bluuebunny/crossref_metadata_2025_split huggingface dataset:
Inhibition of Hb S Polymerization In Vitro by a Novel EF Helix ß73 His Containing Peptide.
Authors: Akbar Mohammed G.K. , Ding Min , Surrey Saul , Adachi Kazuhiko 
Published: October 2019

Abstract
Sickle hemoglobin polymerization involves not only ß6 Val in a largely hydrophobic acceptor pocket but other contact sites. We recently found that the ß73 His Hb S variant (ß6 Val and ß73 His) promoted polymerization compared to deoxy Hb S, while ß73 Leu Hb S (ß6 Val and ß73 Leu) inhibited polymerization like deoxy Hb C-Harlem (ß6 Val and ß73 Asn). In fact, ß73 Asp in Hb S makes a hydrogen bond with ß4 Thr in deoxy Hb S polymers, and this is a unique position to promote or inhibit polymerization by amino acid change [Adachi et al., Biochemistry (2003), 42, 4476]. Kinetics of polymerization, solubility and minimum concentration required for polymerization of the ß73 Hb S variants were affected by ß73 amino acid (inhibition of polymerization: His < <Asp <<Asn < Leu). Inhibition of Hb S ß73 Leu polymerization compared to Hb S may be caused by weakening of the hydrogen bond interaction between the hydroxyl group of ß4 Thr and the ß73 amino acid like Hb C-Harlem. Furthermore, kinetics of polymerization of 1:1 Hb S/Hb A ß73 His mixtures were enhanced compared to AS mixtures, while Hb S/Hb A ß73 Leu mixtures showed inhibitory effects similar to FS mixtures. These results suggest that the Hb A ß73His variant promotes Hb S polymerization almost as efficiently as Hb S ß73 His, and that the ß73 His in Hb A and Hb S variants strengthens the hydrogen bond with ß4 Thr, which helps facilitate formation of domains and 14-stranded fibers.
Based on these results, we chemically synthesized a 15-mer EF helix peptide containing ß73 His (Lys-Lys-Val-Leu-Gly-Ala-Phe-Ser-His-Gly-Leu-Ala-His-Leu-Asp) and evaluated effects of this peptide on Hb S polymerization. DIC analysis in 1.0 M phosphate buffer showed peptide-induced inhibition of fiber elongation rate for deoxy-Hb S which increased linearly with increasing amounts of peptide (e.g., a 5-fold molar excess of peptide resulted in a 6-fold decrease in fiber elongation rate). Solubility of deoxy Hb S increased linearly with increasing peptide (e.g., a 5-fold molar excess of peptide increased solubility by 1.2-fold). The delay time prior to polymerization of deoxy Hb S in a high phosphate buffer also was increased significantly in the presence of this peptide. In contrast, the same 15-mer peptide containing ß73 Leu instead of His showed no effect on solubility or kinetics of polymerization, suggesting that the ß73 His peptide specifically affects the A-helix of Hb S containing ß4 Thr just like the ß73 His Hb S variant. This and possibly other peptides interrupting the ß73-ß4 interaction which result in inhibition of deoxy Hb S polymerization may be good candidates for inhibitors of Hb S polymerization.
A Sickle Transgenic Mouse Model of Acute Chest Syndrome.
Authors: Fabry Mary E. , Alleyne Dawn , Klings Elizabeth S. , Farber Harrison , Zhang Yingyi , Resnick Robert , Aldrich Thomas K. , Steinberg Martin H. , Nagel Ronald L. 
Published: October 2019

