bluuebunny/crossref_metadata_2025_split huggingface dataset:
New Functional Assays to Selectively Quantify APC- and TFPI-Cofactor Activities of Protein S in Plasma
Authors: Alshaikh Nahla , Rosing Jan , Thomassen Stella , Castoldi Elisabetta , Simioni Paolo , Hackeng Tilman M. 
Published: October 2019

Abstract
Background: Protein S is an important anticoagulant in the down-regulation of blood coagulation by acting as cofactor for two main regulators of coagulation. Protein S acts as a cofactor for activated protein C (APC) in the inactivation of the procoagulants FVa and FVIIIa and for TFPI in the formation of the TFPI-FXa encounter complex that subsequently inactivates TF-FVIIa. Several studies have indicated that protein S deficiency is associated with increased risks of venous thrombosis.
Aims: Current protein S functional assays are influenced by plasma determinants such as FVLeiden. The aim of the study was to develop thrombin generation based assays that enable quantification of both the APC- and TFPI-cofactor activities of protein S in plasma whereas the outcome of the assays should not be affected by the presence of FVLeiden or other plasma variables.
Methods: APC- and TFPI-cofactor activities of protein S in plasma were measured using calibrated automated thrombography (CAT) in protein S-depleted plasma supplemented with a small amount of sample plasma either in the presence of anti-TFPI antibodies and APC (APC-cofactor activity) or at excess full-length TFPI without APC (TFPI-cofactor activity). Total and free protein S levels in plasma were measured by ELISA's.
Results: Average APC-cofactor activities of protein S were 113%, 122%, and 87% in plasma from normal individuals (n=10), FV Leiden heterozygotes (n=4), and FV Leiden homozygotes (n=2), respectively, while the average APC-cofactor activity of protein S in plasma from heterozygous protein S-deficient individuals (n=21) was significantly lower (55%). Similar trends were observed for the TFPI-cofactor activity of protein S, with averages of 108%, 112%, and 97% in plasma from individuals with normal protein S levels and different FV Leiden genotypes, and 64% in plasma from protein S-deficient patients. APC-cofactor activities of protein S correlated significantly with free and total protein S antigen levels, while TFPI-cofactor activities correlated less with protein S antigen levels.
Summary/Conclusion: We have developed functional protein S assays that measure both the TFPI- and APC-cofactor activities of protein S in plasma and that are not affected by the FV Leiden mutation or other plasma influences.

Disclosures
No relevant conflicts of interest to declare.
PAR1 Antagonists Development and Clinical Utility
Authors: Coughlin Shaun 
Published: October 2019

Abstract
Thrombin is a potent activator of platelets and other cells. The mechanism by which thrombin, a protease, regulates cellular behaviors like a hormone was revealed by expression cloning of a G protein-coupled receptor now known as protease-activated receptor-1 (PAR1). Thrombin activates PAR1 by cleaving its N-terminal ectodomain to reveal a new N-terminus that then serves as a tethered peptide ligand, binding to the heptahelical domain to trigger G protein activation. A synthetic hexapeptide mimicking this new N-terminus activates PAR1 without receptor cleavage. Point mutations that prevent receptor cleavage render PAR1 unactivatable by thrombin without altering activation by exogenous tethered ligand peptide. Replacement of the thrombin cleavage site with sites for other proteases allows PAR1 to signal in response to those other proteases. Removal of sequence N-terminal to the cleavage site and creation of the new protonated amino group at N-terminal of the tethered ligand sequence is necessary for its function, explaining how the tethered ligand is silent in the intact receptor and activated by receptor cleavage. Thus, PARs are, in essence, peptide receptors that carry their own ligands, which can be unmasked by receptor cleavage. Mammalian cells utilize four PARs to respond to coagulation proteases and other proteases with trypsin-like specificity. Three, PAR1, PAR3 and PAR4 can mediate responses to thrombin, with PAR3 and PAR4 mediating platelet activation by thrombin in mice and PAR1 and PAR4 platelet activation in humans. Inhibition of thrombin signaling via PARs in platelets decreases thrombus formation in animal models of platelet-dependent thrombosis. These and other studies led to development of the PAR1 antagonist vorapaxar (Zontivity), a first-in-class antiplatelet agent approved for secondary prevention of atherothrombotic events in selected patients with prior myocardial infarction and peripheral arterial disease. What was and was not learned from animal and human studies and remaining questions related to clinical utility of PAR inhibition will be discussed. The structure-function studies outlined above support the tethered ligand model, but a structure that reveals how the tethered ligand binds and how binding leads to transmembrane signaling and G protein activation is needed. Vorapaxar provided a tool for stabilizing PAR1 protein during solubilization, purification and crystallization. In collaboration with Brian Kobilka and colleagues, we solved a crystal structure of off-state PAR1 in complex with its antagonist vorapaxar, which explains pharmacological properties of vorapaxar and efforts to solve a complementary on-state structure are ongoing.

