bluuebunny/crossref_metadata_2025_split huggingface dataset:
Toll-Like Receptor Triggering Modulates Factor VIII-Specific Immune Memory in Murine Hemophilia A with Factor VIII Inhibitors.
Authors: Allacher Peter , Hausl Christina , Ahmad Rafi U. , Schwarz Hans Peter , Turecek Peter L. , Reipert Birgit M. 
Published: October 2019

Abstract
The development of inhibitory antibodies against factor VIII (FVIII) is the major complication in the treatment of hemophilia A patients with FVIII products. Immune Tolerance Induction (ITI) therapy using long-term application of high doses of FVIII has evolved as an effective therapy to eradicate the antibodies and induce long-lasting immune tolerance. It is a common observation that infections, particularly central venous catheter infections during ITI cause a rise in anti-FVIII antibody titers that can prolong the course of ITI or possibly even lead to failure of ITI. Based on this observation, we asked the question whether microbial components derived from viruses or bacteria modulate the re-stimulation of FVIII-specific immune memory and disturb the recently described inhibition of memory-B-cell-re-stimulation by high doses of FVIII (Hausl et al.: Blood2005; in press). Microbial components are recognized by toll-like receptors (TLRs) that serve as an important link between innate and adaptive immunity. TLRs can discriminate various microbial components such as lipopeptides derived from bacteria or zymosan derived from yeast (recognized by TLR1/2 or TLR2/6), double-stranded RNA derived from viruses (recognized by TLR3), lipopolysaccharide (LPS) derived from gram-negative bacteria (recognized by TLR4), flagellin derived from bacterial flagella (recognized by TLR5), single-stranded RNA derived from viruses (recognized by TLR7/8) or bacterial DNA containing the unmethylated CpG motif (recognized by TLR9). We analyzed the re-stimulation of FVIII-specific memory-B cells using a murine model of hemophilia A as described previously (Hausl et al.: Blood2004; 104:115–22; Hausl et al.: Blood2005, in press). The following TLR ligands were tested: zymosan for TLR2 (0.1–10,000 ng/ml), poly I:C for TLR3 (1.0–50,000 ng/ml), LPS for TLR4 (0.1–10,000 ng/ml), Flagellin for TLR5 (0.01–1,000 ng/ml), Loxoribine for TLR7 (1.0–50,000 ng/ml) and CpG oligonucleotides for TLR9 (0.1–10,000 ng/ml). Our results indicate that none of the TLR ligands at the concentrations tested induced a significant re-stimulation of FVIII-specific memory B cells in the complete absence of either FVIII or T cells. However, ligands for TLR3, TLR4, TLR7 and TLR9 were able to disturb the inhibition of memory-B-cell-re-stimulation by high doses of FVIII and amplified the re-stimulation induced by low doses of FVIII substantially.
We conclude that triggering of TLRs by microbial components that are present during infections amplify the re-stimulation of FVIII-specific memory B-cells induced by low doses of FVIII and disturb the inhibition induced by high doses of FVIII.
Plasma Levels of High Mobility Group Box Protein in Patients with Organ Failure, Disseminated Intravascular Coagulation or Poor Outcome.
Authors: Wada Hideo , Nobori Tsutomu , Yamada Shingo , Maruyama Ikuro 
Published: October 2019

Abstract
High Mobility Group Box chromosomal protein 1 (HMGB1) is a nuclear DNA-binding protein acting as a proinflammatory cytokine when released in the extracellular space from necrotic cells, activated macrophages and dendritic cells. HMGB1 acts on a specific receptor, RAGE (receptor for advanced glycation end-products), and induces prolonged inflammation, organ failure, septicaemia and death. The aim of the study was to determine the diagnostic value of plasma HMGB1 concentration and its role in the development of organ failure in patients with disseminated intravascular coagulation (DIC).
Plasma HMGB-1 levels were measured in patients with suspected DIC by ELISA and their relationships with DIC, organ failure and clinical outcome were determined. The study group included 201 patients suspected of having DIC (79 females and 122 males, age; 52.1±16.5 years, mean±SD).
The study group represented all such patients admitted to the Second Department of Internal Medicine and Intensive Care Unit (ICU) of Mie University Hospital between January 1, 2002 and December 31, 2003. The underlying diseases were infectious diseases (n=40), hematopoietic diseases (n=58), solid cancers (n=16), trauma (n=21), aneurysm (n=12), autoimmune diseases (n=8), liver diseases (n=7), post-operation (n=32) and other diseases (n=7).
