bluuebunny/crossref_metadata_2025_split huggingface dataset:
Proteomic Analysis for the Identification of Disease Biomarkers and Therapeutic Targets in Natural Killer Cell Lymphoma.
Authors: Lee Hojung , Shereck Evan , Cairo Mitchell S. , Seiler Seiler E , Basrur Venkatesha , Fermin Damian , Elenitoba-Johnson Kojo SJ , Lim Megan S 
Published: April 2020

Abstract
Natural killer cell lymphoma (NKL) is a highly aggressive form of hematolymphoid malignancy that arises from natural killer cells, a critical component of the innate immune system. Early diagnosis of the tumor is critical however there are no biomarkers which are useful currently for the diagnosis and monitoring of patients with NKLs. We hypothesized that comprehensive analysis of differentially expressed proteins by NKL and normal NK cells would be a rational strategy for the discovery of NKL biomarkers and lead to better understanding of pathogenetic mechanisms of NKL. We used total cell lysates obtained from highly enriched peripheral blood NK cells (CD3-, CD56dim) and an Epstein-Barr virus negative, IL-2 dependent cell line (NK-92MI) for quantitative proteomic analysis. Isotopecoded affinity tags (ICATTM)-labeling followed by liquid chromatography-tandem mass spectrometry analyses (LC-MS/MS) identified 114 differentially expressed proteins in NKL compared to the normal NK cells. Out of these, 80 were upregulated 1.5-fold or greater and 34 were downregulated 1.5-fold or greater in NKL. Proteins within diverse cellular functional categories including cell signaling, transcriptional regulation, DNA metabolism, and immune response were differentially expression. Furthermore, known proteins relevant to the function of NK cells such as perforin, granzyme and defensins were downregulated in the NKL cell line. We selected eight proteins to validate the changes identified by ICAT-LC-MS/MS using Western blot analysis. HSP90, PCNA, alpha-enolase, GSTP1, galectin-1, and PARK7 were upregulated, and lactoferrin and annexin A1 were downregulated by western blot analysis demonstrating high concordance with the results of ICAT-LC-MS/MS analysis. We validated the functional significance of HSP90 as a potential therapeutic target in NKL by investigating the effect of inhibitors of HSP90 (geldanamycin) on the survival of an NKL cell line. Inhibition of HSP90 led to significant decrease in cell viability and apoptotic cell death as confirmed by western blotting demonstration of PARP-1 induction and detection of its cleavage products. This study represents the first global quantitative proteomic cataloguing of proteins expressed by NKL and normal NK cells, and reveals potential candidate proteins that may be useful for the diagnosis and function as therapeutic targets in patients with NKL.
Alterations of the CxxC Domain Preclude Oncogenic Activation of Mixed Lineage Leukemia 2 (MLL2).
Authors: Slany Robert K. , Bach Christian , Mueller Dorothee , Buhl Sebastian , Garcia-Cuellar Maria-Paz 
Published: April 2020

Abstract
The mixed lineage leukemia (MLL) family of histone methyltransferases has become notorious for the participation of the founding member MLL in fusion proteins that cause acute leukemia. Despite a high overall structural conservation no other MLL homolog has been found involved in leukemogenic fusion proteins. This surprising fact might be either due to a relative genomic stability of the MLL2 locus that prevents chromosomal translocations or MLL2 might be altogether incapable of contributing to oncogenic fusions because of functional constraints. Here we demonstrate the latter to be true for fusion proteins based on MLL2, the closest relative of MLL. A MLL2-ENL protein constructed in analogy to the highly leukemogenic MLL-ENL was incapable of transforming primary hematopoietic cells. Elaborate swap experiments identified the zinc binding “CxxC”- region of MLL2 and an adjacent “post-CxxC” stretch of basic amino acids as the essential determinants for the observed difference between MLL and MLL2. Surprisingly gel shift experiments indicated that the CxxC and post-CxxC domains of MLL and MLL2 conferred almost indistinguishable in vitro DNA binding properties. However, in vivo these motifs guided MLL-ENL and MLL2-ENL to a largely non-overlapping gene repertoire. Divergent nuclear localization, a distinct potency to activate a MLL model promoter in reporter assays, and measurements of homeobox gene levels in primary cells expressing MLL and MLL2 fusion proteins all argued for a separate target site selection of the two fusions. Therefore, the CxxC domain appears to be largely responsible for target finding and as a consequence this domain is a promising object for therapies aimed at MLL fusion proteins without affecting the general function of other MLL family members.
