bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evaluation of Schistocyte Analysis By a Novel Automated Digital Cell Morphology Application
Authors: Hervent Anne-Sophie , Godefroid Maaike , Cauwelier Barbara , Van Hoof Achiel , Billiet Johan , Emmerechts Jan 
Published: October 2019

Abstract
Introduction: The CellaVision Advanced Red Blood Cell (RBC) Software Application is a new software for advanced morphological analysis of RBC which automatically performs a preliminary characterization and grouping of RBC into 21 morphological categories, including schistocytes. Upon automated classification, the software offers the possibility of reclassification of RBC by the operator. The aim of this study was to evaluate the schistocyte analysis by the CellaVision Advanced RBC Application.
Methods: Schistocyte counts were evaluated comparing the automated count on a CellaVisionTM DM96, both before and after reclassification, with the reference manual microscopic method. 76 samples of both hospitalized patients and healthy donors were analyzed.
Results: Within-run, between-run and between-observer coefficients of variation were lower when counted with the CellaVision compared to the manual microscopic count. The very high sensitivity but rather poor specificity implicates the need for reclassification by the operator, following automated analysis. After reclassification, method comparison studies revealed good agreement with the manual microscopic method, with however slightly higher values of schistocytes for the automated analysis.
Conclusion: The CellaVision Advanced RBC Software Application provides a sensitive, accurate and reproducible measurement of schistocytes in peripheral blood. Furthermore, it is an easy-to-use software and an excellent teaching tool that might contribute to standardization in the investigation of schistocyte-related conditions.

Disclosures
No relevant conflicts of interest to declare.
Could Cobalamin Deficiency Mimic a Clonal Aberration Cytogenetic? a Case Report
Authors: Bellesso Marcelo , Dias Daniela Ferreira , Centrone Renato Torrescasana , Garcia Laura Yolanda Chialanza , Lopes Annelise Correa Wengerkievicz , Santucci Rodrigo , Pinto Ana Paula Almeida Rocha Teixeira , Silva Leniza Pola , Oliveira Veronica Ramos Souza 
Published: October 2019

Abstract
A 32 years old female patient presented to our service complaining asthenia, apathy and loss of her power to work. It was observed, by laboratory test, pancytopenia (Hb: 6.0 g/dL; WBC: 2,100/mm³, platelets 95,000/mm³, Reticulocytes 0.8%) associated with marked elevation of LDH 1,800 U/L (normal range: 240 – 480U/L). Bone marrow aspirate demonstrated morphologic features of megaloblastic anemia. Moreover, low serum concentration of vitamin B12 < 150pg/mL (normal range: 200 – 95-pg/mL), Folic acid 15,9ng/mL (normal range: 3 – 17 ng/mL) confirmed Megaloblastic anemia by cobalamin deficiency. It was not evidenced gastritis. It was initiated the treatment with vitamin B12. However, in the next clinical attendance it was observed an unexpected cytogenetic result: 46,XX,del(9)(q13q22)[3]/76~78,XXX,+1,+3,+4,+9,+11,+12,+17,+20,+21[cp3]/46,XX[34].
It was interpreted that del(9)(q13q22)[3] could be a clonal cytogenetics aberration and the others data due to defects in synthesis of DNA by cobalamin deficiency. It was important, because in diagnosis of Megaloblastic anemia frequently it is not included cytogenetic analysis of bone marrow cells and a new cytogenetic analysis has become necessary due to this data. After two months, a complete hematological recovery was achieved and six months later, bone marrow aspirate and cytogenetic analisys were repeated. Normal morphologic bone marrow cells and normal cytogenetic: 46,XX[20] were evidenced. Therefore, it was really hard to conclude this case. First, cobalamin deficiency could promote this clonal deletion or, as a second hypothesis, a clonal cytogenetic with low proliferative rate was selected due to inefficient hematopoiesis, by vitamin B12 deficiency, and after the recovery of the hematopoiesis, this clonal was suppressed.