Results: Plasma HMGB1 was below the detection limit in normal subjects, but moderately elevated in patients with infectious diseases (4.54 ± 8.18 ng/ml, mean±SD), malignancies (2.15 ± 5.34 ng/ml), and traumas (6.47 ± 13.13 ng/ml). DIC was associated with significantly high plasma HMGB1 (14.05 ± 12.56 ng/ml) in these patients. The highest HMGBI levels were in patients with organ failure (8.29 ± 10.99 ng/ml) and non-survivors (16.58 ± 11.01 ng/ml). HMGB1 plasma levels correlated with the DIC score and sepsis-related organ failure assessment (SOFA) score. Plasma levels of HMGB1 were significantly higher in patients with PAI-I of >50 ng/ml (3.66 ± 6.36 ng/ml) than in those with PAI-I <50 ng/ml (1.38 ± 2.97 ng/ml) (p< 0.05) but there was no significant difference in HMGB1 levels between patients with TNF-a of >1.5 (3.04 ± 5,56 ng/ml) and those with less than 1.5 pg/ml (2.44 ± 5.36 ng/ml).
Our data suggest that HMGB-1 is a potentially suitable prognostic marker of OF or DIC.
Thrombophilia in Patients with Occlusions of Vascular Bypass Grafts.
Authors: Klamroth Robert , Orlovic Marija , Rimpler Hartmut , Landgraf Helmut 
Published: October 2019

Abstract
Background: Causes of vascular bypass occlusions are surgical failure, hyperplastic intima, progression of arterial occlusive disease and thrombophilic disorders. The aim of this registry was to determine the prevalence of thrombophilic risk factors in patients with occlusions of arterial bypass grafts of the lower extremities.
Patients and Methods: From 2001 to 2004 we studied 81 consecutive patients (pts) with a mean age of 60,5 years. 64/81 pts showed occlusions of arterial prosthetic grafts and 17/81 pts occlusions of venous grafts of the lower extremities. Bypass occlusion occured 7,1 months (mean value) after primary operation. 68/81 patients had concomittent risk factors for arteriosclerosis like arterial hypertension, diabetes, hypercholesterinemia or smoking.
The laboratory testing for thrombophilia (performed at least 4 weeks after the bypass occlusion) included antithrombin, protein C, protein S, homocysteine, factor V-G1691A-mutation, prothrombin-G20210A-mutation, lupus anticoagulant, anti-cardiolipin-antibodies, lipoprotein (a), factor VIII, plasminogen activator inhibitor (PAI) and induced platelet aggregation in platelet rich plasma with different concentrations of ADP to evaluate hyperreactive platelets.
Results: Thrombophilia screening revealed abnormalities in 65/81 pts. 36/81 patients showed combined defects. Screening for antiphospholipid antibodies (LA and/or ACA) revealed pathologic values in 31/81 pts, heterozygous factor V- mutation was present in 8/81 pts, heterozygous prothrombin-G20210A-mutation in 2/81. 20/81 pts had elevated homocysteine levels, 16/81 pts elevated lipoprotein (a) levels, 19/81 pts had a persistent factor VIII elevation higher than 200% and 14/81 pts had PAI-levels more than two fold higher than the upper normal value. There was 1/81 patient with protein S deficiency and 20/81 pts had abnormal platelet aggregation fulfilling the criteria of hyperreactive platelets.
Conclusions: In patients with occlusions of arterial bypass grafts of the lower extremities we found a high prevalence of hereditary and acquired thrombophilic risk factors. There seems to be a causal relation between arterial bypass occlusions and especially antiphospholipid antibosies. The evaluation of thrombophilic risk factors in patients with arterial bypass occlusions could lead to an improved antithrombotic treatment after surgical revasculation.
Characterization of Anti-Human GPVI Autoantibodies Mediating Persistent Target Immnunodepletion over a 15-Year Period.
