bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mir-34a as Part of the Chemotherapy Resistance Network in Chronic Lymphocytic Leukemia.
Authors: Zenz Thorsten , Mohr Julia , Eldering Eric , Kater Arnon P , Bühler Andreas , Winkler Dirk , Kienle Dirk , Dürig Jan , Mertens Daniel , Döhner Hartmut , Stilgenbauer Stephan 
Published: April 2020

Abstract
The central role of 17p (TP53) deletion and 11q (ATM) deletion in identifying patients with CLL who respond sub-optimally to chemotherapy suggests a prominent role of the DNA damage pathway in CLL. MiR34s have been shown to be direct p53 targets and mediate some of the p53 dependent effects.
To assess the role of miR-34s in a large cohort of CLL patients (n=60), we studied miR34a and miR34b/c levels together with a detailed assessment of the DNA damage response. While no expression of miR-34b/c could be detected, we found diverse expression levels of miR34a. MiR34a levels in CLL are up-regulated after DNA damage in the presence of functional p53. In contrast, cases with 17p deletion showed low levels of miR-34a and failure to up-regulate miR-34a after irradiation (median basal expression 2.5 (1.0–15.78) vs. 32.8 (0.89–337.8) in cases without 17p deletion (p&lt;0.0001); after irradiation (median 5.8 (1.38–12.9) vs. 121.8 (1.79–608.9) (p&lt;0.0001)). Since we also observed heterogeneous expression in cases without 17p deletion we investigated causes for impaired miR-34a expression in the absence of 17p- and found a strong correlation of impaired miR-34a levels with impaired DNA damage response, TP53 mutations (without 17p deletion) and F-refractory disease (in the absence of 17p deletion). Importantly, decreased miR-34a expression was associated with resistance to apoptosis and the expression levels differentiated subgroups of CLL with different rates of apoptosis after irradiation better than induction of p21/p53 or cytogenetics. In order to assess the relation of miR-34a to other known targets of p53 we compared the miR-34a expression to mRNA levels of p21, Bax and Puma. Up-regulation of miR-34a after irradiation was associated with induction of Bax and p21 but not with PUMA.
Low expression of miR-34a is associated with p53 inactivation but also chemotherapy-refractory disease, impaired DNA damage response and apoptosis, irrespective of 17p deletion/TP53 mutation. The demonstration of decreased miR-34a basal levels in cases with TP53 mutation could be interpreted as an oncogenic function of mutated TP53, where the mutated protein acts to decrease miR-34a levels and thus leads to reduced apoptosis. The mechanism underlying miR-34a regulation and function warrants further study.
Mice Lacking Wasp-Interacting Protein Evolve Anti-Platelet IgAs That Impair Platelet Responses Mediated by the Collagen Receptor GPVI.
Authors: Falet Hervé , Marchetti Michael P. , Simon Delphine , Geha Raif S. , Hartwig John H. 
Published: April 2020

Abstract
Wiskott-Aldrich Syndrome (WAS) is a hematopoietic disorder characterized by immune deficiency, eczema and severe thrombocytopenia with small platelets. However, the role of the WAS protein (WASp), mutated or absent in WAS, is unclear in platelets because platelets that lack WASp function normally. WASp constitutively associates with WIP in resting and activated platelets. We investigated the role of the platelet WIP-WASp complex by using mice that lack either WIP or WASp. WIP knockout (KO) platelets lack WASp and WIP expression is reduced in WASp KO platelets, indicating a requirement for both proteins for each other’s integrity. WIP KO, but not WASp KO mice evolve platelet-associated antibodies of the IgA class, as evidenced by immunoblot analysis and flow cytometry. We present evidence that platelet-associated IgAs specifically target the collagen receptor GPVI in WIP KO mice:
protein tyrosine phosphorylation, including that of phospholipase C-γ2, and calcium mobilization are impaired in IgA-presenting WIP KO platelets stimulated through GPVI, resulting in α-granule secretion, integrin αIIbβ3 activation and actin assembly defects; binding of the anti-GPVI antibody JAQ1 to normally expressed GPVI is reduced in IgA-presenting WIP KO platelets; JAQ1 induces the irreversible loss of GPVI from circulating platelets in WT mice, but not in WIP KO mice that bear anti-platelet IgAs.
Together, the data indicate that platelet-associated IgAs negatively modulate GPVI signaling in WIP KO mice.
The Effect and Underlying Mechanism of Melatonin on Platelet Formation and Survival in a Thrombocytopenic Model.
Authors: Yang Mo , Zhou Min , Ye Jie Yu , Cheung Yiu Fai , Chan Shing , Ha Shau Yin , Chan Godfrey Chifung 
Published: April 2020