Abstract
Acute chest syndrome (ACS), the most common cause of death among adults with sickle cell disease and a significant threat to children, is often triggered by bone marrow embolization, especially among adults. We have recently developed a sickle transgenic mouse model for acute chest that consists of injecting a mixture of bone marrow and mouse fat into the tail vein of a sickle transgenic partial knockout mouse. The mouse used was a NY1 partial KO expressing 100% human α and 55% βS globins. Mice were injected with bone marrow (BM) from two femurs (about 15 mg) and purified peritoneal fat (about 15 mg, as murine BM lacks fat), and examined at 0, 6, 24, and 48 hours post-injection. Creatine kinase (CK), predominantly the MM form, increased above baseline levels at 6, 24, and 48 hours in NY1 partial KO mice; whereas, in control C57Bl mice, there was an initial increase at 6 hours with a return to baseline values at 24 and 48 hours. sVCAM levels were not significantly different between C57Bl and NY1 partial KO mice at baseline: 717±203 in C57Bl mice and 752±147 in NY1 partial KO mice (N=15 and 11, respectively); after injection of fat/BM sVCAM levels nearly doubled in NY1 partial KO mice and remained elevated at 48 hours. In fat/BM injected C57Bl mice, sVCAM levels remained essentially constant from 0 to 48 hours. Mouse lungs were harvested at 48 hours after saline perfusion and inflated with 10% phosphate buffered formalin. The hematoxylin and eosin stained lung sections were normal among the C57Bl control mice examined 48 hours after fat/BM injection, but among NY1 partial KO mice edema, inflammation, and congestion were found with numerous red cells entrapped, despite saline perfusion. In summary, this preparation shows promise as an animal model of ACS. Pulmonary edema and elevated levels of sVCAM are characteristic features of ACS in sickle cell patients and are found, after fat/BM injection, in our sickle mice but not in control mice. The mice survived the procedure, indicating that it will be possible to study the natural course of lung injury and interventions that may ameliorate injury.
Adult Bone Marrow-Derived Cell Fusion with Cardiomyocytes and Purkinje Neurons in Response to Irradiation but Not in Steady State.
Authors: Nygren Jens M. , Stott Simon , Liuba Karina , Breitbach Martin , Röll Willhelm , Fleischmann Bernd , Kirik Deniz , Björklund Anders , Jacobsen Sten Eirik W. 
Published: October 2019

Abstract
Several recent studies have suggested that bone marrow (BM) cells can contribute to non-hematopoietic cell lineages through cell fusion rather than transdiffentiation. As this phenomenon has been observed in multiple organs, including the brain and heart, without prior infliction of organ-specific insults, it has been proposed that BM cells might contribute to replacement of non-hematopoietic cell lineages during steady state, and that BM transplantation might be developed as a therapeutic modality in diseases of these organs. However, as all observations of BM-derived cell fusion in vivo have been made in lethally irradiated mice reconstituted with genetically marked BM cells, we addressed to what degree cell fusion occurs normally and/or in response to whole body irradiation. To be able to distinguish between these possibilities we used c-kit deficient (w41/w41) mice, which unlike wild type mice do not require irradiation-induced myeloablation to facilitate reconstitution of transplanted BM cells. Noteworthy, no BM-derived cell fusion events were observed in the brain (purkinje neurons) or heart (cardiomyocytes) when unconditioned w41/w41 mice were reconstituted with beta actin GFP transgenic BM cells. In striking contrast, following whole body irradiation (875 rad), BM-derived cell fusion was observed in recipient cardiomyocytes and purkinje neurons of all BM transplanted mice. Thus, spontaneous adult BM-derived cell fusion does not occur in steady state but is potently facilitated by irradiation-induced injuries to the organs in which cell fusion occurs.
Decreased Serum Pro-Hepcidin in Thalassaemia Intermedia; a Mechanism for Increased Iron Absortion Mediated Though Ineffective Erthropoiesis and NTBI?.
Authors: Shah Farrukh T. , Rafique Roozina , Evan Patricia , Porter John B. 
Published: October 2019

