New tricks from an old dog: Mitochondrial redox signaling in cellular inflammation

Reactive oxygen species (ROS) such as superoxide (O₂^?) and hydrogen peroxide (H₂O₂) have long been implicated as pro-inflammatory, yet the sources of ROS and the molecular mechanisms by which they enhance inflammation have been less clear. Recent advances in the understanding of the molecular basis of inflammation mediated by the innate immune system have allowed these issues to be revisited. Although the Nox2 NADPH oxidases generate the bulk of ROS for antimicrobial host defense, recent studies have found that NADPH oxidase-dependent ROS production can actually dampen macrophage inflammatory responses to sterile pro-inflammatory stimuli. Instead, production of mitochondrial ROS has emerged as an important factor in both host defense and sterile inflammation. Excess mitochondrial ROS can be generated by either damage to the respiratory chain or by alterations of mitochondrial function such as those that increase membrane potential and reduce respiratory electron carriers. In autoinflammatory diseases, where key components of innate immune responses are activated by genetic mutations or environmental stimuli, inflammation has been found to be particularly sensitive to inhibition of mitochondrial ROS production. These findings have highlighted mitochondrial ROS as a novel generator of pro-inflammatory ROS and a potential therapeutic target in inflammatory diseases.     

Contribution of free radicals to Pseudomonas aeruginosa induced acute pyelonephritis

Pyelonephritis induces an inflammatory process in the renal parenchyma, which may occur as a result of excessive reactive nitrogen intermediates (RNI) and reactive oxygen species (ROS) and/or impaired antioxidant capacity. In the present investigation, contribution of free radicals to the development of acute pyelonephritis induced by planktonic and biofilm cells of Pseudomonas aeruginosa was studied. Increase in production of RNI and ROS in urine, bladder and renal tissue following infection with P. aeruginosa was observed which correlated with bacterial load, neutrophil recruitment and malondialdehyde (MDA). Evaluation of the data revealed that excessive production of free radicals causes tissue damage leading to bacterial persistence in host's tissues. Treatment of mice with N-acetylcysteine (NAC), a potent antioxidant, lead to significant amelioration of oxidative stress and subsequent decrease in bacterial titer, neutrophil influx, MDA as well as tissue pathology highlighting important role of free radicals in P. aeruginosa induced pyelonephritis. Results of the present study bring out that production of RNI and ROS contributes to the pathophysiology of pyelonephritis. These findings may be relevant for the better understanding of host–parasite interactions and may be of clinical importance in the development of preventive intervention against P. aeruginosa induced pyelonephritis.     

Sialic acid residues play a pivotal role in α1-acid glycoprotein (AGP)-induced generation of reactive oxygen species in chemotactic peptide pre-activated neutrophil granulocytes

Objective  

We have recently shown that terminal sialic acid residues are essential for α₁-acid glycoprotein (AGP)-induced Ca²⁺ mobilization in neutrophils. The aim of the present study was to establish the importance of sialic acid residues on AGP in modulating human neutrophil functions, with emphasis on the generation of reactive oxygen species (ROS).     

Modulation of human neutrophil function by fibronectin degradation products isolated from cryoglobulins

The proteolytic fragments of native fibronectin (cryopeptides) complexed to pathological immunoglobulins in cryoglobulins were isolated from the serum of six different patients. Two peptides of 117,000 daltons and 70,000 daltons were purified and characterized byN-termmal amino acid sequencing. The priming effect of the mixture of these two peptides, referred to as cryopeptides, on neutrophil functions, namely the respiratory burst, exocytosis, and Ca²⁺ release, was investigated. Cryopeptides and cryoglobulins assayed separately did not increase neutrophil respiration or cytosol free calcium concentration; however, both of them induced granule enzyme secretion. Sequential exposure of neutrophils to either cryopeptides or the respective cryoglobulins, and then toN-formyl-methionyl-leucyl-phenylalanine or serum opsonized zymosan, resulted in a synergistic stimulation of O₂ uptake compared to the effect ofN-formyl-peptides or opsonized zymosan tested separately; Ca²⁺ release was significantly enhanced by the pretreatment of neutrophils with cryopeptides. These data suggest that cryopeptides and cryoglobulins may play a role in host defense against bacterial infections through neutrophil activation.     

