Priming Signal Research Articles

Learned irrelevance is disrupted in first-episode but not chronic schizophrenia patients

Learned irrelevance (LIrr) is a pre-exposure effect in which uncorrelated presentations of a conditioned stimulus (CS) and an unconditioned stimulus (US) retard subsequent CS-US association. LIrr is closely related to the phenomenon of latent inhibition (LI). LI refers to the retarding effects of inconsequential stimulus pre-exposure on subsequent conditioning to that stimulus, and is considered to reflect the organism's capacity to ignore irrelevant stimuli. LI is disrupted in schizophrenia patients, due to faster learning of the association between the preexposed CS and the US. A new within-subject target-recognition LIrr procedure was applied. The target was either cued by a priming signal or appeared at random, and priming signals were novel or preexposed cues. Schizophrenia patients were compared to age- and sex-matched control subjects. Normal subjects ( n = 24) have shown robust LIrr, namely, faster cue-target associations of novel compared to preexposed cues. Schizophrenia patients at the early stages of their first episode ( n = 7) showed LIrr disruption, namely, cue-target associations to preexposed cues were as fast as for novel cues. Chronic patients during an acute phase ( n = 18) did not show LIrr as they failed to learn the cue-target association. In addition to the LIrr paradigm the same subjects were tested in a covert-orientation task. No differences were observed between the groups on this task. The possible advantages of the new LIrr paradigm are discussed.

Tenascin‐C and SF/HGF produced by myofibroblasts in vitro provide convergent proinvasive signals to human colon cancer cells through RhoA and Rac

Myofibroblasts are present at the invasion front in colon cancer. In an attempt to understand their putative proinvasive activity, we have developed an in vitro model. Myofibroblasts isolated from colon cancer tissue or obtained through transdifferentiation of colon fibroblasts by transforming growth factor (TGF)-beta stimulate invasion of colon cancer cells into collagen type I and Matrigel. We identified two convergent proinvasive agents secreted by myofibroblasts: namely scatter factor/hepatocyte growth factor (SF/HGF) and the TGF-beta-upregulated extracellular matrix glycoprotein tenascin-C (TNC), each of which is necessary though not sufficient for invasion. Myofibroblast-stimulated invasion into collagen type I is characterized by a change from a round, nonmigratory morphotype with high RhoA and low Rac activity to an elongated, migratory morphotype with low RhoA and high Rac activity. RhoA inactivation is determined by the epidermal growth factor (EGF)-like repeats of TNC through EGF-receptor signaling that confers a permissive and priming signal for the proinvasive activity of SF/HGF that activates Rac via c-Met. We confirmed the validity of this mechanism by using pharmacological modulators and dominant negative or constitutive active mutants that interfere with RhoA-Rho kinase and Rac signaling. Our in vitro results point to a new putative proinvasive signal for colon cancer cells provided by myofibroblasts in the tumor stroma.

Cytochalasin B triggers a novel pertussis toxin sensitive pathway in TNF-alpha primed neutrophils

BackgroundCytochalasin B does not directly activate the oxygen-radical-producing NADPH oxidase activity of neutrophils but transfers desensitized G-protein coupled receptors (GPCR) into an active signaling state by uncoupling GCPR from the cytoskeleton. The receptor uncoupling results in respiratory burst activity when signals generated by reactivated formyl peptide receptors trigger the NADPH-oxidase to produce superoxide anions.ResultsTumor necrosis factor alpha (TNF-alpha) primes neutrophils for subsequent activation by cytochalasin B. Pretreatment with TNF-alpha induced mobilization of receptor-storing neutrophil organelles, suggesting that receptor up-regulation significantly contributes to the response, but the receptor mobilization was not sufficient for induction of the cytochalasin B sensitive state. The TNF-alpha primed state resembled that of the desensitized non-signaling state of agonist-occupied neutrophil formyl peptide receptors. The fact that the TNF-alpha primed, cytochalasin B-triggered activation process was pertussis toxin sensitive suggests that the activation process involves a GPCR. Based on desensitization experiments the unidentified receptor was found to be distinct from the C5a receptor as well as the formyl peptide receptor family members FPR and FPRL1. Based on the fact the occupied and desensitized receptors for interleukin-8 and platelet activating factor could not be reactivated by cytochalasin B, also these could be excluded as receptor candidates involved in the TNF-alpha primed state.ConclusionsThe TNF-alpha-induced priming signals could possibly trigger a release of an endogenous GPCR-agonist, amplifying the response to the receptor-uncoupling effect of cytochalasin B. However, no such substance could be found, suggesting that TNF-alpha can transfer G-protein coupled receptors to a signaling state independently of agonist binding.

