31/04 we treated 533 NHL patients with oral busulfan 14mg/kg, VP-16 60 mg/kg, and cyclophosphamide 120 mg/kg followed by ASCT. Busulfan levels were not measured. We have followed these patients for toxicity, survival, and cause of death (COD). Of these 533 patients, 214 have died, and 53 have died without relapse (15%). Most patients (n=329; 62%) had intermediate grade NHL by the IWF system. The median age was 50 years (range, 16–77) and 43% had a history of smoking. Radiotherapy was delivered to 27% and less than 3 chemotherapy regimens were administered to 75% of patients before ASCT. Sixteen patients (3%) died of non-relapse pulmonary complications. The median time to pulmonary mortality was 5 months (range, 2.5 – 21.4 months). We routinely screen patients with pulmonary function tests including DLCO before ASCT and most patients (n=490; 92%) were screened at a median of 45 days after ASCT (range, 7–90 days). For patients for whom data was available, there was no significant decrease in the median pre-ASCT and the post-ASCT DLCO in the either the whole cohort of patients or the patients dying of pulmonary complications. However, a lower baseline DLCO before ASCT predicted for pulmonary mortality after ASCT in univariable and multivariable analysis. Patients with a baseline DLCO &lt;94% predicted, had an increased risk of pulmonary mortality compared to patients with a baseline DLCO ≥94% predicted (p=0.016). Other significant risk factors included older age, lower baseline FEV1, prior radiotherapy, and longer time from diagnosis to ASCT. A history of smoking and disease status at the time of ASCT did not predict for pulmonary mortality. Because patients died of pulmonary complications at other institutions, we could neither determine treatment, nor confirm chemotherapy induced pulmonary toxicity. However, the early onset of pulmonary mortality after ASCT suggests the possibility of a toxic insult. Busulfan is a known pulmonary toxin, but the dose in this series of patients is relatively low and we have not seen similar pulmonary complications in 95 patients (median age 52 years) with myeloma treated with the combination of busulfan 16 mg/kg and cyclophosphamide 120mg/kg. Nonetheless, caution is warranted for older patients being treated with high-dose oral busulfan.
Does Highly Active Anti-Retroviral Therapy (HAART) Delay Myeloid Engraftment after Autologous Stem Cell Transplant (ASCT) in HIV Associated Lymphoma (HIV+Ly) Patients?.
Authors: Diez-Martin Jose L. , Carrion Rafael , Anguita Javier , Balsalobre Pascual , Miralles Pilar , Berenguer Juan , Buno Ismael , Gomez-Pineda Alfonso , Conde Eulogio , Varela Rosario , Ribera Jose M. , Espigado Ildefonso , Escoda Lourdes , Loscertales Javier , Serrano David 
Published: April 2020

Abstract
HAART has improved the outcome of AIDS patients (pts). Thus high-dose chemotherapy followed by ASCT has been used for the treatment of HIV+Ly pts, similarly to HIV-neg-Ly pts. In a recent report we found delayed engraftment after ASCT in HIV+Ly compared with HIV-neg Ly pts (Exp.Hem33;487–494,2005). To test the role played by HAART on engrafment we conducted this study, comparing engraftment results on HIV+Ly pts that received HAART with those in wich HAART was withdrawn after ASCT. Patients and Methods: From June 2000 a total of 19 HIV+Ly pts (17 male) received an ASCT in a multicenter cooperative study; 5 were Hodgkin disease (HD) and 14 non-Hodgkin high-grade malignant lymphoma (NHL). Median age was 43 years (range, 31–61); adjusted IPI (aIPI) was: 0–1 in ten pts and 2–3 in nine more pts; advanced Ann Arbor stage (III–IV) was present in 14 pts.Ten pts were treated with HAART during ASCT (HAART+) and in nine pts HAART was withdrawn due to gastrointestinal toxicity (HAART neg). Both groups were similar for the most relevant clinical characteristics (p value Mann-Whitney test or chi-square test &gt; 0.05): HD/NHL, Ann Arbor stage, aIPI, status at ASCT (CR-1/more than CR-1), age, number of lines of chemotherapy before ASCT, number of cycles of mobilization to obtain an adequate amount of CD34+ cells, number of CD34+ cells infused, day of start of GCSF and duration of GCSF-treatment. Results: Median time to reach PMN&gt;0.5 x 109/L was 12 days (9–17) for HAART neg group and 18 days (9–33) for HAART+ pts (p=0.069). Platelet engraftment (&gt;20x109/L) was achieved after a median of 20 days (11–28) and 26 days (11–455) respectively (p=0.25). Nevertheless, a multivariate regression analysis using as covariates HAART, GCSF (starting day) and amount of CD34+ cells infused, was performed. Accordingly no statisticaly significant results were found to relate with myeloid engraftment. Conclusions: Despite the HAART+ group shows longer interval to reach PMN and platelets engraftment following ASCT, we can not definitively conclude that HAART plays a negative influence on engraftment in this clinical setting. In order to clarify these results further recruitment of patients is needed.
