Multiple G-protein-coupling specificity of beta-adrenoceptor in macrophages.

Adrenergic signalling of the immune system is one of the important modulator pathways of the inflammatory immune response realized via G protein-mediated pathways. The resulted signal depends on the type of the receptor-coupled G-protein (GPCR) that, according to the classical paradigm in the case of beta-adrenergic receptor (beta-AR), is Gs-type. Recently, alternate and/or multiple G protein coupling specificity of GPCRs have been demonstrated including a switch from Gs to Gi binding.

Dual beta-adrenergic modulation in the immune system: stimulus-dependent effect of isoproterenol on MAPK activation and inflammatory mediator production in macrophages.

This is the first study to demonstrate that the interaction between beta-adrenoceptor activation, and the production of inflammatory mediators can be modulated in opposite ways by two inflammatory stimuli, namely, protein kinase C (PKC)-activating phorbol myristyl acetate (PMA) and lipopolysaccharide (LPS). We provided evidence that isoproterenol treatment, when combined with phorbol ester increased the production of tumor necrosis factor-alpha, interleukin-12, and nitric oxide in murine macrophages, as well as in human monocytes and differentiated PLB-985 cells, while in agreement with earlier findings, it decreased inflammatory mediator production in combination with LPS stimulation. The contrasting effect on inflammatory mediator production, shown for the PMA and LPS activated cells was accompanied by parallel changes in activation of ERK1/2 and p38 MAPKs.

Fine-tuning of helper T cell activation and apoptosis by antigen-presenting cells.

The role of antigen-presenting cells (APC) in regulating helper T cell responses and activation-induced cell death (AICD) was investigated in vitro. T cell activation was monitored by measuring the early rise of intracellular free calcium [Ca+]ic, mRNA and cell surface expression of activation and apoptotic molecules, the production of cytokines and the activation of transcription factors. Our results demonstrate that the unique characteristics of a given APC can modify the threshold, kinetics and magnitude of the T cell response.

Calpain as a multi-site regulator of cell cycle.

Calpain has long been implicated in the regulation of cell cycle, mostly based on studies with inhibitors that lack strict specificity toward the enzyme. Further, previous work has primarily focused on one particular point, the G(1) checkpoint, and made no attempt at dissecting the full cycle in terms of calpain action. To extend and complement these findings, we tested the effect of a specific inhibitor, PD 150606, on granulocyte-macrophage-colony stimulating factor (GM-CSF)-stimulated human TF-1 cells by flow cytometry following single- and double labelling by propidium iodide and bromodeoxyuridine.

Effects of intracellular calcium on cell survival and the MAPK pathway in a human hormone-dependent leukemia cell line (TF-1).

Changes in the cytoplasmic calcium concentration ([Ca(2+)](i)) regulate a wide variety of cellular processes. Here we demonstrate that increased [Ca(2+)](i) was able to induce hormone-independent survival and proliferation, as well as to evoke apoptosis in human myelo-erythroid GM-CSF/IL-3 dependent leukemia cells (TF-1). Cellular responses induced by elevated [Ca(2+)](i) depended on the duration and amplitude of the calcium-signal.

Opposite effects of inhibitors of mitogen-activated protein kinase pathways on the egr-1 and beta-globin expression in erythropoietin-responsive murine erythroleukemia cells.

The effect of erythropoietin (Epo) on the expression of mitogen-activated protein kinase (MAPK) target genes egr-1 and c-fos was investigated in Epo-responsive murine erythroblastic cell line ELM-I-1. Epo induced a transient rise in egr-1 mRNA without a similar effect on c-fos expression. The induction of egr-1 correlated with a rapid ERK1/2 phosphorylation and was prevented with MEK1/2 inhibitors PD 98059 and UO126.

Expression and function of Toll-like receptors 2 and 4 in human keratinocytes.

Keratinocytes have the ability to kill pathogenic fungi and bacteria by producing antimicrobial substances. Recent studies suggest that microbial components use signaling molecules of the human Toll-like receptor (TLR) family to transduce signals in various cells. Here we provide evidence that keratinocytes express both TLR2 and TLR4 at the mRNA and protein levels, and show that TLR2 and TLR4 are present in the normal human epidermis in vivo and that their expression is regulated by microbial components.

Calcium induces cell survival and proliferation through the activation of the MAPK pathway in a human hormone-dependent leukemia cell line, TF-1.

Survival and proliferation of cells of a human myelo-erythroid CD34+ leukemia cell line (TF-1) depend on the presence of granulocyte-macrophage colony-stimulating factor or interleukin-3. Upon hormone withdrawal these cells stop proliferating and undergo apoptotic process. In this report we demonstrate that a controlled increase in [Ca2+]i induces hormone-independent survival and proliferation of TF-1 cells.

IL-7 withdrawal induces a stress pathway activating p38 and Jun N-terminal kinases.

IL-7 delivers survival signals to cells at an early stage in lymphoid development. In the absence of IL-7, pro-T cells undergo programmed cell death, which has previously been associated with a decline in Bcl-2 and translocation of Bax from cytosol to mitochondria. A new, earlier feature of IL-7 withdrawal was identified using an IL-7-dependent thymocyte line.

Flow cytometry used for the analysis of calcium signaling induced by antigen-specific T-cell activation.

Background: In this study, the effect of antigen-presenting cells (APC), peptide concentration, and CD28 costimulation on calcium signaling, induced by antigen-specific T-cell activation, was studied by flow cytometry.

Methods: We used two experimental approaches, which differed in their time scale and in the duration of the T cell-APC interaction, to measure the increase of intracellular free calcium levels ([Ca(2+)](i)) in activated T cells: (1) Fluo-3-loaded T cells were activated by cocentrifugation with peptide-loaded APC and the kinetics of fluorescence intensity changes was monitored continuously and (2) peptide-loaded APC and T cells were mixed, cocultured, and the fluorescence intensity was measured at various time intervals.

Results: The calcium signal of T cells was dependent on the APC as demonstrated by the ratio of cells exhibiting high versus low fluorescence intensity and by the magnitude of the calcium signal in the activated population.