bluuebunny/crossref_metadata_2025_split huggingface dataset:
Notch Signaling Induces Megakaryocytic Cell Fate.
Authors: Mercher Thomas , Cornejo Melanie , Sears Christopher , Kindler Thomas , Moore Sandra , Aster Jon , Gilliland D. Gary 
Published: April 2020

Abstract
The Notch pathway regulates a broad range of biological mechanisms including proliferation, border formation and cell fate decisions. In the hematopoietic system, Notch signaling is generally thought to specify T cell lineage fate at the expense of the B cell whereas its role in the myeloid lineage development is unclear. When using heterotypic co-cultures of murine primary hematopoietic stem cells (HSC: Lin-Sca1+Kit+) with OP9 stromal cells, or OP9 cells expressing the Notch ligand Delta1 (OP9-DL1), we unexpectedly observed the development of large cells with cytoplasmic protrusions reminiscent of proplatelet production by megakaryocytes on OP9-DL1 stroma. These cells stained positive for acetylcholinesterase, specific for megakaryocyte, and displayed large polylobated nuclei. Flow cytometric analysis indicated a 10-fold increase in the number of CD41+ cells in OP9-DL1 co-cultures compared to parental OP9 co-cultures. Expression of a constitutively active intra-cellular Notch (ICN) mutant allowed differentiation of HSC into CD41+ cells in parental OP9 co-cultures without DL1 stimulation, whereas expression of a dominant-negative MAML1 (dnMAML1) mutant abrogated this effect in OP9-DL1 co-cultures. In addition, megakaryocyte differentiation in OP9-DL1 co-cultures was blocked by γ-secretase inhibitors treatment and rescued by ectopic expression of ICN. Global gene expression analysis demonstrated engagement of a megakaryopoietic transcriptional program when HSC were co-cultured with OP9-DL1 vs. OP9 stroma or OP9-DL1 stroma treated with γ-secretase inhibitor. Bone marrow transplantation experiments with ICN, resulted in enhanced megakaryopoiesis in vivo with increased MEP numbers and megakaryocyte colony formation. Furthermore, transplantation of bone marrow cells transduced with dnMAML1 significantly impaired megakaryopoiesis in vivo with a 4- to 7-fold decrease in maturing megakaryocytes. These findings demonstrate a positive regulatory role for Notch signaling in specification of megakaryocyte development, and indicate that Notch plays a complex role in cell fate decisions among myeloid progenitors. They suggest the possibility that inhibition of Notch signaling may have therapeutic potential in malignancies of the megakaryocytic lineage. Furthermore, Notch pathway stimulation could be of value in enhancing megakaryocyte growth in clinical contexts associated with severe thrombocytopenia, such as hematopoietic reconstitution following bone marrow transplantation or chemotherapy.
Dyskeratosis Congenita: Low Regimen-Related Toxicity Following Hematopoietic Cell Transplantation (HCT) Using a Reduced Intensity Conditioning Regimen.
Authors: Tolar Jakub , Orchard Paul J. , Miller Jeffrey S. , MacMillan Margaret L. , Wagner John E. , Alter Blanche P. , Baker K.S. 
Published: April 2020

Abstract
Dyskeratosis congenita (DC) is a congenital disorder characterized by very short telomeres. Clinical presentation includes a diagnostic triad (lacey reticular pigmentation, nail dystrophy, and leukoplakia), aplastic anemia (the main cause of premature death), myelodysplastic syndrome/leukemia and solid tumors. Allogeneic HCT is the only curative option for the hematologic complications of DC but has been associated with a high risk of peri-transplant morbidity and early death. Only about fifty HCT for DC have been performed to-date, and the five year survival after related donor HCT has been about 75%, but only approximately 35% when an unrelated donor was used. To improve survival in DC patients by decreasing transplant mortality, we introduced a reduced intensity regimen including cyclophosphamide (50 mg/kg), fludarabine (200 mg/kg), low dose total body irradiation (200 cGy), and (in patients 3 and 4) campath 1H (1 mg/kg). To decrease the risk of graft rejection, grafts were not T-cell depleted. We report outcomes in four consecutive patients, two adults and two children, all of whom engrafted with donor hematopoiesis:
Age (years) Sex Graft and HLA match NC dose (×108/kg) CD34 dose (×108/kg) Follow-up (months) Donor chimerism 24 M URD dUCB 4/6, 4/6 0.55, 0.39 0.5, 0.43 1 (dead*) 83% 29 F REL PBSC 6/6 13.93 4.43 20 (alive) 100% 5 F URD BM 7/8 1.38 1.55 16 (alive) 100% 2 M URD BM 8/8 5.92 2.31 3 (alive) 100%
