scholarly journals Application of a Static Fluorescence-based Cytometer (the CellScan) in Basic Cytometric Studies, Clinical Pharmacology, Oncology and Clinical Immunology

2005 ◽  
Vol 12 (3) ◽  
pp. 187-195 ◽  
Author(s):  
Michal Harel ◽  
Boris Gilburd ◽  
Yael S. Schiffenbauer ◽  
Yehuda Shoenfeld
Keyword(s):  
Clinical Pharmacology ◽  
Lupus Erythematosus ◽  
Cell Biology ◽  
Clinical Immunology ◽  
Laser Scanning ◽  
Lymphocyte Activation ◽  
Living Cells ◽  
Peripheral Blood Lymphocytes ◽  
Laser Scanning Cytometer

The CellScan apparatus is a laser scanning cytometer enabling repetitive fluorescence intensity (FI) and polarization (FP) measurements in living cells, as a means of monitoring lymphocyte activation. The CellScan may serve as a tool for diagnosis of rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) as well as other autoimmune diseases by monitoring FP changes in peripheral blood lymphocytes (PBLs) following exposure to autoantigenic stimuli. Changes in FI and FP in atherosclerotic patients' PBLs following exposure to various stimuli have established the role of the immune system in atherosclerotic disease. The CellScan has been evaluated as a diagnostic tool for drug-allergy, based on FP reduction in PBLs following incubation with allergenic drugs. FI and FP changes in cancer cells have been found to be well correlated with the cytotoxic effect of anti-neoplastic drugs. In conclusion, the CellScan has a variety of applications in cell biology, immunology, cancer research and clinical pharmacology.

Hepcidin: A Clue for a New Role for Lymphocytes.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1999-1999
Author(s):  
Jorge P Pinto ◽  
Vera Dias ◽  
Heinz Zoller ◽  
Pedro N Rodrigues ◽  
Helena Carmo ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Iron Overload ◽  
T Lymphocyte ◽  
Peripheral Blood ◽  
Lymphocyte Activation ◽  
Peripheral Blood Lymphocytes ◽  
Ferric Citrate ◽  
Intracellular Iron ◽  
Hepcidin Expression

Abstract Abstract 1999 Poster Board I-1021 Background: Recent evidence suggests the involvement of lymphocytes in the severity of iron overload disorders, with increased severity of iron overload in Hereditary Hemochromatosis patients and in animal models with lymphocyte number deficiencies. However, no mechanism(s) has been suggested to explain these observations. The aim of this study was to analyze hepcidin expression in lymphocytes. Methods: Expression of hepcidin was analyzed by Real-time PCR in human and mouse Peripheral Blood Lymphocytes (PBLs) and in selected resting lymphocyte populations, in response to holotransferrin and ferric citrate. The effect of hepcidin in the expression of the iron exporter Ferroportin was analyzed by FACS and confocal immunofluorescence. Cellular iron traffic was analyzed by measurement of 55Fe and 125I-TF, cell proliferation assessed by BrdU incorporation and silencing of gene expression in lymphocytes performed with siRNAs. Results: Hepcidin is expressed in PBLs and is up-regulated in response to holotransferrin and ferric citrate. The response to holotransferrin was observed in CD8+ and not in CD4+ lymphocytes, a result confirmed by the failure of lymphocytes from β2-microglobulin-KO mice to respond to holotransferrin, in comparison with Bl6/J controls. Hepcidin up-regulation induced by holotransferrin decreases ferroportin expression, inducing its co-localization with the proteasome marker LMP2. Tumor Necrosis Factor-á (TNF-á) expression in PBLs increases with holotransferrin treatments. siRNA-mediated silencing of TNF-á in PBLs abrogates hepcidin up-regulation by holotransferrin and incubation of PBLs with recombinant TNF-á increases hepcidin expression, suggesting the involvement of this cytokine in the basal and holotransferrin-induced hepcidin expression in these cells. The role of hepcidin in a situation of high iron demand - lymphocyte activation and proliferation - was assessed. Hepcidin expression increases with T-lymphocyte activation and siRNA-mediated silencing of hepcidin in activated T lymphocytes causes a decrease in intracellular iron levels, by increasing ferroportin-mediated iron export. The low intracellular iron levels were associated with impaired T lymphocyte proliferation. Discussion: The ability of PBLs to increase hepcidin expression in response to ferric citrate distinguishes lymphocytes from hepatocytes and places peripheral blood lymphocyte numbers as a first line of response to increases in transferrin saturation and presence of NTBI, characteristic of iron overload disorders. The findings that hepcidin modulates ferroportin expression, intracellular iron levels and cell proliferation in lymphocytes demonstrate the importance of this protein for lymphocyte iron homeostasis. The control of the intracellular iron levels of lymphocytes confers to hepcidin a pivotal role in the postulated ability of circulating lymphocytes to function as “biological iron chelators”. Conclusion: With the demonstration, for the first time, of hepcidin synthesis by peripheral blood lymphocytes, of its regulation by elemental iron and of its involvement in T cell proliferation, the present results put forward a molecular mechanism for the described modifier role of lymphocytes in protection from iron toxicity. Disclosures: No relevant conflicts of interest to declare.