Authors: Takayama Hiroshi , Hosaka Yoshitaka , Nakayama Kazuyuki , Shirakawa Kamon , Kawahara Tetsushi , Naitoh Katsuki , Shinozaki Mikihiko , Honda Motoyasu , Yamakawa Toru , Furusako Shoji 
Published: October 2019

Abstract
The first reported patient with platelet GPVI deficiency (Blood1987;69:1712) had anti-GPVI antibodies (YA-Abs) in the patient’s serum, which contributed to later progress in identification of GPVI as a novel collagen receptor. Although there is now a concept that anti-GPVI Ab induces immunodepletion of GPVI in vivo and may become a promising therapeutic antithrombotic agent, precise nature of anti-human GPVI Ab mediating immunodepletion of GPVI is unknown. Since the patient’s platelets are still lacking GPVI and unresponsive to collagen stimulation, we purified YA-Abs from the patient’s serum kept in 1988 (YA-Abs-88) and in 2003 (YA-Abs-03) using sequential affinity chromatography on protein A-Sepharose and recombinant human GPVI fused to Fc (rhGPVI-Fc)-Sepharose. The amount of YA-Abs-03 in the serum was estimated to be about 3 μg/mL and twenty times less than that of YA-Abs-88. We also generated 75 mouse monoclonal antibodies (mAbs) against the extracellular part of human GPVI. Among those, several mAbs cross-reacted with YA-Abs for the binding to rhGPVI-Fc, indicating oligoclonal nature of YA-Abs. Two representative mAbs with no cross-reactivity, F1201 and F1232, cross-reacted with YA-Abs-88 and with both YA-Abs-88 and -03, respectively. Previous studies identified two distinct collagen binding sites of human GPVI; the one is a sequence including Lys59 of human GPVI which is recognized by 10B12 Ab, (Blood2004;103:903) and the other is a sequence including Gly30, Val34 and Leu36 which involves the binding to 9O12 Ab (J.Biol. Chem.2004;279:52293). Point mutated rhGPVI-Fc at Lys59 (K59E) was recognized by YA-Abs-88, -03, and F1232 but not by F1201. Another mutated rhGPVI-Fc in which human GPVI sequence from Pro31 to Leu36 was substituted for mouse GPVI counter sequence was recognized by any of those Abs. YA-Abs-88 and F1201but not F1232 evoked surface P-selectin expression or aggregation of human platelets, while F1232 inhibited collagen-induced aggregation. Since both F1201 and F1232 were found to react with nonhuman primate platelets, either mAb was intravenously injected into monkeys at a dose of 0.3 mg/kg that reached its maximum plasma concentration of 10 μg/mL. Within 1 to 2 days after injection, their platelets showed the lack of both GPVI and collagen-induced aggregation. Thus, GPVI-deficiency of the patient’s platelets is now maintained by unexpectedly much decreased amount of specific Abs which contain F1232-like Ab but not F1201-like Ab included in YA-Abs-88. Immunodepletion of GPVI can be safely induced by specific Ab like F1232 recognizing different epitope from previously reported collagen binding sites where their cross-linking causes platelet activation.
Immune Thrombocytopenia Mediated by Anti-GPIbα Antibodies May Occur Via an FcR-Independent Pathway: A Potential Explanation for Refractory Cases to IVIG Therapy.
Authors: Webster Michelle L. , Sayeh Ebrahim , Crow Min , Chen Pingguo , Nieswandt Bernhard , Freedman John , Ni Heyu 
Published: October 2019

Abstract
Intravenous immunoglobulin G (IVIG) is used to treat immune thrombocytopenia (ITP). While benefit has been observed in many cases, for unknown reasons some patients are refractory to this therapy. ITP is characterized by platelet clearance mainly mediated by antibodies against platelet surface glycoproteins (GP) GPIIbIIIa and GPIbα. Approximately 70–80% patients have antibodies against GPIIbIIIa, 20–40% patients have antibodies against GPIbα complex, and some patients have antibodies to both. Phagocytotic macrophages activated via their Fc receptors (FcR) are thought to be responsible for the clearance of opsonized platelets. However, anti-GPIbα antibodies may be able to induce ITP in an FcR-independent manner. Since the prevailing view of the mechanism of action of IVIG is Fc receptor blockade, we hypothesized that IVIG may not be effective at preventing thrombocytopenia induced by anti-GPIbα antibodies. Thrombocytopenia was induced in BALB/c mice using monoclonal antibodies against mouse GPIbα (p0p3, p0p4, p0p5, p0p9, and p0p11). Pretreatment with different doses of IVIG (1, 2, and 4g/kg of mouse) failed to prevent ITP in all anti-GPIbα-treated mice, except for p0p4. These results were repeated with p0p3, p0p4, and p0p5 using 2g/kg IVIG in C57BL/6 mice, and using 2g/kg of a different IVIG preparation. It has been shown that F(ab)2’ fragments of the p0p antibodies can induce thrombocytopenia to the same extent as intact antibodies. To confirm that these anti-GPIbα antibodies act in an FcR-independent manner, we demonstrated that p0p3, p0p4, and p0p5 are able to induce thrombocytopenia in mice lacking activating Fc receptors (FcRγ chain −/−). Interestingly, IVIG was still able to prevent thrombocytopenia induced by p0p4 in these FcRγ chain −/− mice. We demonstrated in vitro that there are no anti-idiotype antibodies against any of the anti-GPIbα antibodies used present in IVIG. Thus, the amelioration of thrombocytopenia in p0p4-treated mice is not due to a neutralization of the antibody by IVIG. In contrast to the anti-GPIbα antibodies, anti-GPIIbIIIa monoclonal antibodies (JON1, JON2, and JON3) that induce thrombocytopenia were sensitive to IVIG treatment, which is consistent with earlier studies using other monoclonal antibodies against GPIIbIIIa. Also, JON2 was unable to induce thrombocytopenia in FcRγ chain −/− mice, confirming that this anti-GPIIbIIIa antibody is FcR-dependent. Our results suggest that ITP induced by anti-GPIbα antibodies may be mediated by FcR-independent mechanisms, many of which may be refractory to IVIG therapy. Thus, IVIG may not be as effective in treating patients with predominantly anti-GPIbα antibodies.
