bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of Genetic Polymorphisms Related to Folate and Homocysteine Metabolism In the Etiology of Recurrent Miscarriages
Authors: Guerra-Shinohara Elvira Maria , da Silva Giusti Kelma Cordeiro , Monteiro Nathalia Sierra , Lazaro Robson José , Seino Fernanda Midori , Bueno Carolina Tosin , Carvalho Mário Henrique Burlacchini , Filho Antônio Amorim , Chiattone Carlos , Zugaib Marcelo 
Published: October 2019

Abstract
Abstract 5136

Introduction:
Folic acid deficiency has been associated with obstetric complications such as preeclampsia, placental abruption and recurrent miscarriages (RM). Folic acid is a methyl group donating in various reactions. Low concentrations of 5,10-methylenetetrahydrofolate, the cofactor of enzyme thymidylate synthase, decrease the synthesis of thymidylate, which results in increased ratio deoxyuridylate monophosphate/deoxytimidylate monophosphate (dUMP/dTMP) and increased incorporation of deoxyuridylate triphosphate (dUTP) to DNA. The removal of dUTP by DNA-glycosylase can cause permanent damage to DNA, which may lead to apoptosis or increase the risk of developing cancer. Polymorphisms in genes of enzymes (MTHFR - methylene tetrahydrofolate reductase, MTR - methionine synthase and MTRR - methionine synthase reductase) and also in the gene of the reduced folate carrier (RFC1) were related to reduced folate and increased total homocysteine concentrations and have been associated as risk factors for RM.

Material and Methods:
171 women with a history of three or more recurrent miscarriages and 95 healthy women with no history of abortion and having two or more normal babies were included. Weight and height of women were obtained and the body mass index (BMI) was calculated. The presence of antibodies (ANA and anti-DNA) was evaluated using immunofluorescence kits. The genotypes of the polymorphisms MTHFR c. 677C>T, MTR c. 2756A>G and RFC1 c.80G>A were obtained by PCR-RFLP, while genotyping for polymorphisms MTRR c. 66A>G and MTHFR c. 1298A>C was made by real time PCR. Multivariate logistic regression model (forward conditional) was used to obtain the odds ratio and its 95% confidence intervals of having MR (dependent variable). The independent variables were: quartiles of BMI, age range, positive ANA (titer of 1/40), positive anti-DNA (titre 1/10), genotypes for the MTHFR c. 677C>T, MTHFR c.1298A>C, MTR c. 2756A>G, MTRR c. 66A>G and RFC1 c. 80G>A.

Results:
No differences between the groups were observed for serum total homocysteine or allele frequencies for MTHFR c. 677C>T, MTHFR c. 1298A>C, MTR c. 2756A>G and RFC1 c.80G>A polymorphisms. In a conditional logistic regression analysis the risk of RM was significantly associated with BMI OR [95% CI] = 1.40 [1.02, 1.93] per quartile increase in BMI), positive anti-DNA OR [95% CI] = 7.24 [0.92, 57.25], positive ANA OR [95% CI] = 2.48 [1.21, 5.08], and AA genotype for MTRR c. 66A>G polymorphism (OR [95% CI] = 2.19 [1.16, 4.12].

Conclusion:
The etiology of RM is multifactorial and it is associated with increasing of BMI, presence of autoantibodies and AA genotype for MTRR c. 66A>G polymorphism.

Disclosures:
No relevant conflicts of interest to declare.
Study On Immune Mechanism of Human Marrow Mesenchymal Stem Cells Adjusting Dendritic Cells for Treatment of Aplastic Anemia
Authors: Xiao Yang , Zhang Leqin 
Published: October 2019

Abstract
Abstract 5137

Objective
(1)To explore whether MSC has inhibiting effect on the proliferation ofAA patients' Tcell;(2)To discuss whether MSC affects T cell's proliferation via adjusting the growth of DCs.