The Essential Role of Stem Cell Factor/KIT Signaling in the Proliferation and Survival of Blast Cells from Transient Leukemia in Neonates with Down Syndrome.
Authors: Toki Tsutomu , Kanezaki Rika , Adachi Souichi , Fujino Hisanori , Xu Gang , Sato Tomohiko , Suzuki Kahori , Tauchi Hisamichi , Endo Mikiya , Ito Etsuro 
Published: April 2020

Abstract
Children with Down syndrome (DS) are predisposed to developing leukemia. A leukemoid reaction occurring uniquely in approximately 10% of newborn infants with DS referred to as transient leukemia (TL). This disorder, in most cases, resolves spontaneously within three months after birth. Of all TL patients approximately 20–30% develop myeloid leukemia (ML-DS) within four years. Although treatment with low dose cytarabine is effective in high-risk TL cases, about 20% of severe patients still suffer early death. Improved treatments for TL are necessary for a better long-term prognosis of DS patients. In this study, we demonstrate abundant KIT expression in all 13 TL patients examined, although no significant difference in expression levels was observed between TL and acute myeloid leukemia (AML). Stem cell factor (SCF) mRNA was expressed at extremely low levels in TL cells and there were no significant differences in SCF expression between TL and AML. SCF stimulated the proliferation of the TL cells from all five patients examined and treatment with the tyrosine kinase inhibitor imatinib suppressed the proliferation and KIT phosphorylation effectively in vitro. To investigate the signal cascade, we established the first SCF-dependent, ML-DS cell line, KPAM1. Withdrawal of SCF or treatment with imatinib induced apoptosis of KPAM1 cells. SCF activated the RAS/MAPK and PI3K/AKT pathways, followed by downregulation of the pro-apoptotic factor BIM and upregulation of the anti-apoptotic factor MCL1. Although we found novel missense mutations of KIT in two of 14 TL patients, functional analysis using KPAM1 cells showed that neither mutation led to KIT activation and neither reduced the cytotoxic effects of imatinib. These results suggest the essential role of SCF/KIT signaling in the proliferation of DS-related leukemia and the possibility of therapeutic benefits of KIT-targeting tyrosine kinase inhibitors for TL patients.
Association of Elevated Soluble P-Selectin Levels and Fetal Loss in Women with a History of Venous Thromboembolism.