Hydrogen peroxide enhances phagocytosis of Pseudomonas aeruginosa in hyperoxia

Mechanical ventilation with hyperoxia is a necessary treatment for patients with respiratory distress. However, patients on mechanical ventilation have increased susceptibility to infection. Studies including ours have shown that reactive oxygen species (ROS), generated by exposure to prolonged hyperoxia, can cause a decrease in the phagocytic activity of alveolar macrophages. Hydrogen peroxide (H₂O₂) is a form of ROS generated under hyperoxic conditions. In this study, we examined whether treatment with H₂O₂ directly affects macrophage phagocytic ability in RAW 264.7 cells that were exposed to either 21% O₂ (room air) or 95% O₂ (hyperoxia). Moderate concentrations (ranging from 10 to 250 µM) of H₂O₂ significantly enhanced macrophage phagocytic activity and restored hyperoxia-suppressed phagocytosis through attenuation of hyperoxia-induced disorganization of actin cytoskeleton and actin oxidation. These results indicate that H₂O₂ at low–moderate concentrations can be beneficial to host immune responses by improving macrophage phagocytic activity.     

Development of a device for chemiluminescence determination of superoxide generated inside a dialysis hollow-fiber membrane

Reactive oxygen species (ROS) generated during hemodialysis treatment cause dialysis complications because of the high reactivity of ROS. To prevent dialysis complications caused by oxidative stress, it is important to evaluate the generation and dismutation of ROS during hemodialysis treatment. In this study, our aim was to develop a device to determine superoxide (O₂ ⁻) generated inside a dialysis hollow fiber, and also to examine whether this device could detect O₂ ⁻ separated from plasma using hollow fibers. Experimental apparatus was set up so that hypoxanthine (HX) solution flowed inside the hollow fibers and 2-methyl-6-p-methoxyphenylethynyl-imidazopyrazinone (MPEC) solution flowed outside the hollow fibers. Then, xanthine oxidase (XOD) solution was added to the HX solution to generate O₂ ⁻, and chemiluminescence resulting from the reaction of O₂ ⁻ with MPEC was measured with an optical fiber. Chemiluminescence intensity was measured at different HX concentrations, and the peak area of relative luminescence intensity yielded a first-order correlation with the HX concentration. Based on the relationship between HX and O₂ ⁻ concentrations determined by the cytochrome c reduction method, the relative luminescence intensity measured by this device was linearly dependent on the O₂ ⁻ concentration inside the hollow fibers. After modifications were made to the device, XOD solution injection into plasma including HX resulted in an increase in the relative luminescence intensity. We concluded that this novel device based on chemiluminescence is capable of determining aqueous O₂ ⁻ generated inside a hollow fiber and also of detecting O₂ ⁻ in plasma.     

Iminoboronate Backbone‐Based Hyperbranched Polymeric Micelles with Fenton‐Like Enhanced ROS Response

Compared to normal cells, there is a relatively high level of reactive oxygen species (ROS) in tumor cells caused by defecting ROS scavenging systems. To this issue, developing ROS‐responsive nanocarriers has been a promising way to cancer therapy. However, it is always difficult for a certain ROS‐responsive nanocarrier to be perfectly triggered due to the complexity of cancerous tissues. Thus, it is pressing to improve the response sensitivity of ROS‐responsive nanocarriers. Herein, the Fenton‐like reaction enhanced ROS response of polymeric nanocarriers of an iminoboronate backbone‐based hyperbranched polymer with metallisable 8‐hyroxyquinoline (HQ) moieties is demonstrated. HQ‐Cu catalytic sites can be formed at the prepared nanocarriers via the complexation of Cu(II) ions and HQ moieties. Upon H₂O₂, hydroxyl radicals (·OH) are quickly generated via the Fenton‐like reaction between H₂O₂ and HQ‐Cu. Then, the oxidative cleavage of iminoboronate moieties can be effectively activated to disrupt the nanocarriers, leading to a rapid release of encapsulated drugs. It is proposed that this kind of polymeric nanocarriers with Cu‐complexed catalyzing‐triggered ROS response may achieve highly effective oxidation therapy.     