Materials and Methods
(1) MSC were separated and cultured in vitro. Cell morphology was observed and the cell surface antigen was determined by flow cytometry.(2) Peripheral blood mononuclear cells were extracted from 20 patients suffered AA and then T lymphocytes were separated by nylon fiber column. Flow cytometry was applied to determine the surface antigen and subpopulation of T cell.(3)mononuclear cells were separated from normal human peripheral blood. DCs were prepared under the culture condition of recombination human granulocyte-macrophage colony stimulating factor (GM-CSF) and recombinant human interleukin-4 (IL-4). After acquiring the mature DCs induced by LPS, the phenotype analysis of DCs before and after culture was examined by flow cytometry, respectively.

Results
(1) After co-culture of MSC and T lymphocytes from AA peripheral blood, flow cytometry showed that the ratio of D8+ in T cells reduced significantly from 38.7% to 29.7 % (p < 0.05), whereas the CD4+ ratio increased from 24.9% to 34.9% significantly (p < 0.05). Meanwhile, ELISA analysis indicated that the concentration of IL-2 and IFN-γ were significantly decreased from 38.9 and 38.5 ng/L to 6.8 and 6.6 ng/L, respectively (p < 0.05). However, IL-4 and IL-10 increased from 2.8 and 2.9 to 5.3 and 8.3 ng/L, respectively (p < 0.05).(2) After the induction of immature DCs by LPS, flow cytometry showed that the expression of CD1a increased from 2.4% to 68.4% in the treatment without MSC, while that of CD14+ decreased from 83.6% to 3.5% (p < 0.05).(3) After the co-culture of mature DCs and MSC, the expression of CD14+ increased from 5.8% to 62.8% when the expression of CD1a, CD83 and CD80 decreased from 48.6%, 60.8% and 50.2% to 30.7%,40.9% and 20.3%, respectively.

Conclusions
Our study shows that (1)MSC inhibits the proliferation of T lymphocytes from AA p-atients by regulating CD8+ and CD4+; (2)futher study indicates that MSC can inhibit the growth of DCs and reverse the status of DCs from matureness to immatureness; (3)thus suggests the possible mechanism of MSC's inhibiting effect as follows: MSC decreases the liveness by controlling the growth of DCs and further inhibits its proliferation.

Disclosures:
No relevant conflicts of interest to declare.
Activation Markers of Thrombogenesis and Endothelial Dysfunction In End-Stage Renal Disease
Authors: Sharain Korosh , Davis Rachael , Hoppensteadt Debra , Cunanan Josephine , Litinas Evangelos , Bansal Vinod , Fareed Jawed 
Published: October 2019

Abstract
Abstract 5183

Introduction:
Over 500,000 patients currently have ESRD and it is estimated that another 200,000 will develop ESRD in the next 5 years. Hemostatic activation resulting in cardiovascular disease accounts for over 50% of the mortality seen in this patient population. Common sequelas seen in ESRD are DVT, PE, and ACS. Derangement of biochemical markers associated with these disease states have been observed in ESRD patients. In this study we profiled several markers of thrombogenesis and endothelial dysfunction to determine their levels in the ESRD population.

Materials/Methods:
117 ESRD patients on maintenance dialysis for at least 3 months over the age of 18 were included in this study. Additionally, 50 age matched control samples were obtained from healthy volunteers with no known kidney function. The markers measured include C-Reactive Protein (CRP), D-Dimer (DDMER), and Thrombomodulin (TM) which were detected by biochip array using the Randox Evidence Investigator (Antrim, United Kingdom). Additionally, Plasminogen Activator Inhibitor -1 (PAI-1) and Asymmetric Dimethylarginine (ADMA) were measured using commercially available kits by Diagnostica Stago (Paris, France) and DLD Diagnostika GmbH (Hamburg, Germany), respectively.

Result:
The mean, standard deviation, and p-value for each marker is tabulated below. Additionally, the fold increase of the ESRD samples compared to control samples were calculated and included in the table below. Compared to controls, all markers measured were increased in the ESRD samples, each of which demonstrated statistically significant p-values between the two groups. TM expressed a 5.9 fold increase in ESRD compared to controls, this was the highest fold increase amongst the markers tested. CRP, DDMER, PAI-1, and ADMA were increased at 4.4, 3.0, 1.4, and 1.6 times their control counterparts, respectively.