Authors: Ay Cihan , Kaider Alexandra , Koder Silvia , Husslein Peter , Pabinger Ingrid 
Published: April 2020

Abstract
Pregnancy is a hypercoagulable state with an increased risk of venous thromboembolism (VTE). Furthermore, accumulating evidence support the hypothesis that pregnancy complications such as fetal loss are associated with both inherited and acquired thrombophilic defects which may predispose to thrombosis of the placental vasculature and lead to subsequent fetal loss. In recent studies the cell adhesion molecule P-selectin has been identified to be a strong risk factor for VTE. Interestingly, soluble P-selectin (sP-selectin) plasma levels were previously reported to increase during pregnancy. Whether P-selectin is also associated with fetal loss in women with a history of venous thromboembolism, is not known yet. Therefore, the aim of our present study was to investigate the significance of elevated sP-selectin levels for fetal loss (e.g. miscarriage or stillbirth) in women with a history of VTE. We retrospectively evaluated data on pregnancy-associated complications in 304 women (mean age [+/-SD]: 45.6 [+/-11.5] yrs) with a history of VTE. sP-selectin plasma levels were measured using a sensitive ELISA (sP-selectin Immunoassay, R&D Systems®, Minneapolis, MN, USA). At the time of measurement of sP-selectin none of the women was pregnant and did not have an acute event of VTE. The mean age (±SD) of patients at the time of the VTE event was 31.3 (+/- 8.4) yrs. The prevalence of miscarriage (defined as intrauterine fetal death before the 24th week of gestation or when fetus weighed &lt;500 g) in our study population was 21.8% and the prevalence of stillbirth (defined as intrauterine fetal death at or after the 24th week of gestation) was 4.3%. Median (interquartile range [IQR]) sP-selectin level of the total study population was 38.0 [21.7-44.4] ng/mL. The cut-off point for elevated sP-selectin was set at 44.4 ng/mL, which represents the 75th percentile of sP-selectin levels of the study population. The prevalence of stillbirth was significantly higher in subjects with elevated sP-selectin levels compared to those with lower levels (10.5% vs. 2.6%, p=0.008), whereas no statistically significant difference in prevalence of miscarriage was observed between women with and without elevated sP-selectin (17.1% vs. 22.9%, p=0.303). The odds ratio [95% CI] of elevated sP-selectin for stillbirth was 4.2 [1.5-12.7] and for miscarriage 0.7 [0.4-1.3]. In summary, elevated sP-selectin plasma levels were associated with a 4-fold risk for stillbirth in women with a history of VTE. Our data support a possible role of P-selectin in the aetiology of late pregnancy complications.
Microparticle-Associated Tissue Factor Activity and Venous Thrombosis in Multiple Myeloma.
Authors: Leebeek Frank , Yuana Yuana , Osanto Suzanne , de Maat Moniek , Auwerda Hajo , Bertina Rogier , Sonneveld Pieter 
Published: April 2020

Abstract
Background: Multiple myeloma (MM) is associated with an increased risk of venous thrombo-embolic complications. The pathogenesis of venous thromboembolism (VTE) in MM is not yet clarified. Recently an association between microparticle-associated tissue factor (MP-TF) activity in plasma and thrombosis in patients with adenocarcinoma has been reported. Aim of this study was to measure MP-TF activity in patients with MM before and after treatment with chemotherapy and to investigate whether MP-TF activity is associated with VTE in these patients.
Patients and results: MP-TF activity was assessed in 123 patients with newly diagnosed multiple myeloma who were eligible for combination chemotherapy and in 20 healthy volunteers. MP-TF activity levels were higher in untreated MM patients compared to the healthy volunteers (17.8 fM Xa/min [8.8–32.8] (median [Inter Quartile Range] versus 4.7 fM Xa/min [2.3–6.6], p&lt;0.001). After three courses of chemotherapy the level of MP-TF activity had decreased significantly to 11.8 fM Xa/min [7.0–18.3], but were still higher than in controls (P&lt;0.001). VTE occurred in 12 patients (10%). In MM patients with VTE the level of MP-TF activity remained elevated after induction chemotherapy (17.9 [9.3– 36.3]), in contrast to MM patients not developing VTE (11.1 [6.9–16.6] ). MP-TF levels after chemotherapy were significantly associated with the risk of VTE, with a 4% increase of VTE risk for every fM Xa/ min MP-TF activity increase (95% CI: 1–8%, p=0.03)
Conclusion: MP-TF activity levels in plasma are increased in patients with multiple myeloma and they decrease significantly after induction chemotherapy. Persistent high MP-TF activity is observed in patients who developed VTE during the induction phase chemotherapy, and is associated with VTE risk. Our findings suggest that MP-TF activity levels may have a pathogenetic role in VTE in multiple myeloma patients.