Disclosures
No relevant conflicts of interest to declare.
Experience of a Single Latin-American Institution Using a Colloidal Iron Oxide Coated with a Semisynthetic Carbohydrate (ferumoxytol) for Iron Deficiency Anemia (IDA) in Patients with Intolerance or Treatment Failure with Oral Iron
Authors: Ovilla Roberto , Calles-Payan Dannia , Reynolds-Ocampo Lucia A , Ruiz-Reyes Gabriela , Balderas-Delgado Carolina , Montesinos-Ovilla Andres , Cabrera Garcia Alvaro 
Published: October 2019

Abstract
Tolerance to oral iron and low percentage of absorption in the gut has provoked the Iron Deficiency Anemia patients (IDA) fail to treatment or they have symptoms of anemia even more than 12 weeks until recovery. Intravenous iron is an alternative to minimize risk and treatment-related adverse effects and maximize profits in the short and long term. The number of patients who have access to ferumoxytol is reduced in developing countries, therefore experience in the treatment with this type of iron is limited. Patients received ferumoxytol 1020mg (two 510mg 8 days apart) and the response at 4 weeks of treatment by raising hemoglobin and clinical response was evaluated without transfusion support. From a total of 30 patients 76.6% (19) were women of whom 36.8% (7) were postmenopausal. The average age was 45.3 years (range 21-76 years). Before starting treatment, the mean serum iron, ferritin and rate of transferrin saturation was 35.6 mcg (range 8–174 mcg/dL), 13.8 ng/mL (range 1.6–60 ng/mL), 10.8% (range 2-58%) respectively. The mean initial hemoglobin was 10.5 g / dL (range 7.1-13.7g/dL), reaching an average of 13.03 g dL (range 9.4 - 16 _g/dL) at 4 weeks, raising 2.52 g dL (0.3 - 5.4g / _ g/dL) in 28 days. The percentage of patients who achieved an increase of > 2 g/dL of hemoglobin was 70% (21). The initial symptoms attributed to anemia disappeared in 66.6% within the first 2 weeks and the rest in 4-5 weeks. Adverse effects more frequently than 2%: fatigue 16.6% (5), headache 10% (3), nausea and dizziness 10% (3), peripheral edema 6.6% (2) and hypersensitivity 3.3% (1). All previous were easily controlled with medications. Despite the low number of patients treated due to the difficult accessibility of the drug in our population, ferumoxytol was shown to be an attractive option that rapidly improves symptoms with a satisfactory safety profile.

Disclosures
No relevant conflicts of interest to declare.
CD14 Mediates Phagocytic Activity during the Granulocytic Differentiation Process in Acute Promyelocytic Leukemia Cells
Authors: Hsu Hui-Chi , Tsai Wen-Hui , Lin Yu-Chieh 
Published: October 2019

Abstract
All-trans retinoic acid (ATRA) can induce acute promyelocytic leukemia (APL) cells differentiation into mature granulocytes. CD14 and Toll-like receptor 4 (TLR-4) play an important role in the phagocytic activity of macrophage, however, their role during granulopoiesis is still unclear. In this study, we determined the role of CD14/TLR-4 in the development of phagocytic activity in NB4 APL cells after induction into the process of granulocytic differentiation by ATRA. Flow cytometry analysis demonstrate that, during ATRA treatment for 6 days, the phagocytic activity of NB4 cells in engulfing either fluorescein-latex beads or idarubicin-induced apoptotic cells increased in a time-dependent manner, and the level of CD14 expression on NB4 cells was also significantly increased in a time dependent manner, though its level was only minimally expressed in ATRA-untreated NB4 cells. However, TLR-4 was constitutionally expressed in ATRA-untreated cells and its level did not changed significantly during the first 5 days of ATRA treatment. Further study demonstrates that the phagocytic activity of ATRA-NB4 cells was significantly inhibited by pre-treating cells with antibodies specific to either CD14 or TLR-4 before phagocytosis assay. In exploring the role of CD14/TLR4 associated signal transduction mediators, NF-κB and IRF-3, we further demonstrate that the phagocytic activity of ATRA-NB4 cells in engulfing beads was significantly inhibited when cells were pretreated with either a NF-κB inhibitor (BAY 11-7082) or an IRF-3 inhibitor (SP600125). However, this activity in engulfing apoptotic cells was only significantly inhibited by pretreatment with BAY11-7082, but not by pre-treatment with SP600125. Finally, our results indicate that the level of CD14(+) microparticles (MPs) released by ATRA-NB4 cells was significantly enhanced when those cells were induced into the process of apoptosis by pre-treatment with idarubicin. Moreover, by incubation with MPs derived from apoptotic ATRA-NB4 cells, the phagocytic activity of living ATRA-NB4 cells in engulfing apoptotic cells was significantly enhanced, and this phagocytic activity was also significantly inhibited by pre-treating MPs with antibody specific to CD14 before phagocytic assay. We conclude that CD14 contributes to the phagocytic activity of APL cells during the process of granulocytic differentiation.