The Application of a Minicircle Substrate in the Study of the Coordinated T4 DNA Replication

A reconstituted in vitro bacteriophage T4 DNA replication system was studied on a synthetic 70-mer minicircle substrate. This substrate was designed so that dGMP and dCMP were exclusively incorporated into the leading and the lagging strand, respectively. This design allows the simultaneous and independent measurement of the leading and lagging strand synthesis. In this paper, we report our results on the characterization of the 70-mer minicircle substrate. We show here that the minicircle substrate supports coordinated leading and lagging strand synthesis under the experimental conditions employed. The rate of the leading strand fork movement was at an average of approximately 150 nucleotides/s. This rate decreased to less than 30 nucleotides/s when the helicase was omitted from the reaction. These results suggest that both the holoenzyme and the primosome can be simultaneously assembled onto the minicircle substrate. The lagging strand synthesized on this substrate is of an average of 1.5 kb, and the length of the Okazaki fragments increased with decreasing [rNTPs]. The proper response of the Okazaki fragment size toward the change of the priming signal further indicates a functional replisome assembled on the minicircle template. The effects of various protein components on the leading and lagging strand synthesis were also studied. The collective results indicate that coordinated strand synthesis only takes place within certain protein concentration ranges. The optimal protein levels of the proteins that constitute the T4 replisome generally bracket the concentrations of the same proteins in vivo. Omission of the primase has little effect on the rate of dNMP incorporation or the rate of the fork movement on the leading strand within the first 30 s of the reaction. This inhibition only becomes significant at later times of the reaction and may be associated with the accumulation of single-stranded DNA leading to the collapse of active replisomes.

Priming of eosinophil migration across lung epithelial cell monolayers and upregulation of CD11b/CD18 are elicited by extracellular Ca2+.

In patients with asthma, eosinophils are primed and massively infiltrate lung tissues and migrate across epithelia into airways. Using blocking monoclonal antibodies, we found that eosinophil transmigration across a lung epithelial cell monolayer depended on the functions of alphaMbeta2 integrin CD11b/CD18. To study the role of Ca2+ in eosinophil priming and transepithelial migration, we treated eosinophils with eotaxin or thapsigargin (TG), reagents that increase cytoplasmic free Ca2+ concentrations by receptor- or nonreceptor-mediated mechanisms, respectively. Pretreatment of eosinophils with TG enhanced CD11b/CD18-dependent transmigration across lung epithelium. Within minutes, TG time- and dose-dependently upregulated the expression of CD11b/CD18 but did not upregulate the expression of alphaL (CD11a) or beta1 (CD29) integrin. The upregulation of CD11b/CD18 expression by eotaxin or TG was prevented when Ca2+ entry was blocked. The priming of eosinophil transmigration by TG was also abrogated by the blockade of Ca2+ entry. Our results indicate that induction of Ca2+ entry by the depletion of Ca2+ from intracellular stores upregulates CD11b/CD18 expression on eosinophils and primes eosinophil transmigration across lung epithelium. Both responses are therefore elicited by extracellular Ca2+. We suggest that, as an important priming signal for human eosinophil functional responses, store-operated Ca2+ entry may be one of the underlying mechanisms of eosinophilic inflammation in asthma.

Synergistic action of nitric oxide release from murine macrophages caused by group B streptococcal cell wall and beta-hemolysin/cytolysin.

