Use of COX-2 inhibitors for preventing immunodeficiency

publication date: 2004-04-29; filing date: 2001-07-20The present invention provides a method of treating or preventing a disorder typified by an immunodificiency (e.g. HIV), wherein the patient is administered a COX-2 inhibitor or derivative or pharmaceutically acceptable salt thereof, preferably diisopropylfluorophasphate. L-745337, rofecoxib, NS 398, SC 58125, etodolac, meloxicam, celecoxib or nimesulide, and compositions and products containing the same or use of the same in preparing medicaments and for treatment

Remodeling of secretory lysosomes during education tunes functional potential in NK cells

Inhibitory signaling during natural killer (NK) cell education translates into increased responsiveness to activation;however, the intracellular mechanism for functional tuning by inhibitory receptors remains unclear. Secretory lysosomes are part of the acidic lysosomal compartment that mediates intracellular signalling in several cell types. Here we show that educated NK cells expressing self-MHC specific inhibitory killer cell immunoglobulin-like receptors (KIR) accumulate granzyme B in dense-core secretory lysosomes that converge close to the centrosome. This discrete morphological phenotype is independent of transcriptional programs that regulate effector function, metabolism and lysosomal biogenesis. Meanwhile, interference of signaling from acidic Ca2+ stores in primary NK cells reduces target-specific Ca2+-flux, degranulation and cytokine production. Furthermore, inhibition of PI (3,5) P-2 synthesis, or genetic silencing of the PI(3,5) P-2-regulated lysosomal Ca2+-channel TRPML1, leads to increased granzyme B and enhanced functional potential, thereby mimicking the educated state. These results indicate an intrinsic role for lysosomal remodeling in NK cell education

Defective lung macrophage function in lung cancer +/- chronic obstructive pulmonary disease (COPD/emphysema)-mediated by cancer cell production of PGE2?

In chronic obstructive pulmonary disease (COPD/emphysema) we have shown a reduced ability of lung and alveolar (AM) macrophages to phagocytose apoptotic cells (defective ‘efferocytosis’), associated with evidence of secondary cellular necrosis and a resultant inflammatory response in the airway. It is unknown whether this defect is present in cancer (no COPD) and if so, whether this results from soluble mediators produced by cancer cells. We investigated efferocytosis in AM (26 controls, 15 healthy smokers, 37 COPD, 20 COPD+ non small cell lung cancer (NSCLC) and 8 patients with NSCLC without COPD) and tumor and tumor-free lung tissue macrophages (21 NSCLC with/13 without COPD). To investigate the effects of soluble mediators produced by lung cancer cells we then treated AM or U937 macrophages with cancer cell line supernatant and assessed their efferocytosis ability. We qualitatively identified Arachidonic Acid (AA) metabolites in cancer cells by LC-ESI-MSMS, and assessed the effects of COX inhibition (using indomethacin) on efferocytosis. Decreased efferocytosis was noted in all cancer/COPD groups in all compartments. Conditioned media from cancer cell cultures decreased the efferocytosis ability of both AM and U937 macrophages with the most pronounced effects occurring with supernatant from SCLC (an aggressive lung cancer type). AA metabolites identified in cancer cells included PGE2. The inhibitory effect of PGE2 on efferocytosis, and the involvement of the COX-2 pathway were shown. Efferocytosis is decreased in COPD/emphysema and lung cancer; the latter at least partially a result of inhibition by soluble mediators produced by cancer cells that include PGE2.Francis C. Dehle, Violet R. Mukaro, Craig Jurisevic, David Moffat, Jessica Ahern, Greg Hodge, Hubertus Jersmann, Paul N. Reynolds, Sandra Hodg

Increased Membrane Cholesterol in Lymphocytes Diverts T-Cells toward an Inflammatory Response

Cell signaling for T-cell growth, differentiation, and apoptosis is initiated in the cholesterol-rich microdomains of the plasma membrane known as lipid rafts. Herein, we investigated whether enrichment of membrane cholesterol in lipid rafts affects antigen-specific CD4 T-helper cell functions. Enrichment of membrane cholesterol by 40–50% following squalene administration in mice was paralleled by an increased number of resting CD4 T helper cells in periphery. We also observed sensitization of the Th1 differentiation machinery through co-localization of IL-2Rα, IL-4Rα, and IL-12Rβ2 subunits with GM1 positive lipid rafts, and increased STAT-4 and STAT-5 phosphorylation following membrane cholesterol enrichment. Antigen stimulation or CD3/CD28 polyclonal stimulation of membrane cholesterol-enriched, resting CD4 T-cells followed a path of Th1 differentiation, which was more vigorous in the presence of increased IL-12 secretion by APCs enriched in membrane cholesterol. Enrichment of membrane cholesterol in antigen-specific, autoimmune Th1 cells fostered their organ-specific reactivity, as confirmed in an autoimmune mouse model for diabetes. However, membrane cholesterol enrichment in CD4+ Foxp3+ T-reg cells did not alter their suppressogenic function. These findings revealed a differential regulatory effect of membrane cholesterol on the function of CD4 T-cell subsets. This first suggests that membrane cholesterol could be a new therapeutic target to modulate the immune functions, and second that increased membrane cholesterol in various physiopathological conditions may bias the immune system toward an inflammatory Th1 type response

