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Effect of Oxyhemoglobin on SNAP-Mediated Chemotaxis'
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Nitric oxide (NO) has been shown to be both an intercellular and intracellular messenger. We propose here that exogenous NO induces chemotactic locomotion of human neutrophils. Indeed, when human neutrophils were placed in a gradient of a nitric oxide donor (S-nitroso-N-acetylpenicillamine; SNAP), a directed locomotion was induced, as evidenced by...
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... at a concentration which was previously shown to be optimal ( M) was performed in parallel in the same experimental conditions as a positive control (chemotaxis index: 1,170 5 32 pm; n = In order to determine whether free NO was involved in NO donor-mediated chemotaxis, we added the NO scavenger oxyhemoglobin which inhibits the effects of free NO by increasing NO oxidation ( Ignarro et al., 1993). Addition of oxyhemoglobin ( 300 @ I ) strongly inhibited the SNAP-mediated chemotaxis (Table 1). The inhibitory effect of oxyhemoglobin was observed on the 7; P < 0.001). ...Citations
... Also our metrics offer a unified approach to the empirical study of goal directedness. Measures and theoretical frameworks have been developed in various other contexts, as in the study of chemotaxis (e.g., McCutcheon 1946;Orr and Ward 1978;Beauvais et al. 1995;Lind et al. 1995;Tanaka et al. 2019;Ramirez-Gomez et al. 2019;Varennes et al. 2019), in the study of homeostasis (e.g., Gershenson and Fernandez 2012;Atkin et al. 2005), and in movement ecology (e.g., Marsh and Jones 1988;Benhamou and Bovet 1992;Turchin 1998;Benhamou 2006;Nathan et al. 2008). In these areas, goal directedness has been an implicit component of the conversation, but none of the measures have been developed expressly for studying goal directedness in general. ...
Goal directedness is one of the most commonly observed behavior patterns in biology, exemplified by systems ranging in complexity from cellular migration to human motivations. Philosophers have long tried to understand goal directedness in terms of necessary and sufficient conditions, but no consensus has been reached. Here we take an entirely novel approach to goal directedness, postponing the search for necessary and sufficient conditions, and instead trying to advance understanding by an empirical route. In particular, we introduce quantitative measures of goal directedness, applicable to systems that are generally agreed to be goal directed. The measures allow one to assess two signature properties of goal-directed systems, persistence and plasticity. Persistence is the tendency for an entity that is on a trajectory toward a goal to return to that trajectory following perturbations. Plasticity we understand as the tendency for an entity to find a trajectory toward a goal from a variety of different starting distances. We demonstrate the metrics by applying them to goal-directed behavior in two biological systems, bacteria moving up a chemoattractant gradient and a human following a heat gradient. Our approach reveals goal directedness to be an empirically tractable notion, one that makes possible a variety of comparative studies in biology, including comparing degree of goal directedness in different species, or in one species under different conditions, as well as studying evolutionary trends. More generally, the metrics make it possible to investigate the correlates and causes of goal-directed behavior. Finally, our approach challenges the conventional view of goal directedness as a discrete and unitary property, by showing that it can be treated as continuous, as a matter of degree, and that it can be broken down into at least two, and possibly more, partly independent components.
... Nitric oxide (NO) is a multifunctional signaling molecule that mediates different signaling pathways and regulates various cellular functions, including vasodilatation, neurotransmission, macrophage-mediated immunity, chemotaxis and cell migration (20)(21)(22)(23). NO has been demonstrated to regulate migration in several types of cells, including human neutrophils, endothelial cells, vascular smooth muscle cells, and T cells (24)(25)(26)(27)(28). However, the mechanism of NO in T-ALL migration remains poorly understood. ...
Stromal cell-derived factor-1 (SDF-1) regulates multiple cell signal pathways in a variety of cellular functions, including cell migration, proliferation, survival and angiogenesis. SDF-1-induced chemotaxis is an important step of lymphocyte migration. However, the molecular mechanisms that modulate SDF-1-mediated lymphocyte migration are not well identified. Nitric oxide (NO) has been found to function as a signaling molecule in a number of signaling pathways, including migration. In the present study, the potential role of NO in SDF-1-induced migration and the association between NO and the cytoskeletal changes of Jurkat cells was investigated. The present study demonstrated that Jurkat cells induced the production of NO by SDF-1 stimulation, using Griess reaction method and western blot analysis, and that NO was involved in SDF-1-induced rearrangement and polymerization of the cytoskeleton, using NOS inhibitor L-NMMA. Furthermore, NO was required for the migration of Jurkat cells. The research suggested that NO signaling pathways exerted a critical role in SDF-1-induced cytoskeleton changes and the migration of Jurkat cells. This work provides insight into the migration mechanism of acute lymphoblastic leukemia and provides an effective theoretical basis for therapy strategies for acute lymphoblastic leukemia.
