bluuebunny/crossref_metadata_2025_split huggingface dataset:
Revealing the Immune Population Hierarchy Using Novel Interactive Visualization Tools, a Comparison of Methods: Sunburst, viSNE, and Spade on Mass Cytometry Data
Authors: Chen Tiffany J , Kotecha Nikesh 
Published: October 2019

Abstract
The development of new technologies for high-parameter data has resulted in a critical bottleneck: identification of immune subsets is restricted to expert-based analysis, focusing on post-acquisition characterization of cell populations. Identification of cell subsets in flow cytometry has primarily focused on manual analysis, despite the fact that computational tools have proven useful for high-parameter and cross-sample comparisons. Sharing well-annotated data improves transparency and facilitates vital reproduction of results by external groups. Adoption of these new tools for immune subset discovery requires thorough collaborative investigation and validation of identified cell populations. To this end, in this study we compare the ease of discovery of immune subsets by comparing analysis through the use of three visualization tools: the sunburst hierarchy, the SPADE tree, and dimensionality reduction using viSNE. The sunburst hierarchy is a visual and interactive representation of traditional manual gating, whereas the SPADE tree is a semi-automated clustering and visualization tool for identification of cell subsets. viSNE allows interaction with high parameter data in the context of two-dimensional space where gating can be accomplished. In this study, we demonstrate the ability to automatically elucidate many immune subsets using Cytobank via an iterative analytic approach, combining computational tools (viSNE and SPADE) to recapitulate manually derived cell subsets.

Disclosures
Chen: Cytobank, Inc: Employment, Equity Ownership. Kotecha:Cytobank, Inc: Employment, Equity Ownership.
Histone Demethylase LSD1-Mediated Repression of GATA-2 Is Critical for Erythroid Differentiation
Authors: Guo Yidi , Li Yue , Huang Tianrui , Liang Dongxue , Li Zhe , Hu Xin 
Published: October 2019

Abstract
GATA-2 transcription factor is predominantly expressed in hematopoietic stem and progenitor cells (HS/PCs), and counteracts erythroid specific transcription factor GATA-1 to modulate proliferation and differentiation of hematopoietic cells. During hematopoietic cell differentiation, GATA-2 exhibits dynamic expression patterns, regulated by collaborations between GATA-1 and epigenetic regulators. Here, we show that histone specific demethylase1 (LSD1) regulates the expression of GATA-2 gene during erythroid differentiation. Knockdown of LSD1 leads to increased GATA-2 expression and inhibits induced MEL and embryonic stem cell differentiation. Furthermore, we demonstrate that LSD1 binds at the 1S promoter of GATA-2 gene locus, and suppresses GATA-2 expression through histone demethylation. Thus, our data reveals that LSD1 mediates erythroid cell differentiation by epigenetic modification of GATA-2 gene locus.

Disclosures
No relevant conflicts of interest to declare.
Hemogenic Endothelium-like Changes Were Derived to an Adult Human Dermal Fibroblast By Electrical Stimulation
Authors: Matsuo Takuji , Shirasaki Ryosuke , Yoko Oka , Yamamoto Tadashi , Ooi Jun , Matsumoto Kennsuke , Shirafuji Naoki 
Published: October 2019