Abstract
Thrombocytopenia is a major treatment-related problem for cancer patients on chemo- or radiotherapy. In clinical studies, melatonin has been shown to be able to lower the frequency of thrombocytopenia in these cancer patients. However, the actual effects and underlying mechanism of melatonin on megakaryocytopoiesis, and platelet formation/survival have not been formally investigated. Our previous studies have demonstrated that serotonin can stimulate hematopoiesis and megakaryocytopoiesis (Yang et al, Stem Cells, 2007). Melatonin is an acetylated product of serotonin. To delineate the effect of melatonin on platelet production, we hypothesize that melatonin has therapeutic effects on thrombocytopenia by promoting megakaryocytopoiesis and enhancing the recovery of platelets via activation of Akt/Erk signaling. The effect of melatonin on thrombopoiesis was investigated in an irradiated mouse model (4 Gy). Eighteen mice were randomly divided into three groups (6 in each). Group 1 (normal control, N) received no irradiation or melatonin; Group 2 (model control, C) and Group 3 (melatonin, M) received 4 Gy total body irradiation. After irradiation, melatonin (10mg/kg.d) was injected intraperitoneally into Group M for consecutively 21 days. Peripheral blood platelets, white blood cells (WBC), and red blood cells (RBC) were analyzed from the three groups on day 0, 7, 14, and 21.Our results showed that melatonin enhanced the recovery of platelets and WBC counts. We also investigated the in-vitro effect of melatonin on CFU-MK formation at different doses (0–-500 nM). The results showed that melatonin significantly promoted CFU-MK formation at a dose dependant manner. The maximal concentration was at 200 nM (P&lt;0.01, n=6). The size of CFU-MK with melatonin treatment was much larger and each MK cell was more mature. Melatonin also stimulated the formation of bone marrow CFU-F. Using megakaryocytic cell CHRF-281, we further found that melatonin exerted protective effect on serum-free induced apoptosis of CHRF cells. The cell viability was significantly increased with the treatment of melatonin. Flow cytometric dot-plot analysis demonstrated that the population of apoptotic cells (Annexin V/PI staining) was significantly decreased with melatonin treatment (200nmol/L) for 72 hrs (n=4). It also showed trends of amelioration of caspase-3 expression and the proportion of cells containing JC-1 monomers (indicating a trend in the drop of mitochondrial membrane potential). The population of phospho-Akt and Erk1/2 were significantly increased after treatment by melatonin. Our results showed that melatonin had a promoting effect on platelet formation and survival in a thrombocytopenic model. The underlying mechanisms may be involved in directly stimulating megakaryocytopoiesis and having anti-apoptotic effect in megakaryocytes via the activation of Akt/Erk signaling.
Integrin-Independent Role of CalDAG-GEFI in Neutrophil Chemotaxis.
Authors: Carbo Carla , Goerge Tobias , Hattori Hidenori , Duerschmied Daniel , Cifuni Stephen M. , Luo Hongbo R. , Wagner Denisa D. 
Published: April 2020