Proinflammatory and immunoregulatory roles of eicosanoids in T cells

Eicosanoids are inflammatory mediators primarily generated by hydrolysis of membrane phospholipids by phospholipase A2 to ω-3 and ω-6 C20 fatty acids that next are converted to leukotrienes (LTs), prostaglandins (PGs), prostacyclins (PCs), and thromboxanes (TXAs). The rate-limiting and tightly regulated lipoxygenases control synthesis of LTs while the equally well-controlled cyclooxygenases 1 and 2 generate prostanoids, including PGs, PCs, and TXAs. While many of the classical signs of inflammation such as redness, swelling, pain, and heat are caused by eicosanoid species with vasoactive, pyretic, and pain-inducing effects locally, some eicosanoids also regulate T cell functions. Here, we will review eicosanoid production in T cell subsets and the inflammatory and immunoregulatory functions of LTs, PGs, PCs, and TXAs in T cells

Phosphoproteomics-based characterization of prostaglandin E2 signaling in T cells

Prostaglandin E2 (PGE2) is a key lipid mediator in health and disease and serves as a crucial link between the immune response and cancer. With the advent of cancer therapies targeting PGE2 signaling pathways at different levels, there has been increased interest in mapping and understanding the complex and interconnected signaling pathways arising from the four distinct PGE2 receptors. Here, we review phosphoproteomics studies that have investigated different aspects of PGE2 signaling in T cells. These studies have elucidated PGE2’s regulatory effect on T cell receptor signaling and T cell function, the key role of protein kinase A in many PGE2 signaling pathways, the temporal regulation of PGE2 signaling, differences in PGE2 signaling between different T cell subtypes, and finally, the crosstalk between PGE2 signaling pathways elicited by the four distinct PGE2 receptors present in T cells

A Cell-Based High-Throughput Assay for Gap Junction Communication Suitable for Assessing Connexin 43-Ezrin Interaction Disruptors Using IncuCyte ZOOM

Connexin 43 (Cx43), the predominant gap junction (GJ) protein, directly interacts with the A-kinase-anchoring protein (AKAP) Ezrin in human cytotrophoblasts and a rat liver epithelial cells (IAR20). The Cx43-Ezrin–protein kinase (PKA) complex facilitates Cx43 phosphorylation by PKA, which triggers GJ opening in cytotrophoblasts and IAR20 cells and may be a general mechanism regulating GJ intercellular communication (GJIC). Considering the importance of Cx43 GJs in health and disease, they are considered potential pharmaceutical targets. The Cx43-Ezrin interaction is a protein-protein interaction that opens possibilities for targeting with peptides and small molecules. For this reason, we developed a high-throughput cell-based assay in which GJIC can be assessed and new compounds characterized. We used two pools of IAR20 cells, calcein loaded and unloaded, that were mixed and allowed to attach. Next, GJIC was monitored over time using automated imaging via the IncuCyte imager. The assay was validated using known GJ inhibitors and anchoring peptide disruptors, and we further tested new peptides that interfered with the Cx43-Ezrin binding region and reduced GJIC. Although an AlphaScreen assay can be used to screen for Cx43-Ezrin interaction inhibitors, the cell-based assay described is an ideal secondary screen for promising small-molecule hits to help identify the most potent compounds. © 2017 SAGE Publication

Spatiotemporal regulation of cAMP signaling controls the human trophoblast fusion

During human placentation, mononuclear cytotrophoblasts fuse to form multinucleated syncytia ensuring hormonal production and nutrient exchanges between the maternal and fetal circulation. Syncytial formation is essential for the maintenance of pregnancy and for fetal growth. The cAMP signaling pathway is the major route to trigger trophoblast fusion and its activation results in phosphorylation of specific intracellular target proteins, in transcription of fusogenic genes and assembly of macromolecular protein complexes constituting the fusogenic machinery at the plasma membrane. Specificity in cAMP signaling is ensured by generation of localized pools of cAMP controlled by cAMP phosphodiesterases (PDEs) and by discrete spatial and temporal activation of protein kinase A (PKA) in supramolecular signaling clusters inside the cell organized by A-kinase-anchoring proteins (AKAPs) and by organization of signal termination by protein phosphatases (PPs). Here we present original observations on the available components of the cAMP signaling pathway in the human placenta including PKA, PDE, and PP isoforms as well as AKAPs. We continue to discuss the current knowledge of the spatiotemporal regulation of cAMP signaling triggering trophoblast fusion