Abstract
Background: Pro-hepcidin is 10 kDa peptide localized at the basolateral membrane of hepatocytes which is also present in plasma and is believed to be a hepcidin precursor (Kulaksis et al, Gut2004;53). Experimental hepatic iron loading in mice up-regulates hepcidin mRNA while iron deficiency has the reverse effect (Pigeon et al, JBC2001;276). Paradoxically mice with a thalassaemia intermedia phenotype had reduced liver hepcidin mRNA. Furthermore, presentation of non-transferrin bound iron (NTBI) to human liver cells (Nemeth, Blood2003; 101)(Rafique, 2004 in preparation) decreased hepcidin mRNA. Patients and methods: 3 groups of patients were compared (thalassaemia major (TM), thalassaemia intermedia (TI) and sickle cell anaemia (HbSS)) to examine if variations in pro-hepcidin expression could account for the differences in iron loading. Pro-hepcidin measurements were performed at 1:1 dilutions in duplicate with a Pro-hepcidin ELISA (DRG diagnostics) using a previously published method (Kulakis, 2004). Iron status was evaluted by liver CMR , serum ferritin and NTBI; C reactive protein (CRP) was measured to exclude inflammation. All patients with a raised creatinine were excluded from the analysis. Results: TI patients (n=20) had a significantly lower serum pro-hepcidin (77 ± 40ng/ml) ( p<0.001) compared to HbSS (n=26) (111 ± 53 ng/ml), TM (n=27) (107 ± 44.5ng/ml) or control subjects (n=9) (119 ± 45ng/ml). The difference in serum pro-hepicidin between TI and HbSS persisted when patients were matched for serum ferritin (p = 0.02) or liver iron (p < 0.001). NTBI was significantly lower in HbSS (1.06 ± 0.27umol/l) than TI (2.49 ± 2.01 umol/l) even when patients were matched for serum ferritin (p <0.001) or for liver iron (p <0.05). Hb values were not significantly different between TI (8.6 g/dl) and HbSS patients (8.9g/dl). When all patients with a Hb <8g/dl were compared to those with a Hb > 10g/dl serum pro-hepcidin was not significantly different. No correlation between serum pro-hepcidin and liver iron, serum ferritin, CRP or Hb in any of the three groups was seen. Conclusion: Pro-hepcidin is significantly lower in TI than TM or HbSS patients and this cannot be accounted for by levels of anemia. We propose that NTBI, present early in TI secondary to high ineffective erythropoiesis, may lead to a similar paradoxical decrease in hepcidin synthesis reported in model systems and that this is reflected by reduced serum pro-hepcidin. Although levels of NTBI and liver iron did not differ significantly in TI and TM , transfusion and chelation therapy in TM may overide the NTBI mediated down regulation of pro-hepcidin. In HbSS, NTBI is lower than in TI or TM at matched levels of iron loading, resulting in less inhibition of pro-hepcidin synthesis and less duodenal iron absorption than in TI.
Early Hepatitis C Viral Response (EVR) to Peginterferon Alfa 2a and Ribavirin in Patients with β Thalassemia.
Authors: Harmatz Paul , Olivieri Nancy F. , Vichinsky Elliott , Kwiatkowski Janet , Giardina Patricia , Shah Farrukh , Elizabeth Wright C. , Cunningham Melody , Maureen Jonas 
Published: October 2019

Abstract
Hepatitis C virus (HCV) infection is an important cause of liver failure and death in patients with thalassemia who received frequent blood transfusions before HCV screening became routine. Severe transfusion-related iron overload may accelerate the progression of liver disease, influence response to therapy, and increase the risk of antiviral treatment in these patients. An open-label, single arm trial of peginterferon alfa 2a (180 ug weekly) and ribavirin (1200 mg/day for > 75 kg, 1000 mg/day 50 to 75 kg, 800 mg for < 50kg) was initiated in August, 2003 in the NIH-sponsored Thalassemia Clinical Trials Network to examine safety and efficacy in adult patients. Although the study is still enrolling patients, we are presenting viral response data for the first 16 patients (mean age: 33 yrs, range 22 to 44, 50% female, 75% genotype 1). Iron burden was assessed by chemical analysis of liver biopsy and HCV RNA levels were measured using the Roche COBAS HCV Tests (qualitative and quantitative). These analyses were performed by a central laboratory. HCV levels were measured at baseline, weeks 1, 2, 3, 4, 12 and 24.
Early Virologic Responses
Week 1 Week 2 Week 3 Week 4 Week 12 Week 24 *number with result available; **non-detectable; ***greater than 2 log decrease N* 16 16 16 16 11 6 Non-d** 1 2 2 3 7 5 >2-log 5 6 7 7 3 1 % >2 log 38% 50% 56% 63% 91% 100%
Four patients discontinued between weeks 4 through 12 (1 death, 1 persistent cough, 1 extreme fatigue, and 1 pregnancy (spouse). These patients had all achieved > 2 log drop by week 4. The mean liver iron at baseline was 10.1 mg/gm dry wt (range: 1.1 to 21.2) and the mean log HCV RNA level was 5.8 (range 4.0 to 6.8). The median time to > 2 log decrease was 1 week for patients with genotype 2 and 4 weeks for patients with genotype 1 (p=0.004). There was no correlation either between baseline iron or baseline HCV level and time to > 2 log decrease. Transfusion requirement increased 52% as assessed at 12 weeks and did not predict earlier viral response. In summary, there is EVR (>2 log drop) in 63% at 4 weeks and 91 % at 12 weeks, comparable to those seen in patients without thalassemia. Quantitative hepatic iron concentration at baseline was not a predictor of EVR. Completion of the study is planned to determine whether the EVR predicts sustained viral response and whether the iron overload or increased transfusion requirements during therapy pose significant safety issues.
Hamster Prions Are a Suitable Model for Partitioning of Human CJD Prions during Plasma Processing Steps.
Authors: Groener Albrecht , Schäfer Wolfram , Baron Henry , Vey Martin 
Published: October 2019