Conclusion:
The results in the table demonstrate that ESRD patients have a significant upregulation in markers of thrombogenesis and endothelial dysfunction. The upregulation of CRP, TM, D-DDMER, PAI-1, and ADMA is consistent with the increased thrombotic events detected in ESRD. The upregulation of the nitric oxide inhibitor ADMA points to disturbances in vascular lumen size regulation while CRP and TM suggest significant endothelial damage. Additionally, the upregulation of PAI-1 and DDimer indicates an activation of coagulation in ESRD. The combined disruption of endothelial regulation and endothelial damage along with the upregulation of thrombogenic markers helps to explain the significant prevalence of thrombosis and cardiovascular disease in ESRD. These markers should also be evaluated for their roles in kidney disease risk stratification and prognosis.

Disclosures:
No relevant conflicts of interest to declare.
Daunorubicin Induces Procoagulant Activity of Cultured Endothelial Cells through Phosphatidylserine Exposure and Microparticles Release
Authors: Fu Yueyue , Li Huibo , Li Wen , Dong Xiushuai , Hou Jinxiao , Xie Rui , Gao Chunyan , Yang Xue , Zhou Jin , Gilbert Gary E. , Shi Jialan 
Published: October 2019

Abstract
Abstract 5185
Administration of various chemotherapeutic agents is associated with an increased risk of thrombotic events. Although vascular endothelium plays a predominant role in blood coagulation and fibrinolysis, the effect of cytotoxic drugs on the procoagulant activity (PCA) of endothelial cells has not been well evaluated. Our study aims to investigate the possibility that daunorubicin, a chemotherapeutic agent, exerts prothrombotic effect on endothelial cells. We tested the impact of daunorubicin on phosphatidylserine (PS) exposure, endothelial microparticles (EMPs) release and consequent PCA. Human umbilical vein endothelial cells (HUVECs) were treated with daunorubicin (0.1, 0.2, 0.5, 1, 2 μ M) for 24 h. PCA of HUVECs was measured using clotting time and purified coagulation complex assays. Counts and PCA of EMPs were evaluated by flow cytometry and clotting time assay, respectively. Lactadherin was used as a novel probe for detection of PS exposure and EMPs release. We found that daunorubicin dose-dependently increased the PS exposure and consequent PCA of HUVECs. Moreover, daunorubicin treatment also enhanced the release of EMPs which were highly procoagulant. This increment was especially significant at 0.2 μ M of daunorubicin or over. Blockade of PS with lactadherin inhibited over 90% of HUVECs and EMPs PCA. However, anti-TF antibody had no significant inhibition effect. Our results demonstrate that daunorubicin treatment enhanced PCA of HUVECs through PS exposure and shedding of procoagulant EMPs. Lactadherin acts as an efficient anticoagulant in this process.

Disclosures:
No relevant conflicts of interest to declare.
Interleukin-32, α1 Anti-Trypsin (AAT-1) and Graft-Versus-Host-Disease
Authors: Marcondes A. Mario Q. , Tabellini Laura , Hansen John A. , Dinarello Charles A. , Deeg H. Joachim 
Published: October 2019