Protein Kinase Cδ Differentially Regulates Platelet Functional Responses.
Authors: Chari Ramya , Getz Todd , Nagy Bela , Bhavaraju Kamala , Mao Yingying , Bynagari Yamini Saraswathy , Murugappan Swaminathan , Nakayama Keiko , Daniel James , Kunapuli Satya P. 
Published: April 2020

Abstract
Protein Kinase Cδ (PKCδ), a novel PKC isoform is expressed and activated in platelets downstream of PARs and GPVI receptors. In the current study, the role of PKCδ in regulating platelet functional responses was investigated using a pharmacological inhibitor, (δV1-1)TAT (a PKCδ inhibitor) in human platelets. These studies were further confirmed by a knockout approach using PKCδ+/+ and PKCδ−/− mice. In both human and murine platelets, PAR4-mediated dense granule secretions were inhibited, whereas GPVI-mediated dense granule secretions were potentiated. Furthermore, α-granule secretions and thromboxane A2 (TXA2) generation were differentially regulated in murine platelets.. These data suggest a differential role for this isoform in regulating dense granule secretion, α-granule secretion and TXA2 generation. Previous studies have shown that PAR-mediated fibrinogen receptor activation is regulated by a Calcium-dependent and a PKC-dependent pathway. The contribution of PKCδ to PAR-mediated fibrinogen receptor activation was studied by pretreating human and murine platelets with BAPTA. Our results showed a inhibition of AYPGKF-induced aggregation in human and murine platelets in the presence of BAPTA and fibrinogen. These results suggest a small contribution of PKCδ to PAR-4- mediated platelet aggregation and aIIbb3 activation. The in vivo significance of PKCδ was tested using a FeCl3 injury model. While the wildtype mice occluded in 7 minutes, PKCδ −/− mice occluded after 4 minutes of injury with 10 % FeCl3. Therefore, we conclude that PKCδ regulates platelet functional responses such as dense, α-granule secretions, TXA2 generation downstream of both PARs and GPVI receptors, contributes to PAR-4-mediated fibrinogen receptor activation ex vivo and plays a critical role in the thrombus formation in vivo. This study is supported by predoctoral fellowships to Ramya Chari and Swaminathan Murugappan from American Heart Association, Great Rivers affiliate.
The Role of Livin, An Inhibitor of Apoptosis Protein, in Thrombopoiesis.
Authors: Abd-Elrahman Ihab , Deutsch Varda , Hershko Klilah , Noyman Tsahi , Lazar Itay , Nachmias Boaz , Perlman Riki , Ben-Yehuda Dina 
Published: April 2020

Abstract
The exact mechanism of platelet production is poorly understood. A relationship between activation of the apoptotic cell machinery and the formation of pro-platelets has been established. In this study we show that Livin plays a major role in this process. Livin is a member of the Inhibitor of Apoptosis Protein (IAP) family, a novel family of intracellular anti-apoptotic proteins that act by binding and inhibiting caspases. We found that Livin mRNA and protein are expressed in platelets and Livin was also clearly detected by immunohistochemistry staining in normal mature bone marrow megakaryocytes (MK). Overexpression of Livin was also demonstrated in MK of patients with various hematological diseases such as ITP, MDS, Hodgkin’s disease, ET and PV. An in vitro model was established to evaluate the potential role of Livin in thrombopoiesis. The human erythroleukemic cell line, LAMA-84, was induced by a phorbol ester (PMA) to differentiate to MK. Differentiation was characterized by upregulation of the MK marker CD41 from 6% to &gt;60% and downregulation of the erythroid cell marker CD71 from 97% to 70%. Increased cell size, ploidy, and DNA synthesis, all markers of MK differentiation, were detected by flow cytometry. Upon differentiation induced by PMA, LAMA-84 cells formed pro-platelets and produced functional platelets capable of aggregation. This thrombopoiesis was accompanied by Livin overexpression. Moreover, overexpression of Livin by transfection decreased the percentage of the undifferentiated CD71+ cells to 53 % compared to 73 % in control cells (p= 0.007) and increased the production of platelets by two folds compared to control cells. Overexpression of Livin reduced caspase 3 activity and inhibited MK apoptosis. Our results show that anti-apoptotic Livin plays a role in thrombopoiesis possibly by protecting the maturing MK from apoptosis, thus affording more time for efficient platelet production by longer living pro-platelets. We are currently using this new cell line model to further characterize the molecular relationship between apoptosis and thrombopoiesis.