Abstract
Prion removal evaluation of plasma processing procedures is one important basis to assess the margin of safety of plasma protein therapeutics. Currently, in these evaluation studies to assess the removal capacity of selected manufacturing steps for human prions mainly prions derived from scrapie-infected hamsters or mice are used in spiking studies. In order to test the validity of hamster prions instead of different human prion strains as spiking reagents, we compared the partitioning of these prion preparations at two purification steps common to the manufacturing of several human plasma-derived therapeutic proteins at ZLB Behring. The glycine precipitation (inherent in the manufacture of e.g., vWF/Factor VIII) and the 25% ethanol precipitation step (inherent in the manufacture of e.g., alpha-1-proteinase inhibitor and albumin) were down scaled to mimic the production process. A microsomal membrane preparation as well as purified full-length pathogenic prion (PrPSc; without membrane association) prepared (a) from hamster brains infected with the model scrapie strain Sc237, (b) from brains of transgenic mice infected with human sporadic CJD prions, and (c) from the brain of a patient who died of vCJD were used for spiking experiments. The different prion preparations were spiked to the starting material of the manufacturing steps studied and the manufacturing step was performed in a dedicated laboratory. The amount of PrPSc in all fractions was determined quantitatively utilizing an ELISA-formatted Conformation Dependent Immunoassay [Safar et al., Nat Med1998; 41157–1165]. The 25% ethanol precipitation removed very effectively all prion preparations equally regardless of their origin (reduction factor ≥ 3 log10). The glycine precipitation removed the microsomal as well as the purified prion preparations from all species equally whereby prions of the purified prion preparation were removed to a considerably higher degree than the membrane preparation (reduction factor ≥ 3 log10 vs. 1.6 to 1.8 log10). In parallel, prion infectivity of each sample is currently tested in animal bioassays involving transgenic mice susceptible for the tested prion strain. It can be concluded, up to now based on immunological data, that spiked hamster prions are a reliable model to confirm the safety of human plasma-derived products also for human derived prions.
GPIbα Is Essential for Platelet Adhesion during Thrombus Formation: Studies with Mutant Mice Deficient in the Extracellular Domain of GPIbα.
Authors: Bergmeier Wolfgang , Piffath Crystal L. , Ruggeri Zaverio M. , Ware Jerry L. , Wagner Denisa D. 
Published: October 2019

Abstract
GPIbα binding to von Willebrand factor (vWf) exposed at a site of vascular injury is the first step in the formation of a hemostatic plug. In addition, GPIbα binding to platelet-bound vWf has previously been shown to play a key role in the incorporation of circulating platelets into a growing thrombus. However, studies in vWf-deficient mice demonstrated delayed but not absent arterial thrombus formation, suggesting that GPIbα may bind a ligand other than vWf to facilitate platelet adhesion, or that a platelet surface receptor other than GPIbα can mediate platelet adhesion in arterioles in the absence of vWf. Here we studied thrombus formation in transgenic mice expressing GPIbα in which the extracytoplasmic sequence has been replaced by an isolated domain of the human interleukin 4 receptor (IL4R-tg mice). Early platelet adhesion to ferric chloride-treated mesenteric arterioles in IL4R-tg mice was decreased by >98% when compared with controls. As a consequence, thrombus formation was completely inhibited in all of the mutant mice. To study the role of GPIbα in platelet adhesion to already adherent platelets, we studied thrombus formation in wild-type mice infused with wild-type platelets labeled with calcein-green and IL4R-tg platelets labeled with calcein-orange/red. Upon ferric chloride-induced injury, wild-type but not IL4R-tg platelets incorporated into the growing thrombus. A similar result was observed with wild-type platelets treated with O-sialoglycoprotein endopeptidase to remove the 45 kD N-terminal domain of GPIbα. In summary, our studies in GPIbα mutant mice provide additional evidence that under arterial flow conditions GPIbα is the only receptor expressed on the platelet surface that mediates initial platelet adhesion to the subendothelium as well as to already adherent platelets. Our data further suggest that a ligand other than vWf may contribute to GPIbα-dependent platelet adhesion to subendothelium and to a growing thrombus.
Polynitroxyl Albumin Prevents Vaso-Occlusion in Transgenic Sickle Mice.
Authors: Mahaseth Hemchandra , Belcher John D. , Welch Thomas E. , Sonbol Khalid M. , Bowlin Paul R. , Hebbel Robert P. , Ma Li , Hsia Carleton J.C. , Vercellotti Gregory M. 
Published: October 2019