Legend: M, male; F, female; NC, nucleated cell; URD, unrelated donor; REL, related donor; BM, bone marrow; dUCB, double umbilical cord blood; PBSC, peripheral blood stem cells; *Patient had autologous recovery after the first dUCB and died of sepsis 1 month after the second dUCB; HLA matching is reported for antigen level HLA-A, B and allele level DRB1 for cord blood, and allele level typing for HLA-A, B, C, DRB1 for PBSC or BM. The most recent donor chimerism is reported. To decrease the risk of graft rejection and prevent graft versus host disease (GvHD) patients received cyclosporine and mycophenolate mofetil. Patient 2 developed limited chronic GvHD and patient 4 developed grade III skin acute GvHD. Both were treated successfully with systemic and topical steroids. Our data suggest that this conditioning regimen results in a low rate of transplant related complications without compromising engraftment. Critically, early fatal pulmonary and vascular complications, common in post-transplant courses in DC patients, were not observed. This highlights the need to avoid drugs that are associated with pulmonary toxicity such as busulfan, and to limit radiation to the lung in patients with DC. This new less intensive conditioning regimen appears to result in a low rate of transplant related complications, and yet has adequate immunosuppressive activity to permit engraftment from alternative donors in DC patients.
Systematic Genomic Screen for Tyrosine Kinase Mutations in CLL.
Authors: Brown Jennifer R. , Levine Ross , Raderschall Elke , Thompson Christina , Gilliland D. Gary , Freedman Arnold S. 
Published: April 2020

Abstract
Genomic screens have identified activating somatic mutations in tyrosine kinases in a spectrum of acute and chronic hematologic malignancies. These mutations lead to increased cellular proliferation and/or impairment of apoptosis. Chronic lymphocytic leukemia is a common chronic leukemia of adults that is thought to result at least in part from failure of apoptosis. Recent data suggest that CLL cell turnover is higher than previously thought and may correlate with disease progression, suggesting that somatic mutations that activate signal transduction may contribute to CLL pathogenesis and/or progression. The purpose of this study was to look systematically for activating mutations in protein tyrosine kinases in CLL using high-throughput DNA resequencing. 95 CLL patients were selected to encompass the range of prognostic groups defined by somatic hypermutation of the immunoglobulin heavy chain gene, ZAP-70 status and FISH cytogenetics. DNA was isolated from Ficoll separated peripheral blood mononuclear cells, and germline DNA was obtained from buccal swabs and/or saliva samples. 70 tyrosine kinases were selected for resequencing based on their involvement in lymphocyte biology, known altered gene expression in CLL, or location at a site of chromosomal gain, loss or mutation in CLL. All coding exons of ZAP-70 were included. Nested M13 tailed primers were designed to amplify and sequence the exons encoding the activation loops and JM domains. Bidirectional sequence analysis was performed using Mutation Surveyor version 2.28 (SoftGenetics). Candidate mutations were reamplified and sequenced from the original CLL DNA sample for verification; in parallel, the same exon was amplified and resequenced from the paired germline sample to assess whether non-synonymous mutations not known to be SNPs were present in the germline. A total of 178 exons were sequenced per sample. Analysis of 6.21 megabases of sequence data identified nine novel non-synonymous alterations in seven different genes; at least seven of these nine alterations are present in the germline, with the other two presently undergoing analysis. Although these non-synonymous alleles may have functional relevance to the pathogenesis of CLL, these data indicate that a significant proportion of non-synonymous alleles identified in high-throughput genomic screens are non-synonymous polymorphisms. These findings further suggest that the rate of acquired somatic mutation in the tyrosine kinome in CLL is low, and demonstrate the importance of matched germline DNA to allow assessment of candidate non-synonymous mutations in human cancer samples.
Prognostic Assessment of Three Single Nucleotide Polymorphisms (BCL2–938C&gt;A, MTHFR 677C&gt;T, GNAS1 T393C) in Chronic Lymphocytic Leukemia.