Disclosures
Coughlin: Merck: Research Funding; Novartis: Consultancy; Portola Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.
The Impact of 3D Chromosomal Topology in Acute Leukemia
Authors: Aifantis Iannis 
Published: October 2019

Abstract
Acute myeloid leukemia (AML) is the most common adult leukemia characterized by excessive proliferation of abnormal myeloid progenitors. AML continues to have a dismal survival rate amongst all subtypes of leukemia (<50% five-year overall survival rate), which can largely be attributed to limited advances in treatment regimens that, for the last decades, have relied on the use of two non-targeted cytotoxic drugs: cytarabine and anthracycline. Large-scale sequencing efforts have shed new light on genetic and epigenetic determinants of AML. Interestingly, these studies identified a frequent co-occurrence of somatic mutation between genes encoding cohesin complex subunits (such as STAG2, SMC1A, RAD21 and SMC3) and well-characterized AML oncogenic triggers, such as FLT3-ITD, TET2, and NPM1. Recent work has demonstrated an important role for the cohesin complex in normal stem/progenitor self-renewal and differentiation, gene regulation, and suppression of myeloproliferative neoplasms and AML, despite the precise mechanisms underlying these functions remaining poorly understood. It is believed that cohesin may suppress tumor formation by regulating chromatin looping at loci critical for self-renewal and myeloid progenitor differentiation. Utilizing established models of murine and human AML, this we will focus on the molecular mechanisms of cohesin-dependent myeloid tumor-suppression, with an emphasis on understanding novel treatment approaches that can exploit these functions. Using established protocols for identifying genome-wide changes in chromatin topology and gene expression, we propose undertaking an extensive characterization of cohesin-regulated chromatin changes driving AML. Furthermore, we investigate the application of targeted agents in cohesin-deficient AML whilst extensively mapping the mechanisms-of-action underlying these specific treatments. Ultimately, our work aims to generate novel, pre-clinical disease models of cohesin-mutated AML with strong mechanistic insights into the tumor-suppressive function of this complex.

Disclosures
No relevant conflicts of interest to declare.
Genomic Applications for Transfusion Support
Authors: Chou Stella 
Published: October 2019

Abstract
Red blood cell (RBC) and platelet antigen determination by DNA methods (genotyping) has become an important part of the practice of transfusion medicine over the past decade.The results are reliable and highly correlated with serologic phenotyping results, and are superior in some situations. Application of RBC genotyping to provide more precise transfusion therapy for various patient populations will be discussed, with a particular emphasis on those requiring chronic RBC therapy or with complex serologic reactivity. Current DNA arrays can test for a large number of blood group antigens, but routine typing of the ABO and Rh systems is not currently feasible due to the large number of genetic mutations.
Patients with sickle cell disease (SCD) are one of the most challenging populations to safely transfuse, in part due to high rates of alloimmunization against Rh antigens despite prophylactic matching for D, C, and E antigens. Genotyping of the RH loci has revealed that RHD and RHCE variants are prevalent in African Blacks. Nearly 90% of patients with SCD have at least one altered RH gene and contributes to Rh alloimmunization in patients with SCD. The role of RH variation and alloimmunization in SCD will be reviewed, and the feasibility and approach to genetic RBC matching of patients and donors will be discussed. The use of targeted DNA arrays, whole exome sequencing, or whole genome sequencing to identify common and variant RH alleles will be addressed, including limitations of each method.