... The production of NO is mediated by the inducible nitric oxide synthase (iNOS) which converts L-arginine to L-citrulline. NO is important for immune function, vasculare tone, cell proliferation and apoptosis ( Kubes et al., 1991;Beauvais et al., 1995;Papapetropoulos et al., 1996;Saeij et al., 2000;Ferret et al., 2002;Dash et al., 2003). ...
The inducible nitric oxide synthase (iNOS) plays a central role in the inflammatory reactions that follow infection or tissue damage. Induction of nitric oxide (NO) synthesis by bacterial lipopolysaccharide (LPS) depends on activation of G protein-coupled receptors in mammals. Thus, it was our intention to evaluate whether similar mechanisms are involved in iNOS activation in fish leukocytes. Therefore, the participation of membrane-bound receptors which activate effectors via G proteins has been confirmed using the G protein inhibitor suramin. Furthermore, the NO produced by iNOS performs both beneficial and detrimental actions. It is thus conceivable that regulatory mechanisms exist which control the timing and intensity of NO production by iNOS in order to outweigh protective effects against detrimental ones. The second messenger cAMP produced by adenylyl cyclases (ACs) plays a key role in the regulation of many cellular functions. Since cAMP signaling inhibits numerous immunological reactions, studies have been carried out to determine whether cAMP-dependent pathways could inhibit NO production by carp leukocytes as well. To measure cellular responses such as NO production by carp leukocytes derived from head and trunk kidneys treatments were performed with the cAMP elevating agents forskolin and dibutyryl-cAMP (db-cAMP) prior to stimulation with Aeromonas hydrophila. Pharmacological studies in stimulated kidney leukocytes showed that increased intracellular cAMP levels lead to reduced NO formation. This reduction of NO production was not due to decreased cell numbers, since a tetrazolium dye-based assay revealed no reduction of cell viability by cyclic nucleotide elevating agents. Thus, our data provide evidence that the AC/cAMP signaling pathway is well established in carp leukocytes. Cyclic AMP leads to type II immune response. We provide evidence that the predominant AC in fish leukocytes is a particulate enzyme due to its sensitivity to forskolin. Treatment of leukocytes with agents increasing intracellular cAMP gave clear evidence for participation of this cyclic nucleotide in immune signaling.
... Regulation of neutrophil function by NO is not straightforward however. In apparent conflict to the above, the NO synthase (NOS) inhibitor N G -monomethyl-L-arginine (L-NMMA) and the NO scavenger carboxy-PTIO attenuate neutrophil chemotaxis in vitro, 16 and exogenously generated NO has been reported to both induce 17 and inhibit 16 neutrophil chemotaxis, supporting both pro-and anti-inflammatory activities of this mediator. The complex-ities of the effects of NO on inflammatory responses were highlighted in a recent investigation using inducible (i)NOS-deficient mice. ...
The role of nitric oxide (NO) in regulating neutrophil migration has been investigated. Human neutrophil migration to interleukin (IL)-8 (1 nmol/L) was measured after a 1-hour incubation using a 96-well chemotaxis plate assay. The NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (L-NAME) significantly (P < 0.001) enhanced IL-8-induced migration by up to 45%. Anti-CD18 significantly (P < 0.001) inhibited both IL-8-induced and L-NAME enhanced migration. Antibodies to L-selectin or PSGL-1 had no effect on IL-8-induced migration but prevented the increased migration to IL-8 induced by L-NAME. L-NAME induced generation of neutrophil-derived microparticles that was significantly (P < 0.01) greater than untreated neutrophils or D-NAME. This microparticle formation was dependent on calpain activity and superoxide production. Only microparticles from L-NAME and not untreated or D-NAME-treated neutrophils induced a significant (P < 0.01) increase in IL-8-induced migration and transendothelial migration. Pretreatment of microparticles with antibodies to L-selectin (DREG-200) or PSGL-1 (PL-1) significantly (P < 0.001) inhibited this effect. The ability of L-NAME-induced microparticles to enhance migration was found to be dependent on the number of microparticles produced and not an increase in microparticle surface L-selectin or PSGL-1 expression. These data show that NO can modulate neutrophil migration by regulating microparticle formation.