Abstract
Neutrophil chemotaxis and transmigration towards a source of inflammation are two crucial processes for host defense against infection that rely on integrin function. Recently, integrin-independent migration of dendritic cells to the lymph node has been brought to light, although neutrophil migration in the presence of EDTA was reported many years ago. Ca2+ and diacylglycerol-regulated guanine nucleotide exchange factor I (CalDAG-GEFI), is a small signaling protein that plays a key role in the activation of beta-1, beta-2, and beta-3 integrins in platelets and neutrophils by activating the small GTPase Rap1. We explored the role of CalDAG-GEFI in integrin-independent chemotaxis in neutrophils. Here we report that CalDAG-GEFI−/− neutrophils have impaired chemotaxis that is independent of integrin function. In a chemotaxis transwell assay towards LTB4 and in the presence of 10mM EDTA, CalDAG-GEFI−/− neutrophils had a 50% reduction in transmigration over 60 minutes compared to wild-type (WT) neutrophils (p&lt;0.05). In separate experiments we confirmed that the transwell assay is independent of integrins using either CD18−/− neutrophils or WT neutrophils plus a blocking anti-CD18 monoclonal antibody. We previously showed that LTB4 signaling upstream of CalDAG-GEFI was not affected in CalDAG-GEFI−/− neutrophils, as assessed by intracellular calcium flux measurements. Using videomicroscopy to visualize the live migrating neutrophils in a horizontal plate in the presence of 10mM EDTA, we found that the reason CalDAG-GEFI−/− neutrophils fail to reach the chemotactic stimulus (10 pg/mL LTB4) is because they have a significantly reduced migration speed compared to WT neutrophils (16 um/sec vs. 23 um/sec, p&lt;0.05), and also because they have an abnormal chemotactic directionality, with a directionality index (the distance between the start and finish points of a migrating neutrophil/total distance covered by the migrating neutrophil) of 0.84 vs 0.94 in WT neutrophils, p&lt;0.05. We investigated whether the observed differences in chemotaxis between CalDAG-GEFI−/− and WT neutrophils could be explained by differences in F-actin polymerization. Using fluorescence microscopy, we found that the percentage of CalDAG-GEFI−/− neutrophils with F-actin pseudopodia after LTB4 stimulation was significantly lower compared to WT neutrophils (22% vs. 56.7%, p&lt;0.05), suggesting that CalDAG-GEFI−/− neutrophils have a defect in F-actin polymerization. Overall, our studies suggest that CalDAG-GEFI plays a role in the mechanisms that regulate both the migration speed and direction of neutrophils during chemotaxis, independent of its established role in integrin activation.
Comprehensiveness of Quality of Life Instruments in Capturing Concerns Related to Chemotherapy-Induced Neutropenia
Authors: Nina Lathi A , Isogai Pierre , Mittmann Nicole , DeAngelis Carlo , Cheung Matthew , Sandra Knowles , Piliotis Eugenia , Shear Neil , Walker Scott 
Published: April 2020

Abstract
Neutropenia is a serious hematologic consequence of cancer chemotherapy that can lead to further complications such as febrile neutropenia (FN). FN is potentially life threatening and often requires hospitalization. Few studies have evaluated the impact of neutropenia on quality of life (QoL). This study quantified QoL using two nonneutropenia-specific instruments, the EQ-5D questionnaire, a generic tool used to measure health-related QoL, and the Functional Assessment of Cancer Therapy - General (FACT-G) questionnaire, and a neutropenia-specific instrument, the Functional Assessment of Cancer Therapy - Neutropenia (FACT-N) questionnaire. The FACT-G is a 27-item questionnaire that examines QoL in patients with cancer using four subscales. A neutropenia-specific subscale (NSS) has been developed for use with the FACT-G; this combined questionnaire is the FACT-N. Data were collected from patients, who provided informed consent, and who were admitted to Sunnybrook Health Sciences Centre, Toronto, Canada, for the treatment of chemotherapy-induced FN. Linear regression models were fitted to examine the relationship of scores from the neutropenia-specific instrument with those obtained from the other instruments. Two models were fitted using the NSS as the response variable. Predictors for the regression models were the FACT-G scores for each of the subscales (physical, emotional, social and functional wellbeing) and the five domains of the EQ-5D (mobility, self-care, usual activity, pain/discomfort and anxiety/depression) along with the visual analog scale (VAS) component of this tool. The physical and emotional wellbeing subscales of the FACT-G had a strong relationship to the NSS (p &lt; 0.05); the social and functional well-being subscales had a much weaker relationship (p &gt; 0.5). For the EQ-5D, the pain/discomfort domain had the strongest relationship to the NSS (p=0.18); the remaining domains, with or without the VAS, all demonstrated a weaker relationship (p &gt; 0.5). Model fit was assessed by the adjusted R2 statistic; it was 0.54 when FACT-G subscales were used as the predictors compared to −0.04 for the EQ-5D domains indicating that the FACT-G was a better predictor of neutropenia-related concerns. Neutropenia concerns appear to be more closely related to cancer specific QoL compared to general quality of life as demonstrated by the stronger relationship of the NSS to the FACT-G than to the EQ-5D. This may be due to the comprehensiveness of the FACT-G questionnaire where a possible score anywhere from 0 to 24 or 28 can be obtained in each of the subscales, compared to three-point descriptive system for each of the domains of the EQ-5D.
Enforced Nr4a2 expression Drives HSCs into Quiescence.
Authors: Sirin Olga , Conneely Orla M , Goodell Margaret A. 
Published: April 2020