Disclosures
No relevant conflicts of interest to declare.
Transcription Factor Gene Fluorescent Reporters Track Lineage Fate in Lymphoid Commitment
Authors: Rothenberg Ellen 
Published: October 2019

Abstract
The transition from multipotency to lineage commitment can be followed with particular clarity for T cell precursors. In this lineage, the role of environmental signals can be clearly separated from the role of intrinsic fate programming in individual cells and the cells' developmental responses to changing conditions and can be tracked in real time. T cell precursors are still multipotent when they first enter the thymus, and if they are removed from the thymic microenvironment at this stage they can give rise to non-T cells including dendritic cells and myeloid cells. For multiple cell divisions, they preserve this multipotency and are only kept in line to become T cells conditionally, by Notch signaling from the thymic stroma. Then at a specific point of no return, the cells become unable to give rise to anything except T cells regardless of environment, and this is the point of commitment. Commitment is clearly the readout of a change in internal transcriptional regulatory state. To determine how this is controlled, we and others have charted transcription factor expression changes across this interval and changes in chromatin modification and DNA accessibility that accompany the transition, and we have been able to use functional perturbation tests to narrow down the key regulators that catalyze and enforce this transition. A particularly important commitment factor is encoded by the Bcl11b gene, which is released from previously repressed chromatin and sharply activated at the transcriptional level just as the cells become committed. The Bcl11b gene product is required in all alpha beta and most gamma delta T cells to enable the commitment process to occur. These properties make it highly illuminating as an indicator of the regulatory state in individual differentiating T-cell precursors. We have generated a series of knock-in Bcl11b fluorescent reporter alleles to probe the correlation of Bcl11b expression with changes in specific target genes, to determine the transcription factor requirements for Bcl11b gene activation in the gene regulatory network controlling commitment in single cells, and to measure the role of epigenetic modification of the Bcl11b locus on the kinetics of transition from uncommitted to committed states. These results and their implications will be presented. Importantly, the use of live-cell reporters reveals a level of all-or-none, stochastic regulation in the responses of individual cells to combinatorial transcription factor action at this developmental watershed1.
Reference:
1. Kueh HY, Yui MA, Ng KKH, et al. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment. Nature Immunology. 2016.17, 956-965.

Disclosures
No relevant conflicts of interest to declare.
Marrow Infiltrating Lymphocytes and Vaccines in Myeloma
Authors: Borrello Ivan 
Published: October 2019

Despite several decades of research with limited clinical successes, immunotherapy is now establishing a permanent presence in the clinical arena for several malignancies including multiple myeloma. The ability to understand the mechanisms mediating tumor-induced tolerance as well as the availability of reagents to target them have facilitated the clinical development of several immune-based strategies. The efficacy of adoptive cell therapies has already been demonstrated by the clinical benefit observed with allogeneic bone marrow transplantation where the immune-mediated graft vs. tumor effect accounts for the deeper and more durable clinical responses when compared to autologous stem cell transplantation. Recently, adoptive T cell approaches have focused on ex vivo manipulation of cells with the goal of increasing their tumor specificity. To date, the approaches can be divided into three groups: unmodified cells; T cells expressing transgenic T cell receptors; and cells modified with chimeric antigen receptors (CARs). Each has its own benefits. Historically, adoptive T cell therapy utilized peripheral blood lymphocytes in large part because of their ease of access. Early studies in myeloma demonstrated their ability to augment immune reconstitution and effectively transfer vaccine-specific responses. However, their lack of tumor specificity has prompted our group to evaluate the role of marrow infiltrating lymphocytes (MILs). These cells possess a broad endogenous tumor specificity and are enriched for central memory T cells that make them a more suitable source of cells for adoptive T cell approaches. The properties and potential benefits of this approach will be discussed.
Another immunotherapeutic approach with potential appeal is the use of myeloma vaccines to address the issue of minimal residual disease. In light of the fact that patients with myeloma rarely have long-term sustained remissions, it is reasonable to integrate a novel approach that is capable of eradicating the remaining disease burden by generating a broad, myeloma-specific, systemic immune response. We have developed a cellular, GM-CSF-based, myeloma vaccine which has been administered in the combination with the immunomodulatory agent, lenalidomide, to patients in a near complete remission (negative M-spike but with a positive immunofixation). We have demonstrated priming of a tumor-specific immune response which correlates with a durable clinical benefit.
With the ever-increasing development of immune-based approaches, combining them with various agents in different disease setting to improve the overall outcomes of myeloma patients.