Abstract
Abstract 522
Graft-versus-host disease (GVHD) is an important complication of allogeneic hematopoietic cell transplantation (HCT). The role of various cell populations, cytokines and chemokines, present pre and post-transplantation, in the development of GVHD has been studied extensively.We investigated the potential role of Interleukin (IL)-32 in alloreactivity and GVHD. IL-32 is the protein product of the NK4 transcript first reported in IL-2 activated T lymphocytes and natural killer cells. IL-32 has pro-inflammatory and pro-apoptotic properties and induces expression of TNF-α in several cell targets. We used one-way mixed lymphocyte cultures (MLC) as a simple in vitro model of GVHD to determine IL-32 expression upon alloactivation. The α and γ isforms of (IL)-32 protein were 2-fold upregulated in allogeneic MLC compared to autologous controls (n=4, p=0.037). Concurrently, the concentrations of TNF-α, IL-6 and IL-8 in the allogeneic MLC supernatants were, as expected, upregulated significantly. This finding led us to evaluate IL-32 expression as a potential marker for GVHD after allogeneic HCT in 45 patients with either active acute GVHD or chronic GVHD, and 16 patients who did not show clinical evidence of GVHD. IL-32 mRNA levels in peripheral blood unsorted white blood cells, correlated with acute GVHD (RT-PCR values expressed as mean +/− SEM of the ratio of expression of IL-32/GUS-B = 1.01, p=0.011, vs control values IL-32/GUS-B = 0.23) but not with chronic GVHD, (IL-32/GUS-B = 0.26, p=0.16) (Figure 1). As the serine protease neutrophil proteinase 3 (PR3) has been shown to serve as activator of IL-32, and to process several inflammatory cytokines, including IL-32, TNF-α and IL-8, we postulated that the addition of the serine protease inhibitor α-1 anti-trypsin (AAT), would interfere with the processing of IL-32 by PR3, and as a result would lead to decreased proliferation of cells in MLC, and reduced cytokine production. To test this hypothesis, MLCs were treated with AAT at concentrations of 1–20 ug/ml and expression of IL-32 and PR3 were determined. In MLCs treated with AAT at the optimal concentration of 5 ug/ml, added to cultures on alternate days for a period of 7 days T lymphocyte proliferation was suppressed when compared to vehicle-treated MLC, (mean CPM=33.000 versus CPM=67.000; p=0.012). Concurrently there was a 2.5 fold decrease in IL-32 and PR3 protein levels (n=4, p=0.023). CONCLUSION: IL-32 is upregulated in patients with acute GVHD. Determination of IL-32 in patients with suspected GVHD may support the diagnosis and the decision to treat.AAT interferred with T cell activation and the release of cytokines, including IL-32, suggesting that administration of AAT may have therapeutic potential in patients with acute GVHD.

Disclosures:
No relevant conflicts of interest to declare.
Genome-Wide Analysis Identifies Regulators of Hematologic Parameters
Authors: Gieger Christian , Soranzo Nicole 
Published: October 2019

Abstract
Abstract SCI-10
Recent genome-wide association scans published by ours and other groups have revealed > 50 genetic loci associated with hematological parameters, including the count and volume of red and white blood cells and platelets. These variants map to known and novel regulators of hematopoiesis, and provide evidence for a high degree of connectivity guiding differentiation of the different blood elements. I will review progress in the field as well as ongoing efforts within the HaemGen consortium, where we have used statistical analyses, animal models and genome-wide functional annotation to address the in-depth genetic exploration of such parameters.

Disclosures:
No relevant conflicts of interest to declare.
Prevalence and Clinical Effect of TET2 Mutations in Cytogenetically Normal Acute Myeloid Leukemia (CN-AML)
Authors: Markus Birgit , Thol Felicitas , Morgan Michael A , Wagner Katharina , Heuser Michael , Krauter Juergen , Ganser Arnold , Damm Frederik 
Published: October 2019