Authors: Zenz Thorsten , Duehrsen Ulrich , Dohner Harald , Siffert Winfried , Stilgenbauer Stephan , Nuckel Holger 
Published: April 2020

Abstract
A number of single nucleotide polymorphisms (SNP) have been implicated to impact on the disease course in chronic lymphocytic leukemia (CLL). Nonetheless, there have been few polymorphisms for which replications of association were done in different studies. We genotyped three recently described SNPs (BCL2 –938C&gt;A, MTHFR 677C&gt;T, GNAS1 T393C) in a large group of CLL patients (n=273) with mature follow up and detailed analysis of molecular genetics. The cohort had a median follow-up of over 150 months and the known poor risk molecular genetics (17p–, 11q– and unmutated VH-status) were associated with poor prognosis. The allele frequencies for all three SNPs were in Hardy Weinberg equilibrium and we could not identify imbalances when we stratified for subgroups (genomic aberrations, VH status, stage according to Binet). Moreover, the polymorphisms did not affect clinical outcome (time to first treatment and overall survival) in univariate or multivariate analysis in the current cohort. Comparing CLL patients with over 100 healthy controls we found no significant differences in allele frequencies or genotype distributions for all three polymorphisms. While there have been reports showing the molecular effects and clinical consequences of the polymorphisms we studied, we did not find SNP genotype-dependent effects in a large cohort of CLL patients. Potential reasons for the discrepancy of the results are numerous. Because of the advancement of genotyping technologies, a large number of SNPs can be evaluated (multiple testing). The sample sizes of the reported association studies are often too small and frequently only one patient cohort is investigated. Moreover, negative results are often not published (publication bias). One major reason for the different results is the different genetic background of the populations due to ethnicity (selection bias). Summarized, this study supports the importance of confirmation of initial findings in an independent data set. However, results of genome-wide association studies will soon be reported with additional novel genes identified and will help making great strides in identifying the genetic determinants of CLL risk.
Screening for Copy Number Alterations and Loss of Heterozygosity in Chronic Lymphocytic Leukemia - A Comparative Study of Four Differently Designed, High Resolution Microarray Platforms.
Authors: Gunnarsson Rebeqa , Staaf Johan , Jansson Mattias , Ottesen Anne Marie , Göransson Hanna , Liljedahl Ulrika , Ralfkiaer Ulrik , Mansouri Mahmoud , Smedby Karin Ekström , Hjalgrim Henrik , Syvänen Ann-Christine , Borg Åke , Isaksson Anders , Jurlander Jesper , Juliusson Gunnar , Rosenquist Richard 
Published: April 2020

Abstract
Screening for copy number alterations (CNA) has improved by applying genome wide microarrays, where SNP-arrays also allow analysis of loss of heterozygosity (LOH). Currently, comparisons of high resolution microarray platforms are few, thus we performed a study to evaluate the power of differently designed microarrays for copy number analysis and LOH. We here analyzed 10 diagnostic chronic lymphocytic leukemia (CLL) samples (five IGVH mutated and five IGVH unmutated) using four different high-resolution platforms: BAC-arrays (32K), oligonucleotide-arrays (185K, Agilent), and two SNP-arrays (250K, Affymetrix and 317K, Illumina). Comparison of copy number data showed that the platforms are concordant in terms of detecting large CNA, including the known recurrent alterations. Mono-allelic and bi-allelic loss of 13q14 (3 and 1 sample, respectively), mono-allelic loss of 11q (1 sample), trisomy 12 (2 samples) and mono-allelic loss of 17p (2 samples) were concordant in all platforms. These aberrations were validated with FISH, which in addition identified subclones with mono-allelic loss of 13q14 in two cases, only detected with the BAC platform, rendering a cut-off for the power of detecting subclones to approximately 25% of investigated cells. As expected, all poor prognostic aberrations were detected in patients carrying unmutated IGHV genes whereas four of five mutated samples were detected with mono-allelic loss of 13q14, as the only recurrent alteration. Furthermore, detection of small CNA were in many cases discordant between platforms. Therefore, we defined alterations identified by at least two platforms and identified 47 losses and 31 gains using this criterion. We are currently validating the presence of a number of these alterations using other techniques. Evaluation of LOH showed concordance for 86 regions between the Illumina and Affymetrix platforms. Of these regions 12 LOH coincided with CNA, leaving the remaining 74 as copy-neutral LOH. In conclusion, all platforms investigated are powerful tools for screening of CNA, however, since non-overlapping CNA were detected by individual platforms, we emphasize the importance of validating findings. Also, there is a cut-off for detecting subclones, here estimated to 25%. Genomic arrays will improve the detection of new recurrent aberrations, which may potentially refine the prognostic hierarchy established by FISH.
Clinical and Outcomes Findings for Thrombotic Thrombocytopenic Purpura among 467 Persons with Severely Versus Not Severely Deficient ADAMTS-13 Levels.