Disclosures
No relevant conflicts of interest to declare.
Dysregulated Choline Metabolism in T-Cell Lymphoma and Its Relationship to Oncogenesis
Authors: Xiong Jie , Wang Li , Zhao Weili 
Published: October 2019

Abstract
T-cell lymphoma (TCL) is the malignant proliferation of CD3+ lymphocytes and represents the main type of non-Hodgkin's lymphoma. TCL patients are generally resistant to conventional chemotherapy and have poor clinical outcome. Therefore, biomarker commonly expressed and closely related to tumor progression needs to be further investigated in TCL, helping to develop targeted therapeutic approaches and to eventually improve prognosis of the patients.
Growing evidences show that metabolic enzymes regulate key oncogenic signaling pathways and play an essential role on tumor progression. Here serum metabolomic analysis was performed in 45 patients with T-cell lymphoma (TCL) and 50 healthy volunteers, and the results showed that dysregulation of choline metabolism occurred in TCL. (Figure 1A-B) Further verified by real-time PCR, choline kinase-¦Á (CHKA), the main regulatory enzyme of choline metabolism, was overexpressed in patients with TCL, but without significant difference in the TCL subtypes. (Figure 1C-D)
In T-lymphoma cells, pharmacological and molecular silencing of Chok¦Á significantly decreased Ras-GTP activity, AKT and ERK phosphorylation, MYC oncoprotein expression (Figure 2A-C), leading to induction of tumor cell apoptosis and necropotosis (Figure 2D-H). Clinically, gene expression profile analyzed in tissue samples of TCL patients showed that altered genes were enriched in PI3K and MAPK pathways according to CHKA expression of the TCL cases (Figure 2I).
In a T-lymphoma xenograft murine model, Chok¦Á inhibitor CK37 remarkably retarded tumor growth (Figure 3A), and SUV intensity of the CK37-treated tumors was significantly decreased on the figure of 18F-FDG small-animal PET/CT (Figure 3B). As in vitro study, comparing with the untreated group, p-AKT, p-ERK, MYC expression, and activity of Ras-GTP were significantly decreased, while in situ cell apoptosis/necropotosis increased in the tumor of the CK37 group. (Figure 3C-F)
To our knowledge, our report is the first time to analyze serum metabolomic profile in TCL patients, and provides direct evidence of aberrant choline metabolism in this disease. Importantly, dysregulation of choline metabolism was present in all histological types of TCL studied, consistent with acute myeloid leukemia that metabolomic status is independent on morphological classification or genetic feature. Therefore, manifested with a heterogeneous group of disease, TCL may share the similar profile of disturbed choline metabolism and this metabolic pathway can be exploited as a source of biomarkers and a potential therapeutic target in TCL.
Figure 1 Figure 1. Figure 2 Figure 2. Figure 3 Figure 3.

Disclosures
No relevant conflicts of interest to declare.
Increasing TCR Zeta Expression and Maintaining the Clonality of T Cells from AML Patients after IL-2, IL-7 and IL-12 Induction
Authors: Shi Li , Chen Shaohua , Yang Lijian , Lu Yuhong , Luo Gengxin , Zhu Kanger , Li Yangqiu 
Published: October 2019

Abstract
T-cell immunodeficiency is a common feature in patients with leukemia, lower activation of T cells was one of reasons. The TCR zeta chain has emerged as a key subunit of the T-cell antigen receptor, which plays a central role in the signal-transducing events leading to T cell activation. The proliferation and activation of T cells may be inducted by T cell related cytokines. In this study, we explored the change of TCR zeta gene expression and the clonality of T cells after induction with different immune cytokines, including IL-2, IL-7 or IL-12. CD3+ T cells sorted from peripheral blood of 4 cases with AML were induced with different immune cytokines, including IL-2, IL-7, IL-12, anti-CD3 and anti-CD28 antibodies in vitro. The expression levels of TCR zeta gene and related genes in T cells before and after induction were then analyzed by fluorescence quantitative RT-PCR. The distribution and clonality of TCR Vβ subfamily T cells were analyzed by RT-PCR and Genescan techniques. Increasing expression levels of TCR zeta gene and zap-70 (TCR zeta chain associated-protein) gene in CD3+T cells from AML patients were found after induction with single stimulating factor or the combination with different cytokines, while the expression of FcεRIγ (TCR ζ gene complementary factor) was down-regulated. We further compared the T cell clonality in CD3+T cells from AML patients after cytokine induction, eight to 22 TCR V β subfamilies could be detected in T cells from AML cases, most of them displayed polyclonal expansion. The number of expressed TCR Vβ subfamilies was increased without the change of clonality in T cells induced by CD3+CD28+IL7. In conclusion, TCR zeta gene and its related genes could be upregulated through induction with different cytokine combination such as IL-2, IL-7 and IL-12, therefore to improve the T cell activation in patients with AML. And the main effect of cytokines might to maintain the T cell clonality and nonspecific amplification of T cell clones. Further investigation can be designed to amplify the specific anti-AML TCR Vβ clones using AML associated antigens and such cytokine combination.

