bluuebunny/crossref_metadata_2025_split huggingface dataset:
Whole Liver Iron Overload Measurement by Magnetic Iron Detector (MID). A Non Cryogenic Bio-Susceptometer.
Authors: Marinelli Mauro , Beruto Piergiorgio , Gianesin Barbara , Lavagetto Antonella , Lamagna Martina , Oliveri Eraldo , Saccone Maria Luigia , Sobrero Giuliano , Terenzani Laura , Luca Gian 
Published: October 2019

Abstract
Accurate assessment of body-iron accumulation is essential for diagnosis and therapy of iron-overload in diseases such as thalassemia, hereditary hemochromatosis and other forms of severe congenital or acquired anemias. At present, the gold standard to determine liver-iron concentration (LIC) is the invasive liver needle biopsy. This technique might lead to large error, in assessing iron burden, due to the heterogeneous distribution of iron deposition in the liver. SQUID bio-susceptometer and MRI are currently the only non-invasive validated methods for LIC measurements. The susceptometer presented herein, named Magnetic Iron Detector (MID), measures directly the iron overload in the whole liver. All of its components operate at room temperature. Since February 2005 about 150 patients and 90 healthy volunteers have been measured and the measures were obtained in blind. The local Ethics Committee approved the study and all subjects gave informed consent. The result of correlations with the LIC measurements by SQUID susceptometry (Dr. A. Piga, Turin) in 43 patients showed a R 0.86 (Fig 1). In 2 patients, affected by Congenital Hemocromatosis, we correlated the LIC measurement by MID with the assessment of the expected iron depletion obtained with the phlebotomy therapy R 0.94 (Fig 2). All the measurements were correlated with the serum-ferritin concentration values R 0.72. We obtained correlation with the LIC measurement by liver biopsy in 7 patients R 0.89, further measures are in progress. The reproducibility of the iron overload of the same patients, measured after a relatively short lapse of time, is better than 0.5g. In conclusion the data obtained shows that MID is a reliable instrument for the diagnosis of the liver iron overload and for the follow-up of the chelation therapy. It is simpler to operate being manageable directly in the Clinical Center and more affordable than competing techniques.
Fig. 1 LIC measured by MID vs LIC measured by SQUID Fig. 1. LIC measured by MID vs LIC measured by SQUID Fig. 2 The iron reduction of two hemochromatosis patients, under phebotomy therapy, compare with the reduction measured by the MID. Fig. 2. The iron reduction of two hemochromatosis patients, under phebotomy therapy, compare with the reduction measured by the MID.
Thyroid Function in Thalassemic Patients Treated with Combined Chelation Therapy.
Authors: Farmaki Kalistheni , Angelopoulos Nicholas , Anagnostopoulos George , Goula Anastasia , Pappa Christina 
Published: October 2019