... In our study, inhibition of allergeninduced bronchial hyperresponsiveness was coincident with inhibition of allergen-induced neutrophilia. This observation is consistent with the observation that inhibitors of nitric oxide synthase attenuate human neutrophil chemotaxis in vitro (Belenky et al., 1993), and that nitric oxide may be involved in the chemotaxis of neutrophils (Beauvais et al., 1995). It is also possible that neutrophils may be important for the expression of allergen-induced bronchial hyperresponsiveness, but the role of eosinophils is still not excluded. ...
Chronic inflammation in asthmatic airways can lead to characteristic airway smooth muscle (ASM) thickening and pathological changes within the airway wall.
We investigated the long-term effects of repeated allergen exposure.
Brown-Norway (BN) rats sensitized to ovalbumin (OVA) were exposed to OVA or saline aerosol every third day on six occasions and studied 24 h, 7 days and 35 days after the final exposure. We measured airway inflammation, ASM cell proliferation (by incorporation of bromodeoxyuridine; BrdU) and bronchial responsiveness to acetylcholine.
At 24 h, in OVA-exposed rats, we detected elevated OVA-specific serum IgE, increased numbers of macrophages, eosinophils, lymphocytes and neutrophils in the bronchoalveolar lavage (BAL) fluid and increased numbers of MBP+ (major basic protein) eosinophils and CD2+ T cells within the bronchial submucosa. This coincided with increased numbers of ASM cells expressing BrdU and with bronchial hyper-responsiveness (BHR). At 7 days, BHR was detected in OVA-exposed rats, coincident with increased numbers of macrophages and lymphocytes in BAL fluid together with increased numbers of CD2+ T cells within the bronchial submucosa. This coincided with increased numbers of ASM cells expressing BrdU. By day 35, the number of ASM cells expressing BrdU remained elevated in the absence of cellular infiltration and BHR.
Repeated OVA-challenge results in persistent ASM cell proliferation in the absence of bronchial inflammation and BHR, which lasts for at least 1 week following cessation of exposure.
... For example, NO can activate cyclooxygenases (Salvemini et al. 1993), which can increase prostaglandin production and inflammation. NO can also function as a chemoattractant to inflammatory cells (Beauvais et al. 1995). However, NO can also induce apoptosis in inflammatory cells (Albina et al. 1993;von Knethen & Brune, 1997) and inhibit diapedesis (Akimitsu et al. 1995;Fukumura et al. 1997). ...
Nitric oxide (NO) can function as either a pro-inflammatory or anti-inflammatory molecule, depending upon its concentration and the microenvironment in which it is produced. We tested whether muscle-derived NO affects muscle inflammation and membrane lysis that occur in modified muscle use. Transgenic mice with muscle-specific over-expression of neuronal NO synthase (nNOS) were generated in which transgene expression was driven by the human skeletal muscle actin promoter. Transgenic mice and non-transgenic littermates were subjected to hindlimb muscle unloading followed by reloading, which causes muscle inflammation and membrane lysis. NOS expression decreased in transgenic and non-transgenic mice during muscle unloading. Muscle inflammation was assessed by immunohistochemistry after 24 h of muscle reloading following 10 days of unloading. Soleus muscles of non-transgenic mice showed significant increases in the concentrations of neutrophils (4.8-fold) and macrophages (11.3-fold) during reloading, compared to mice that experienced unloading only. Muscles of transgenic mice showed 51 % fewer neutrophils in reloaded muscles than those of non-transgenic mice, but macrophage concentrations did not differ from non-transgenic mice. Muscle membrane damage was determined by measuring influx of an extracellular marker dye. Significantly more membrane damage occurred in muscles of non-transgenic mice experiencing reloading than in ambulatory controls. However, membrane damage in the reloaded muscles of transgenic mice did not differ from that in ambulatory mice. In vitro cytotoxicity assays confirmed that mouse neutrophils lyse muscle cell membranes, and showed that inhibition of NOS in muscle and neutrophil co-cultures significantly increased neutrophil-mediated lysis of muscle cells. Together, these data show that muscle-derived NO can function as an anti-inflammatory molecule in muscle that experiences modified loading, and that NO can prevent neutrophil-mediated damage of muscle cell membranes in vivo and in vitro.