Serial Analysis of Whole Blood Cell and CD3+ T-Lymphocyte Chimerism Following Allogeneic Stem Cell Transplantation with Alemtuzumab Containing Reduced-Intensity-Conditioning (RIC).
Authors: Tauro S. , Lovell R. , Begum G. , McMullan D. J. , Griffiths M. J. , Clark F. J. , Cook M. A. , Mahendra P. , Milligan D. W. , Craddock C. F. 
Published: April 2020

Abstract
Serial analysis of haemopoietic chimerism can be used to predict outcome after allogeneic SCT using a RIC regimen and guide post-transplant intervention. Although alemtuzumab is increasingly used as a component of RIC regimens, there have been few studies of its impact on chimerism status post-transplant. We have therefore measured chimerism following allogeneic SCT in whole blood/marrow nucleated cells (WB) and magnetically selected CD3+ T-lymphocytes in 64 patients with lymphoid (8 high-grade NHL, 18 low-grade NHL, 5 myeloma, 3 mantle cell lymphoma, 9 Hodgkin’s lymphoma) or myeloid malignancy (14 AML, 3 MDS, 1 myelofibrosis, 3 CML) conditioned with alemtuzumab and either fludarabine with melphalan (n=46), BEAM (carmustine, etoposide, cytarabine and melphalan) (n=16) or fludarabine/busulphan (n=2). Forty-seven patients received a transplant from an HLA compatible sibling donor and 17 from a matched unrelated donor. All patients achieved neutrophil and platelet engraftment. Donor chimerism was quantified within the first year post-allograft as well as following donor lymphocyte infusions (DLI) by FISH or PCR-based analysis of polymorphic microsatellite regions. 85% of patients demonstrated full donor chimerism (FDC, defined as ≥ 95% cells of donor origin) in WB within the first 90 days post-transplant. By contrast FDC was only present in the CD3+ compartment of 45% of patients. The proportion of patients with WB FDC declined to 64% by 12 months post transplant whilst the proportion of patients with CD3+ FDC remained constant. Thirteen patients received DLI using escalating CD3+ doses for management of mixed chimerism (MC) including 6 with evidence of disease relapse. Following DLI, 7 patients achieved FDC in WB and CD3+ compartments and 4 failed to switch to FDC. Seven patients developed acute GVHD post-DLI. Acquisition of FDC in the CD3+ compartment within 90 days post-transplant correlated with the presence of acute GVHD (p=0.003). Sixteen patients relapsed of whom 13 exhibited MC in WB or CD3+ compartments. Three patients relapsed despite the presence of FDC in WB and CD3+ cells. There was a trend towards improved disease free survival in patients who achieved FDC in WB within 90 days of transplantation compared to patients with MC (median 30 months v 11 months respectively). These data define a different pattern of WB and CD3+chimerism after alemtuzumab regimens compared with T-replete RIC regimens and confirm a correlation between chimerism status and outcome post-transplant.
Secondary Long-Term Prophylaxis in Severe Patients with von Willebrand’s Disease: An Italian Cohort Study.
Authors: Federici Augusto B. , Gianniello Francesca , Canciani Maria T. , Mannucci Pier M. 