Abstract
Background and Aims: We previously reported that when an adult human dermal fibroblast (HDF) was cultured with interleukin (IL)-1-beta (b) and erythropoietin (EPO), hematopoiesis-related molecules expressed. And, lymphatic duct-neogenesis genes also expressed, and vascular endothelial growth factor (VEGF)-A, and -C were produced. When anti-human VEGF-C antibody (Ab) was added to the cultures, hematopoiesis-related genes expressed; however, morphologic changes were not observed (54th ASH, 18th EHA). Reported findings on nuclear transfer reveal that electrical stimulation induces nuclear fusion and changes cell-fate. We observed the effect of electrical stimulation to IL-1-b-stimulated HDF.
Materials and Methods: HDF was cultured with IL-1-b, EPO, VEGF-A, and anti-human VEGF-C Ab for 14 days. Then, cells were suspended in electrical stimulation-buffer (0.25 M d-sorbitol, 0.1 mM Ca-acetate, 0.5 mM Mg-acetate, 1 mg/mL fatty-acid-free BSA, 0.5 mM HEPES), and incubated on ice for 10 minutes. Cells were stimulated at 110V 20mA (0.2 cm electrode, gap), and were further cultured with k/o DMEM containing 20% KSR and SCF, IL-6, FL, IGF-2, and VEGF-A. Morphological changes and expressions of vascular endothelial cell-related genes, and hematopoiesis-related ones were observed.
Results: When HDF was stimulated electrically, a few cells showed vascular endothelial cell-morphology after two days. When cells were further cultured in a hematopoietic stem cell-culturing condition, a part of endothelial cell-morphology changed to that of non-adherent hematopoietic cells or blastic colonies, in which increased expression-levels of SCL, CD41, GATA-2, CD34, and CD45 were accepted with significant statistical difference.
Discussion: Recent reports reveal that a kind of vascular endothelial cells, called hemogenic endothelium, can convert to a hematopoietic cell, and works an important role in hematopoietic stem cell-generation. We hypothesized that when cells expressed a kind of specific transcripts, some stimulation triggerred to convert morphologic changes and cell-fate as are observed in nuclear fusion. And, our observations indicated that when IL-1-b-stimulated HDF, expressing hematopoiesis-related transcripts, were stimulated electrically, cellular morphology changed. Currently, we are precisely analyzing electrically stimulated HDF’s biological characteristics.

Disclosures
No relevant conflicts of interest to declare.
Hypomethylation and Overexpression of CD70 in CD4+ T Cells of the Patients with Immuno-Related Pancytopenia
Authors: Shao Zonghong , Ren Yue , Fu Rong 
Published: October 2019

Abstract
Objective To study the expression of CD 70 and the methylation level of CD70 promoter in immuno-related pancytopenia (IRP) patients, and explore the role of CD70 in the pathogenesis of IRP.
Methods Thirty-five IRP patients and fifteen healthy donors were enrolled in this study. Peripheral blood mononuclear cells were isolated from venous blood by density gradient centrifugation and CD4+ T cells were isolated by immunomagnetic beads.The mRNA level and the percentage of CD70 of CD4+ T cells were measured by real-time quantitative polymerase chain reaction (RT-PCR) and flow cytometry respectively. Methylation-Specific PCR (MSP) was performed to determine the methylation level of CD70 promoter.
Results The percentage of CD70 expression on CD4+ T cells of untreated IRP patients[(7.46±1.51)%] was significantly higher than that of recovered IRP patients [(5.95±1.34) %] and normal controls [(1.83±0.6)%] and the result of recovered IRP patients was significantly higher than that of normal controls (P<0.05).The relative expressions of CD70 mRNA in CD4+ T cells were [2.314(0.200~6.084)]A[1.021 (0.135~3.434)]A [0.353 (0.008~2.258)] in three groups respectively. The differences between untreated IRP patients,recovered IRP patients and normal controls were significant (P<0.05). The CD70 promoter methylation level in CD4+ T cells of all IRP patients was significantly lower than that of normal controls (p<0.05). The expression of CD70 positively correlated to the ratio of CD5+ B cells (r=0.533, p<0.001).
Conclusions The overexpression of CD70 may lead to immunologic disarrangement and patients and it may play important role in the pathogenesis of IRP.

Disclosures
No relevant conflicts of interest to declare.
Chromatin State Shaping the Gene Expression Profiling in the Inducible Differentiation of Promyelocytic Leukemia Cells
Authors: Song Chunhua , Chung Lisa , Pan Xiaokang , Zhanjun Li , Ding Yali , Sachdev Mansi , Muthusami Sunil , Gowda Chandrika S. , Ge Zheng , Steffens Sadie , Tan Bi-Hua , Kendziorski Christina , Newton Michael , Dovat Sinisa 
Published: October 2019

