bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Commitment to Lineage
Authors: Glimcher Laurie H. , Lazarevic Vanja , Ermann Joerg , Garrett Wendy 
Published: October 2019

Abstract
Abstract SCI-22
The transcription factor T-bet, isolated in our laboratory a decade ago, is a master regulator of Type 1 immunity in cells of both the adaptive and innate immune system. In adaptive immunity, T-bet instigates genetic programs in T helper 1 (Th1) cells and is required for production of the hallmark Th1 cytokine IFNg. It simultaneously represses the differentiation of T helper 2 cells and the profibrotic cytokines IL-13 and TGFb. We have recently determined that T-bet is also a repressor of the Th17 genetic program and have established the molecular mechanisms that underpin that function. T-bet also controls the optimal differentiation and function of the cytolytic CD8 cell, and is required for the development of the natural killer T cell. T-bet deficient animals are largely protected from autoimmune/inflammatory diseases such as multiple sclerosis, systemic lupus, type 1 diabetes and inflammatory arthritis, but are susceptible to type 2 driven diseases such as asthma and scleroderma. An exception to this overall rule is our recent discovery that in the absence of an adaptive immune system, the majority of mice lacking T-bet develop a spontaneous ulcerative colitis that progresses to colonic dysplasia and rectal adenocarcinoma. This colitis and inflammation associated colorectal cancer are MyD88 independent, driven by colitogenic flora and ameliorated by treatment with TNF blockade, antibiotics, and transfer of T regulatory cells. This phenotype maps to the T-bet deficient dendritic cell that drives this pro-inflammatory program; selective over-expression of T-bet in DCs was sufficient to reduce colonic inflammation and prevent the progression to neoplasia. The molecular pathogenesis of TRUC colitis and colitis-associated colorectal (caCRC) shares several key features with human caCRC. This model of colitis and colitis-associated colorectal cancer provides opportunities to further understand host-microbial relationships in inflammation and neoplasia and test preventative and therapeutic strategies pre-clinically. The function and mechanism of action of T-bet in the pathogenesis of immune system driven diseases will be discussed.

Disclosures:
Glimcher: Merck: Consultancy, Patents & Royalties, Research Funding; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.
Th17/Treg Imbalance in Sickle Cell Disease and Hydroxyurea Therapy
Authors: Gilli Simone , Pericole Fernando V , Fernandes Luis Gustavo , Ribeiro Karina A. R. , Castilho Lilian , Costa Fernando Ferreira , Saad Sara T Olalla 
Published: October 2019

Abstract
Abstract 1050

Background:
Tregs and proinflammatory Th17 cells have been described as two distinct subsets from Th1 and Th2 cells and have opposite effects in autoimmunity and inflammation. Th17/Treg balance controls inflammation and is important in the pathogenesis of autoimmune, infectious and chronic diseases. In view of growing evidence linking sickle cell disease (SS) and chronic inflammation and the potential anti-inflammatory role of Tregs cells, we hypothesized that SS is associated with decreased Treg subpopulation and consequent T cell dysfunction. Furthermore, the effects of hydroxyurea (HU) in this balance are not understood. To assess whether the Th17/Treg balance is impaired in patients with sickle cell disease, we investigated the peripheral blood Th17 and Treg frequencies and the expression of TGFβ1 and RORγT, a master regulator of Th17 differentiation, in sickle cell patients under HU therapy (SS-HU) and without HU (SSw/oHU). Methods. Thirty-eight patients with steady-state sickle cell disease (mean age 43 ± 10.5 yo; range 19–57 yo; 21 female/17 male), 19 SS-HU, 19 SSw/oHU and 20 healthy blood donors were evaluated. Tregs and Th17 quantification was performed by flow cytometry analysis and the expression of TGFβ1 and RORγT by q-PCR. Results. We found significant decrease in the Treg frequency (Foxp3+CD25+CD4+ population) comparing total SS patients and healthy controls (p=0.001) and this difference was more expressive between SS-HU and healthy controls (P=0.0006). Moreover, we found statistically increase of Th17 in SS-HU (p=0.02) and in SSw/oHU (p=0.01), comparing both with healthy controls. The TGFβ1 expression was increased in SS-HU (p=0.04) and SSw/oHU patients (p=0.01) when compared to control group. Interestingly, RORγT expression was significantly elevated only in SSw/oHU vs controls (p=0.03). Conclusion: Our results demonstrated a numerical imbalance of Th17/Treg in the peripheral blood of patients with SS disease. HU has no role mediating Th17/Treg plasticity and even SS patients under HU therapy remains Treg depleted. However, HU is able to modulated RORγT, corroborating the importance of other regulating factors during Th17 differentiation. Moreover, the higher expression of TGFβ1 may be an important positive instructive signal for Th17 differentiation. Th17/Treg imbalance might impact the disease presentation and phenotype and HU therapy may be involved in this issue.