Authors: Bennett Charles L. , Luu Thanh Ha , Zakarija Anaadriana , Kwaan Hau C. , Bandarenko Nicholas , Pandey Dilip K. , Cursio John F. , McKoy June M. , Buffie Charlie , Winters Jeffrey L. , Ortel Thomas L. , Sarode Ravindra , Zheng X. Long , Raife Thomas J. , Matsumoto Masanori , Fujimura Yoshihiro , Rock Gail A. 
Published: April 2020

Abstract
Background: Thrombotic thrombocytopenic purpura (TTP) is a rare disorder that presents with microangiopathic hemolytic anemia and thrombocytopenia, fevers, renal insufficiency and neurologic features. We reviewed clinical, laboratory, and outcome data for TTP cases with severely deficient versus non-severely deficient ADAMTS13 activity levels.
Methods: Mean and median data were from the Surveillance, Epidemiology and Risk Factors for TTP (SERF-TTP) study group for idiopathic TTP cases, the Canadian Apheresis Group (CAG), and five published series (Zheng 2004, Raife 2004, Vesely 2003 (Oklahoma TTP-HUS Registry), Matsumoto 2004 (Japan Referral Center), Bennett 2007).
Results: Compared to TTP cases with near-normal ADAMTS13 activity levels (n= 282), TTP cases with severe ADAMTS13-deficiency (n=185) were more likely to have severe thrombocytopenia, normal renal function and neutralizing ADAMTS13 antibodies. Severe ADAMTS13 deficient TTP cases have better overall survival after therapeutic plasma exchange (TPE) but are more likely to relapse. TTP patients with severe ADAMTS13 deficiency were primarily categorized as idiopathic or ticlopidine-associated, while TTP patients with non-severely deficient ADAMTS13 activity levels were frequently categorized as idiopathic, secondary to drugs (clopidogrel, quinine), stem cell transplantation, or cancer.
Conclusions: Severe ADAMTS13 deficiency is most commonly idiopathic, has better survival following TPE, and a 35–40% spontaneous relapse rate. By contrast, non-ADAMTS13 deficient TTP cases are usually associated with an underlying disorder or external insults. Amongst this cohort, four series have 47–62% survival rates and three series, which contain mostly idiopathic cases, have 83–90% survival rates following TPE. From this, we propose that TTP may occur by three possible mechanism; ADAMTS13-deficient (antibody-mediated), an immunologic mediated pathway independent of ADAMTS13 (i.e. quinine) that is responsive to TPE, and endothelial injury related TTP that is unresponsive to TPE.
Platelet count mean (x10^9/L) Creatinine mean (mg/dl) ADAMTS13 neutralizing antibodies (%) Survival % Relapse % * &lt;15% ADAMTS13 activity cutoff Severe ADAMTS13 Deficiency (&lt;10–15%) SERF-TTP (n=30) 19 1.3 83 97 41 Zheng (n=16) 19 1.6 44 81 38 Bennett (n=26) 15 85 Oklahoma (n=18) 12 1.8 94 81 38 Raife (n=50) * 13 1.2 92 35 Japan (n=34) 35 91 Canada (n=11) 16 2.4 82 Not Severely Deficient ADAMTS13 Activity (&gt; 15%) SERF-TTP (n=22) 57 3.9 35 90 0 Raife (n=57) * 44 2.7 83 9 Canada (n=17) 57 4.1 88 Zheng (n=13) 40 3.0 0 54 Bennett (n=13) 62 Japan (n=66) 9 62 Oklahoma (n=94 ) 23 47 3
Effects of Serotonin on Proplatelet Formation and F-actin Reorganization in Human Megakaryocytes.
Authors: Cheng Yuan Shan , Liu Yuan Sheng , Chan Godfrey ChiFung , Ye Jie Yu , Lau Yu Lung , Yang Mo 
Published: April 2020

Abstract
We previously reported that serotonin (5-HT) is a growth factor for hematopoietic stem cells and megakaryocytic progenitor (Yang et al, Stem Cells, 2007). We further proposed a possible role of serotonin on megakaryocyte differentiation and platelet formation. The effect of serotonin on proplatelet formation and F-actin reorganization in human megakaryocytes (MKs) was investigated in this study. Our results showed that:
There was a stimulating effect of serotonin on proplatelet formation in human bone marrow megakaryocytes. Human BM MK progenitors cultured in serum free medium with either 5-HT (200nM) or TPO (50 ng/ml) had more proplatelet bearing MKs than the control group (5-HT(11.33% ± 4.93) vs. control (6% ± 3.60), P=0.026; TPO (14.66% ± 1.53) vs. control, P=0.043; n=3). The 5-HT treatment group showed more mature and more in the final stage MK cells as compared to TPO group; The effect of serotonin on proplatelet formation in Meg-01 cells were via 5-HT2 receptors. Meg-01 cells strongly expressed 5-HT 2A, 2B, 2C receptors by using western blot method. 5-HT also promoted proplatelet formation in these cells and this effect was reduced by 5-HT2 receptor inhibitor ketanserin (KE); and Serotonin acted on cytoskeleton reorganization in human megakaryocytes via 5-HT2 receptors and ERK1/2 pathway.
