Exchange protein directly activated by cAMP 1 (Epac1) is expressed in human neutrophils and mediates cAMP-dependent activation of the monomeric GTPase Rap1

Epac1 and Epac2 bind cAMP and mediate cAMP-dependent activation of Rap1. cAMP is produced in neutrophils in response to many chemoattractants. This second messenger plays a key role in the regulation of the functions of neutrophils. However, it is still not known whether Epacs are expressed in human neutrophils. We found that stimulation of PLB-985 cells differentiated into neutrophil-like cells, human neutrophils with 8CPT-2Me-cAMP (a selective activator of Epacs), or FK (a diterpene that augments the intracellular level of cAMP) led to GTP-loading of Rap1. Epac1 mRNA was expressed in UND and DF PLB-985 cells, but Epac1 protein was only detected in DF PLB-985 cells. In human neutrophils, the Epac1 transcript was present, and Epac1 protein could be detected by Western blot analysis if the cells had been treated with the serine protease inhibitor PMSF. FK induced adhesion of PLB-985 cells and human neutrophils on fibrinogen, a ligand for beta 2 integrins. Interestingly, in DF PLB-985 cells, but not in human neutrophils, 8CPT-2Me-cAMP induced beta 2 integrin-dependent adhesion. The failure of 8CPT-2Me-cAMP to induce beta 2 integrin-dependent human neutrophil adhesion could be explained by the fact that this compound did not induce a switch of the beta 2 integrins from a low-affinity to a high-affinity ligand-binding conformation. We concluded that Epac1 is expressed in human neutrophils and is involved in cAMP-dependent regulation of Rap1. However, the loading of GTP on Rap1 per se is not sufficient to promote activation of beta 2 integrins. J. Leukoc. Biol. 90: 741-749; 2011.

Cell survival responses after exposure to modulated radiation fields.

In the present study survival responses were determined in cells with differing radiosensitivity, specifically primary fibroblast (AG0-1522B), human breast cancer (MDA-MB-231), human prostate cancer (DU-145) and human glioma (T98G) cells, after exposure to modulated radiation fields delivered by shielding 50% of the tissue culture flask. A significant decrease (P < 0.05) in cell survival was observed in the shielded area, outside the primary treatment field (out-of-field), that was lower than predicted when compared to uniform exposures fitted to the linear-quadratic model. Cellular radiosensitivity was demonstrated to be an important factor in the level of response for both the in- and out-of-field regions. These responses were shown to be dependent on secretion-mediated intercellular communication, because inhibition of cellular secreted factors between the in- and out-of-field regions abrogated the response. Out-of-field cell survival was shown to increase after pretreatment of cells with agents known to inhibit factors involved in mediating radiation-induced bystander signaling (aminoguanidine, DMSO or cPTIO). These data illustrate a significant decrease in survival out-of-field, dependent upon intercellular communication, in several cell lines with varying radiosensitivity after exposure to a modulated radiation field. This study provides further evidence for the importance of intercellular signaling in modulated exposures, where dose gradients are present, and may inform the refinement of established radiobiological models to facilitate the optimization of advanced radiotherapy treatment plans.

Feedback via Ca²⁺-activated ion channels modulates endothelin 1 signaling in retinal arteriolar smooth muscle

PURPOSE: To investigate the role of feedback by Ca²?-sensitive plasma-membrane ion channels in endothelin 1 (Et1) signaling in vitro and in vivo. Methods. Et1 responses were imaged from Fluo-4-loaded smooth muscle in isolated segments of rat retinal arteriole using two-dimensional (2-D) confocal laser microscopy. Vasoconstrictor responses to intravitreal injections of Et1 were recorded in the absence and presence of appropriate ion channel blockers using fluorescein angiograms imaged using a confocal scanning laser ophthalmoscope. Results. Et1 (10 nM) increased both basal [Ca²?](i) and the amplitude and frequency of Ca²?-waves in retinal arterioles. The Ca²?-activated Cl?-channel blockers DIDS and 9-anthracene carboxylic acid (9AC) blocked Et1-induced increases in wave frequency, and 9AC also inhibited the increase in amplitude. Iberiotoxin, an inhibitor of large conductance (BK) Ca²?-activated K?-channels, increased wave amplitude in the presence of Et1 but had no effect on frequency. None of these drugs affected basal [Ca²?](i). The voltage-operated Ca²?-channel inhibitor nimodipine inhibited wave frequency and amplitude and also lowered basal [Ca²?](i) in the presence of Et1. Intravitreal injection of Et1 caused retinal arteriolar vasoconstriction. This was inhibited by DIDS but not by iberiotoxin or penitrem A, another BK-channel inhibitor. Conclusions. Et1 evokes increases in the frequency of arteriolar Ca²?-waves in vitro, resulting in vasoconstriction in vivo. These responses, initiated by release of stored Ca²?, also require positive feedback via Ca²?-activated Cl?-channels and L-type Ca²?-channels.