Abstract
Recently introduced chelation regimens that combine deferoxamine (DFO) and deferiprone (DFP) have been shown to have greater efficacy in promoting iron excretion than either chelator alone and have been associated with rapid reduction of the iron load in the heart and liver, and with reversal of cardiac dysfunction. It is unclear whether this combined therapy could be associated with a decline in the severity of iron-induced endocrinopathies. The primary endpoint of the present study was to investigate the effects of this therapy on the thyroid function in thalassemic patients with subclinical hypothyroidism (SH) or normal thyroid function. Starting in January 2001, 42 patients with b-tlalassaemia major, previously maintained on subcutaneous DFO only, were switched to combined treatment with DFO and DFP. Before the initiation of combined therapy 14 patients had overt hypothyroidism and were treated with thyroxin substitution. The thyroid function of the remaining patients with normal fasting levels of both FT4 and FT3 was further assessed with TRH test. TSH was measured at 0, 30, 60 and 90 minutes after IV injection of 200 mcg of TRH. 15 patients with normal TSH responses (7 males, 8 females, age 28.86 ± 2.20 years, mean ± SEM), and 13 patients (6 males, 7 females, age 31.15 ± 1.85 years), who were considered suffering from SH were finally enrolled. Criteria for the diagnosis of SH was an elevated basal TSH concentration (>5 TSH μIU/ml) or an increment of the TSH levels during the test more than 20 μIU/ml from the basal value. Combination therapy markedly decreased ferritin levels (585 ± 457 vs. 2124 ± 456 μg/l, P < 0.001 in SH group and 868 ± 339 vs. 2877 ± 552 μg/l, P < 0.001 in eythyroidal group). At the time of reassessment (June 2006), the levels of TSH were decreased at all times during the second TRH test in patients with SH: Basal TSH: 4.12 ± 0.63 vs. 6.27 ± 1.08, P=0.01. TSH At 30′ mins: 22.13 ± 2.18 vs. 34.06 ± 4.75, P=0.005. TSH At 60′ mins: 15.89 ± 1.13 vs. 25.69 ± 3.72, P=0.002. TSH At 90′ mins: 11.83±1.26 vs. 19.44±3.27, P=0.001. TSH quantitative secretion, calculated as the area under the curve, was also significantly decreased with combined therapy (1380±118) compared with the initial assessment (2178±312, P=0.004), while no change occurred in basal FT4 (1.06 ± 0.04 vs. 1.15 ± 0.08 in 2001, normal range 0.71–1.85 ng/ml) and FT3 levels (1.55± 0.07 vs. 1.59± 0.08 in 2001, normal range 1.45–3.48 pg/ml). Nevertheless, 7 patients with previous SH exhibited normal results in the reassessment. In patients with previous normal thyroid function TSH response to TRH stimulus was significantly improved only at 60′ mins (9.78±0.73 vs. 11.49±1.28, P=0.03). This study showed that the combination of DFO and DFP followed by an intensive iron chelation might be associated with an improvement in thyroid function in the early stages of hypothyroidism.
Mac-1 and F4/80 Surface Markers Are Present on Murine Hematopoietic Stem Cells.
Authors: Zomorodian Toska J. , Greer Debbie , Wood Kyle , Foster Bethany , Demers Delia , Johnson Kevin , Quesenberry Peter J. , Abedi Mehrdad 
Published: October 2019

Abstract
Transplanted bone marrow donor cells with tissue specific phenotypes have been found in the brain, liver, heart, skin, lung, kidney, and gut of transplanted humans and mice. Such observations have led to the controversial hypothesis that hematopoietic stem cells (HSC) might be intrinsically plastic, and through transdifferentiation or fusion lead to the repair of damaged tissues throughout the body. Alternately, it is suggested that fusion of macrophages to the recipient cells may explain this phenomenon. We have shown recently that purified HSC are the cells responsible for GFP positive donor-derived muscle fibers in the recipient mice post bone marrow transplantation. However, further studies sorting for macrophage markers Mac-1 and F4/80 also resulted in donor-derived muscle fibers in the host. To address this discrepancy, we investigated subpopulations of Mac-1 and F4/80 positive cells, in the presence or absence of stem cell markers (Sca-1 and C-kit). We demonstrate that only the subpopulations of Mac-1 and F4/80 positive cells harboring stem cell markers, Sca-1 or c-kit, were capable of contributing to the regenerating muscle post transplantation. Furthermore, these same subpopulations demonstrated single cell High Proliferative Potential (HPP) (6–26%) in a 7 factor cytokine cocktail, compared to the Mac-1 or F4/80 cells with no stem cell markers (0%). Additionally, they demonstrated long-term engraftment in all three lineages at 1-year (average chimerism of 55% versus 0% in stem cell marker negative groups). These subpopulations were also evaluated for morphology using Hematoxylin/Eosin (H/E), Wright-Giemsa, and Nonspecific Esterase staining. In the Mac-1 and F4/80 positive groups, those negative for stem cell markers resembled differentiated cells of the myeloid origin (macrophages, granulocytes), while those with positive stem cell markers demonstrated stem cell characteristics. We did not observe any engraftability, donor-derived muscle fibers, or HPP potential for CD14 or cfms positive cells coexpressing stem cell markers, indicating that these markers are more appropriate for identifying macrophages. In conclusion, our studies demonstrate that both Mac-1 and F4/80 surface markers are present on HSC and therefore caution must be taken in the interpretation of data using these macrophage markers. It is reasonable to believe that the use of Mac-1 and/or F4/80 surface markers in a lineage depletion process may result in the loss of a subpopulation of stem cells, and other markers such as CD14 or c-fms may be more appropriate for eliminating differentiated macrophages.
Hematopoietic and Endothelial Progenitor Cells from Mouse Adult Testis Derived Stem Cell Lines.
Authors: Schreiter Jessica , Guan Kaomei , Nayernia Karim , Chapuy Bjoern , Truemper Lorenz , Hasenfuss Gerd , Engel Wolfgang , Wulf Gerald G. 
Published: October 2019