Abstract
Background Epigenetic changes in DNA and chromatin are regarded as emerging major players for hematopoietic stem cells development and lineage differentiation. Epigenetic deregulation of gene expression leads to leukemia and reversibility of epigenetic modifications makes DNA and chromatin changes attractive targets for therapeutic intervention. The promyelocytic leukemia HL-60 cell can differentiate into microphage, granulocyte and monocyte with stimulation of phorbol myristic acid (PMA), dimethylsulfoxide (DMSO), and Vitamin D3, respectively, by affecting the gene expression on cell cycle and cell differentiation. However, how epigenetic regulation on gene expression in the inducible differentiation is still undetermined.
Methods and Results We did the microarray analysis to identify the genome-wide gene expression profile of HL-60 cells in various time points(0h, 6h, 1d,2d,4d and 6d) under the treatment of PMA, DMSO and D3 and found that around 3000 genes are significantly altered commonly in the cells upon the 3 treatments. The percentage of down-regulated genes in the all commonly altered genes is significantly higher than that of up-regulated genes, and the significantly altered genes showed significantly physical clustering on chromosome loci, indicating the epigenetic regulation involved in the regulation of the gene expression. We did observed the expression changes of epigenetic enzymes in the process and further did the ChIP-on-ChIP analysis by using the customer array tiling the selected changed genes and 5 genomic loci hybridizing with ChIP’d DNA of histone markers (H3K4me3, H3K9me3 and H3K27me3). We found that the obvious decrease of H3K4me3 binding and increase of H3K9me3 and H3K27me3 are observed in the promoter of the commonly down-regulated genes. These data indicated histone H3K4, H3K9 and H3K27 marker play important roles in shaping the chromatin state and regulating gene expression.
Conclusion We identified the signature genes controlling the inducible differentiation of promyelocytic leukemiacells and found the epigenetic mechanism regulating the gene expression in the process.

Disclosures
No relevant conflicts of interest to declare.
Leukemia Cell Differentiation upon Targeted Therapy Revealed By a Systems Biology Approach
Authors: Li Yulin , Seita Jun , Felsher Dean W , Dill David 
Published: October 2019

Abstract
Accurately determining the differentiation status of hematopoietic cells is crucial for understanding tumorigenesis and developing novel therapies for human leukemia. However, most current methods are subject to various biases. In this paper, we have developed a novel systems biology method, called the Lineage Maturation Index, to define the changes in differentiation state of hematopoietic cells based on comprehensive gene expression profiles. The changes in gene expression profiles of a specific hematopoietic lineage are shown asthe “lineage vector” from a point representing the gene expression of a hematopoietic stem cell (HSC) to another point representing a mature cell. By projecting the gene expression profile of a specific hematopoietic cell type onto the “lineage vector”, we can compute the LMI and define the relative differentiation state of this cell type. A shift in LMI from HSCs towards mature cell types indicates differentiation. We have confirmed the LMI method can track the differentiation and transdifferentiation of normal hematopoietic cells. Applying this method, we have shown that differentiation is a major consequence of various targeted therapies against leukemia irrespective of the driving oncogenes. Furthermore, we have used the method to screen a drug response library and discovered several drugs currently in clinical use for other purposes have potent differentiation activities in leukemia cells. Therefore, the LMI method is a robust computational approach to characterize differentiation of the hematopoietic system and discover novel differentiation therapy for leukemia.

Disclosures
No relevant conflicts of interest to declare.
Novel FLT3 Mutation Shows Dominant Negative Effects on the Wild-Type FLT3 Receptor
Authors: Bauer Julia , Sandhöfer Nadine , Hiddemann Wolfgang , Spiekermann Karsten 
Published: October 2019