Impaired Activation of Platelets Lacking Protein Kinase C Theta (PKCΘ) Isoform.
Authors: Nagy Bela , Bhavaraju Kamala , Getz Todd , Bynagari Yamini Saraswathy , Kim Soochong , Kunapuli Satya P. 
Published: April 2020

Abstract
Protein kinase C (PKC) has been implicated in platelet functional responses, but the contribution of individual isoforms has not been directly evaluated. PKCΘ is activated by glycoprotein VI (GPVI) and protease-activated receptor (PAR) agonists, but not by ADP. In human platelets, PKCΘ-selective receptor for activated C kinase (RACK) antagonistic peptide inhibited agonist-induced aggregation and secretion. Consistently, in murine platelets lacking PKCΘ, GPVI- or PAR-mediated aggregation and secretion were also impaired. Previously, fibrinogen receptor has been shown to be activated independently by calcium and PKC pathways. In the presence of dimethyl BAPTA, AYPGKF-induced platelet aggregation was inhibited by PKCΘ antagonistic RACK peptides, suggesting a role for this isoform in PKC-dependent fibrinogen receptor activation. In addition, the levels of thromboxane A2 (TXA2) release measured in GPVI and PAR-mediated activation of PKCΘ −/− murine platelets, were significantly lower compared to WT platelets. Moreover, agonist-induced extracellular-signal regulated kinase (ERK) phosphorylation was also significantly decreased in PKCΘ −/− murine platelets, which could be contributing to decreased TXA2 levels. PKCΘ −/− mice displayed unstable thrombus formation and prolonged arterial occlusion in the FeCl3 in vivo thrombosis model versus WT mice. In conclusion, PKCΘ isoform plays a significant role in platelet functional responses downstream of GPVI and PARs.
Role of Glycoprotein Ibα Mobility in Platelet Function.
Authors: van der Wal Dianne E , Verhoef Sandra , Schutgens Roger EG , Peters Marjolein , Akkerman Jan-Willem 
Published: April 2020

Abstract
For preservation of platelet concentrates, storage at 0°C might be superior to current storage procedures at 22°C since cold storage reduces bacterial growth and might better preserve platelet functions. A major problem is that cold damages the glycosylation of the von Willebrand Factor (VWF) receptor Glycoprotein (GP) Ib&alpha, exposing N-acetyl- D-glucosamine (GlcNAc) residues and inducing receptor clustering. We investigated the effect of dispersed and clustered conformations of GPIb-alpha on the haemostatic functions of the receptor. We deduced the glycosylation state from binding of succinylated wheat germ agglutinin (sWGA) and clustering from binding of AN-51 antibody directed against aa 1–35 which differed between dispersed and clustered GPIb-alpha. We compared different GPIb-alpha conformations with anchorage to the membrane- and cytoskeleton and VWF/ristocetin (VWF*)-induced platelet functions and confirmed specificity for GPIb-alpha and VWF effects by control studies in GPIb-alpha- and VWF-deficient platelets. VWF*- and cold-treatment released GPIb-alpha from the membrane skeleton, inducing sWGA-binding, clustering and anchorage to the cytoskeleton. Addition of GlcNAc restored dispersed GPIb-alpha after cold but not after VWF*-treatment. VWF*- induced aggregation and adhesion to coated VWF under flow were little affected by changes in GPIb-alpha conformation but agglutination, spreading and formation of stress fibers strongly depended on clustered GPIb-alpha. Following cold-induced clustering of GPIb-alpha subsequent incubation at 37°C triggered formation of thromboxane (Tx)A2 in a GPIb-alpha-dependent manner but independent of added or endogenous VWF. GlcNAc strongly inhibited VWF*-induced agglutination, spreading on coated VWF and VWF-independent TxA2 formation. We conclude that incubation at 0°C triggers clustering of GPIb-alpha which activates platelets independent of its ligand VWF.