SPECTRALLY MODULATED 2ND-HARMONIC EMISSION FROM LASER-PLASMA FILAMENTS

Spectra of forward emitted second harmonic light from laser interaction with filamentary plasmas have been experimentally studied. Rather regular modulations in the frequency domain have been observed into overall red-shifted spectra. The observed spectral features are consistent with self-phase-modulation of the intense laser light in growing filaments. A model accounts for this effect.

Pharmacy Students’ Views of Faculty Feedback on Academic Performance

Objective: To investigate students' views on and satisfaction with faculty feedback on their academic performance.

Methods: A 41-item survey instrument was developed based on a literature review relating to effective feedback. All pharmacy undergraduate students were invited via e-mail to complete the self-administered electronic questionnaire relating to their views on feedback, including faculty feedback received to date regarding their academic performance.

Results: A response rate of 61% (343/561) was obtained. Only 32.3% of students (107/331) agreed that they were satisfied with the feedback they received; dissatisfaction with examination feedback was particularly high. The provision of faculty feedback was perceived to be variable in terms of quality and quantity.

Conclusions: There are some inconsistencies relating to provision of feedback within the MPharm degree program at Queen's University Belfast. Further work is needed to close the gap between student expectations and the faculty's delivery of feedback on academic performance.

The Escherichia coli transcriptional regulator MarA directly represses transcription of purA and hdeA

The Escherichia coli MarA protein mediates a response to multiple environmental stresses through the activation or repression in vivo of a large number of chromosomal genes. Transcriptional activation for a number of these genes has been shown to occur via direct interaction of MarA with a 20-bp degenerate asymmetric "marbox" sequence. It was not known whether repression by MarA was also direct. We found that purified MarA was sufficient in vitro to repress transcription of both purA and hdeA. Transcription and electrophoretic mobility shift experiments in vitro using mutant promoters suggested that the marbox involved in the repression overlapped the -35 promoter motif and was in the "backward" orientation. This organization contrasts with that of the class II promoters activated by MarA, in which the marbox also overlaps the -35 motif but is in the "forward" orientation. We conclude that MarA, a member of the AraC/XylS family, can act directly as a repressor or an activator, depending on the position and orientation of the marbox within a promoter.

Hänsch--Couillaud locking of Mach--Zehnder interferometer for carrier removal from a phase-modulated optical spectrum

We describe and analyse the operation and stabilization of a Mach--Zehnder interferometer, which separates the carrier and the first-order sidebands of a phase-modulated laser field, and which is locked using the H\"ansch--Couillaud method. In addition to the necessary attenuation, our interferometer introduces, via total internal reflection, a significant polarization-dependent phase delay. We employ a general treatment to describe an interferometer with an object which affects the field along one path, and we examine how this phase delay affects the error signal. We discuss the requirements necessary to ensure the lock point remains unchanged when phase modulation is introduced, and we demonstrate and characterize this locking experimentally. Finally, we suggest an extension to this locking strategy using heterodyne detection.

Nonlinear Modulated Envelope Electrostatic Wavepacket Propagation in Plasmas

A brief review of the occurrence of amplitude modulated structures in space and laboratory plasmas is provided, followed by a theoretical analysis of the mechanism of carrier wave (self-) interaction, with respect to electrostatic plasma modes. A generic collisionless unmagnetized fluid model is employed. Both cold-(zero-temperature) and warm-(finite temperature) fluid descriptions are considered and compared. The weakly nonlinear oscillation regime is investigated by applying a multiple scale (reductive perturbation) technique and a Nonlinear Schrödinger Equation (NLSE) is obtained, describing the evolution of the slowly varying wave amplitude in time and space. The amplitude’s stability profile reveals the possibility of modulational instability to occur under the influence of external perturbations. The NLSE admits exact localized envelope (solitary wave) solutions of bright (pulses) or dark (holes, voids) type, whose characteristics depend on intrinsic plasma parameters. The role of perturbation obliqueness (with respect to the propagation direction), finite temperature and — possibly — defect (dust) concentration is explicitly considered. The relevance of this description with respect to known electron-ion (e-i) as well as dusty (complex) plasma modes is briefly discussed. © 2004 American Institute of Physics