Disclosures
Borrello: Celgene: Honoraria, Research Funding, Speakers Bureau; BMS: Honoraria, Research Funding; WindMIL Therapeutics: Equity Ownership, Patents & Royalties, Research Funding.
The Effects of Splenectomy on the Red Blood Cells of the Dog with Particular Emphasis on the Reticulocyte Response
Authors: LORBER MORTIMER 
Published: October 2019

Abstract
In splenectomized dogs the following observations on the erythroid elements were made:
1. There was no unusual postoperative depression of erythrocyte, hemoglobin or hematocrit values, and regeneration occurred at the same rate as in the control dogs.
2. A significant increase in the number of circulating reticulocytes was noted, which persisted for a longer duration than the associated thrombocytosis. There is a suggestion that the reticulocytes circulating during regeneration were more immature than those of the control animals.
3. Varying numbers of target cells accompanied by increased osmotic resistance were present in all the splenectomized dogs. Most also developed normoblastemia, particularly during active regeneration, which persisted thereafter in some of the animals. Howell-Jolly bodies were rare.
4. No significant bone marrow changes were noted.
It is suggested that:
1. In nonsplenectomized animals, most erythrocytes released from the bone marrow in response to blood loss are mature red cells rather than reticulocytes.
2. Postsplenectomy reticulocytosis is not dependent on a fall in hematocrit and is not a reflection of hemolysis or even of increased erythropoiesis, except, perhaps, in part during regeneration from the blood loss incurred at surgery.
3. Postsplenectomy reticulocytosis and normoblastemia are unaccompanied by alterations in the myeloid-erythroid ratio of the bone marrow and are probably largely due to a diminution of the normal inhibition of erythroid release from the marrow, permitting their earlier entry into the circulation.
4. The aforementioned changes are due specifically to the removal of the spleen itself, rather than merely to the removal of splenic blood from the portal circulation.
Modification of X. radiation Lethality by Autologous Marrow Infusion in Dogs
Authors: ALPEN EDWARD L. , BAUM S. J. 
Published: October 2019

Abstract
Dogs have been treated after x-irradiation with doses of 400 r and 600 r by injection of either homologous or autologous bone marrow. The bone marrow was obtained by preirradiation biopsy and was reinjected either into the animal from which it was drawn (autologous) or into an alternate recipient (homologous). Injection of 2 x 109 autologous nucleated cells successfully protected all dogs given either 400 r or 600 r. Dogs given 1 x 109 autologous nucleated cells after 600 r failed to survive. No protection was provided by injection of 2 x 109 homologous nucleated cells after 400 r nor by injection of 2 x 109 or 5 x 109 homologous nucleated cells after 600 r. It is proposed that a differential species sensitivity to foreign cell implants accounts for the failure of homologous marrow infusions in dogs in contradistinction to the protection provided by homologous or heterologous marrow implants in rats and mice, and by homologous implants in monkeys.
Hematopoietic and Nitrogen-Sparing Effect of Preparations from Hog Gastric Mucosa with High Content of Blood Group Substances
Authors: CRESSERI ANGELO , CANTONE AURELIO , PICCININI FRANCO , CAPRARO VITTORIO 
Published: October 2019

Abstract
A mucin fraction with high content of A and H blood group substances was obtained from hog gastric mucosa. This fraction administered per os together with vitamin B12 in rats made anemic by phenylhydrazine injection shows a significant effect on the hematologic response in comparison with the animal treated with vitamin B12 alone.
The same fraction administered per os in cortisone-treated rats has also a significant nitrogen sparing action.
The possible mechanisms and relationships of the observed biologic effects are briefly discussed.
An In Vitro Technic for Quantitating and Studying the Dynamics of Leukocyte Agglutination
Authors: HARTMAN JOHN D. 
Published: October 2019