Disclosures
Shi: National Natural Science Foundation of China (no. 81100353, 81270604), the Fundamental Research Funds for the Central Universities (No. 21611447, 21612116), And Medical Science Foundation of Guangdong Province(A2011325). : Research Funding.
Treatment Reality of Patients with Immune Thrombocytopenia (ITP) in Routine Care
Authors: Weide Rudolf , Feiten Stefan , Friesenhahn Vera , Heymanns Jochen , Kleboth Kristina , Thomalla Jörg , van Roye Christoph , Köppler Hubert 
Published: October 2019

Abstract
Introduction: New treatment options like thrombopoietin receptor agonists (TRAs) and rituximab have been introduced into the clinic which have found their way into national and international treatment guidelines. The aim of this study was to answer the following questions concerning diagnosis and treatment of patients with ITP in routine care: How are patients diagnosed and treated?Which sequences of therapy are applied?How many patients die due to bleeding complications?
Methods: All patients with ITP diagnosed between 06/1995-06/2014 in a community-based oncology group practice in Germany were analyzed retrospectively.
Results: 402 patients with a median age of 55 (7-90) were evaluated. 57% were female and 43% male. 357 (89%) were classified as primary ITP and 311 (77%) as having chronic ITP. In 234 patients (58%) a bone marrow biopsy was part of the diagnostic work up. Only 191 patients (48%) needed therapy. First line therapy (n=191) were steroids in 81%, intravenous immunoglobulins (ivIgG) in 12% and ivIgG plus steroids in 6%. Second line therapy (n=102) were ivIgG in 49%, steroids in 23%, ivIgG plus steroids in 17%, other immunosuppressive agents in 11% and splenectomy in 5%. Third line therapy (n=63) was splenectomy in 22%, other immunosuppressive agents in 27%, steroids in 19%, ivIgG in 16%, 11% combination therapy, rituximab in 10% and TRAs in 5%. Fourth line therapy (n=38) consisted of steroids in 26%, splenectomy in 26%, immunosuppressive agents in 34%, ivIgG in 13%, rituximab in 11% and TRAs in 3%. Patients received a median of 2 lines (1-10) of therapy. Treatment modalities most frequently used were steroids in 93%, immunoglobulins in 56%, splenectomy in 21% and other immunosuppressive agents in 21% of patients. Rituximab and TRAs were used in 10% and 5% only. 75% of patients received a durable remission (complete or partial) after their last therapy. 10% showed no response, in 15% remission couldn't be evaluated due to external treatments. 146 patients (76%) are off treatment. 1 patient (0.2%) died due to bleeding complications.
Conclusions: Bone marrow biopsy is used as a diagnostic procedure in 58% of patients. Treatment modalities most frequently used are steroids, immunoglobulins, splenectomy and other immunosuppressive agents. Rituximab and TRAs are used infrequently. A high percentage of ITP-patients achieve durable remissions and ITP-related mortality is low.