Vasoactive intestinal peptide dampens formyl-peptide-induced ROS production and inflammation by targeting a MAPK-p47phox phosphorylation pathway in monocytes

Reactive oxygen species (ROS) produced by the phagocyte NADPH oxidase (NOX2) are required for microbial clearance; however, when produced in excess they exacerbate inflammatory response and injure surrounding tissues. NOX2 is a multicomponent enzyme composed of membrane-associated cytochrome b588 and cytosolic components p47^phox, p67^phox, p40^phox, and rac1/2. We investigated whether vasoactive intestinal peptide (VIP), an endogenous immune-modulatory peptide, could affect ROS production by NOX2 in primary human phagocytes. VIP did not modulate basal ROS production by phagocytes, but it inhibited monocyte and not neutrophil ROS production in response to the bacterial peptide N-formyl-methionyl-leucyl-phenylalanine (fMLF). The action of VIP was essentially mediated by high-affinity G-protein coupled receptors VPAC1 as its specific agonist, [ALA^11,22,28]VIP, mimicked VIP-inhibitory effect, whereas the specific VPAC1 antagonist, PG97-269, blunted VIP action. Further, we showed that VIP inhibited fMLF-induced phosphorylation of ERK1/2 (extracellular signal–regulated kinase 1/2), p38MAPK (p38 mitogen-activated protein kinase) pathways, and phosphorylation of p47^phox on Ser345 residue. Also, VIP exerted an anti-inflammatory effect in a model of carrageenan-induced inflammation in rats. We thus found that VIP exerts anti-inflammatory effects by inhibiting the “MAPK–p47^phox phosphorylation–NOX2 activation” axis. These data suggest that VIP acts as a natural anti-inflammatory agent of the mucosal system and its analogs could be novel anti-inflammatory molecules.     

Intracellular free calcium concentration / force relationship in rabbit inferior vena cava activated by norepinephrine and high K+

The changes in isometric force and the underlying fluctuations in intracellular free calcium concentration ([Ca²⁺]_i) were monitored simultaneously in thin sheets of rabbit inferior vena cava loaded with the fluorescent Ca²⁺ indicator fura-2. In resting tissues bathed in physiological saline solution, the estimated [Ca²⁺]_i was approximately 105 nM. The -adrenergic agonist norepinephrine (10 M) caused an initial rise in [Ca²⁺]_i to 264 nM during force development, which dropped to 216 nM during force maintenance. The maintained norepinephrine-induced increase in force and [Ca²⁺]_i was reversed in Ca²⁺-free (2 mM EGTA) solution. Membrane depolarization by high K⁺ (80 mM) significantly increased [Ca²⁺]_i to 234 nM. Compared to norepinephrine, high K⁺ caused about the same steady-state increase in [Ca²⁺]_i, but a smaller increase in force. [Ca²⁺]_i/force curves were constructed at different concentrations of extracellular Ca²⁺, with either norepinephrine or high K⁺ as a stimulant. The curve generated with norepinephrine was located to the left of that generated with high K⁺.     

Contribution of free radicals to Pseudomonas aeruginosa induced acute pyelonephritis

Pyelonephritis induces an inflammatory process in the renal parenchyma, which may occur as a result of excessive reactive nitrogen intermediates (RNI) and reactive oxygen species (ROS) and/or impaired antioxidant capacity. In the present investigation, contribution of free radicals to the development of acute pyelonephritis induced by planktonic and biofilm cells of Pseudomonas aeruginosa was studied. Increase in production of RNI and ROS in urine, bladder and renal tissue following infection with P. aeruginosa was observed which correlated with bacterial load, neutrophil recruitment and malondialdehyde (MDA). Evaluation of the data revealed that excessive production of free radicals causes tissue damage leading to bacterial persistence in host's tissues. Treatment of mice with N-acetylcysteine (NAC), a potent antioxidant, lead to significant amelioration of oxidative stress and subsequent decrease in bacterial titer, neutrophil influx, MDA as well as tissue pathology highlighting important role of free radicals in P. aeruginosa induced pyelonephritis. Results of the present study bring out that production of RNI and ROS contributes to the pathophysiology of pyelonephritis. These findings may be relevant for the better understanding of host–parasite interactions and may be of clinical importance in the development of preventive intervention against P. aeruginosa induced pyelonephritis.     