Published: April 2020

Abstract
Background and Objectives. Patients with severe forms of von Willebrand’s disease (VWD) may have frequent episodes of mucocutaneous bleeding and also of hemarthrosis or hematomas. However, little retrospective or prospective data on secondary long term prophylaxis in VWD are available. Aim of this study was to evaluate the efficacy and safety of fixed regimens of prophylaxis with factor VIII/VWF concentrates in our cohort of VWD patients with recurrent joint and gastrointestinal (GI) bleeds. Design and Methods. This is a cohort study on 452 VWD patients regularly followed up at our Center for at least three years. 89/452 cases (20%) were treated with FVIII/VWF concentrates during the last two years because of one or more bleedings and 11/89 (12%) were included in a long term prophylaxis program because of frequent recurrence of bleeds at the same sites. All patients were characterized by a bleeding severity score derived from a detailed history of 11 symptoms. Since concentrates available in Italy are still labelled in FVIII IU, patients were given 40 FVIII IU/Kg of high- (Alphanate, Fanhdi) or intermediate-purity (Haemate-P) concentrates, two times a week (joint bleeds) or every other day (GI bleeds) to maintain FVIII/VWF levels higher than baseline during prophylaxis. Effectiveness of prophylaxis was based on resolution/reduction of bleeding as well as on numbers of transfused packed red blood cells (PRBC) and days of hospitalization. Safety was measured by monitoring side effects and FVIII levels before and after every injection during the first three weeks of prophylaxis. Results. All the 11 patients were severe, as shown by high bleeding scores (&gt; 20) and VWF:RCo baseline levels &lt;10 U/dL. Prophylaxis was started because of GI bleeds in 7 patients with VWD type 3 (n=1), 2A (n=4), 2M (n=1) and 1 (n=1) and for joint bleeds only in VWD type 3 (n= 4). Prophylaxis could stop bleeding in 8 patients and largely reduced hospitalization for PRBC transfusions in the remaining 3. When prophylaxis was compared with previous on demand regimen in all 11 cases, the annual total FVIII IU (x 1000) of concentrate (a) as well as number of PRBC used (b) and days of hospitalization (c) were significantly reduced (mean ± SD, with * p &lt; 0.01): a * = 239±207 versus 385±247; b* = 9±11 versus 3±4; c* = 18±13 versus 4±3. As far as safety, FVIII levels were always &lt;180 U/dL in all VWD and no side effects, including thrombosis, were observed. Interpretations and Conclusions. Secondary long-term prophylaxis by high-and intermediate purity FVIII/VWF concentrates is effective and safe in severe forms of VWD. Cost-effectiveness of these prophylaxis regimens versus on demand therapy should be further investigated in large prospective studies.
Antigen Levels of Thrombospondin-1 and ADAMTS13 Are Reduced in Parallel in TTP.
Authors: Liu Fang , Feys Hendrik B. , Dong Ningzheng , Bai Xia , Vanhoorelbeke Karen , Hans Deckmyn , Ruan Changgeng 
Published: April 2020

Abstract
Background Thrombospondin-1 (TSP-1) is considered to control the von Willebrand factor (VWF) multimer size by reducing interchain disulfide bridges, whereas ADAMTS13 is the major VWF cleaving protease. However the exact contribution of each of these to VWF size control is not known. In this study we investigate the relationship between ADAMTS13 antigen levels and TSP-1 levels in diseased plasmas.
Methods Antigen levels of TSP-1 and ADAMTS13 were determined in 21 plasma samples derived from 11 patients with TTP, and compared to normal control, 20 ITP samples and 20 SLE samples. TSP-1 antigen was measured by a commercial kit (Cytimmune, Rockville, MD), which can detect both bound and free TSP-1 in plasma. ADAMTS13 antigen was measured in a newly developed sandwich ELISA using 3 monoclonal antibodies to ADAMTS13. All donors were Chinese.