The HPA-15 (Gov) Platelet Alloantigen System: Allele Frequencies in the Canadian Population.
Authors: Chamberlain Kristen , Smith James W. , Moffatt Diana , Moore Jane , Arnold Donald , Kelton John G. 
Published: October 2019

Abstract
Human platelet antigens (HPAs) are genetically defined polymorphisms expressed on plasma membrane proteins. Alloimmunization following exposure to the incompatible antigen during pregnancy, or following transfusion or transplantation, can cause severe thrombocytopenia and bleeding. The Gov-a/Gov-b biallelic system is expressed on CD109, a 175kD glyco sylphosphatidylinositol-anchored glycoprotein found on platelets, endothelial cells, and activated T-cells (Kelton, 1990; Smith, 1995). Alloimmunization to Gov-a/b is implicated in neonatal alloimmune thrombocytopenia (NAT) and post-transfusion purpura (PTP), and anti-Gov antibodies have been found in polytransfused patients. The Gov-a/Gov-b alleles are encoded by an A-C single nucleotide polymorphism (SNP) at position 2108, resulting in a Tyr682Ser substitution in CD109. Sequence analysis determined that CD109 is a member of the alpha 2 macroglobulin/C3, C4, C5 family of thioester-containing proteins (Lin, 2002). The Gov antigen frequencies are expressed almost equally in the population, and European and Asian studies determined that the Gov-b allele was expressed slightly more often than the Gov-a. Based on these studies the Gov-a/Gov-b antigens were assigned to the HPA-15 system (HPA-15a/15b = Gov-b/Gov-a). South American studies found similar gene frequencies in the general population, but significantly higher frequencies for HPA-15a (0.78) in aboriginal populations (Cardone, 2004). No larger North American studies have been reported since the original report of the Gov allele frequencies based on a limited number of platelet donors (Gov-a: 0.532; Gov-b: 0.468, n=33). We used the polymerase chain reaction with sequence-specific primers (PCR-SSP) to investigate the HPA-15 allele frequencies in the Canadian population. DNA from 273 blood donors was tested using PCR-SSP. The frequency of genotypes HPA-15a/15a, 15a/15b, 15b/15b was found to be 27.84, 49.45, and 22.71 respectively. Thus the allele frequencies for the Canadian population were determined to be 0.517 for HPA-15a and 0.483 for HPA-15b. This data is similar to the frequencies reported among some European studies, and supports evidence that the HPA-15a (Gov-b) allele is expressed slightly more frequently in the general population. Initial reports investigating the Gov alloantigens localized the polymorphism to a novel platelet protein, CD109, and lack of sequence data prevented the use of genotyping studies. The original studies of the Gov antigens, which share almost equal distribution, were limited by available alloantisera and complex screening technologies. The results from this report support the relative allele frequencies for the HPA-15a/15b (Gov-b/Gov-a) antigen system, and emphasize the importance of using genotyping studies to determine allele frequencies.
Direct Injection of Allogeneic Multipotent Adult Progenitor Cells Improves Cardiac Function after Myocardial Infarct.
