Influencing the fates of CD4 T cells on the path to memory: lessons from influenza

In the face of emerging infectious diseases caused by rapidly evolving and highly virulent pathogens, such as influenza, we are challenged to develop innovative vaccine strategies that can induce lasting protection. Since CD4 T cells are needed to generate and maintain protective B-cell and CD8 T-cell immunity, and can also mediate additional protective mechanisms, vaccines should ideally elicit efficient CD4 T cell, in addition to CD8 T and B-cell responses. We outline here the process of CD4 T-cell differentiation from na?ve to effector and from effector to memory with an emphasis on how exposure to microbial products and variables in antigen presentation can impact the functional quality and heterogeneity of activation-based CD4 T-cell subsets in vitro and in vivo. We discuss the impact of different phases of antigen recognition, the inflammatory milieu, acute versus chronic antigen presentation, and the contribution of residual antigen depots on CD4 T-cell effector differentiation and the formation and maintenance of CD4 T-cell memory. We propose that novel vaccine strategies, which incorporate both microbial products and antigen targeting, may provide a flexible and long-lived memory CD4 T-cell pool.     

Fibroblast growth factor receptor-1 signaling in pancreatic islet beta-cells is modulated by the extracellular matrix

Maintenance of pancreatic beta-cell mass depends on extracellular stimuli that promote survival and proliferation. In the islet, these stimuli come from the beta-cell microenvironment and include extracellular matrix deposited by associated vascular endothelial cells. Fibroblast growth factor receptor-1 (FGFR1) has recently been implicated as a signaling pathway that is important for normal beta-cell function. We would like to understand how extracellular matrix and FGFR1 signaling interact to promote beta-cell survival and proliferation. To examine beta-cell-specific receptor responses, we created lentiviral vectors with rat insulin promoter-driven expression of Venus fluorescent protein-tagged full-length (R1betav) and kinase-deficient (KDR1betav) FGFR1. Significant FGF-1-dependent activation of ERK1/2 was observed in betaTC3 cells, dispersed beta-cells, and beta-cells in intact islets. This response was enhanced by R1betav expression and reduced by KDR1betav expression. Plating-dispersed beta-cells on collagen type IV resulted in enhanced expression of endogenous FGFR1 that was associated with sustained activation of ERK1/2. Conversely, plating cells on laminin reduced expression of FGFR1, and this reduction was associated with transient activation of ERK1/2. Addition of neutralizing antibodies to inhibit beta-cell attachment to laminin via alpha(6)-integrin increased high-affinity FGF-1-binding at the plasma membrane and resulted in sustained ERK1/2 activity similar to cells plated on collagen type IV. These data show that the FGF-stimulated beta-cell response is negatively affected by alpha(6)-integrin binding to laminin and suggest regulation associated with vascular endothelial cell remodeling.     

Involvement of sphingosine kinase in plant cell signalling

In mammalian cells sphingosine-1-phosphate (S1P) is a well-established messenger molecule that participates in a wide range of signalling pathways. The objective of the work reported here was to investigate the extent to which phosphorylated long-chain sphingoid bases, such as sphingosine-1-phosphate and phytosphingosine-1-phosphate (phytoS1P) are used in plant cell signalling. To do this, we manipulated Arabidopsis genes capable of metabolizing these messenger molecules. We show that Sphingosine kinase1 (SPHK1) encodes an enzyme that phosphorylates sphingosine, phytosphingosine and other sphingoid long-chain bases. The stomata of SPHK1-KD Arabidopsis plants were less sensitive, whereas the stomata of SPHK1-OE plants were more sensitive, than wild type to ABA. The rate of germination of SPHK1-KD was enhanced, whereas the converse was true for SPHK1-OE seed. Reducing expression of either the putative Arabidopsis S1P phosphatase (SPPASE) or the DPL1 gene, which encodes an enzyme with S1P lyase activity, individually, had no effect on guard-cell ABA signalling; however, stomatal responses to ABA in SPPASEDPL1 RNAi plants were compromised. Reducing the expression of DPL1 had no effect on germination; however, germination of SPPASE RNAi seeds was more sensitive to applied ABA. We also found evidence that expression of SPHK1 and SPPASE were coordinately regulated, and discuss how this might contribute to robustness in guard-cell signalling. In summary, our data establish SPHK1 as a component in two separate plant signalling systems, opening the possibility that phosphorylated long-chain sphingoid bases such as S1P and phytoS1P are ubiquitous messengers in plants.     

1-Benzylbenzimidazoles: the discovery of a novel series of bradykinin B(1) receptor antagonists

The design, synthesis, and structure-activity studies of a novel series of BK B(1) receptor antagonists based on a 1-benzylbenzimidazole chemotype are described. A number of compounds, for example, 38g, with excellent affinity for the cynomolgus macaque and rat bradykinin B(1) receptor were discovered.     

Discovery of small molecule benzimidazole antagonists of the chemokine receptor CXCR3

High-throughput screening identified a low molecular weight antagonist of CXCR3 displaying micromolar activity in a membrane filtration-binding assay. Systematic modification of the benzimidazole core and tethered acetophenone moiety established tractable SAR of analogs with improved physicochemical properties and sub-micromolar activity across both human and murine receptors.     

SAR profiles of spirocyclic nicotinamide derived selective HDAC1/HDAC2 inhibitors (SHI-1:2)

A potent family of spirocyclic nicotinyl aminobenzamide selective HDAC1/HDAC2 inhibitors (SHI-1:2) is profiled. The incorporation of a biaryl zinc-binding motif into a nicotinyl scaffold resulted in enhanced potency and selectivity versus HDAC3, but also imparted hERG activity. It was discovered that increasing polar surface area about the spirocycle attenuates this liability. Compound 12 induced a 4-fold increase in acetylated histone H2B in an HCT-116 xenograft model study with acute exposure, and inhibited tumor growth in a 21-day efficacy study with qd dosing.     

Environmental distribution and persistence of Quahog Parasite Unknown (QPX)

Quahog Parasite Unknown (QPX) is the cause of mass mortality events of hard clams Mercenaria mercenaria from Virginia, USA, to New Brunswick, Canada. Aquaculture areas in Massachusetts, USA, have been particularly hard hit. The parasite has been shown to be a directly infective organism, but it is unclear whether it could exist or persist outside of its clam host. We used molecular methods to examine water, sediment, seaweeds, seagrass and various invertebrates for the presence of QPX. Sites in Virginia and Massachusetts were selected based upon the incidence of QPX-induced clam die-offs, and they were monitored seasonally. QPX was detectable in almost all of our different sample types from Massachusetts, indicating that the parasite was widely distributed in the environment. Significantly more samples from Massachusetts were positive than from Virginia, and there was a seasonal pattern to the types of samples positive from Massachusetts. The data suggest that, although it may be difficult to completely eradicate QPX from the environment, it may be possible to keep the incidence of disease under control through good plot husbandry and the removal of infected and dying clams.