Abstract
Hematopoietic stem cells (HSCs) are used as a paradigm for understanding somatic stem cell biology, yet the factors that regulate their self-renewal and differentiation are poorly understood. Our lab has recently discovered that within the hematopoietic lineage, Nr4a2 expression is restricted to the stem and progenitor cell compartment. Furthermore, we have observed a dramatic decrease in its expression when the bone marrow cells are exposed to 5-fluorouracil which forces quiescent HSCs into cycle. We therefore hypothesize that Nr4a2 may be involved in maintaining stem cell quiescence.
To better understand the role of Nr4a2 in hematopoiesis, we have performed gain-of-function studies in which Nr4a2 is overexpressed in HSCs. Bone marrow transplantation (BMT) experiments revealed that Nr4a2 overexpressing cells contributed significantly less to the peripheral blood compared to the control (1% vs 35%, see fig). This suggests that enforced Nr4a2 expression negatively affects HSC regeneration potential. We also found that Nr4a2 overexpression has a negative effect on the overall proliferation capacity with an in vitro methocult assay. Together, these results support our hypothesis that the high level expression of Nr4a2 regulates HSC quiescence by inhibiting HSC self-renewal and/or differentiation.
Cell cycle analysis demonstrates that enforced expression of Nr4a2 causes an increase in the number of cells in quiescence. These data are supported further by analysis at a molecular level revealing that overexpression of Nr4a2 increases expression of cell cycle inhibitors such as p18 and p19, both of which are important for Go to G1 entry. By uncovering the role of Nr4a2 in hematopoiesis we aim to achieve not only a better understanding of the regulation of stem cell self-renewal and differentiation but also of the pathways involved in HSC cell cycle regulation.
Figure Figure
C/Ebpβ Inhibits Apoptosis in Hematopoietic Cells Via Interaction with NF-κb p50.
Authors: McFall Rebecca , Paz-Priel Ido , Friedman Alan D. 
Published: April 2020

Abstract
The CCAAT/enhancer binding protein (C/EBP) family of proteins bind DNA via their C-terminal basic region – leucine zipper (BR-LZ or bZIP) domains and activate transcription via N-terminal trans-activation domains. We previously demonstrated that C/EBPα inhibits apoptosis when the immature hematopoietic Ba/F3 cell line is withdrawn from IL-3 and that a C/EBPα BR mutant that does not bind NF-κB p50 but retains the ability to bind NF-κB p65 is inactive. We have now sought to extend these findings to C/EBPβ. We first created two point mutations in C/EBPβ, R240G and ΔK253. The C/EBPβ(R240G) is analogous to the C/EBPα(R300G) BR variant that does not bind NF-κB p50 or inhibit apoptosis, and C/EBPβ(ΔK253) is analogous to the C/EBPα(ΔK313) LZ variant that does not bind DNA but retains the ability to interact with NF-κB p50 and inhibit apoptosis. In co-ip experiments using transiently transfected 293T extracts, both C/EBPβ and C/EBPβ(ΔK253) interacted with NF-κB p50, whereas C/EBPβ(R240G) did not, as predicted. In contrast, all three isoforms bound NF-κB p65. And in gel shift assay, C/EBPβ bound a consensus C/EBP site, whereas neither C/EBPβ(R240G) nor C/EBPβ(ΔK253) bound DNA, again as expected. Ba/F3 cell lines were then generated stably expressing C/EBPβ-ER, C/EBP(R240G)-ER, or C/EBPβ(ΔK253)-ER proteins, via transduction with pBabePuro retroviral vectors and puromycin selection of pooled populations. Western blot analysis confirmed similar expression of each transgene. Ba/F3 lines were removed from IL-3 and cultured with or without estradiol, to activate the ER fusion proteins. Remarkably, both CEBPβ-ER and C/EBPβ(ΔK253)-ER prolonged survival of the cells during the first 48 hrs after IL-3 withdrawal, similar to the activity of C/EBPα-ER, whereas C/EBPβ(R240G)-ER was ineffective. Current efforts focus on identifying genetic targets of C/EBPβ:NF-κB p50 cooperation relevant to the observed inhibition of apoptosis, on assessing activities of C/EBPβ and it variants in normal hematopoietic cells isolated from adult marrow, and on determining the activities of the long and short (LAP and LIP) C/EBPβ translational isoforms. Overall, the finding that C/EBPβ inhibits apoptosis in cooperation with NF-κB p50 is significant due to the expression of both C/EBPβ and activated NF-κB in several human lymphoma subsets and in solid tumors such as breast cancer. Targeting this interaction with small molecules may provide a novel therapeutic approach for these malignancies.
JunB Limits Hematopoietic Stem Cell (HSC) Functions as a Protective Mechanism against Initiation of Myeloid Malignancy
Authors: Santaguida Marianne T , Schepers Koen , King Bryan C , Passegue Emmanuelle 
Published: April 2020