Using an immunofluorescence microscope with F-actin specific binder rhodamine-phalloidin staining, the polymerized actin level was lower in the control group (serum free) than the 5-HT group and actin distributed diffusely throughout the cytoplasm. In contrast, polymerization actin level was higher in 5-HT group. Adding ketanserin and ERK1/2 inhibitor PD98059 to 5-HT treatment, the fluorescence intensity was correspondingly reduced (5HT vs. Control, P=0.006; 5-HT vs.5-HT plus KE, P=0.014; n=6). Our data also demonstrated that ERK1/2 was activated in MK cells treated with 5-HT for 30 minutes (21.76% ± 7.42). Our studies showed that serotonin had a stimulating effect on proplatelet formation and F-actin reorganization in human megakaryocytes and this effect involved the 5-HT2 receptors and the activation of ERK1/2 pathway.
Differential Expression of ERG Isoforms in Acute Myeloid and T-Lymphoblastic Leukemia Is Regulated by DNA-Methylation.
Authors: Baldus Claudia D. , Schlee Cornelia , Thibaut Julia , Heesch Sandra , Bohne Arend , Thiel Eckhard , Hofmann Wolf K. 
Published: April 2020

Abstract
The oncogenic ETS transcription factor ERG is involved in various cellular pathways including developmental regulation, proliferation, and differentiation. In hematopoiesis ERG plays a specific role during normal T-cell differentiation showing high expression levels in stem cells and down regulation in the progenitor compartment. In this regard, it is intriguing that aberrant expression of ERG was found in a subset of patients with acute T-lymphoblastic leukemia (T-ALL) and was associated with an inferior outcome. Furthermore, high level ERG expression was of adverse prognostic significance in patients with newly diagnosed acute myeloid leukemia (AML), thus highlighting ERG’s potential role in myeloid as well as T-lineage leukemogenesis. ERG3 (NM_182918) and ERG2 (NM_004449) represent the main isoforms and show abundant expression in myeloid and lymphoid hematopoietic progenitor cells. The expression pattern of specific ERG isoforms in acute leukemias has yet to be investigated. To further elucidate the nature of aberrant ERG expression we have determined the existence and transcriptional regulation of ERG isoforms in pretreatment bone marrow samples of adult T-ALL (n=21) and AML (n=20) patients as well as in normal CD34+ hematopoietic cells of healthy volunteers (n=5). 5′RACE revealed the presence of a new ERG isoform (ERG3Δex12) characterized by expression of exon 5 and absence of exon 12. Expression of ERG3Δex12 was verified by RT-PCR in AML, T-ALL, and CD34+ cells. In addition, real-time RT-PCR showed concomitant expression of the two main isoforms ERG2 and ERG3 in AML and normal CD34+ cells. In contrast, T-ALL patients lacked expression of ERG isoforms harboring exon 4 (ERG2). Promoter analyses of ERG2 and ERG3 revealed the presence of two CpG islands in the ERG2 promoter region, whereas no CpG island was predicted in the ERG3 promoter. Bisulfit conversion of genomic DNA and sequencing of cloned PCR products revealed a significantly higher degree of methylation of CpG island 2 in T-ALL samples (median: 86.4%, range: 16.0 – 98.8%) as compared to AML (median: 38.1%, range: 10.9 – 60.7%; P-value=0.0002 - two sided T-test). As for CpG island 1, CD34+ cells had the lowest rate of methylation in CpG island 2 (median: 7.7%, range: 2.4 – 20.7%). Thus, the differential expression of ERG isoforms is mediated by epigenetic silencing of exon 4 containing transcripts in T-ALL. In conclusion, the identification of the new ERG isoform (ERG3Δex12) suggests the association with different partners as the central exons, including exon 12, guide the interaction with different proteins. Furthermore, the distinct expression of specific ERG transcripts controlled by methylation adds to the complexity of ERG directed downstream pathways in different leukemic subtypes.