... We have previously demonstrated how nitric oxide (NO), an endogenously produced gaseous mediator, is involved in neutrophil adhesion and migration in vivo (3). A role for NO in neutrophil migration and activation is also supported by data derived from numerous in vitro experiments (4)(5)(6)(7)(8). Carbon monoxide (CO) is another endogenously produced gas with similar physiological properties as NO. ...
... In the aspect of chemotaxis and random migration, neutrophilic cells seem to respond much in the same way when exposed to CO as to NO; an initial stimulatory effect is replaced with an inhibitory effect at higher concentrations. This similarity might further increase the validity of the comparison of these two gaseous mediators (4)(5)(6)(7)37). ...
Carbon monoxide (CO), is an endogenously produced gas, generated by the rate-limiting enzyme heme oxygenase (HO), present in man throughout the respiratory tract. CO can elicit important physiological responses like bronchial relaxation and vasodilation. Both HO expression and CO levels in the airways increase in response to hypoxic challenge and to a wide variety of inflammatory stimuli, such as intermittent allergic rhinitis, asthma and upper respiratory tract infections. A role for CO in airway regulation and inflammation has therefore been suggested. However, information about CO-induced effects on cells involved in airway inflammation is scarce. The present study was designed to investigate if the HO substrate analog hemin could affect neutrophil random migration, and N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) induced chemotaxis.
Hemin was added to and incubated with whole blood and the effects of the anticipated CO production were then evaluated on isolated neutrophils using a chemotaxis chamber.
A biphasic dose-response curve emerged for both the neutrophil spontaneous random migration and the fMLP-induced chemotaxis. Low concentrations of hemin (10(-11) m to 10(-9) m) enhanced the migratory response, whereas higher concentrations (10(-7) m and 10(-5) m) inhibited migration. The inhibition induced by hemin on fMLP-induced migration was abolished after pre-treatment with Rp-8Br-cyclicGMPS, an inhibitor of cyclicGMP.
The present data indicate that endogenously produced CO can affect both spontaneous and stimulated neutrophil migration, partly via a cyclicGMP-related process, hence strengthening the idea of a role for CO in airway inflammation.
... In our study, inhibition of allergeninduced bronchial hyperresponsiveness was coincident with inhibition of allergen-induced neutrophilia. This observation is consistent with the observation that inhibitors of nitric oxide synthase attenuate human neutrophil chemotaxis in vitro (Belenky et al., 1993), and that nitric oxide may be involved in the chemotaxis of neutrophils (Beauvais et al., 1995). It is also possible that neutrophils may be important for the expression of allergen-induced bronchial hyperresponsiveness, but the role of eosinophils is still not excluded. ...
The role of nitric oxide (NO) in allergic inflammation and bronchial hyperresponsiveness is unclear. We studied a selective prodrug nitric oxide synthase (NOS)-2 inhibitor, L-N(6)-(1-iminoethyl)lysine 5-tetrazole amide (SC-51). In ovalbumin-sensitized and challenged rats, exhaled NO levels increased by 3 h following challenge (3.73 +/- 0.74 ppb; P < 0.05), peaking at 9 h (11.0 +/- 2.75; P < 0.01) compared to saline controls (1.87 +/- 0.26; P < 0.05 and 2.81 +/- 0.18; P < 0.01). Immunoreactive lung NOS2 expression was increased in ovalbumin-challenged rats compared with ovalbumin-sensitized, saline-challenged rats at 8 h post-challenge. SC-51 (10 mg/kg; p.o.) inhibited allergen-induced increase in exhaled NO levels to 1.3 +/- 0.17 ppb. SC-51 inhibited bronchial hyperresponsiveness in ovalbumin-sensitized and challenged rats (P < 0.05). In sensitized non-exposed rats, SC-51 increased bronchial responsiveness (P < 0.05). SC-51 reduced the allergen-induced increase in bronchoalveolar lavage neutrophils, but caused a nonsignificant reduction in bronchial mucosal eosinophil numbers. NO generated through NOS2 contributes to allergen-induced bronchial hyperresponsiveness but not to bronchial eosinophilia, indicating that these are independently expressed.