Abstract
Background
The FMS-like tyrosine kinase-3 (FLT3) gene encodes for a receptor tyrosine kinase playing an important role in hematopoiesis. In acute leukemias it is one of the most frequently mutated genes. In this study we functionally characterized a novel frameshift deletion mutation of FLT3 found in a relapsed patient with acute myeloid leukemia (AML). The frameshift leads to a premature stop codon resulting in a truncated form of the receptor lacking most of the intracellular domains.
Material and Methods
FLT3 cDNA was expressed in the IL-3 dependent pro-B cell line Ba/F3 via a retroviral expression vector. The transduced cell lines were sorted by fluorescence-activated cell sorting (FACS). Stable expression of the receptor was confirmed on mRNA level by polymerase chain reaction (PCR) and on protein level by western blotting and cell surface expression of the receptor by flow cytometry. Cell proliferation assays were performed in presence or absence of IL-3 or FLT3-ligand. By western blotting receptor activation and its downstream signaling were analyzed. Ligand-binding of the receptor was shown via biotinylated and fluorescent FLT3-ligand-receptor-complexes by flow cytometry.
Results and Discussion
Stable expression of FLT3 Q569Vstop does not enable Ba/F3 cells to grow IL-3-independent. FLT3-ligand can still be bound by the mutated receptor but is not able to stimulate receptors´ signaling and growth of the cells. Furthermore coexpression of wild-type (WT) and mutant FLT3 receptor also abolishes the ability to stimulate the WT receptor with its ligand. This is confirmed by analyzing downstream signaling in the cells as MAPK is less phosphorylated in the FLT3-WT/Q569Vstop coexpressing cells than in FLT3-WT expressing cells alone.
Conclusion
Most of the FLT3 mutations are activating mutations leading to a constitutive activation of the receptor and ligand-independent growth. In our study we characterized a novel FLT3 mutation found in an AML patient which has not been described before. The resulting truncated receptor is still integrated in the plasma membrane and binds its ligand but its ability to be fully activated is completely lost. Furthermore coexpressed with a FLT3-WT receptor, it even prevents stimulation and activation of the WT receptor, thus acting in a dominant negative manner. How the truncated form of the receptor contributes to progression of acute leukemia is of great interest and will be further investigated.

Disclosures
No relevant conflicts of interest to declare.
Decreased SATB1 Expression in ALT and Promoted Invasion of Jurkat Cell in Vitro
Authors: Xu Lihua , Tan Huo , Yang Shaojiang 
Published: October 2019

Abstract
BACKGROUND: Special AT-rich sequence-binding protein-1 (SATB1) is critical for genome organizer that reprograms chromatin organization and transcription profiles, and associated with tumor growth and metastasis in several cancer types cancer. However, their expression patterns and function values for adult T-cell leukemia (ALT) are still largely unknown.
METHODS: Expression of SATB1 in normal and ALT peripheral blood leucocytes (n=50 for normal, n=30 for ALT) were evaluated by RT-qPCR in mRNA level. Expression of protein SATB1 in leukemia cell lines were evaluated by Weston Blot.SATB1-shRNA was constructed in psuper-retro-puro retrovirus vector. Jurkat-SATB1-sh and control stable cell lines were established after virus infection and puromycin selection. Cellular proliferation and invasion of Jurkat-ctr and SATB1 knockdown Jurkat cells were evaluated by CCK-8 kit and transwell matrigel invasion assay, respectively.
RESULTS: SATB1 mRNA expression was decreased in ALT (p<0.001) compared with normal peripheral blood leucocytes cells. SATB1 protein expression in Jurkat cell was higher than other leukemia cell lines. SATB1-shRNA was constructed successfully. Weston Blot show Jurkat-SATB1-sh and control stable cell lines were established successfully. Knockdown of SATB1 expression significantly enhanced invasion of Jurkat cell in vitro but no effect in proliferation
CONCLUSIONS: Decreased expression of SATB1 in ALT was associated with cell invasion.

Disclosures
No relevant conflicts of interest to declare.
Multidrug Resistance Gene (MDR1) Polymorphism and Response to Nilotinib in Egyptian Patients with Chronic Myeloid Leukemia (CML)
Authors: Mattar Mervat , Ellithy Hend , Elnahas Yasser , Azzazi Mohamed osman 
Published: October 2019

Abstract
Introduction: Inappropriate expression of the multidrug resistance (MDR1) gene I in chronic phase chronic myeloid leukemia cases (CML-CP) encodes P glycoprotein (Pgp)that may cause resistance to second generation tyrosine kinase inhibitors (TKIs)
Patients and methods: Thirty-one upfront CML-CP patients, planned to receive nilotinib, were included. Detection of MDR1 gene polymorphism C3435T, using PCR Restriction Fragment Length Polymorphisms (PCR-RFLP) was done initially for every patient. We prospectively followed up the patients between February 2012 and February 2014 with PCR for BCR-ABL1 transcripts every 3 months. The molecular response to nilotinib, according to the level of BCR-ABL1 by PCR, was compared to the different MDR1 3435 genotypes.
Results: The majority of the patients carried the MDR1 3435CC genotype of Molecular response was optimal in 56%, 60% and 80% of the patients at month 3, 6 and 12 respectively.There was no statistically significant difference between MDR- C3435T genotypes and the molecular response to treatment with nilotinib.