scholarly journals CD8+DR+ T-Cells and C3 Complement Serum Concentration as Potential Biomarkers in Thrombotic Antiphospholipid Syndrome

2014 ◽  
Vol 2014 ◽  
pp. 1-8
Author(s):  
Elizabeth Sarmiento ◽  
Jonathan Dale ◽  
Mauricio Arraya ◽  
Antonio Gallego ◽  
Nallibe Lanio ◽  
...  
Keyword(s):  
T Cells ◽  
Antiphospholipid Syndrome ◽  
Lupus Erythematosus ◽  
Neuropsychiatric Symptoms ◽  
Lymphocyte Activation ◽  
Immunological Parameters ◽  
Clinical Criteria ◽  
Clinical Complications ◽  
Potential Biomarkers

Purpose. To assess complement factors and T lymphocyte activation subset abnormalities in patients with thrombotic antiphospholipid syndrome (APS) as potential biomarkers for development of clinical complications.Methods. We assessed C3, C4, factor B concentrations (nephelometry), complement haemolytic functional activity (CH100, radial immune diffusion), and the activation status of CD4+ and CD8+ T-cells (three-colour flow cytometry) in patients with thrombotic APS. Antiphospholipid (aPL) positive patients without APS-related clinical criteria, systemic lupus erythematosus (SLE) patients, and healthy individuals were evaluated as controls. A clinical followup was performed to assess the potential relationship between the immunological parameters and development of APS-related complications.Results. Lower concentrations of C3 and higher levels of CD8+DR+ cells were risk factors for development of APS-related complications during followup, including rethrombosis and neuropsychiatric symptoms. Patients with diagnosed thrombotic APS had significantly lower levels of C3, C4, and CH100 as well as higher percentages of activated CD4+DR+ and of CD8+DR+ T-cells than healthy controls but similar to that observed in autoimmune disease controls.Conclusion. Lower C3 and C4 complement levels and higher percentages of CD8+DR+ T-cells were observed in thrombotic APS patients. The potential role of these abnormalities as biomarkers of clinical outcome warrants further evaluation in a multicenter study.

scholarly journals Activation and tyrosine phosphorylation of p72syk as well as calcium mobilization after hydrogen peroxide stimulation in peripheral blood lymphocytes

1995 ◽  
Vol 308 (1) ◽  
pp. 347-352 ◽  
Author(s):  
S Qin ◽  
T Inazu ◽  
H Yamamura
Keyword(s):  
Hydrogen Peroxide ◽  
Tyrosine Phosphorylation ◽  
Peripheral Blood ◽  
Lymphocyte Activation ◽  
Peripheral Blood Lymphocytes ◽  
Calcium Mobilization ◽  
Cellular Proteins ◽  
Blood Lymphocytes ◽  
Cell Lysates

