ABCA1 is believed to affect macrophage inflammatory responses, but the mechanism by which ABCA1 may impact cytokine secretion in macrophages has yet to be fully defined. We observed that the induction of ABCA1 expression in three different cell lines, namely BHK, RAW 264.7 macrophages, and primary bone marrow derived macrophages (BMDMs), results in a significant increase in phosphorylated CREB, a known protein kinase A (PKA) substrate. In RAW macrophages, induction of ABCA1 expression by the LXR-agonist T0901317 is correlated with a decrease in LPS-stimulated secretion of proinflammatory cytokines IL-6 and TNF-α. Additionally, the secretion of anti-inflammatory cytokine IL-10 was increased upon ABCA1 induction. A similar trend was observed in BMDMS: ABCA1-expressing BMDMs released less TNF-α and more IL-10 compared to ABCA1-knockout BMDMs. We speculated that the inflammation modulating effects of ABCA1 in macrophages could be a result of PKA activation. Indeed, we found that the LXR-induced ABCA1 phenotype can be mimicked by cAMP in macrophages. 8-bromo-cAMP, a PKA activator, dose-dependently suppressed inflammatory cytokine secretion while promoting IL-10 release in the absence of ABCA1 expression. Finally, we found that the T0901317-induced ABCA1 expression is correlated with higher expression levels of MKP-1, a downstream target of PKA known to suppress inflammatory responses. Together, our results suggest that ABCA1 expression may activate PKA and CREB and that such activation may contribute to the inflammatory modulating effects of ABCA1.

ABCA1 is believed to affect macrophage inflammatory responses, but the mechanism by which ABCA1 may impact cytokine secretion in macrophages has yet to be fully defined. We observed that the induction of ABCA1 expression in three different cell lines, namely BHK, RAW 264.7 macrophages, and primary bone marrow derived macrophages (BMDMs), results in a significant increase in phosphorylated CREB, a known protein kinase A (PKA) substrate. In RAW macrophages, induction of ABCA1 expression by the LXR-agonist T0901317 is correlated with a decrease in LPS-stimulated secretion of proinflammatory cytokines IL-6 and TNF-α. Additionally, the secretion of anti-inflammatory cytokine IL-10 was increased upon ABCA1 induction. A similar trend was observed in BMDMS: ABCA1-expressing BMDMs released less TNF-α and more IL-10 compared to ABCA1-knockout BMDMs. We speculated that the inflammation modulating effects of ABCA1 in macrophages could be a result of PKA activation. Indeed, we found that the LXR-induced ABCA1 phenotype can be mimicked by cAMP in macrophages. 8-bromo-cAMP, a PKA activator, dose-dependently suppressed inflammatory cytokine secretion while promoting IL-10 release in the absence of ABCA1 expression. Finally, we found that the T0901317-induced ABCA1 expression is correlated with higher expression levels of MKP-1, a downstream target of PKA known to suppress inflammatory responses. Together, our results suggest that ABCA1 expression may activate PKA and CREB and that such activation may contribute to the inflammatory modulating effects of ABCA1.

Regensburg, Univ., Diss., 2008.

No description available.

ABCA1 is believed to affect macrophage inflammatory responses, but the mechanism by which ABCA1 may impact cytokine secretion in macrophages has yet to be fully defined. We observed that the induction of ABCA1 expression in three different cell lines, namely BHK, RAW 264.7 macrophages, and primary bone marrow derived macrophages (BMDMs), results in a significant increase in phosphorylated CREB, a known protein kinase A (PKA) substrate. In RAW macrophages, induction of ABCA1 expression by the LXR-agonist T0901317 is correlated with a decrease in LPS-stimulated secretion of proinflammatory cytokines IL-6 and TNF-α. Additionally, the secretion of anti-inflammatory cytokine IL-10 was increased upon ABCA1 induction. A similar trend was observed in BMDMS: ABCA1-expressing BMDMs released less TNF-α and more IL-10 compared to ABCA1-knockout BMDMs. We speculated that the inflammation modulating effects of ABCA1 in macrophages could be a result of PKA activation. Indeed, we found that the LXR-induced ABCA1 phenotype can be mimicked by cAMP in macrophages. 8-bromo-cAMP, a PKA activator, dose-dependently suppressed inflammatory cytokine secretion while promoting IL-10 release in the absence of ABCA1 expression. Finally, we found that the T0901317-induced ABCA1 expression is correlated with higher expression levels of MKP-1, a downstream target of PKA known to suppress inflammatory responses. Together, our results suggest that ABCA1 expression may activate PKA and CREB and that such activation may contribute to the inflammatory modulating effects of ABCA1.

Cells process a wide range of signals by means of multi-component receptors that span the plasma membrane. Our knowledge about the individual proteins involved in these signaling cascades has grown considerably over recent years. However, critical information about the detailed mechanisms of receptor activation, and the quantitative relationships between stimulus and biological response, is still missing. Here, I used the RET receptor tyrosine kinase (RTK), together with its glycosylphosphatidylinositol-coupled co-receptor GFRα3 and their activating growth factor artemin (ART), as a model system to investigate the quantitative and mechanistic features of receptor activation and signaling. I used a set of anti-RET agonist antibodies to induce different extents of receptor clustering on the cell surface, and studied how this factor affects the amplitude and kinetics of membrane-proximal and downstream signaling events, as well as the biological response of neurite outgrowth. Using simulations of the RET-GFRα3-ART system, I studied the effect of co-receptor involvement in the activation mechanism, as well as the importance of the specific activation pathway for the RET system’s response to variations in the expression levels of different components. The principal findings of my work include the following: 1) Higher order receptor clustering is required for full RET activation, as well as for the biological response of neurite outgrowth. 2) The activated forms of the receptor brought about by the agonist antibodies and by ART plus GFRα3 are identical with respect to the ability to activate the transient extracellular signal-regulated kinase (ERK) and Akt responses, but the antibodies show a reduced ability to induce sustained activation of ERK, Akt or c-Jun N-terminal kinase (JNK). 3) The involvement of GFRα3 co-receptor in the activation mechanism of RET provides cells with the ability to regulate their sensitivity to ligand without affecting the maximum amplitude of the pRET response. 4) This effect is limited if the co-receptor GFRα3 is pre-dimerized. Overall, my work aims to elucidate broad principles that underlie the quantitative relationships between RET activation, signaling, and the resulting cellular functional response, that can be applied to other receptor systems.

No description available.

No description available.

No description available.

Ballistic movements have been shown to be controlled differently by the central nervous system than slow, ramp actions. It has been suggested that the cerebellum is involved primarily with ballistic actions, while the basal ganglia primarily control slower movements. These command and control differences have been shown to manifest in unique ways at the neuromuscular level. Ballistic actions evidence high firing rates, brief contraction times, and high rates of force development. A characteristic triphasic agonistantagonist-agonist burst pattern presents itself during ballistic movement, wherein the amount and intensity of antagonist co-activation is variable. In conditions of low-grade tonic muscular activity, a premovement depression (P:MD; or silent period, PMS) can occur in both agonist and antagonist muscles prior to ballistic contraction. The agonist P:MD period may serve to potentiate the force and velocity of the following contraction A selective activation of fast twitch motor units may occur in ballistic contractions under certain movement conditions. Finally, high velocity, ballistic training induces specific neuromuscular adaptations that are representative of the underlying neurophysiological mechanisms that sub serve ballistic movement. / Thesis / Master of Science (MS)