Disclosures:
No relevant conflicts of interest to declare.
CD8+ CD28null T Cell Expansion Impairs Lenalidomide Immunomodulatory Function in Myelodysplastic Syndrome
Authors: McDaniel Jessica M. , Zou JianXiang , List Alan F. , Epling-Burnette P.K. 
Published: October 2019

Abstract
Abstract 1117
Lenalidomide, a thalidomide analog, is known to induce high rates of transfusion independence in Myelodysplastic Syndrome (MDS) patients harboring a commonly deleted region on chromosome 5q (del(5q)). Erythroid response is also seen in 20–30% of low-risk, non-del(5q) patients, although the mechanism of response is incompletely understood. Lenalidomide is a potent immunomodulating agent and hematopoietic response has been linked to increased lymphocyte infiltration in the bone marrow and improved T cell and NK cell proliferation/function. Lenalidomide's effect on the erythroid compartment is well documented, but the effect of lenalidomide on the immune compartment, and the relationship between the T cell compartment and erythroid response in vivo in MDS is not known. We therefore examined 23 T cell parameters before lenalidomide treatment and correlated them to hematologic response. We found that patients who fail lenalidomide therapy had higher CD8+ Terminal Effector Memory (TEM) T cells than did responders (p=0.02). TEM cells express CD45RA, have variable expression of the CD45RO memory marker and loss of L-selectin (CD62L) lymph node homing receptor. Interestingly, T cells within the TEM compartment are uniformly CD28 deficient (CD28null). Lenalidomide has been shown by us and others to increase the proliferation and function of T cells by providing co-stimulation (LeBlanc et al. 2004, Blood) through CD28, so CD28 expression may be important for immunomodulatory response. We found that MDS patients non-responsive to lenalidomide had an overall increase in CD4 (p=0.0195) and CD8 (p=0.0317) CD28null T cells, as well as an increase in CD28null cells (p=0.0159) within the TEM compartment compared to hematologic responders. To determine if CD28 expression is necessary for lenalidomide action in T cells, we sorted healthy donor T cells into CD8+CD28+ and CD8+CD28− populations, and found that lenalidomide-induced proliferation and interleukin-2 (IL-2) production were completely ablated within the CD28null subset. Because CD28null T cells displayed less proliferation (p<0.001) and produced less IL-2 (p<0.001) after stimulation, possibly related to excessive proliferative history in vivo, we therefore used shRNA to knockdown CD28 in healthy (CD28+) T cells and determined the functional response of these cells to lenalidomide. With CD28 knockdown, lenalidomide produced significantly less IL-2 compared to CD28+ controls (p<0.001), indicating the necessity of the immunotyrosine-based activation motifs (ITAMs) on the intracellular domain of the CD28 receptor for lenalidomide action. These results indicate that lenalidomide-mediated immune modulation requires CD28 expression on T cells and expansion of CD8+ CD28 deficient T cells may represent a predictor of erythroid non-response in non-del(5q) low-risk MDS patients.

Disclosures:
List: Celgene: Consultancy. Epling-Burnette:Celgene: Research Funding.
The Disulfide Isomerase Erp57 Mediates Platelet Aggregation and Hemostasis
Authors: Wu Yi , Ahmad Syed S. , Zhou Junsong , Wang Lu , Cully Matthew P , Essex David W 
Published: October 2019

Abstract
Abstract 1129
A close homologue to protein disulfide isomerase (PDI) called ERp57 forms disulfide bonds in glycoproteins in the endoplasmic reticulum and is expressed on the platelet surface. We generated two rabbit antibodies to ERp57. One antibody strongly inhibited ERp57 in a functional assay and strongly inhibited platelet aggregation. There was minimal cross reactivity of this antibody with PDI by Western blot or in the functional assay. Using flow cytometry this antibody substantially inhibited activation of the alphaIIbbeta3 fibrinogen receptor. Furthermore, adding ERp57 to platelets potentiated aggregation. In contrast, adding a catalytically inactive ERp57 inhibited platelet aggregation. When the inactive ERp57 was infused into mice the tail bleeding time was prolonged. We generated two IgG2a monoclonal antibodies that reacted with ERp57 by immunoblot. One of these antibodies inhibited both ERp57 activity and platelet aggregation. The other antibody did not inhibit ERp57 activity or platelet aggregation. When the antibodies were infused into mice the tail bleeding time was prolonged by the inhibitory antibody, but not by the non-inhibitory antibody. Finally, we found that a commonly used monoclonal antibody to PDI also inhibited ERp57 activity. We conclude that a glycoprotein specific member of the PDI family, ERp57, is required for platelet aggregation and hemostasis.