Genetic Modulators: Prevalence and Their Effect on the Phenotype of HbE/β Thalassemia Patients.
Authors: Sharma Vineeta , Saxena Renu 
Published: April 2020

Abstract
HbE, a β-globin chain variant occurs due to GAG to AAG i.e. Glu-Lys substitution at codon 26.In heterozygous and homozygous state it usually remain asymptomatic but the compound heterozygous state with β thalassemia (HbE/β thalassemia) results in a variable and often severe anemia, with the phenotype ranging from transfusion dependence to a complete lack of symptoms. The basis of the interaction and the cause of the variability remains unexplained. Some of the possible explanations for the observed variable clinical severity are coinheritance of mild β mutations, Effect of alpha gene number and increased production of HbF. Aims of the study are to determine the frequency of primary (β mutations) and secondary modulators (α deletion and α triplication, XmnI polymorphism, HPFH-3 and Asian-Indian Inv/DelGγ(Aγδbeta;)0 deletion) in HbE/β thalassemia patients and to study their effect on the phenotype of the Patients. Subjects were 180 HbE/β thalassemia patients diagnosed by HPLC and confirmed by PCR-RFLP. Patients were divided into three subgroups according to a scoring system based on seven clinical criteria as mild (score 0–3.5)[Gp1], moderate (score 4–7)[Gp2] and severe (score 7.5–10) [Gp3]. β-mutations were studied by ARMS PCR, XmnI polymorphism was studied by PCR-RFLP while α deletions, HPFH-3 and Asian-Indian Inv/DelGγ (Aγ δβ)0 deletions were studied by GAPPCR and alpha triplication by a modified single tube PCR. The frequency of β mutation was almost similar in all the groups (IVS 1–5 G-C (58.8%) was the predominant one). α deletion was found in 19 (10.5%) out of which 13 (12 αα/−α3.7 & 1 αα /--SA) were from Gp1 and 6 (αα/-α3.7) were from Gp2. α triplication was found in 11(6.1%) out of these 11 patients 8 were (αα/αααanti3.7) from Gp3 and 3 were (αα/αααanti3.7) from Gp2. XmnI was found in 112(62.2%), out of which 98 were heterozygous (+/−) and 14 were homozygous (+/+).XmnI +/+ was present in 10 Gp1 & 4 Gp2 patients, while XmnI +/− was present in 36 Gp1 ,43 Gp2 & 33 Gp3 patients.HPFH-3 deletion was present in 8 (4.4%) out of which 5 were from Gp1 and 3 were from Gp2. Asian-Indian Inv/DelGγ (Aγδβ)0 deletion was present in 2(1.1%) patients one from Gp2 and another from GP3. Primary modulator (β mutation) is not the actual factor responsible for the disease heterogeneity. Patients with coexisting α deletion required lesser transfusions and had less severe phenotype while patients with α triplication were on frequent transfusions and had severe phenotype. XmnI polymorphism in homozygous state was observed to alleviate the severity while in heterozygous state it had no effect on the disease severity except the delay of the onset of disease up to some extent. Patients having HPFH3 deletions and Asian-Indian Inv/DelG γ (Aγδβ)0 deletion showed mild to moderate disease severity. Thus it can be concluded that secondary modulators play an important role in the disease heterogeneity of HbE/βthalassemia.