Disclosures
No relevant conflicts of interest to declare.
Targeted DNA Sequencing for Hemophilia A, B, and von Willebrand Disease: Analytical Precision in Clinical Laboratories and Improved Disease Characterization
Authors: Severino Patricia , Oliveira Liliane Santana , Torres Natalia , Guerra Joao Carlos , Hamerschlak Nelson , Rezende Suely M 
Published: October 2019

Abstract
Hemophilia A, B, and von Willebrand disease correspond to more than 90% of all inherited bleeding disorders associated with coagulation factor deficiencies. Symptoms between these deficiencies may vary greatly and yet are often phenotypically similar. Bleeding episodes can range from mild to severe, at times with life threatening hemorrhages. Currently, biochemical assays are performed to assess the function of each coagulation factor, but diagnosis remains cumbersome and prone to multiple sources of variability between laboratories. Genetic evaluation allows for the examination of multiple coagulation factor genes simultaneously and may quickly identify possible causes to the disease. Additionally, genetic testing should be more reproducible and readily comparable between clinical laboratories. In this work we evaluate the potential use of targeted sequencing of three coagulation factors genes – F8, F9 and VWF – for the concurrent diagnosis and characterization of hemophilia A, B, and von Willebrand disease samples. For targeted DNA sequencing we selected specific DNA probes using genomic coordinates spanning the complete intronic and exonic regions of the three genes, as well as flanking gene sequences. Eleven hemophilia A samples and four hemophilia B samples, clinically characterized and submitted to Sanger sequencing for F8 and F9 genes coding regions, respectively, were included in this study. Our results indicate that even though DNA quality may be ideal for traditional DNA sequencing, enrichment techniques require more intact fragments, as reflected by variations in sequencing coverage between samples: quadruplicate results per sample showed 100X coverage varying from 80% of sequenced regions to less then 20%. Point substitutions found in F9 genes by Sanger sequencing were confirmed by targeted sequencing, but results for F8 gene were less satisfactory, in agreement with probe design limitations at this point. Of interest for hemophilia A patients, four samples possessed, in addition to the alterations in F8, point mutations in VWF. Probe design and sequencing parameters did not allow for the identification of F8 intron 1 and intron 22 inversions, frequent alterations in hemophilia A, but optimization procedures are currently underway. We conclude that targeted sequencing approach may be a viable and more complete solution for the diagnosis and management of hemophilia A, B and von Willebrand disease.

Disclosures
No relevant conflicts of interest to declare.
The PUP-GCP Clinical Trial: A Low Inhibitor Rate in Previously Untreated Patients with Severe Hemophilia a Treated with Octanate
Authors: Knaub Sigurd , Klukowska Anna , Komrska Vladimir , Laguna Pawel , Vladimir Vdovin , Jansen Martina 
Published: October 2019

Abstract
Background :
Octanate is a highly purified, double virus inactivated, human plasma-derived factor VIII (FVIII) concentrate with all coagulation FVIII bound to its natural stabilizer VWF in a VWF:RCo/FVIII:C ratio of approximately 0.4. A prospective clinical trial has been initiated in 2000 in order to assess the immunogenicity, efficacy and tolerability of Octanate in previously untreated patients (PUPs).
Materials and Methods:
Patients with severe hemophilia A without previous exposure to FVIII or FVIII-containing products were enrolled. Inhibitor screening, using the modified Bethesda method, was performed prior to treatment, starting every 3-4 EDs (ED 1-20), and afterwards every 10 EDs (ED 21-100), but at least every three months. Efficacy and tolerability were assessed by a 4-point verbal rating scale.
Results:
The study is clinically completed, after fifty-one subjects with severe haemophilia A, at screening 0,01-5,61 years old (median 0,65 years; median 0,99 years at treatment start) have been enrolled. Three of them developed clinically relevant high titer inhibitors over the course of the study. Another two displayed transient inhibitors that disappeared spontaneously without changing the dose or dosing interval. All inhibitors developed under on-demand treatment and before ED 50. For 4 inhibitor patients an intron 22 inversion, for 1 inhibitor patient a large deletion of exons 7 – 12 was found.
From 51 subjects, 45 exceeded 50 EDs. Octanate was well-tolerated and the adverse event profile was consistent with the population studied. The haemostatic efficacy in prophylaxis and treatment of bleeding episodes was generally rated as “excellent” and no complication was reported for any surgical treatment.
Conclusion:
Despite frequent inhibitor testing and predominant on-demand treatment, the data indicate a low overall inhibitor rate for Octanate in patients who exceeded 50 EDs (5/45) of which only 3 (6.7%) were clinically relevant.

Disclosures
Knaub: Octapharma AG: Employment. Klukowska:Octapharma AG: Investigator Other. Komrska:Octapharma AG: Investigator Other. Laguna:Octapharma AG: Investigator Other. Vladimir:Octapharma AG: Investigator Other. Jansen:Octapharma AG: Employment.