Abstract
Adult mouse testis was recently found to harbour multipotent adult germline stem cells (maGSC) with the differentiation capacity along several mesodermal and epithelial lineages. We here addressed the hematopoietic potential from three multipotent germline stem cell lines in vitro. In the undifferentiated state, these cells expressed stem cell markers including Sca1, Thy1 and c-kit, but lacked markers of definitive hematopoiesis such as CD45. On exposure to in vitro culture conditions favouring hematopoietic differentiation, these cells gave rise to both primitive erythropoiesis with nucleated red blood cells expressing ζ-globin, and definitive hematopoiesis, forming β-globin positive erythroid and myeloid colonies in colony assays. On OP9 stroma cells, the maGSC lines formed mesoderm-like colonies, from which progenitor cell populations for either endothelial or hematopoietic differentiation were isolated. The differentiation patterns and efficiencies were similar to the results obtained from the mouse embryonic stem cell line MPI2. In summary, adult germline derived stem cell lines can be induced to recapitulate early embryonic hemangioblastic development and may thus serve as tools for studies on hematopoietic and angioblastic development as well as applications in regenerative medicine.
Cleavage at Arg740 Appears Essential for Thrombin-Catalyzed Cleavage at Arg372 during the Activation of Factor VIII.
Authors: Newell Jennifer , Fay Philip J. 
Published: October 2019

Abstract
Factor VIIIa serves as an essential cofactor for the factor IXa-catalyzed activation of factor X during the propagation phase of coagulation. The factor VIII procofactor is converted to factor VIIIa by thrombin-catalyzed proteolysis of three P1 positions at Arg372 (A1–A2 junction), Arg740 (A2–B junction), and Arg1689 (a3–A3 junction). Cleavage at Arg372 exposes a cryptic functional factor IXa-interactive site, while cleavage at Arg1689 liberates factor VIII from von Willebrand factor and contributes to factor VIIIa specific activity, thus making both sites essential for procofactor activation. However, cleavage at Arg740, separating the A2–B domainal junction, has not been rigorously studied. To evaluate thrombin cleavage at Arg740, we prepared and stably expressed two recombinant factor VIII mutants, Arg740His and Arg740Gln. Results from a previous study examining proteolysis at Arg372 revealed substantially reduced cleavage rates following substitution of that P1 Arg with His, whereas replacing Arg with Gln at residue 372 yielded an uncleavable bond at that site (Nogami et al., Blood, 2005). Specific activity values for the factor VIII Arg740His and Arg740Gln variants as measured using a one-stage clotting assay were approximately 50% and 18%, respectively, that of the wild type protein. SDS-PAGE and western blotting following a reaction of factor VIII Arg740His with thrombin showed reduced rates of cleavage at His740 as well as at Arg372 relative to the wild type. Alternatively, factor VIII Arg740Gln was resistant to thrombin cleavage at Gln740 and showed little, if any, cleavage at Arg372 over an extended time course. The mutant proteins assayed in a purified system by factor Xa generation showed a slight increase in activity for the Arg740His variant compared with the Arg740Gln variant in both the absence and presence of thrombin, and the activities for both variants were reduced compared with wild type factor VIII. These results suggest that cleavage at residue 740 affects subsequent cleavage at Arg372 and generation of the active cofactor factor VIIIa. Preliminary results obtained evaluating proteolysis of these mutants by factor Xa, which cleaves the same sites in factor VIII as thrombin, also revealed slow proteolysis at the P1 His and no cleavage at the P1 Gln. However, subsequent cleavage at Arg372 exhibited less dependence on initial cleavage at residue 740. These observations may explain the higher than predicted specific activity values obtained for the two variants and suggest a different mechanism of action for the two activating proteinases. Overall, these results support a model whereby cleavage of factor VIII heavy chain by thrombin is an ordered pathway with initial cleavage at Arg740 required to facilitate cleavage at the critical Arg372 site to yield the active cofactor.
The Effect of an Arg/Trp Polymorphism in α2-Antiplasmin on Fibrinolysis.
Authors: Christiansen Victoria J. , Jackson Kenneth W. , Lee Kyung N. , Chissoe Dini , McKee Patrick A. 
Published: October 2019

