21 research outputs found
Identification and Characterization of a Critical CP2-binding Element in the Human Interleukin-4 Promoter
No full textSpecificities and Cell Distribution of Transcription Factors Binding To the Interleukin-4 (IL-4) Negative Regulatory Element (NRE)
No full textVEGF-A stimulates ADAM17-dependent shedding of VEGFR2 and crosstalk between VEGFR2 and ERK signaling
No full textVascular endothelial growth factor (VEGF)-A and the VEGF receptors are critical for regulating angiogenesis during development and homeostasis and in pathological conditions, such as cancer and proliferative retinopathies. Most effects of VEGF-A are mediated by the VEGFR2 and its coreceptor, neuropilin (NRP)-1. Here, we show that VEGFR2 is shed from cells by the metalloprotease disintegrin ADAM17, whereas NRP-1 is released by ADAM10. VEGF-A enhances VEGFR2 shedding by ADAM17 but not shedding of NRP-1 by ADAM10. VEGF-A activates ADAM17 via the extracellular signal-regulated kinase (ERK) and mitogen-activated protein kinase pathways, thereby also triggering shedding of other ADAM17 substrates, including tumor necrosis factor alpha, transforming growth factor alpha, heparin-binding epidermal growth factor-like growth factor, and Tie-2. Interestingly, an ADAM17-selective inhibitor shortens the duration of VEGF-A-stimulated ERK phosphorylation in human umbilical vein endothelial cells, providing evidence for an ADAM17-dependent crosstalk between the VEGFR2 and ERK signaling. Targeting the sheddases of VEGFR2 or NRP-1 might offer new opportunities to modulate VEGF-A signaling, an already-established target for treatment of pathological neovascularization
Impaired endothelial barrier function in apolipoprotein M-deficient mice is dependent on sphingosine-1-phosphate receptor 1
No full textApolipoprotein M (ApoM) transports sphingosine-1-phosphate (S1P) in plasma, and ApoM-deficient mice (Apom(-/-)) have ∼50% reduced plasma S1P levels. There are 5 known S1P receptors, and S1P induces adherens junction formation between endothelial cells through the S1P1 receptor, which in turn suppresses vascular leak. Increased vascular permeability is a hallmark of inflammation. The purpose of this study was to explore the relationships between vascular leakage in ApoM deficiency and S1P1 function in normal physiology and in inflammation. Vascular permeability in the lungs was assessed by accumulation of dextran molecules (70 kDa) and was increased ∼40% in Apom(-/-) mice compared to WT (C57Bl6/j) mice. Reconstitution of plasma ApoM/S1P or treatment with an S1P1 receptor agonist (SEW2871) rapidly reversed the vascular leakage to a level similar to that in WT mice, suggesting that it is caused by decreased plasma levels of S1P and reduced S1P1 stimulation. In a carrageenan-induced model of inflammation, Apom(-/-) mice had increased vascular leakage compared with that in WT mice. Adenoviral overexpression of ApoM in Apom(-/-) mice decreased the vascular leakage compared to adenoviral overexpression of green fluorescent protein. The study suggests that vascular leakage of albumin-sized particles in ApoM deficiency is S1P- and S1P1-dependent and this dependency exacerbates the response to inflammatory stimuli.-Christensen, P. M., Liu, C. H., Swendeman, S. L., Obinata, H., Qvortrup, K., Nielsen, L B., Hla, T., Di Lorenzo, A., Christoffersen, C. Impaired endothelial barrier function in apolipoprotein M-deficient mice is dependent on sphingosine-1-phosphate receptor 1.</p
Stimulation of Platelet-derived Growth Factor Receptor β (PDGFRβ) Activates ADAM17 and Promotes Metalloproteinase-dependent Cross-talk between the PDGFRβ and Epidermal Growth Factor Receptor (EGFR) Signaling Pathways
No full textDevelopment and initial validation of the comprehensive HIV adherence with treatment scale
No full text2021 Spring.Includes bibliographical references.HIV remains a significant public health concern despite decreasing rates of transmission in the U.S. (Centers for Disease Control and Prevention, 2020c). Contributing factors include low rates of treatment adherence (de Bruin et al., 2010) and high rates of comorbidities with other medical and mental health conditions (Bing et al., 2001; Gallant et al., 2017; Lerner et al., 2020). Antiretroviral therapy (ART) has significantly improved HIV health outcomes and reduced AIDS diagnoses and AIDS-related mortality (Crum et al., 2006; Glass et al., 2008; Ickovics & Meade, 2002; Paterson et al., 2000; Stone, 2001; World Health Organization, 2015). Because of ART's effectiveness, HIV is considered a chronic rather than terminal health condition for people adherent with treatment (Aberg, 2006; Swendeman, Ingram, & Rotheram- Borus, 2009). Treatment for HIV as a chronic health condition includes several pro-health behaviors in addition to ART adherence to support overall wellness. To support future research and treatment recommendations, the current study developed a measure of adherence with pro- health behavior and conducted an initial analysis of the measure's psychometric properties with a sample of 118 people living with HIV. Structural equation modeling explored relations among antecedents (personality, treatment self-efficacy, treatment information, and treatment motivation) and health outcomes of pro-health behaviors and ART adherence. Regularly assessing engagement in, as well as antecedents and outcomes of, treatment behaviors can enhance communication between providers and people living with HIV, reinforce HIV's status as a manageable chronic condition, and link people living with HIV to appropriate interventions