Group B streptococcus (GBS) is the leading cause of sepsis in neonates. Nitric oxide (NO) release plays a role in the hypotension that characterizes septic shock. It has been shown that GBS beta-hemolysin/cytolysin (beta-h/c) stimulates the transcription of inducible NO synthase (iNOS) in murine macrophages via intracellular pathways similar to those that mediate lipopolysaccharide-induced iNOS activation. Here, it is demonstrated that the GBS cell wall and beta-h/c act synergistically to induce iNOS in interferon (IFN)-gamma-primed [corrected] RAW 264.7 murine macrophages. In nonprimed macrophages, combined activation by the GBS cell wall plus beta-h/c is necessary to induce an NO response, which indicates that both virulence factors cooperate to substitute for the priming signal typically provided by IFN-gamma [corrected].

Leishmania priming of human dendritic cells for CD40 ligand-induced interleukin-12p70 secretion is strain and species dependent.

A major question in the study of leishmaniasis is what dictates clinical disease expression produced by different Leishmania species, i.e., cutaneous versus systemic and healing versus nonhealing. Animal models using a Leishmania species associated with self-limiting cutaneous disease (L. major) have revealed that protective immunity requires CD40/CD40 ligand (CD40L)-dependent, interleukin-12 (IL-12)-driven Th1 responses. We recently showed that L. major can prime human dendritic cells (DCs) for CD40L-triggered IL-12p70 secretion and that these cells can drive a Th1 response in autologous T cells from sensitized individuals. Here we show that in contrast to L. major, Leishmania species responsible for visceral disease (L. donovani), as well as species associated with persistent, cutaneous lesions and occasional systemic disease (L. tropica), did not induce CD40L-dependent IL-12p70 production, despite comparable levels of uptake by DCs. Up-regulated surface expression of CD40 did not correlate with IL-12p70 production, and appreciable CD40L-induced IL-12p40 secretion was observed in uninfected as well as infected DCs, regardless of species. Reverse transcription-PCR analysis confirmed that the production of heterodimeric IL-12 was limited by expression of IL-12p35 mRNA, which was dependent on both a microbial priming signal and CD40 engagement for its high-level induction. The intrinsic differences in the ability of Leishmania species to prime DCs for CD40L-dependent IL-12p70 secretion may account, at least in part, for the evolution of healing and nonhealing forms of leishmanial disease.

Chemokine-induced leishmanicidal activity in murine macrophages via the generation of nitric oxide.

This study explored the role of the proinflammatory chemokines macrophage inflammatory protein (MIP)-1alpha and macrophage chemoattractant protein (MCP)-1 for development of antileishmanial activity. There was substantial inhibition in nitrite generation in Leishmania donovani-infected macrophages. A marked elevation of nitrite generation and induction of inducible nitric oxide (NO) synthase (iNOS) mRNA was found in chemokine-primed parasite-infected macrophages. Tumor necrosis factor-alpha, which is the priming signal for NO production, was also up-regulated under similar experimental conditions. The priming with chemokine inhibited the multiplication of L. donovani amastigotes within the intramacrophageal milieu. The antileishmanial effect of chemokines was almost completely abrogated when the macrophages were preincubated with l-N-monomethyl arginine, the specific inhibitor of iNOS. The results of this investigation suggest that the CC chemokines MIP-1alpha and MCP-1 orchestrate an antileishmanial armamentarium via the induction of an NO-mediated regulatory mechanism to control the intracellular growth and multiplication of the Leishmania protozoan.

Suppression of IL-12 production by soluble CD40 ligand: evidence for involvement of the p44/42 mitogen-activated protein kinase pathway.

IL-12 is a key cytokine in skewing immune responses toward Th1-like reactions. Human monocytes/macrophages produce high amounts of bioactive IL-12 when a priming signal (IFN-gamma or GM-CSF) precedes a second signal (e.g., LPS). We and others have previously shown that preincubation with LPS before this stimulation procedure can efficiently and selectively suppress the production of IL-12 by human monocytes. In this study, we show that an almost complete suppression of IL-12 production can also be observed after preincubation of monocytes with costimulatory cell surface molecules that bind to members of the TNFR superfamily (CD40 ligand, TNF-related activation-induced cytokine (TRANCE)). The suppression of IL-12 was observable on the mRNA and protein levels and was not due to endogenous production of known IL-12 antagonists (i.e., IL-10, IL-4, and PGE(2)), to an increased number of cells undergoing apoptosis, nor to down-regulation of the IFN-gamma or CD40 receptor. Cell surface expression of the costimulatory molecules CD80 and CD86 was not reduced by the preincubation procedure, and only a moderate reduction of IL-6 production was observed. Several studies have identified signal transduction pathways that are activated by CD40 signaling, including activation of mitogen-activated protein kinases. The presence of the extracellular signal-related kinase-specific mitogen-activated protein kinase kinase 1/2-specific inhibitors PD98059 and U0126 abrogated suppression induced by sCD40 ligand or other second signals. This indicates that activation of extracellular signal-regulated kinase 1/2 contributes to the underlying mechanism of IL-12 suppression. This mechanism may be relevant in other inflammatory responses and may help to develop therapeutic strategies in Th1-mediated diseases.