Abstract
Sickle cell disease is characterized by excessive oxidative stress. Sickle patients have enhanced rates of endogenous reactive oxygen species (ROS) production and impaired antioxidant defense mechanisms. We hypothesize that excessive production of ROS promotes activation of vascular endothelium and vaso-occlusion in sickle cell disease. Intravascular ROS production has been shown to trigger the activation of vascular endothelium and increase leukocyte-endothelium interactions. We investigated whether inhibition of intravascular ROS production by treatment with polynitroxyl albumin (PNA), a superoxide dismutase and catalase mimetic agent, could modulate endothelial cell activation and vaso-occlusion in S+S Antilles transgenic sickle mice after hypoxia-reoxygenation. Nuclear factor-kappa B (NF-kB), an oxidant sensitive transcription factor critical for endothelial cell activation, was elevated in lungs and livers of sickle mice and was reduced 2.5 (p=0.043) and 1.5 (p=0.043) fold respectively, by treatment with PNA after hypoxia-reoxygenation. Similarly, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) were elevated 3 to 4.7 fold in the same organs (p<0.05). After treatment with PNA, VCAM-1 expression was reduced 2.3 fold in lungs (p=0.014) and 1.5 fold in livers (p=0.006), ICAM-1 expression decreased 1.7 fold in lungs (p=0.014) and 2 fold in livers (p=0.007). Control studies using human serum albumin had no effect on NF-kB activation or adhesion molecule expression in sickle mice. These anti-inflammatory effects of PNA on endothelium corresponded with changes in blood flow in dorsal skin venules. Intravital microscopy revealed that PNA significantly inhibited hypoxia-reoxygenation-induced leukocyte rolling (3.8 fold inhibition, p<0.001) and completely inhibited vaso-occlusion (p<0.001) in venules in the dorsal skin of transgenic sickle mice. We speculate that therapies that reduce ROS will result in improved organ perfusion in sickle cell disease.
Childhood Paroxysmal Nocturnal Hemoglobinuria (PNH), a Report of 11 Cases in the Netherlands.
Authors: Van Den Heuvel-Eib M.M. , Bredius R.G.M. , te Winkel M.L. , Tamminga R. , de Kraker J. , Meeteren N. Schouten-van , Bruin M. , Korthof E.T. 
Published: October 2019

Abstract
PNH, a rare disease in children, is caused by an acquired somatic mutation of the phosphatidyl-inositol glycan (GPI)class A (PIG-A) gene, followed by a survival-advantage of the PNH clone. This results in a deficiency of GPI-anchored proteins on the hematopoietic cells, making them more susceptible for attacks of the complement system, causing hemolysis and pancytopenia. Binding-impairment of coagulation-system-regulators increases thrombosis-risk. Until now, world-wide only 40 cases of pediatric PNH have been reported. Clinical data from Dutch cases from 1983–2003 were collected nation-wide and studied retrospectively. Diagnosis was confirmed by a decrease of GPI-linked anchor proteins on granulocytes and/or erythrocytes (CD24, CD66b/67, CD59, CD16).
All 11 PNH patients (4 boys, 7 girls), with a median age of 11 years (range 6–14 years) presented with pancytopenia-related symptoms. Two cases showed clinical hemoglobinuria, and one developed thrombosis during the course of the disease. BM biopsies showed hypoplasia/aplasia (AA)(n=7) or MDS (n=4). All cases showed decreased percentages of the GPI-linked proteins, 3/8 showed positive acid-Ham tests, 5/6 showed positive sucrose-lysis tests. Seven patients received prednisolone, in 5 combined with androgens and in 4 with ATG. After initial treatment, 3 patients reached complete remission (CR), and 2 showed partial response (PR), 1 subsequently relapsed and was transplanted. Three died of disease related mortality, one showed spontaneous recovery of blood counts.In total 5 patients underwent BMT (5MUD/2MSD). Conditioning regimens were Busulphan/Melphalan/Cyclophosphamide (n=2) in MDS, and Cyclophosphamide/ low dose TBI (n=3) in AA. Three patients received additional anti-Tcell antibody therapy. Of the 5 BMT patients 4 in CCR (median follow-up: 6 years, 10 months (range 7 months-14 years)). One patient died post-BMT of intracranial hemorrhage. PNH, a rare but serious disease in children, should be considered in every child with aplastic anemia/hypoplastic MDS. Hemoglobinuria is not an obligatory presenting symptom. GPI-linked anchor protein analysis provides the most reliable diagnostic tool. If prednisolone/ATG/androgens is not effective, BMT should be considered even if no matched sibling donor is available.