Abstract
α2-antiplasmin (α2AP) is a serine proteinase inhibitor with plasmin as its primary target. Plasmin plays a critical role in fibrin proteolysis and tissue remodeling and is highly regulated to prevent excessive proteolysis. α2AP is cleaved by antiplasmin-cleaving enzyme (APCE) between Pro12-Asn13 to yield a shortened version with Asn as the amino-terminus. Asn-α2AP is crosslinked into fibrin ~13 X faster than the full-length Met-α2AP. Clot lysis is slowed in direct proportion to the ratio of Asn-α2AP to Met-α2AP present during fibrin formation. The cloned gene for Met-α2AP contained either a CGG codon resulting in Arg as the sixth amino acid or TGG, resulting in Trp at that position. Both polymorphic forms of Met-α2AP, Arg6 and Trp6, have been found in human plasma. In the present study we determined the genotype frequency of the polymorphism and whether it affected cleavage of Met-α2AP to Asn-α2AP, and ultimately whether fibrinolytic rates were affected by one polymorphic form as opposed to the other. In our normal population, the genotype frequencies were found to be RR 60.9%; RW 35.8%; WW 3.4%. The R allele had a frequency of 78.8% and the W allele had a frequency of 21.2%. A primate DNA panel revealed that the polymorphism follows an evolutionary split with Old World monkeys having the WW genotype and Great Apes having the RR genotype. We found that the polymorphism affected the ratio of Asn-α2AP to Met-α2AP in plasma with WW having the highest percentage of Met-α2AP (56.4%), RR having the least (23.6%) and RW falling in between (40.6%). We examined whether this was due to Met-α2AP(Arg6) being a better substrate for APCE than Met-α2AP(Trp6). When purified α2AP(Arg6) or α2AP(Trp6) were digested with APCE, comparisons of reaction rates based on linear regression analysis of early time points revealed that APCE cleaved Met-α2AP(Arg6) ~8-fold faster that Met-α2AP(Trp6). This phenomenon was confirmed in whole plasma, by incubating plasma from subjects with either the RR, RW or WW genotype and determining the Asn-α2AP/Met-α2AP ratio at 0, 24 and 48 hours. When APCE was removed from the plasma by a specific antibody bound to chromatography beads, the ratio of Asn-α2AP to Met-α2AP did not change over the same time period. Since Asn-α2AP/Met-α2AP ratios are in direct proportion to clot lysis times, we determined whether the polymorphism also impacts and correlates with correspondent plasma clot lysis times. The first tertile of plasma clot lysis times contained all of the WW persons; the longer lysis times in the second and third tertiles contain only RR and RW persons. We conclude that the Arg6Trp polymorphism is functionally significant in that it clearly affects conversion of Met-α2AP to Asn-α2AP, and as a consequence, the rate of Asn-α2AP incorporation into fibrin. Based on these data, the Arg6Trp polymorphism may play a significant role in governing the long-term deposition/removal of intravascular fibrin.
Identification of a Thrombin-Interactive Site within Factor VIII A2 Domain Responsible for the Cleavage at Arg372.
Authors: Nogami Keiji , Shima Midori , Saenko Evgueni L. , Takeyama Masahiro , Tatsumi Kohei , Fujita Yuri , Tanaka Ichiro , Yoshioka Akira 
Published: October 2019