Abstract
Abstract 1448
Mutations in the TET2 gene were recently identified in a variety of myeloid neoplasms including acute myeloid leukemia (AML). To investigate the frequency and clinical relevance of TET2 mutations in cytogenetically normal AML (CN-AML), the largest cytogenetic subgroup of adult AML, the total coding region of TET2 was analyzed by direct sequencing in 215 adult CN-AML patients younger than 60 years. All patients were uniformly treated on the AML SHG 0295 or AML SHG 0199 treatment protocols, both including double induction and intensive consolidation regimes. To test the prognostic value of TET2 mutations in the context of other prognostic molecular markers, mutational analyses for CEBPA, DNMT3A, FLT3-ITD, IDH1/2, MLL-PTD, ND4, NPM1, NRAS and WT1 and expression analyses for BAALC, ERG, EVI-1, IDH1, MLL5, MN1 and WT1 were performed. At least 1 sequence variation in TET2 was found in 16 out of 215 patients (7.4%). In 3 of these patients the somatic origin of the mutation could not be validated, so these patients were considered wildtype in all subsequent analyses. Mutations were detected throughout the entire gene with the highest frequency found in exon 3 (n=6). Patients harboring TET2 mutations showed higher platelet counts (P=.041), a higher frequency of concomitant NPM1 (11 of 13, P=.047) and DNMT3A (10 of 13, P=.001) mutations and tended to be older (P=.078) when compared to TET2 wildtype patients. The recently reported mutational exclusiveness of IDH1/2 and TET2 was validated in the present cohort (P=.042). No differences for complete remission (CR) rate (P=.76), relapse-free survival (RFS; P=.63) or overall survival (OS; P=.59) was observed based on the TET2 mutation status. When restricting the analyses to CN-AML patients assigned to the European LeukemiaNet (ELN) favorable-risk or intermediate1-risk groups (ELN-favorable risk-group: CN-AML patients with mutated CEBPA and/or mutated NPM1 without FLT3 -ITD, n=96; ELN-intermediate-1-risk group CN-AML with wild-type CEBPA and wild-type NPM1 and/or FLT3-ITD; n=106), the TET2 mutation status again did not impact on CR rate, RFS or OS. We conclude that in younger CN-AML patients, TET2 mutations occur with a lower prevalence compared to other myeloid malignancies such as MDS or CMML, and have no prognostic effect. Therefore, TET2 mutation analysis at present is not required for risk-stratified protocols in AML.

Disclosures:
No relevant conflicts of interest to declare.

Disclosures:
No relevant conflicts of interest to declare.
JAK2 mutations and CRLF2 rearrangements in Down Syndrome-Associated Acute Lymphoblastic Leukemia in Japan
Authors: Hanada Isamu , Terui Kiminori , Toki Tsutomu , Kudo Ko , Sato Tomohiko , Kamio Takuya , Sasaki Shinya , Takahashi Yoshihiro , Hayashi Yasuhide , Sugita Kanji , Kojima Seiji , Koike Kenichi , Kosaka Yoshiyuki , Kobayashi Masao , Ito Etsuro 
Published: October 2019

Abstract
Abstract 1451
Children with Down syndrome (DS) have a 10- to 20-fold increased risk of developing acute lymphoblastic leukemia (ALL). In DS-associated ALL (DS-ALL), the chromosome aberrations which are generally common in childhood ALL, such as hyperdiploidy and t(12;21), are less frequent. Recent studies have shown that activating JAK2 mutations and overexpression of cytokine receptor-like factor 2 (CRLF2) gene are identified in approximately 20% and 50–60% of DS-ALL in Western countries, respectively. Most of the patients with CRLF2 overexpression have been reported to be associated with interstitial deletions of the pseudoautosomal region 1 (PAR1) of the sex chromosomes and the P2RY8-CRLF2 fusion gene. In addition, one report showed that the activating CRLF2 F232C mutation was identified in about 10% of DS-ALL. However, there have been no studies to determine the incidence of these genetic aberrations in Asian patients with DS-ALL. In this study, 23 patients with DS-ALL in Japan were screened for mutations in the pseudokinase domain of the JAK2 gene, the P2RY8-CRLF2 fusion gene, and the CRLF2 F232C mutation by PCR/RT-PCR and direct sequencing. Fourteen patients, whose bone marrow RNAs were available, were also screened for CRLF2 overexpression by real-time quantitative RT-PCR. We identified the JAK2 R683G mutation in 2 patients (9%) and the P2RY8-CRLF2 fusion gene in 4 patients (17%). The CRLF2 F232C mutation was not detected in any patient. CRLF2 overexpression was observed in 2 of 14 patients examined (14%). Although bone marrow RNA was available in only 1 of 4 patients positive for P2RY8-CRLF2, high-level expression of CRLF2 was confirmed in this patient. The other patient with CRLF2 overexpression was negative for P2RY8-CRLF2, indicating the involvement of the other type of CRLF2 rearrangement, IGH@-CRLF2 in this patient. We also performed a preliminary study on JAK1, JAK3, and Interleukin-7 receptor-α (IL7R) mutations, and 14, 11, and 12 patients were screened for mutations in the pseudokinase domain of JAK1, JAK3, and exon 5 and 6 of IL7R, respectively. However, no mutations were identified in any patient. Our results show the lower incidence of CRLF2 rearrangements in DS-ALL patients in Japan than that in Western countries. Gene alterations other than CRLF2 rearrangements may contribute to leukemogenesis in Japanese patients with DS-ALL. To clarify if the incidences of the mutations in JAK1-3, CRLF2, and IL7R are also lower in DS-ALL patients in Japan than those in Western counties, more patients need to be studied.