Induction of apoptotic death in cells via Bad gene expression by infectious pancreatic necrosis virus infection.

A Bcl-2 related family member, Bad, promotes cell death, and its function is regulated by phosphorylation. In this study, we show how the IPNV elicits the induction of Bad gene expression and promotes host apoptotic death. Anti-IPNV-E1S polyclonal and anti-VP3 monoclonal antibodies are used to neutralize the virus that blocks the prime death signal via the virus receptor. In the viability assay, each antibody could also enhance cell viability during IPNV infection. We tested tyrosine kinase inhibitors on IPNV-infected cells in order to assess their effect on blocking the death signal. With 100 microg/ml genistein treatment, Bad-like gene expression was blocked, either by rescuing the IPNV-infected CHSE-214 cells or by blocking internucleosomal DNA cleavage; but the tyrphostin group did not block Bad expression. For CHSE-214 cells, treatment with the protein synthesis-inhibitor, cycloheximide (1microg/ml), blocked new protein synthesis via activated tyrosine kinase during IPNV infection. We found that Bad protein expression could be blocked, and apoptotic death prevented. Together, these results demonstrate that the IPNV exerts up-regulation of a pro-apoptotic death gene (Bad), the expression of which serves to trigger apoptotic cell death. Our data also suggests that the IPNV induces apoptotic death via a viral receptor which triggers death effector Bad gene expression, possibly through a tyrosine kinase-dependent pathway.

Shedding Cytokines

Interleukin-1β (IL-1β) is a cytokine released from monocytes, the release of which requires two signals: a priming signal that stimulates the synthesis of IL-1β precursor and a second signal that triggers release and activation of the precursor through cleavage to an active form. Adenosine triphosphate (ATP) acting through purinergic P2X 7 receptors is one of two known physiological signals that trigger release of active IL-1β. IL-1β is unusual for a secreted protein in that it lacks a secretory signal and is not released by classic calcium-stimulated secretion through fusion of vesicles formed during synthesis through the secretory pathway (endoplasmic reticulum to Golgi to regulated secretory vesicle). MacKenzie et al. show that human embryonic kidney (HEK) cells transfected to express the P2X 7 receptors or monocytes, which endogenously express P2X 7 receptors, undergo rapid microvesicle shedding and reversible exposure of the inner leaflet lipid phosphatidylserine upon exposure to ATP. In monocytes primed with lipopolysaccharide to stimulate IL-1β production, treatment with ATP led to release of precursor and cleaved active IL-1β in microvesicles shed from the membranes of the cells in a process that required extracellular calcium. Microvesicles were visualized by microscopy, and membrane loss was detected by a decrease in membrane capacitance. Medium containing the IL-1β microvesicles was capable of activating IL-1 receptors in two separate assays, suggesting that the microvesicles are a physiological delivery system for this cytokine. A. MacKenzie, H. L. Wilson, E. Kiss-Toth, S. K. Dower, R. A. North, A. Surprenant, Rapid secretion of interleukin-1β by microvesicle shedding. Immunity 8 , 825-835 (2001). [Online Journal]

Translation of chloroplast psbA mRNA is regulated by signals initiated by both photosystems II and I.