Results The average TSP-1 antigen level in normal plasma was 18.34±7.24 μ g/ml (mean ± SD, n=30) and the mean ADAMTS13 level in Chinese normals (CN) was 601±145 mU/ml (n=26) compared to the level set at 1 U/ml in pooled (n=20) normal human plasma from Caucasian origin. In TTP plasma, before plasma exchange (pre-PE), 5.79±2.35 μg/ml TSP-1 was detected (n=11, p&lt;0.01 versus CN), which upon PE increased to 9.06±6.52 μ g/ml (n=10, p&gt;0.05 versus pre-PE). The mean antigen level of ADAMTS13 was 98.7±82.8 mU/ml (n=11, p&lt;0.01, versus CN) and 449±232 mU/ml (n=10, p&lt;0.05 versus pre-PE), before and after PE, respectively. Furthermore TSP-1 antigen levels and ADAMTS13 titres in these patients plasma correlated linearly with an R=0.7 (n=21). To determine whether this significant decrease in both TSP-1 and ADAMTS13 antigen levels is restricted to TTP, plasma from ITP and SLE patients was analyzed next, however no significant decrease of TSP-1 was found. In contrast, the average antigen level of ADAMTS13 was raised to 830±229 mU/ml (n=20, p&lt;0.01 versus CN) and 721±192 mU/ml (n=20, p&lt;0.05 versus CN), for ITP and SLE plasmas, respectively.
Conclusion Both ADAMTS13 and TSP-1 antigen levels are significantly reduced in TTP plasma and hence the combined effect may be responsible for the presence of ultra-long VWF multimers in TTP plasma. Interestingly ADAMTS13 antigen level appears to be increased in SLE/or ITP parients, in contrast to previous reports that showed reduced activity, whereas no changes in TSP-1-levels were found in those patients.
Pharmacokinetics and Dose Linearity of von Willebrand Factor /Factor VIII Concentrate (Humate-P®) in Elective Surgery in von Willebrand Disease Patients.
Authors: DiPaola J. , Gill J. , Bernstein J. , Leissinger C. , Valentino L. , Friedman C. , Knaub S. 
Published: April 2020

Abstract
The VWF/FVIII concentrate Humate-P® (ZLB Behring) was studied in a prospective, uncontrolled, open-label clinical trial in VWD patients undergoing elective surgery. An interim analysis was performed after 24 patients (18 female, 6 male) with the following types of VWD had been included into the study: type 1 (n=11), type 2A (n=2), type 2B (n=4), type 2M (n=2) and type 3 (n=5). All patients underwent the pre-operative PK assessment to individualize the loading and maintenance doses of Humate-P® and 18 patients had completed surgery. The primary objective of the study - to demonstrate safety and hemostatic efficacy - was achieved in 100% of the patients. All patients received at least two infusions: a single infusion of 60 IU VWF:RCo per kg b.w. for a pharmacokinetic (PK) assessment prior to surgery and a PK guided loading dose before start of surgery. In addition, all patients received more infusions for further treatment after surgery. PK results: VWF:RCo: The overall median IVR was 2.1 [% per IU/kg] (range 1.1–4.2). The overall median response was 75.3% (range 37.0–169.4). Type 3 VWD patients had a slightly lower median response of 73.4% (range 63.7–96.1). Median value of Cmax was 142 IU/dL (range 84–330). Median effective half-life was 6.7 h (range 0.2–74.9), median clearance 3.6 [mL/kg/h] (range 1.0–16.6). FVIII:C: The median IVR was 2.8 [% per IU/kg] (range 1.4–4.9). Type 3 VWD patients had a median IVR of 3.4 [% per IU/kg] (range 1.7–4.9). The median response was 96.4% (range 63.0–164.8). Type 3 VWD patients had a median response of 104.5% (range 63.0–155.3). Dose-linearity: Graphical presentation of the relation between IVR and dose administered demonstrated the dose linearity of the VWF concentrate within a wide dose range.
IVR of VWF:RCo versus the amount of VWF:RCo administered (all doses) IVR of VWF:RCo versus the amount of VWF:RCo administered (all doses)
Statistical modeling using all doses (loading dose and maintenance infusions) supported this finding. Using the log-transformed IVR plotted against the dose administered the slope was 0.0004 (95% confidence interval: −0.0045 to 0.0053. Conclusion: The PK results are comparable to PK data from other clinical studies with Humate-P®/Haemate P® and given the interindividual variability underline the importance of pharmacokinetic studies prior to surgical interventions. Dose linearity of VWF:RCo with Humate-P® has been shown over a wide range of dosages (10–135 IU/kg b.w.). These results increase our confidence in the treatment of VWD patients with Humate-P®.