Disclosures
No relevant conflicts of interest to declare.
Synergistic Effect of Quinacrine with Chemotherapeutics or TRAIL in Hematopoietic Malignant Cells
Authors: Wang Wenge , Lulla Amriti R. , Hernandez-Borrero Liz J. , Dicker David T. , Teye Emmanuel K. , Talekar Mala K. , Dolloff Nathan G. , Pu Jeffrey J. , El-Deiry Wafik S. 
Published: October 2019

Abstract
Quinacrine is a bioactive acridine derivative which has been used for treatment of malaria, giardiasis, systemic lupus erythematosus, and rheumatoid arthritis. In searching for p53 pathway activating agents for cancer therapy, we found that quinacrine stabilizes p53 and induces p53-dependent and p53-independent tumor cell death. Quinacrine also induces expression of TRAIL Death Receptor 5 (DR5) and reduces expression of anti-apoptotic Mcl-1 in tumor cells. These activities predict synergies with TRAIL (tumor necrosis factor-related apoptosis inducing ligand) and chemotherapeutic agents in inducing extrinsic and intrinsic pathway mediated apoptosis. In addition, quinacrine suppresses NFkB activity in tumor cells. Clinical trials have been ongoing for treatment of solid tumors including colon cancer, renal cancer, prostate cancer, and non-small cell lung cancer with quinacrine in combination with chemotherapy or tyrosine-kinase inhibitors, however, the therapeutic potential of quinacrine in blood cancer cells has not been established. We tested quinacrine on hematopoietic malignant cells, which included cell lines of myeloid leukemia, lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, and multiple myeloma. We found that quinacrine induces massive cell death in the cell lines tested, at concentrations from as less as 1 microM to 5 microM, 2-10 times lower than required to induce solid tumor cell death. Quinacrine synergizes with TRAIL in inducing cell death of TRAIL-sensitive cells and reverses resistance in TRAIL-resistant cells. Quinacrine also synergizes with chemotherapeutic agents, such as antimetabolites, alkylating agents, and tyrosine kinase inhibitors, in inducing apoptosis of hematopoietic cancer cell lines. Our work supports translational efforts to advance the use of quinacrine from bench to clinic and provides rationale for combination chemotherapeutic regimes for treatment of hematopoietic malignancies.

Disclosures
No relevant conflicts of interest to declare.
Influence of Adriamycin and Wogonin on the Expression of Mir-155 in Burrkit Lymphoma in Vitro
Authors: Wu Xue , Wang Fei , Xia Guohua , Chen Yanping , Chen Baoan 
Published: October 2019

Abstract
Objective:
In this study, we investigated the influence of adriamycin and wogonin to the expression of miR-155 in Burrkit lymphoma in vitro. We focused on exploring the efficiency and pathway of wogonin to induce the apoptosis of lymphoma cells through the transcription and translation level of miR-155 compared with ADM and seeking for the evidence of the synergistic effect between wogonin and ADM.
Methods:
(1) Use drugs to interfere the growth of the Raji cells (Adriamycin (ADM, chemical name:doxorubicin), wogonin, ADM+wogonin), observe the cells through the microscope.
(2) The apoptosis of Raji cells was tested by FCM.
(3) The transcription of miRNA-155 of Raji cells was tested by real-time PCR.
(4) The translation of miRNA-155 of Raji cells was tested by Western blot.
Results:
(5) After being disposed for 24 hours,the experimental groups developed to apoptosis.The statistic significance existed between groups of experiment and control and groups of each experiment.The effect of inducing apoptosis from strong to weak was group of ADM+wogonin,ADM and wogonin.
(6) After being disposed for 24 hours,the transcription level of miRNA-155 of the experimental groups was lower than the control group.Compared with ADM,the transcription level of miRNA-155 of wogonin group seemed to be stronger,while the combination of the two drugs was the weakest.
(7) After being disposed for 24 hours,the translation level of p85α of the experimental groups was lower than the control group. Compared with ADM, the translation level of p85α of wogonin seemed to be stronger, while the combination of the two drugs was the weakest.
Conclusion:
The Chinese materia medica preparation wogonin could induce the apoptosis of lymphoma cells and enhance the effect of ADM through PI3K/AKT pathway.

Disclosures
No relevant conflicts of interest to declare.