Light controls the translation of several mRNAs in fully developed chloroplasts via at least two regulatory pathways. In the first, the light signal is transduced as a thiol-mediated signal that modulates translation in parallel to light intensity. The second light-controlled pathway, termed priming, is a prerequisite to the thiol-mediated regulatory pathway. Light regulation is rapid and requires intrachloroplast photoreceptor(s). To delineate the signaling pathways controlling each of these regulatory events, we assayed the effect of photosynthetic inhibitors and electron donors on the translation of chloroplastic psbA mRNA. We show that the thiol-mediated signal is generated by photosystem I and transduced by vicinal dithiol-containing proteins. We also found that the priming signal probably initiates on reduction of plastoquinone. These findings suggest that translation of chloroplast psbA mRNA is controlled by both linear photosynthetic electron transport, exerted by the reduction of the ferredoxin-thioredoxin system, and the relative activities of photosystems I and II, signaled by the redox state of the plastoquinone pool. These data underscore the function of the light-capturing reactions of photosynthesis as chloroplast photoreceptors.

Lateral Asymmetries in Attentional Priming during a Semantic and a Visuospatial Task

This study tests the hypothesis that lateral cueing of attention activates the contralateral hemisphere and also that performance on a semantic and a visuospatial task will differentially be affected depending on the visual field (right or left) to which attention was drawn. In an experimental setting, 68 subjects performed both a semantic and a visuospatial discriminating task, whereby visual priming signals unbeknownst were presented to either the left or the right visual field. For the semantic task (which is believed to engage primarily the left hemisphere) priming signals enhanced performance more when they were presented to the right visual field. For the visuospatial task, no difference was found between priming signals presented in either visual field. While there may be several alternative explanations for the observed difference in the effects of the priming signals presented to different visual fields, the importance of separating the components inherent in the tasks is stressed. While the process of decision-making may involve different relative activation of the two hemispheres, depending on whether the stimuli are semantic or visuospatial in nature, performance on the task also involves manual coordination which will depend especially on motor activation in the left hemisphere.

Bcl-xL Expression Correlates with Primary Macrophage Differentiation, Activation of Functional Competence, and Survival and Results from Synergistic Transcriptional Activation by Ets2 and PU.1

Depriving primary bone marrow-derived macrophages of colony-stimulating factor-1 (CSF-1) induces programmed cell death by apoptosis. We show that cell death is accompanied by decreases in the expression of anti-apoptotic Bcl-x(L) protein and the Ets2 and PU.1 proteins of the Ets transcription factor family. Macrophages require both priming and triggering signals independent of CSF-1 to kill neoplastic cells or microorganisms, and this activation of macrophage competence is accompanied by increased expression of bcl-x(L), ets2, and PU.1. Furthermore, we show that only Ets2 and PU.1, but not Ets1, function in a synergistic manner to transactivate the bcl-x promoter. The synergy observed between PU.1 and Ets2 is dependent on the transactivation domains of both proteins. Although other transcription factors like Fos, c-Jun, Myc, STAT3, and STAT5a are implicated in the activation of macrophage competence or in CSF-1 signaling, no synergy was observed between Ets2 and these transcription factors on the bcl-x promoter. We demonstrate that the exogenous expression of both Ets2 and PU.1 in macrophages increases the number of viable cells upon CSF-1 depletion and that Ets2 and PU.1 can functionally replace Bcl-x(L) in inhibiting Bax-induced apoptosis. Together, these results demonstrate that PU.1 and Ets2 dramatically increase bcl-x activation, which is necessary for the cytocidal function and survival of macrophages.