Authors: Hof Wouter van’t , Mal Niladri , Zhang Ming , Forudi Farhad , Deans Robert , Penn Marc 
Published: October 2019

Abstract
Multipotent adult progenitor cells (MAPC) are an early stem cell population identified in the bone marrow of adults. They have been shown to be capable of differentiating into cell populations derived from all 3 germ layers. We hypothesized that delivery of MAPC in the peri-infarct period could lead to improved cardiac function. MAPC were isolated from the bone marrow of Sprague-Dawley rats and stably labeled by using a lentiviral construct encoding green fluorescent protein (GFP). In-vitro differentiation assays confirmed that the GFP-tagged MAPCs had retained the potential for tri-lineage differentiation. Myocardial infarction was induced in Lewis rats by direct LAD ligation. Lewis rats received Sprague-Dawley MAPC either immediately after LAD ligation by directly injecting into the infarct border zone (5 injections of 400,000 cells per injection) or 24 h after MI by intravenous infusion (2 million cells). Immunohistochemistry was performed to identify MAPC (GFP positive cells) within the infarct zone 2 weeks after MI. We identified MAPC in the infarct border zone of all animals that received direct injection of MAPC, 2 weeks after MI, but did not identify any MAPC following intravenous infusion. Echocardiography was performed to determine if the delivery of MAPC to the myocardium at the time of MI led to improvement in cardiac function. At 2 weeks post-MI, we observed a significant increase in shortening fraction in those animals that received direct injection of MAPC compared to control (13.9 ± 2.2% (n=4) vs. 24.0 ± 6.6% (n=7), p=0.018). These data demonstrate that MAPC delivery to acutely infracted myocardium may offer significant benefit. These data further suggest that MAPC are capable of surviving in the myocardial tissue even when used in an allogeneic strategy without inducing a significant increase in inflammation or rejection.
Cobalamins and Cobinamides Inhibit Nitric Oxide Synthase Enzymatic Activity.
Authors: Weinberg J. Brice , Chen Youwei C. , Beasley Bethany E. , Ghosh Dipak K. 
Published: October 2019

Abstract
Cobalamins (Cbl) serve as important co-factors for methionine synthase and methylmalonyl-coA mutase. They also can block the activities of nitric oxide (NO), as the cobalt binds NO and quenches its bioactivity. NO plays very important roles in normal physiology and in various pathologic processes. NO is synthesized by NO synthases (NOS), enzymes that convert L-arginine to L-citrulline and NO. We postulated that various corrin derivatives (e.g., cobalamins or cobinamides) would inhibit the enzymatic activity of NO synthase (NOS) by directly interacting with NOS. We used purified, recombinant neuronal NOS (NOS1) and inducible NOS (NOS2) in in vitro assays measuring conversion of 14-C-L-arginine to 14-C-L-citrulline. Results showed that hydroxocobalamin (OH-Cbl), cobinamide (Cbi), and dicyanocobinamide (CN2Cbi) were potent NOS1 and NOS2 inhibitors, while cyanocobalamin had much less activity. In general, NOS1 was inhibited more actively than was NOS2, but both were inhibited over 80% by the agents. Methylcobalamin and adenosylcobalamin had very little ability to inhibit NOS activity, but light illumination of these two compounds (a process that liberates the methyl and adenosyl groups from cobalamin, generating OH-Cbl) “activated” them so that they had inhibitory activity comparable to native OH-Cbl
Dose (uM) for 50% inhibition of purified NOS activity (ID50) OH-Cbl Cbi CN2Cbi CN-Cbl AdoCbl (dark) Ado-Cbl (light) NOS1 5.7 15.0 0.2 72.0 498.0 6.4 NOS2 21.1 50.0 2.3 1620.0 2148.0 60.0
We examined the interactions of NOS2 with cobalamins and cobinamides by spectroscopy. Under reduced conditions, oxygen binds to the NOS heme iron, a step that is obligatory for the NO synthesis catalytic cycle. Carbon monoxide (CO, which serves as an oxygen mimic) causes a marked change in the NOS spectrum under reduced conditions. CN2Cbi markedly diminished the CO-induced spectral changes in NOS2, indicating that CN2-Cbi likely blocks binding of CO to the NOS2 oxygenase domain under reduced conditions. This suggests that CN2Cbi-mediated inhibition of NOS activity is due to blocking of oxygen binding to heme, a step required for the NOS catalytic cycle. When we incubated OH-Cbl or CN2Cbi with cells of the mouse macrophage cell line J774 activated with interferon-gamma and endotoxin to produce NO, there was a dose-dependent inhibition of NO production. While certain cobalamins and cobinamides have been previously noted to bind and quench/scavenge the actions of preformed NO, we show here for the first time that cobalamins and cobinamides can potently inhibit the enzymatic function of NOS and synthesis of NO. These agents (or their derivatives) might serve as NOS inhibitors for treatment of various conditions. Also, native tissue cobalamins and cobinamides might act as important regulators of NOS activity in normal and pathological conditions in vivo.