Abstract
The JunB/AP-1 transcription factor plays a crucial role in controlling homeostasis of the hematopoietic stem cell (HSC) compartment and functions as a tumor suppressor in mice. We have previously demonstrated that mice lacking JunB accurately recapitulate important clinical aspects of human myeloid malignancies, including chronic myelogenous leukemia(CML). Using this model, we have shown that leukemia-initiating stem cells (LSCs) can arise from the hematopoietic stem cell (HSC) compartment. Now, we have identified the mechanisms by which JunB regulates HSC functions and discovered how inactivation of this key transcription factor leads to HSC transformation into LSCs that overproduce myeloid progenitors and drive myeloproliferative disease (MPD) development in vivo. We found that JunB normally controls the proliferation of the long-term HSCs (LT-HSCs) by regulating the expression levels of key cell cycle regulators, including the CKI p57 and the polycomb group gene Bmi1, and limits the rate at which LT-HSCs produce myeloid progenitors by maintaining appropriate responsiveness to both Notch and TGF-β signaling pathways. These functions place JunB at the center of a complex network of intrinsic and extrinsic regulations that ensure coordinate regulation of HSC proliferation and differentiation, independently of LT-HSC maintenance and self-renewal activity. These results provide a mechanism for disease initiation in a broad range of human myeloid malignancies, including those in which JunB inactivation has already been reported. Moreover, they expand our understanding of the normal biology of the HSC compartment and provide a mechanism to explain how mutations that increase the proliferation of a single LT-HSC can become dominant and lead to the expansion of an aberrant clone within the otherwise normal HSC compartment. Understanding clonal dominance of LT-HSCs is clearly at the root of designing new therapeutic treatments for a broad spectrum of hematological malignancies.
Human Hematopoietic Stem Cells and Leukemic Cells Form Cadherin-Catenin Based Junctional Complexes with Mesenchymal Stromal Cells
Authors: Wuchter Patrick , Saffrich Rainer , Wagner Wolfgang , Wein Frederik , Schubert Mario Stephan , Eckstein Volker , Ho Anthony D. 
Published: April 2020

Abstract
The interaction between human hematopoietic stem cells (HSC) and their niche plays a key role in regulating maintenance of “stemness” and differentiation. We have demonstrated that a feeder layer of human mesenchymal stromal cells (MSC) can serve as a surrogate model for the niche for human HSC. We could also show, MSC are intimately connected to one another by a novel kind of adhering junction, consisting of villiformto-vermiform cell projections (processus adhaerentes). With this background, we have analyzed the intercellular junctional complexes between HSC and MSC. In comparison, we also studied the cell-cell contacts between leukemia cells (LC) and MSC.
MSC were derived from bone marrow aspirates from healthy voluntary donors. HSC were isolated from umbilical cord blood. Leukemia cells that were CD34+ were obtained from bone marrow aspirates from patients suffering from acute myeloid leukemia at the time point of initial diagnosis. After 24–48 hours of co-cultivation, we stained the cellular contacts with a panel of antibodies specific for various components of tight, gap and adherens junctions. Using advanced confocal laser scanning microscopy in combination with deconvolution and volume rendering software, we were able to produce 3D-images of intercellular junctions between HSC/MSC as well as between LC/MSC. To examine the specific function of N-cadherin, we analyzed the effect of siRNA knock down of N-cadherin in MSC upon co-cultures of HSC and MSC.
Intercellular connections between HSC and MSC are mainly characterized by podia formation of the HSC linking to the adjacent MSC. At the intimate contact zone to the MSC, we have identified the cytoplasmic plaque proteins alpha- and beta-catenin, co-localized with the transmembrane glycoprotein N-cadherin. Additionally, we compared these findings with a similar setting consisting of human LC co-cultured with feeder-layer of MSC. Our results demonstrated that in comparison to HSC, the proportion of leukemia cells adherent to the feeder-layer is significantly lower and podia formation is less frequent (ratio 1:3). However, the mechanism of adhesion through cadherin-catenin-complex has remained the same. At a functional level, we found that siRNA knock down of N-cadherin in MSC resulted in decreased adhesion of HSC to MSC and in a reduction of cell divisions of HSC. These results confirm that direct cellular contact via N-cadherin-based junctions is essential for homing and adhesion of HSC to the cellular niche and subsequently for the regulation of self-renewal versus differentiation in HSC.