Searching for actions

Suppose that you are searching for a teapot in your kitchen, which is crowded with objects. Current theories of attention assume that it is the perceptual properties of the sought-for object (such as its shape, colour or texture), which are stored in long-term memory and temporarily retrieved in working memory, that guide your selection. However, in a recent report, Humphreys and Riddoch 1xDetection by action: neuropsychological evidence for action-defined templates in search. Humphreys, G.W and Riddoch, M.J. Nat. Neurosci. 2001; 4: 84–88Crossref | PubMed | Scopus (85)See all References1 suggest that the potential action an object affords might also provide an effective memory template for attentional selection. They used a visual search task to compare perceptual versus action cues (‘affordances’) in a patient, ‘MP’, with hemispatial neglect following damage to the right hemisphere. Results were clear-cut. Across the different experiments, MP was significantly faster and more accurate in detecting contralesional targets defined by the action they afforded (e.g. ‘an object you would use to pour from’) than detecting targets cued by colour (‘a brown item’) or name (‘a teapot’). This held regardless of whether the action cue was given through a verbal description or a gesture made by the experimenter. The effect disappeared, however, if words instead of objects were used as targets (words do not afford a limb action), or when objects were oriented so as to make them less suited for action (handles faced away from the patient).Interestingly though, two other patients with neglect showed the opposite pattern of results, being better at detecting targets defined by their names. This suggests that action-based and perceptual-based selections may depend on functionally separated neural mechanisms. Facilitation of visual processing by action cues could operate through top-down priming signals from a network of areas in the dorsal stream, in particular the ventral premotor cortex and inferior parietal cortex. Neurons in these areas encode the relationship between visual objects and the appropriate movements required to grasp them, rather than the perceptual properties of objects per se.This study confirms the importance of action in visual processing and the highly integrated nature of visual and motor representations. As such, it fits nicely with Duncan's integrated-competition hypothesis 2xCompetitive brain activity in visual attention. Duncan, J et al. Curr. Opin. Neurobiol. 1997; 7: 255–261Crossref | PubMed | Scopus (339)See all References2, which suggests that many different biases, both visual and non-visual, influence hemispatial neglect, and with Rizzolatti et al.'s premotor theory of attention 3xReorienting attention across the horizontal and vertical meridians: evidence in favor of a premotor theory of attention. Rizzolatti, G et al. Neuropsychologia. 1987; 25: 31–40Crossref | PubMed | Scopus (989)See all References3, which maintains that preparation to act on an object enhances perceptual processing of that object. Experiments like these bring the understanding of the relationship between perception and action a step closer.

LATERAL ASYMMETRIES IN ATTENTIONAL PRIMING DURING A SEMANTIC AND A VISUOSPATIAL TASK

This study tests the hypothesis that lateral cueing of attention activates the contralateral hemisphere and also that performance on a semantic and a visuospatial task will differentially be affected depending on the visual field (right or left) to which attention was drawn. In an experimental setting, 68 subjects performed both a semantic and a visuospatial discriminating task, whereby visual priming signals unbeknownst were presented to either the left or the right visual field. For the semantic task (which is believed to engage primarily the left hemisphere) priming signals enhanced performance more when they were presented to the right visual field. For the visuospatial task, no difference was found between priming signals presented in either visual field. While there may be several alternative explanations for the observed difference in the effects of the priming signals presented to different visual fields, the importance of separating the components inherent in the tasks is stressed. While the process of decision-making may involve different relative activation of the two hemispheres, depending on whether the stimuli are semantic or visuospatial in nature, performance on the task also involves manual coordination which will depend especially on motor activation in the left hemisphere.

Macrophage activation by antiviral acyclic nucleoside phosphonates in dependence on priming immune stimuli

Acyclic nucleoside phosphonates (ANPs) are potent broad-spectrum antivirals, also effective against immunodeficiency viruses and hepatitis viruses. Effects of several ANPs on in vitro cytokine gene expression and nitric oxide (NO) production by murine peritoneal macrophages were investigated. Included in the study were 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA; Adefovir), 9-( R)-[2-(phosphonomethoxy)propyl]adenine [( R)-PMPA; Tenofovir], 9-( S)-[2-(phosphonomethoxy)propyl]adenine; ( S)-PMPA), 9-[2-(phosphonomethoxy)ethyl]-2,6-diaminopurine (PMEDAP), 9-( R)-[2-(phosphonomethoxy)propyl]-2,6-diaminopurine (PMPDAP), and 9-[2-(phosphonomethoxy)ethyl]guanine (PMEG). Some of them, i.e. ( R)-PMPA, ( S)-PMPA, and PMEG, stimulate secretion of TNF-α and IL-10 in a concentration-dependent manner, and enhance the IFN-γ-induced secretion of TNF-α. Although unable to activate production of nitric oxide (NO) on their own, these compounds substantially augment NO formation induced by IFN-γ. Analysis of the expression of inducible NO synthase mRNA indicates that the NO-enhancing effect of ANPs is mediated posttranscriptionally. In contrast to IFN-γ, production of NO triggered by lipopolysaccharide (LPS) alone, or synergistically by LPS+IFN-γ, remains unaltered by ANPs. The immunomodulatory effects have been differentially expressed in distinct genotypes of inbred strains of mice, including the low NO-responders Balb/c and high NO-responders C3H/HeN. Although less effectively, PMEG and ( R)-PMPA also increase production of TNF-α and NO by the IFN-γ- but not LPS-co-stimulated macrophages from C3H/HeJ mice, which are otherwise hypo-responsive to major immune stimuli provided by IFN-γ and LPS. It can be concluded that the expression of immunomodulatory properties of ANPs depends on the immune state of cells and its activation by distinct priming signals.