Disclosures:
No relevant conflicts of interest to declare.
Markers of Thrombogenesis Activation and Endothelial Dysfunction in End Stage Renal Disease
Authors: Bansal Vinod , Sharain Korosh , Hoppensteadt Debra , Litinas Evangelos , Lewis Bruce E , Fareed Jawed 
Published: October 2019

Abstract
Abstract 1151

INTRODUCTION
The pathogenesis of ESRD is complex involving inflammatory, thrombotic, and calcification processes. The leading cause of morbidity and mortality in ESRD is cardiovascular disease (CVD), accounting for 50% of all deaths. Current studies suggest that chronic inflammation is a major risk factor in ESRD. This investigation profiles markers of inflammation, thrombogenesis, and vascular dysfunction in patients with ESRD. METHODS Samples from ESRD patients (n=117) over the age of 18 on maintenance hemodialysis for at least 3 months and aged matched control samples (n=50) obtained from healthy volunteers were included in the study. The Randox Evidence Investigator (Antrim, United Kingdom) was used to measure IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, Vascular Endothelial Growth Factor (VEGF), Interferon-γ (INF-γ), Tumor Necrosis Factor-α (TNF-α), Monocyte Chemotactic Protein-1 (MCP-1), Epidermal Growth Factor (EGF), C-Reactive Protein (CRP), D-Dimer, and Thrombomodulin (TM). Plasminogen Activator Inhibitor -1 (PAI-1) and Asymmetric Dimethylarginine (ADMA) were measured using ELISA kits from Diagnostica Stago (Paris, France) and DLD Diagnostika GmbH (Hamburg, Germany), respectively. RESULTS IL-6 levels in ESRD were 4.3 times the controls. VEGF and INF-γ were both elevated at 2.6 times the controls. MCP-1, IL-8, IL-10, EGF, TNF-α, and IL-1α were elevated at 2.0, 1.6, 1.4, 1.4, 1.3, 1.2 times the controls, respectively. IL-4, IL-2, and IL-1 β levels were reduced at 0.9, 0.9, and 0.5 times the controls. TM, CRP, DDMER, PAI-1, and ADMA were increased at 5.9, 4.4, 3.0, 1.4, and 1.6 times the controls, respectively, each demonstrating statistically significant p-values. DISCUSSION The elevation of the interleukins, INF-γ, TNF-α and MCP-1 reveals intense inflammatory activation of leukocytes. The increase in these cytokines also demonstrates the vascular and tissue damage that occurs in ESRD. VEGF and EGF are both elevated in the ESRD samples indicating a state of cellular proliferation, possibly in an attempt to replace damaged tissue caused by the heightened inflammatory state. These results also demonstrate that ESRD patients have a significant upregulation in markers of thrombogenesis and endothelial dysfunction. 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 D-Dimer indicates an activation of coagulation in ESRD. The combined disruption of endothelial regulation and endothelial damage along with the upregulation of thrombogenic and inflammatory markers helps to explain the significant prevalence of DVT, PE and CVD in ESRD. These markers should be evaluated for their roles in kidney disease risk stratification and prognosis.

Disclosures:
No relevant conflicts of interest to declare.
NEW FINDINGS From the FRENCH COHORT of MYH9-RELATED DISORDERS: PHENOTYPE PECULIARITIES and GENOTYPE NOVELTIES on Behalf of the National French Reference Center on Inherited Platelet Disorders (CRPP)
Authors: Saposnik Béatrice , Binard Sylvie , Nurden Alan T. , Nurden Paquita , Schlegel Nicole 
Published: October 2019