To determine the regulatory role of p72syk in lymphocyte activation, peripheral blood lymphocytes were treated with 10 mM hydrogen peroxide. Hydrogen peroxide induced a rapid elevation of p72syk activity (4-6-fold) and a dramatic increase in tyrosine phosphorylation of a number of cellular proteins, including phospholipase C gamma 1 (PLC gamma 1) and p72syk. Monoclonal antibodies to PLC gamma 1 co-precipitated p72syk from hydrogen peroxide-stimulated cell lysates, but not from unstimulated cell lysates. Furthermore, we observed a rise in intracellular Ca2+, corresponding to the combination of extracellular Ca2+ influx and the release from intracellular Ca2+ stores. Extracellular Ca2+ influx was necessary for the sustenance of p72syk activity, but not for the initiation of p72syk activation induced by hydrogen peroxide. Taken together, these data suggested that one possible role of p72syk was to activate PLC gamma 1, at least in part through tyrosine phosphorylation, and then to trigger calcium mobilization in pig peripheral blood lymphocytes in response to hydrogen peroxide stimulation.

scholarly journals The Role of Gamma Delta T Cells in Autoimmune Rheumatic Diseases

Cells ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 462 ◽  
Author(s):  
Ilan Bank
Keyword(s):  
T Cells ◽  
T Cell ◽  
Rheumatic Diseases ◽  
Lupus Erythematosus ◽  
Cell Biology ◽  
Γδ T Cells ◽  
Immune Functions ◽  
Autoimmune Rheumatic Diseases ◽  
New Therapeutic Approaches

Autoimmune rheumatic diseases (ARDs), affecting ~1–1.5% of all humans, are associated with considerable life long morbidity and early mortality. Early studies in the 1990s showed numerical changes of the recently discovered γδ T cells in the peripheral blood and in affected tissues of patients with a variety of ARDs, kindling interest in their role in the immuno-pathogenesis of these chronic inflammatory conditions. Indeed, later studies applied rapid developments in the understanding of γδ T cell biology, including antigens recognized by γδ T cells, their developmental programs, states of activation, and cytokine production profiles, to analyze their contribution to the pathological immune response in these disorders. Here we review the published studies addressing the role of γδ T in the major autoimmune rheumatic diseases, including rheumatoid arthritis, juvenile idiopathic arthritis, ankylosing spondylitis, systemic lupus erythematosus and scleroderma, and animal models thereof. Due to their unique properties spanning adaptive and innate immune functions, the ever deeper understanding of this unique T cell population is shedding new light on the pathogenesis of, while potentially enabling new therapeutic approaches to, these diseases.

Application of FRAP and digitized fluorescence microscopy to problems in cell membrane dynamics

1988 ◽  
Vol 46 ◽  
pp. 118-119
Author(s):  
K. Jacobson ◽  
A. Ishihara ◽  
B. Holifield ◽  
F. Zhang
Keyword(s):  
Fluorescence Microscopy ◽  
Cell Biology ◽  
Extended Period ◽  
Dynamic Structure ◽  
Living Cells ◽  
Membrane Dynamics ◽  
Molecular Cell Biology ◽  
Image Averaging ◽  
Single Living Cells ◽  
Single Living

Our laboratory is concerned with understanding the dynamic structure of the plasma membrane with particular reference to the movement of membrane constituents during cell locomotion. In addition to the standard tools of molecular cell biology, we employ both fluorescence recovery after photo- bleaching (FRAP) and digitized fluorescence microscopy (DFM) to investigate individual cells. FRAP allows the measurement of translational mobility of membrane and cytoplasmic molecules in small regions of single, living cells. DFM is really a new form of light microscopy in that the distribution of individual classes of ions, molecules, and macromolecules can be followed in single, living cells. By employing fluorescent antibodies to defined antigens or fluorescent analogs of cellular constituents as well as ultrasensitive, electronic image detectors and video image averaging to improve signal to noise, fluorescent images of living cells can be acquired over an extended period without significant fading and loss of cell viability.

Sign in / Sign up

Export Citation Format

Share Document