Abstract
Factor VIII is activated by limited proteolytic cleavage at Arg372, Arg740, and Arg1689 by thrombin or factor Xa. Recently, it has been reported that the C2 domain and the acidic region comprising residues 389–394 in the A2 domain interacted with thrombin to facilitate cleavage at Arg1689 and Arg740, respectively. However, a thrombin-interactive site(s) responsible for cleavage at Arg372 remain to be determined. In the current study, several approaches were employed to examine the thrombin-interactive site(s). Thrombin-catalyzed factor VIII activation was inhibited by the addition of one anti-A2 monoclonal antibody 413 with an epitope of residues 484–509 in a dose-dependent manner using one-stage clotting assay. SDS-PAGE analysis revealed that cleavage rate at Arg372 in the heavy chain by thrombin selectively decreased by ~25% in the presence of the antibody. However, cleavage at Arg740 was not affected. Direct binding experiments by both surface plasmon resonance-based assay and ELISA using anhydro-thrombin (Ah-thrombin), catalytically inactive derivative of thrombin in which the active-site serine was converted to dehydroalanine, showed that anti-A2 antibody 413 inhibited the A2 subunit binding to Ah-thrombin. A synthetic peptide corresponding to the 484–509 residues also inhibited the A2/Ah-thrombin binding by ~40%. Mutant A2 molecules where the basic residues in the A2 domain were converted to alanine were evaluated for the binding of Ah-thrombin. Among the tested mutants, K466A mutant possessed ~330-fold lower affinity than the wild type A2 (Kd; 18400 nM, 55 nM, respectively), and the affinities of R471A, R484A, R489A, R490A, H497A, K499A, and K523A were decreased by 2–5-fold (Kd; 259, 212, 183, 163, 123, 255, and 165 nM, respectively). These findings demonstrate that clustered basic residues, in particular Lys466, encompassed the 484–509 region play a key role of thrombin-binding site within the A2 domain, responsible for thrombin-catalyzed factor VIII activation by cleavage at Arg372.
Absence of CD81 Paradoxically Results in a Hyper-IgM and IgG Response to T-Independent Antigens.
Authors: Sanyal Mrinmoy , Shoham Tsipi , Fernandez Rosemary , Levy Shoshana 
Published: October 2019

Abstract
The tetraspanin CD81 is required for numerous biological functions including fertilization, infection, cell migration and cellular interactions in the nervous and immune systems. In B cells CD81 is a component of the CD19/CD21 signaling complex. CD81 was shown to facilitate the redistribution of the B cell receptor (BCR) complex and CD21 into lipid rafts in response to co-engagement, and to modulate BCR signaling. In addition, CD81-deficient mice express low levels of cell surface CD19, thereby potentially altering signaling by the CD19/CD21 co-receptor complex. Interestingly, the onset of CD81 expression coincides with the onset of CD19 expression during B cell development. The foregoing observations suggest that CD81 might reduce the in vivo response of B cells to antigenic stimulation. To test this hypothesis we compared the response of CD81-deficient and wild type mice to T-independent (TNP-LPS) and T-dependent (TNP-KLH) antigens. Surprisingly, CD81-deficient mice mounted significantly higher IgM responses against both types of antigens. Moreover, the IgG response of CD81-deficient mice was stronger and persistent in response to T-independent antigen. We further found that CD81-deficient mice have an increase in bone marrow perisinusoidal B cells (IgM+IgD+). These cells are primarily responsible for mounting T-independent immune responses against blood-borne pathogens. In addition, CD81-deficient spleenic B cells have an intrinsic ability to produce higher amounts of IgM. These surprising results suggest that CD81 is involved in modulating B cell activation, particularly in response to infection.
Enhanced IL-6 Production by Bone Marrow Derived Dendritic Cells from Lupus-Prone Mice Inhibits CD4+CD25+ T Cell Mediated Suppression.
Authors: Wan Suigui , Xia Changqing , Morel Laurence 
Published: October 2019

Abstract
The B6.Sle1.Sle2.Sle3 triple congenic mouse (B6.TC) is a model of lupus due to the co-expression of the three major NZM2410-derived susceptibility loci on a C57BL/6 background. B6.TC mice produce high titers of anti-nuclear nephrogenic autoantibodies and a highly penetrant glomerulonephritis. Previous studies have shown the Sle1 locus is associated with a reduced number of regulatory T cells (Treg), and that Sle3 results in intrinsic defects in myeloid cells that hyperactivate T cells. Here, we show that B6.TC dendritic cells (DCs) accumulate in lymphoid organs and present a defective maturation process, in which bone-marrow derived DCs, plasmacytoid and myeloid DCs express a significantly lower level of CD80, CD86 and class II MHC than B6 controls. B6.TC DCs also induce a higher level of proliferation in CD4+ T cells than B6 DCs, and B6.TC DCs block the suppressive activity of Treg. B6.TC DCs over-produce IL-6, which is necessary for the blockade of Treg activity, as shown by anti-IL-6 neutralizing antibody in the suppression assays. The over-production of IL-6 by DCs and the blockade of Treg activity maps to Sle1, which therefore not only confers a reduced number of Treg, but also blocks their ability to regulate autoreactive T cells. Taken together, these results provide a genetic and mechanistic evidence for systemic autoimmunity resulting from an impaired regulatory T cell compartment both in number and function, and for Sle1-expressing DCs playing a major role in the latter defect though their production of IL-6.