... Studies suggest that NO, derived from an inducible isoform of NO synthase, may act as an endogenous modulator of the inflammatory response by inhibiting adhesion of leukocytes to endothelial cells (41)(42)(43). It is also known that exogenous NO facilitates neutrophil chemotaxis in vitro (44) and that inhibition of NO synthase decreases neutrophil migration in vitro (45,46). Furthermore, the addition of NO to the inhaled air reduces the cell surface expression of CD11/CD18 on circulating neutrophils, whereas the incubation of isolated neutrophils with L-NMMA, an inhibitor of NO synthase, increases their expression of CD11/CD18 (47,48). ...
Neutrophils are signaled to sites of infection and inflammation by different chemotactic stimuli. In order to reach the airways they have to adhere to, and then migrate through, the endothelium of pulmonary vessels. Carbon monoxide (CO) is a gaseous mediator, endogenously produced in the human airways. Increased CO production has been demonstrated during airway inflammation and CO as well as hemin, a substrate for CO producing enzymes, has been shown to affect neutrophil migration. Our objective was to investigate if the neutrophil cell surface expression of CD11b, CD66b and CD63 was changed during intermittent allergic rhinitis and to establish whether CO could affect the expression of these markers of cellular activation.
Blood from 10 healthy volunteers was drawn and incubated with different concentrations of hemin. Blood from 12 other healthy volunteers and from 12 patients with intermittent allergic rhinitis was also drawn during grass pollen season. Neutrophils were then isolated from all these three sets, and their expression of CD antigens measured using flow cytometry.
Patients with symptomatic intermittent allergic rhinitis exhibited lower levels of CD11b and CD66b on the neutrophil cell surface. Incubation with hemin decreased the expression of CD11b and CD66b. CD63 was generally weakly expressed and not significantly affected by hemin incubation.
Our results demonstrate that expressions of neutrophil cell surface glycoproteins are changed during the season in patents with intermittent allergic rhinitis and that hemin, a substrate for CO production, may act as an inhibitor of neutrophil activation. This indicates a possible role for CO in the immune defense system.
... In fact, the undegraded form of rhein, like other naturally occurring quinones, interferes with the respiratory chain (4,14) and an altered electron flow can explain the formation of RNS documented by the nitrate-accumulation experiments. Reactive nitrogen species have already been shown to mediate separately intestinal ion secretion (7), human PMN chemotaxis (16), and enterocyte apoptosis (17). Intriguingly, our data clearly indicate that rheingenerated RNS modulate all three of these biologic effects of rhein, although with different time courses. ...
Senna laxatives are used worldwide. However, their misuse can lead to chronic mucosal inflammation with the accumulation of pigment-laden leukocytes and may cause colon cells to undergo apoptosis. This study explores the mechanisms by which rhein, an active component of senna, acts on a human intestinal cell line to induce ion secretion, apoptosis, and indirect chemotaxis of polymorphonuclear leukocytes.
Human colonic adenocarcinoma (CaCo-2) monolayer cells, in the presence or in the absence of rhein, were used to monitor the production of reactive nitrogen species using the Griess reaction. Modified Ussing chambers were used to study electrolyte secretion. The capacity to recruit human polymorphonuclear leukocytes was evaluated using masked well chemotaxis chambers. Rhein-induced apoptosis was investigated by counting apoptotic nuclei stained with Hoechst 33258 dye.
Rhein caused a dose-dependent increase in short-circuit current that was abolished in chloride-free bathing buffer or by preincubating with 100 micromol/L NG-nitro-L-arginine (L-NAME) methyl ester. The concentration that maximally stimulated intestinal secretion, 50 micromol/L rhein, induced nitrate production. Supernatants obtained from CaCo-2 cultures after incubation with 50 micromol/L rhein stimulated a time-dependent polymorphonuclear leukocytes chemotaxis that was significantly decreased with 100 micromol/L L-NAME, whereas rhein per se was not active. Neutralizing antibodies anti-interleukin-8 (IL-8) and anti-ENA78 also inhibited chemotaxis. Overnight rhein incubation produced an increased number of apoptotic cells in the culture supernatant that was significantly decreased by preincubation with 100 micromol/L L-NAME. Light-degraded rhein had no effects on CaCo-2 monolayers.
The integrity of rhein is crucial to generating nitric oxide, which mediates, with different time courses, ion secretion, chemotaxis, and apoptosis of human-derived cells.
