Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 3 de 3
Filter
Add more filters










Database
Language
Publication year range
1.
Analyst ; 140(13): 4558-65, 2015 Jul 07.
Article in English | MEDLINE | ID: mdl-26040325

ABSTRACT

Phospholipids are important membrane components involved in diverse biological activities ranging from cell signaling to infection by viral particles. A thorough understanding of protein-phospholipid interaction dynamics is thus crucial for deciphering basic cellular processes as well as for targeted drug discovery. For any specific phospholipid-protein binding experiment, various groups have reported different binding constants, which are strongly dependent on applied conditions of interactions. Here, we report a method for accurate determination of the binding affinity and specificity between proteins and phospholipids using a model interaction between PLC-δ1/PH and phosphoinositide phospholipid PtdIns(4,5)P2. We developed an accurate Force Distance Spectroscopy (FDS)-based assay and have attempted to resolve the problem of variation in the observed binding constant by directly measuring the bond force. We confirm the FDS findings of a high bond strength of ∼0.19 ± 0.04 nN by Surface Plasmon Resonance (SPR) data analysis, segregating non-specific interactions, which show a significantly lower K(D) suggesting tight binding.


Subject(s)
Blood Proteins/chemistry , Blood Proteins/metabolism , Microscopy, Atomic Force/methods , Phosphatidylinositol 4,5-Diphosphate/metabolism , Phosphoproteins/chemistry , Phosphoproteins/metabolism , Surface Plasmon Resonance/methods , Kinetics , Protein Binding , Protein Structure, Tertiary
2.
Am J Transplant ; 15(7): 1768-81, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25727073

ABSTRACT

The loss of a functional microvascular bed in rejecting solid organ transplants is correlated with fibrotic remodeling and chronic rejection; in lung allografts, this pathology is predicted by bronchoalveolar fluid neutrophilia which suggests a role for polymorphonuclear cells in microcirculatory injury. In a mouse orthotopic tracheal transplant model, cyclosporine, which primarily inhibits T cells, failed as a monotherapy for preventing microvessel rejection and graft ischemia. To target neutrophil action that may be contributing to vascular injury, we examined the effect of a neutrophil elastase inhibitor, elafin, on the microvascular health of transplant tissue. We showed that elafin monotherapy prolonged microvascular perfusion and enhanced tissue oxygenation while diminishing the infiltration of neutrophils and macrophages and decreasing tissue deposition of complement C3 and the membrane attack complex, C5b-9. Elafin was also found to promote angiogenesis through activation of the extracellular signal-regulated kinase (ERK) signaling pathway but was insufficient as a single agent to completely prevent tissue ischemia during acute rejection episodes. However, when combined with cyclosporine, elafin effectively preserved airway microvascular perfusion and oxygenation. The therapeutic strategy of targeting neutrophil elastase activity alongside standard immunosuppression during acute rejection episodes may be an effective approach for preventing the development of irreversible fibrotic remodeling.


Subject(s)
Cyclosporine/pharmacology , Drug Synergism , Elafin/pharmacology , Graft Rejection/prevention & control , Microvessels/pathology , Organ Transplantation/adverse effects , Trachea/transplantation , Animals , Apoptosis/drug effects , Blotting, Western , Cell Movement/drug effects , Cells, Cultured , Chemotaxis/drug effects , Complement C3/metabolism , Drug Therapy, Combination , Endothelium, Vascular/drug effects , Female , Graft Rejection/etiology , Graft Rejection/pathology , Graft Survival/drug effects , Human Umbilical Vein Endothelial Cells , Humans , Immunosuppression Therapy , Leukocyte Elastase/metabolism , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Microcirculation , Microvessels/drug effects , Perfusion , Protease Inhibitors/pharmacology , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Wound Healing/drug effects
3.
Proc Natl Acad Sci U S A ; 106(29): 12145-50, 2009 Jul 21.
Article in English | MEDLINE | ID: mdl-19581601

ABSTRACT

A number of distinct beta-amyloid (Abeta) variants or multimers have been implicated in Alzheimer's disease (AD), and antibodies recognizing such peptides are in clinical trials. Humans have natural Abeta-specific antibodies, but their diversity, abundance, and function in the general population remain largely unknown. Here, we demonstrate with peptide microarrays the presence of natural antibodies against known toxic Abeta and amyloidogenic non-Abeta species in plasma samples and cerebrospinal fluid of AD patients and healthy controls aged 21-89 years. Antibody reactivity was most prominent against oligomeric assemblies of Abeta and pyroglutamate or oxidized residues, and IgGs specific for oligomeric preparations of Abeta1-42 in particular declined with age and advancing AD. Most individuals showed unexpected antibody reactivities against peptides unique to autosomal dominant forms of dementia (mutant Abeta, ABri, ADan) and IgGs isolated from plasma of AD patients or healthy controls protected primary neurons from Abeta toxicity. Aged vervets showed similar patterns of plasma IgG antibodies against amyloid peptides, and after immunization with Abeta the monkeys developed high titers not only against Abeta peptides but also against ABri and ADan peptides. Our findings support the concept of conformation-specific, cross-reactive antibodies that may protect against amyloidogenic toxic peptides. If a therapeutic benefit of Abeta antibodies can be confirmed in AD patients, stimulating the production of such neuroprotective antibodies or passively administering them to the elderly population may provide a preventive measure toward AD.


Subject(s)
Aging/immunology , Alzheimer Disease/immunology , Amyloid beta-Peptides/chemistry , Amyloid beta-Peptides/immunology , Antibodies/immunology , Neuroprotective Agents/immunology , Peptides/immunology , Aging/drug effects , Alzheimer Disease/blood , Alzheimer Disease/complications , Alzheimer Disease/pathology , Amyloid beta-Peptides/toxicity , Animals , Antibodies/blood , Antibodies/cerebrospinal fluid , Cytoprotection/drug effects , Dementia/complications , Dementia/immunology , Disease Progression , Genes, Dominant , Immunization , Immunoglobulin G/blood , Mice , Molecular Weight , Neurons/cytology , Neurons/drug effects , Peptides/chemistry , Primates/immunology , Protein Processing, Post-Translational/drug effects , Protein Structure, Quaternary
SELECTION OF CITATIONS
SEARCH DETAIL
...