Abstract
1. An in vitro technic for quantitating leukocyte agglutination has been described.
2. With this technic the dynamics of leukocyte agglutination and the response of leukocytes to antigenic agents in both sensitive and nonsensitive systems have been studied.
3. The relationship of this study to the problem of agranulocytosis is discussed.
Enhanced Urinary Excretion of Co60 Vitamin B12 Produced by Delayed Release Capsules
Authors: JOHNSON P. C. , BERGER E. S. 
Published: October 2019

Abstract
1. Patients without known hematologic disease were given 8 µg. (0.2 µc. Co60 vitamin B12 as a water solution or in various types of delayed release capsules.
2. All groups showed approximately equal fecal excretion. However, the cobalt60-labeled vitamin B12 delayed release capsules showed a greater urinary excretion after a loading dose of stable B12 than did the water solution of labeled vitamin B12.
Observations on the Anemia of Cryptogenetic Splenomegaly. I. Hemolysis
Authors: MCFADZEAN A. J. S. , TODD DAVID , TSANG K. C. 
Published: October 2019

Abstract
Excessive hemolysis has been shown to occur in patients with cryptogenetic splenomegaly. This is corrected by splenectomy.
The survival of normal red cells transfused to such patients is greatly reduced. Following splenectomy normal red cells so transfused survive normally.
The survival of red cells from such patients upon transfusion to healthy recipients, including a splenectomized recipient, is greatly reduced. Following splenectomy the survival of red cells from such patients upon transfusion to healthy recipients is significantly increased, although still significantly less than normal. However, upon transfusion into splenectomized but otherwise healthy recipients the survival time is normal.
Study of osmotic fragility before splenectomy demonstrates the existence of a mixed population showing both increased and decreased fragility to hypotonic saline. Following splenectomy there emerges a population with increased resistance to hypotonic saline.
It is concluded that prior to splenectomy the pathologic spleen exerts an effect upon circulating erythrocytes including transfused normal red cells which results in marked reduction in their survival. Following splenectomy a second defect in the erythrocytes becomes apparent and this also results in reduction in life-span, although the presence of the spleen would appear necessary for this reduction to occur. It is suggested that this second defect develops as the result of liver damage.
Calcium-Lipid Complexes in Human Platelets
Authors: WALLACH DONALD F. HOELZL , SURGENOR DOUGLAS M. , STEELE BARBARA B. 
Published: October 2019

Abstract
Carefully washed human platelets contain calcium in an unusual state of chemical combination. Platelet calcium is non-ionic and is not exchangeable with ionic Ca45. On extraction of platelets with lipid solvents, the calcium separates with the phospholipids. The calcium-lipid complex thus obtained does not undergo any exchange with radioactive calcium. Study of other properties of platelets reveals that the clot-promoting activity of platelets (activated by antihemophilic factor and plasma thromboplastin component) also separates into the lipid fraction. Preliminary observations are reported on the nature of platelet lipids as revealed by paper chromatography and paper electrophoresis in solvents of low dielectric constant. Small amounts of proteolipid have been isolated from the lipid extracts. The unique calciumlipid complex is thought to originate in the cell membrane of the platelet.
The Effect of Heparin on Components of the Human Fibrinolytic System
Authors: VON KAULLA KURT N. , MCDONALD TOM S. 
Published: October 2019

Abstract
It was demonstrated with human urokinase, human fibrinolytic euglobulins and human pyrogen plasma as enzyme source, and with bovine fibrin plates and human plasma plates as substrate, that high heparin concentrations inhibit fibrinolysis of preformed fibrin, in contrast to small ones which may enhance this phenomenon, provided albumin is present.
No interference could be observed in the absence of plasma, serum or albumin fraction. Albumin or substances associated with the albumin fraction are required for interference of heparin with fibrinolysis. This interference probably takes place with the action of plasmin on fibrin.