Blood
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Patients given unfractionated heparin (UFH) or low-molecular-weight heparin (LMWH) for prophylaxis or treatment of thrombosis sometimes suffer serious bleeding. We showed previously that peptides containing 3 or more tandem repeats of heparin-binding consensus sequences have high affinity for LMWH and neutralize LMWH (enoxaparin) in vivo in rats and in vitro in citrate. We have now modified the (ARKKAAKA)(n) tandem repeat peptides by cyclization or by inclusion of hydrophobic tails or cysteines to promote multimerization. ⋯ These peptides (1 mg/300 g rat) neutralized 1 U/mL anti-Factor Xa activity of enoxaparin in rats within 1 to 2 minutes. Direct blood pressure and heart rate measurements showed little or no hemodynamic effect. These heparin-binding peptides, singly or in combination, are potential candidates for clinical reversal of UFH and LMWH in humans.
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SHP-1, an src homology 2 (SH2) domain containing protein tyrosine phosphatase, functions as a negative regulator of signaling downstream of cytokine receptors, receptor tyrosine kinases and receptor complexes of the immune system. Dephosphorylation of receptors and/or receptor-associated kinases has been described as the mechanism for the function of SHP-1. Here we demonstrate a novel mechanism by which SHP-1 down-regulates the Janus kinase-2 (Jak2)/signal transducer and activator of transcription-5 (Stat5) pathway downstream of the prolactin receptor (PRLR) and the erythropoietin receptor (EPOR) in a catalytic activity-independent manner. ⋯ Our results further indicate that these tyrosine residues, via recruitment of the adaptor protein Grb-2, are required for targeting the inhibitory protein suppressor of cytokine signaling-1 (SOCS-1) to Jak2 kinase. Finally, loss of SOCS-1 expression in SOCS-1(-/-) mouse embryonic fibroblast (MEF) cells led to attenuation in SHP-1 function to down-regulate PRL-induced Stat5 activation. All together, our results indicate that SHP-1 inhibits PRLR and EPOR signaling by recruitment and targeting of SOCS-1 to Jak2, highlighting a new mechanism of SHP-1 regulation of cytokine-receptor signaling.
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Deficiency of granulocyte-macrophage colony-stimulating factor (GM-CSF) in mice results in pulmonary alveolar proteinosis (PAP) from impaired surfactant catabolism by alveolar macrophages (AMs). Recently, we have shown that neutralizing anti-GM-CSF autoantibodies develop specifically in patients with idiopathic pulmonary alveolar proteinosis (iPAP). Analogous to murine PAP models, it is plausible that the autoantibodies reduce GM-CSF activity, resulting in AM dysfunction and surfactant accumulation. ⋯ Although target epitopes varied among patients, GM-CSF amino acids 78 to 94 were consistently recognized. Thus, autoantibodies bind GM-CSF with high specificity and high affinity, exist abundantly in the lung, and effectively block GM-CSF binding to its receptor, inhibiting AM differentiation and function. Our data strengthen the evidence associating anti-GM-CSF autoantibodies with the pathogenesis of this disease.
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Case Reports Comparative Study
Congenital afibrinogenemia: identification and expression of a missense mutation in FGB impairing fibrinogen secretion.
Congenital afibrinogenemia is a rare autosomal recessive disorder characterized by complete absence of detectable fibrinogen. We previously identified the first causative mutations for this disease: a homozygous deletion of approximately 11 kb of the fibrinogen alpha-chain gene (FGA). ⋯ In this study, a young boy with afibrinogenemia was found to be a compound heterozygote for 2 mutations in FGB: an N-terminal nonsense mutation W47X (exon 2) and a missense mutation (G444S, exon 8). Coexpression of the FGB G444S mutant cDNA in combination with wild-type FGA and FGG cDNAs demonstrated that fibrinogen molecules containing the mutant beta chain are able to assemble but are not secreted into the media, confirming the pathogenic nature of the identified mutation.
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Eph receptor tyrosine kinases and their ligands, the ephrins, have been primarily described in the nervous system for their roles in axon guidance, development, and cell intermingling. Here we address whether Eph receptors may also regulate dendritic cell (DC) trafficking. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that DCs derived from CD34+ progenitors, but not from monocytes, expressed several receptors, in particular EphA2, EphA4, EphA7, EphB1, and EphB3 mRNA. ⋯ Interestingly, ephrin expression was detected in epidermal keratinocytes and also in DCs. Adhesion of CD34+-derived DCs to fibronectin, but not to poly-l-lysine, was increased in the presence of ephrin-A3-Fc, a ligand of EphA2, through a beta1 integrin activation pathway. As such, EphA2/ephrin-A3 interactions may play a role in the localization and network of Langerhans cells in the epithelium and in the regulation of their trafficking.