Disclosures:
No relevant conflicts of interest to declare.
New and Highly Efficient Therapy for Treatment NPM-ALK Associated Lymphomas
Authors: Kenner Lukas , Laimer Daniela , Dolznig Helmut , Vesely Paul , Kollmann Karoline , Schiefer Ana-Iris Iris , Sexl Veronika , Staber Philipp B , Hoefler Gerald , Penninger Josef , Inghirami Giorgio , Ulrich Jaeger 
Published: October 2019

Abstract
Abstract 1659
Anaplastic Large Cell Lymphoma (ALCL) is a non-Hodgkin lymphoma of children and young adults with poor survival rates. ALCLs often occur with a chromosomal translocation that results in expression of the NPM-ALK oncoprotein. The molecular mechanism of NPM-ALK resulting in lymphoma growth are not fully understood. In a murine NPM-ALK lymphoma-model with T-cell specific deletion of cJun and JunB we could drastically delay lymphoma development and metastases. We identified PDGFRB as a direct transcriptional target of cJun and JunB. Blockage of the PDGFRβ activity significantly prolonged survival of NPM-ALK transgenic mice and severely reduced the growth of xenografted NPM-ALK tumors. In addition, we demonstrate that PDGFRβ is upregulated in human ALCL tumor specimens. Inhibition of the PDGFRβ in a terminal stage patient with NPM-ALK expressing ALCL which was refractory to any conventional therapy resulted in full remission within 10 days of treatment. We herwith certify the PDGFRB as a novel cJun/JunB target essential for ALCL development. Our data for the first time unravel a highly effective targeted therapy with the outlook to make ALCL a curable disease.

Disclosures:
Off Label Use: Imatinib for treatment of ALCL.
Plasma Membrane Loss in Maturing Human Reticulocytes Occurs Through Endocytosis of Large Glycophorin A- Containing Vacuoles Which Fuse with Autophagosomes Before Exocytosis
Authors: Griffiths Rebecca E , Kupzig Sabine , Cogan Nicola , Betin Virginie , Mankelow Tosti J , Lane Jon , Parsons Steve F , Anstee David J. 
Published: October 2019

Abstract
Abstract 177
The erythrocyte is one of the best characterized human cells. However, studies of the process whereby human reticulocytes mature to erythrocytes have been hampered by the difficulty of obtaining sufficient numbers of cells for analysis. We have developed an in vitro culture system permitting the production of millilitre quantities of functional mature human adult reticulocytes from peripheral blood CD34+ cells. Analysis of reticulocytes obtained from culture using confocal microscopy on permeabilized cells reveals that the final stage of reticulocyte maturation occurs by a previously undescribed mechanism in which large Glycophorin A-containing vacuoles forming at the cytosolic face of the plasma membrane are internalised and fuse with autophagosomes (defined by reactivity with anti-LC3) before expulsion from the cell. Our results constitute the first demonstration of linkage between plasma membrane remodelling and autophagy, and suggest reticulocyte plasma membrane remodelling occurs in two stages. Early maturation is characterized by the selective elimination of unwanted plasma membrane proteins (CD71, CD98, β1 integrin) through the endosome-exosome pathway. In contrast, final maturation is characterized by the generation and expulsion of large Glycophorin A-decorated vacuoles of autophagic origin, thereby simultaneously permitting optimisation of membrane:skeleton interactions to yield the more stable and deformable membrane of mature erythrocytes and eliminating unwanted residual organelles.

Disclosures:
No relevant conflicts of interest to declare.