Abstract
Abstract 1155
MYH9-Related Diseases (RD) are inherited platelet disorders combining thrombocytopenia and giant platelets with various associations of leukocyte inclusions, deafness, nephritis and cataracts. Mutations in the MYH9 gene, that encodes the nonmuscle myosin heavy chain IIA (NMMHC-IIA), are the hallmark of these disorders. Our cohort includes a total of 161 subjects (90 women, 71 men): 76 propositi (37 isolated, 39 family index cases) with MYH9 mutations and 85 family members (45 women, 40 men) belonging to the 39 unrelated families. The cases with an identified mutation were classified according to the location of the mutation either in the motor domain (MD patients (P): “MDP”) or in the tail domain (TD patients (P): “TDP”) of NMMHC-IIA. MYH9 mutations were found in a total of 109 subjects: the 76 propositi (45 women, 31 men) and 33/85 (38.8%) family members (19 women, 14 men, from 22 families). Among the 41 different mutations detected, the majority were missense mutations (33 = 80.5%) and new (25 = 61%). Two substitutions in intron 39 were discovered for the first time: 1 in the donor splice site (corresponding to a classical MYH9-RD phenotype) and the other in position minus 3 of the acceptor splice site (moderate thrombocytopenia only without giant platelets). A total of 15 different exons were identified as having mutations, 4 exons remained unaffected. Mutations were more frequently found in exons 1, 16, 26, 30, 38 and 40, as already published. The majority of patients (102 = 93.6%) were heterozygous for one mutation but 7 patients (6.4%) were compound heterozygous for 2 mutations. Significant differences between MDP and TDP were found for bleeding symptoms (65% of MDP, 33% of TDP, p=0.0037), nephropathy (31.5% of MDP, 11.6% of TDP, p=0.015), age (years, mean) at onset for nephropathy (21 for MDP, 48 for TDP, p=0.0003), deafness (54% of MDP, 23.7% of TDP, p=0.0025), and association bleeding/extra-hematological symptoms (68.2% of MDP, 27.7% of TDP, p=0.01). Patients with bleeding had a lower platelet count than non-bleeders (mean= 42G/L versus 62G/L, p=0.0024) but there were no significant differences in the percentage of giant platelets. ITP (splenectomy for 7) was the initial misdiagnosis for 29/69 (42%), (51.7% of MDP, 35% of TDP, p= 0.165). Cataracts occurred in 4 patients only. Altogether these results showed that the clinical phenotype of MYH9-RD was more severe for MDP than TDP. New mutations were discovered and were widespread along the exons and in some cases two mutations were identified in the same patient. The sequencing of the full gene and all exons-introns boundaries should be mandatory for each patient whatever the initial result.

Disclosures:
No relevant conflicts of interest to declare.
Thrombin Generation Inhibition by Newer Anti Xa and Anti IIa Drugs in Prothrombin Complex Concentrates: Relevance to Endogenous Modulation of Thrombogenesis
Authors: Gray Angel , Litinas Evangelos , Hoppensteadt Debra , Cunanan Josephine , Lewis Bruce E , Iqbal Omer , Fareed Jawed 
Published: October 2019

Abstract
Abstract 1196
Several anti-Xa agents (apixaban, otamixaban, rivaroxaban and edoxaban) and anti-IIa agents (dabigatran and melagatran) have been developed for clinical use. These agents are strong inhibitors of factor Xa and IIa and their potency is usually expressed in terms of the inhibitory parameters of these enzymes. These synthetic agents can also modulate thrombin and factor Xa generation in complex systems such as the platelet rich and platelet poor plasma and whole blood. The order of potency differs depending upon the type of matrix used due to cellular and protein binding and interaction with generated proteases and protein fragments. In order to compare these agents for their relative thrombin generation inhibitory profile, prothrombin complex based assay systems are developed utilizing 5 units/mL of non activated prothrombin complex such as Konyne® and Preconativ. Thrombin generation is triggered by tissue factor and monitored using a thrombin specific florometric substrate based assay (Technoclone, Vienna, Austria). All agents were tested in a concentration range of 0–1000 ng/mL. The relative inhibition of thrombin generation was calculated in terms of IC50. The factor Xa inhibitors were found to be relatively potent inhibitors of thrombin generation exhibiting an IC50 value of 25–96 ng/mL with the following order of potency- Rivaroxaban>Otamixaban>Apixaban. The anti IIa agents exhibited potency in the range of 125–225 ng/mL. Melagatran was more potent than dabigatran. In the plasma based anticoagulant assays such as PT, aPTT, heptest, the anti IIa agents were more potent than factor Xa drugs. Assay based differences were noted in all agents which did not follow the same patterns as thrombin generation inhibition in the prothrombin complex systems. These observations suggest that although considered to be single target, the newer anticoagulant drugs produce variable inhibitory effects on the protease network in complex systems such as plasma and prothrombin complexes. These effects may be related to the relative safety and efficacy profile of these agents and should be included in the preclinical profiling of these agents.