CD40 Triggering of Heterodimeric IL-12 p70 Production by Dendritic Cells In Vivo Requires a Microbial Priming Signal

CD40 ligation triggers IL-12 production by dendritic cells (DC) in vitro. Here, we demonstrate that CD40 cross-linking alone is not sufficient to induce IL-12 production by DC in vivo. Indeed, resting DC make neither the IL-12 p35 nor IL-12 p40 subunits and express only low levels of CD40. Nevertheless, after DC activation by microbial stimuli that primarily upregulate IL-12 p40 and augment CD40 expression, CD40 ligation induces a significant increase in IL-12 p35 and IL-12 p70 heterodimer production. Similarly, IL-12 p70 is produced during T cell activation in the presence but not in the absence of microbial stimuli. Thus, production of bioactive IL-12 by DC can be amplified by T cell–derived signals but must be initiated by innate signals.

Terminal steps in JH biosynthesis in the honey bee ( Apis mellifera L.): developmental changes in sensitivity to JH precursor and allatotropin

Juvenile hormone (JH) is considered the prime endogenous signal for the induction of queen development in honey bees ( Apis mellifera L.). At the beginning of the last (5th) larval stadium, worker corpora allata synthesize less JH than queen corpora allata as a consequence of a limited production of JH precursors and a caste- and stage-specific block of the terminal step in JH biosynthesis. As previously shown, the Manduca sexta allatotropin stimulates JH biosynthesis in honey bee corpora allata in a dose-dependent and reversible manner, but can not overcome the stage-specific block in the terminal step of JH biosynthesis that is typical for worker early 5th instars. In experiments with M. sexta allatotropin and with the JH precursor farnesoic acid, we found characteristic stage-specific differences in their effects on JH biosynthesis. From the end of the spinning stage on, corpora allata could be stimulated by farnesoic acid to a much higher extent than in earlier developmental stages, suggesting a sudden increase in epoxidase activity. Manduca sexta allatotropin, however, stimulated corpora allata activity until the end of the spinning stage, at which time the corpora allata become suddenly insensitive. These data suggest that in worker larvae, important changes in the regulation of the terminal enzymatic steps in JH biosynthesis occur at the transition from the spinning stage to the prepupal stage. However, the analysis of in vitro activities of the involved enzymes, O-methyltransferase and methyl farnesoate epoxidase, remained inconclusive.

Ionizing radiation potentiates the induction of nitric oxide synthase by interferon-gamma and/or lipopolysaccharide in murine macrophage cell lines. Role of tumor necrosis factor-alpha.

Macrophages respond to infection or injury by changing from a "resting" cellular phenotype to an "activated" state defined by the expression of various cytotoxic effector functions. Regulation of the transition from a resting to an activated state is effected by cytokine and/or pathogenic signals. Some signals do not directly induce activation, but instead "prime" the macrophage to respond more vigorously to a second signal. One example of this priming phenomenon involves induction of nitric oxide (NO) synthesis by the enzyme nitric oxide synthase (NOS2). Our experiments indicate that low doses (1-5 Gy) of ionizing radiation can enhance the induction of enzymatically active NOS2 by IFN-gamma or LPS in J774.1 and RAW264.7 murine macrophage cell lines. Radiation alone did not produce this induction, rather, it was effective as a priming signal; cells exposed to radiation produced more NO when a second signal, either IFN-gamma or LPS, was applied 24 h later.