ADAM33: a cellular model of over-expression. From full length to soluble
No full textPolymorphic variation in the A Disintegrin And Metalloprotease 33 (ADAM33) genehas been associated with asthma and bronchial hyperresponsiveness. The ADAMfamily of proteins has a multi-domain structure and has diverse biological functionswhich can include growth factor shedding, cell migration and cell adhesion.Evidence for a soluble form of ADAM33 has been found in BAL fluid of asthmaticpatients and there is evidence that ADAM33 has a function in angiogenesis.To determine the possible function(s) of the full length ADAM33 protein, it wasover expressed in HEK293 that do not normally express ADAM33 and thephenotypic consequences analyzed. Cells stably expressing full length ADAM33under the control of a CMV promoter were obtained by selection with the antibiotic,G418. The expression of ADAM33 was confirmed by western blot analysis. Controlcells were transfected with an empty vector control encoding G418 resistance only.Cells were examined for changes in cell phenotype and ability to produce solubleADAM33.Cells stably expressing ADAM33 grew more quickly than mock transfected cellsbut this effect was lost over time. However, processed (ie. active) ADAM33 proteincould not be detected by western blot. In order to determine whether asubpopulation of cells was able to produce active ADAM33, the cells were clonedand re-cloned by limiting dilution. This allowed isolation of a number of ADAM33clones that produced processed protein. Furthermore, these experiments revealedthat ADAM33 expression increased the clonogenic potential of the cells.Supernatant from cloned cells was pulled down with ConA and samples analysed bywestern blot which showed a band at 50-55kDa in the ADAM33 clones using anantibody to the metalloproteinase domain. This soluble ADAM33 was active andpromoted angiogenesis using a HUVEC capillary tube forming assay. SolubleADAM33 was upregulated in the presence of TGF-?2 and its shedding is unliky tobe autocatalytic.Over expression of ADAM33 in HEK293 cells alters the clonogenic potential andsurvival of HEK293 cells in low density cultures. This property may contribute tothe behaviour of mesenchymal cells in asthmatic airways. Soluble ADAM33 isreleased into culture media and has functional activity. Its release is up regulated inthe presence of TGF-?2
Characterization of the catalytic properties of the membrane-anchored metalloproteinase ADAM9 in cell-based assays
No full textADAM9 (A Disintegrin And Metalloprotease 9) is a membrane-anchored metalloproteinase that has been implicated in pathological retinal neovascularization and in tumor progression. ADAM9 has constitutive catalytic activity in both biochemical and cell-based assays and can cleave several membrane proteins, including epidermal growth factor and Ephrin receptor B4; yet little is currently known about the catalytic properties of ADAM9 and its post-translational regulation and inhibitor profile in cell-based assays. To address this question, we monitored processing of the membrane-anchored Ephrin receptor B4 (EphB4) by co-expressing ADAM9, with the catalytically inactive ADAM9 E &gt; A mutant serving as a negative control. We found that ADAM9-dependent shedding of EphB4 was not stimulated by three commonly employed activators of ADAM-dependent ectodomain shedding: phorbol esters, pervanadate or calcium ionophores. With respect to the inhibitor profile, we found that ADAM9 was inhibited by the hydroxamate-based metalloprotease inhibitors marimastat, TAPI-2, BB94, GM6001 and GW280264X, and by 10 nM of the tissue inhibitor of metalloproteinases (TIMP)-3, but not by up to 20 nM of TIMP-1 or -2. Additionally, we screened a non-hydroxamate small-molecule library for novel ADAM9 inhibitors and identified four compounds that selectively inhibited ADAM9-dependent proteolysis over ADAM10- or ADAM17-dependent processing. Taken together, the present study provides new information about the molecular fingerprint of ADAM9 in cell-based assays by showing that it is not stimulated by strong activators of ectodomain shedding and by defining a characteristic inhibitor profile. The identification of novel non-hydroxamate inhibitors of ADAM9 could provide the basis for designing more selective compounds that block the contribution of ADAM9 to pathological neovascularization and cancer.</jats:p
Sphingosine 1-Phosphate Receptor 1 Signaling Maintains Endothelial Cell Barrier Function and Protects Against Immune Complex-Induced Vascular Injury
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