Disclosures:
No relevant conflicts of interest to declare.
Curcumin Prevents Cytokine-Mediated Tissue Factor Pathway Inhibitor Down-Regulation in Human Endothelial Cells
Authors: Maruyama Keiko , Morishita Eriko , Goto Yukie , Sekiya Akiko , Asakura Hidesaku , Ohtake Shigeki , Nakao Shinji 
Published: October 2019

Abstract
Abstract 1202

Objective:
Curcumin (diferuloyl methane), an active component of the spice turmeric, has been shown to exhibit anti-inflammatory and antioxidant activities in addition to an anticartinogenic activity in vitro and in vivo. Furthermore, we reported that curcumin inhibited the induction of tissue factor (TF) expression in human umbilical vein endothelial cells (HUVECs) at 52nd ASH 2010. Therefore, curcumin may ameliorate hyper-coagulable state associated with inflammation or oxidative stress. On the other hand, tissue factor pathway inhibitor (TFPI) which is expressed by endothelial cells plays a crucial role in hemostasis by regulating TF-induced initiation of coagulation. This study examined whether curcumin modulates the expression of TFPI in HUVECs. Methods: HUVECs were pretreated with curcumin at the concentration of 20 μM for 3h, washed and stimulated with tumor necrosis factor-alpha (TNF-alpha, 10 ng/ml) for additional 12 or 24h. The mRNA and protein levels of TFPI in the cultured HUVECs were determined by reverse transcriptase polymerase chain reaction (RT-PCR) and western blotting, respectively. To determine whether curcumin affects the MAPK signaling pathways, the phosphorylation of p38 mitogen-activated protein kinase (p38MAPK), extracellular signal-regulated kinase1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK) in the HUVECs were analyzed with western blotting. Additionally, to determine whether curcumin affects nuclear factor-kappa B (NF-kB) pathway, nuclear and cytoplasmic fractions were extracted and protein levels were determined by western blotting for NF-kB (p65), p-IkB and IkB. Results: After TNF-alpha stimulation, TFPI mRNA levels were approximately decreased by 40% compared to the control (p<0.05; Figure 1B). Similarly to the mRNA expression, TFPI protein levels were decreased (Figure 1A). On the other hand, pretreatment of HUVECs with curcumin significantly suppresses TNF-alpha-induced TFPI mRNA and protein down-regulation (p<0.05; Figure 1A, B). Curcumin inhibited TNF-alpha-induced activation of p38MAPK, ERK1/2, and JNK. Moreover, curcumin inhibits TNF-alpha-induced IkB activation in HUVECs. And, translocation of NF-kB from the cytosol into the nucleus by TNF-alpha was inhibited by curcumin. Conclusions: These results indicate that curcumin may suppress the TNF-alpha-induced TFPI down-regulation via NF-kB pathways. Thus, curcumin may offer a novel antithrombotic option for treatment of the hypercoagulable state associated with inflammation.

Disclosures:
No relevant conflicts of interest to declare.
Identification of Unique Biomarkers in Sepsis Associated Coagulopathy
Authors: Sharma Gautam , Bijlani Rahul , Litinas Evangelos , Hoppensteadt Debra , Cunanan Josephine , Khan Hussein , Sharma Gautum , Kaul Inder , Fareed Jawed 
Published: October 2019

Abstract
Abstract 1237

Introduction:
Disseminated Intravascular Coagulation represents a complex multi-factorial pathophysiological process in which protease disregulation along with other pathologic processes play an important role. The uncontrolled protease activities result in generation of several mediators which are not fully characterized. Biomarker profiling using protein chip array technology has been used in the identification of unique mediators in various diseases. Surface Enhanced Laser Desorption Ionisation(SELDI) is an ionization method in mass spectrometry that is used for the analysis of protein mixtures and correspondingly in the profiling of biomarkers in various diseases. Purpose: The purpose of this study was to profile and identify unique biomarkers in hospitalized patients with sepsis associated coagulopathy. Study design: There were 385 plasma samples included in this study. Baseline protein chip array profile in Molecular weight range of 0–200 kDa was carried out using SELDI technique employing gold chips. Plasma samples from normal humans (n=100) and pooled preparation from both the normal (NHP) and suspect liver disease/coagulopathy were utilized as controls. Results: Of the 385 baseline samples analyzed 371 patients (96.86%) exhibited the presence of a unique biomarker at 11.6 kDa. In addition down regulation of a biomarker at 56.4 kDa was also noted in 218 of patients (56.74%) In the normal human pooled plasma the 11.6 kDa peak was nearly absent. However in Pathological pooled plasma a distinct biomarker at 11.6 kDa with a much lower intensity was noted. Additional markers in baseline samples in 1–10 kDa range were also noted. Conclusions: These observations suggest that sepsis associated coagulopathy results in the generation of specific unique biomarker at 11.6kDa which requires further characterization. The downregulation of biomarker at 56.4 kDa and the apparent reduction levels during hospitalization require further investigations.

Disclosures:
No relevant conflicts of interest to declare.