Expression of Intracellular Reactive Oxygen Species in Hematopoietic Stem Cells Correlates with Time to Neutrophil and Platelet Engraftment in Patients Undergoing Autologous Bone Marrow Transplantation

Reactive oxygen species (ROS) play important roles in hematopoiesis and regulate the self-renewal, migration, and myeloid differentiation of hematopoietic stem cells (HSCs). This study was conducted to determine whether ROS levels in donor HSCs correlate with neutrophil and platelet engraftment in patients after bone marrow transplantation.Cryopreserved HSC samples from 51 patients who underwent autologous transplantation were studied. Levels of intracellular ROS were assessed by flow cytometry using 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFDA) in the CD45⁺/CD34⁺ HSC population. Colony forming unit assays were performed on HSCs isolated from the ROS^high and ROS^low populations to assess the differentiation potential of these 2 cell subsets.Distinct populations of ROS^high and ROS^low cells were evident in all patient samples. The median percentage of ROS^high expressing HSCs in the study cohort was 75.8% (range, 2% to 95.2%). A significant correlation was identified between the percentage of ROS^high stem cells present in the hematopoietic progenitor cells collected by apheresis product infused and the time to neutrophil engraftment (P < .001, r = –.54), as well as time to plt20, plt50, and plt100 (P < 0.001; r?=?–.55, –.59, and –.56 respectively). The dose of CD34⁺/ROS^high/kg infused also inversely correlated with a shorter time to neutrophil engraftment; time to engraftment for patients receiving > or ≤3?×?10⁶ cells/kg was 11.5 days (range, 9 to 23) versus 14 days (range, 10 to 28), respectively (P?=?.02). The dose of ROS^high HSCs delivered did not correlate with platelet engraftment.Collectively, these data suggest that the dose of ROS^high stem cells delivered to patients may predict time to neutrophil engraftment after autologous transplantation.     

Luminol-independent chemiluminescence by phagocytes is markedly enhanced by dexamethasone,not by other glucocorticosteroids

The effect of several glucocorticosteroids on the generation of reactive oxygen species (ROS) was examined. The ROS assessed were O ₂ ^– , H₂O₂, OH·, and chemiluminescence (CL) (determined in the presence or absence of luminol), generated by both opsonized zymosan-stimulated neutrophils or monocytes and by the xanthine-xanthine oxidase system. Except for luminol-independent CL, only high concentrations (10^–4 M) of steroids could decrease each ROS. In contrast, luminol-independent CL generation in the phagocyte system was increased in a dose-dependent manner by the addition of dexamethasone, but not by any other steroid. Further, in lymphocyte cultures stimulated with Con A for four days, luminol-independent CL generation was demonstrated and enhanced by the addition of dexamethasone, although CL generation was not detected in the absence of dexamethasone. These findings provide evidence that CL does not always represent light specific to ROS, and they suggest the possibility that dexamethasone induces emission of light at sites of inflammation.     

Effects of α1‐acid Glycoprotein Fucosylation on its Ca2+ Mobilizing Capacity in Neutrophils

We recently showed that the acute‐phase protein α₁‐acid glycoprotein (AGP) induces rises in cytosolic calcium concentration, [Ca²⁺]_i, in neutrophils through sialic acid dependent interactions with the neutrophil receptors siglec‐5 and/or siglec‐14. Whereas both siglec‐5 and siglec‐14 have a relatively broad specificity for sialylated oligosaccharide structures, including both structures with terminal α2–3 or α2–6 linked sialic acid, there is a markedly reduced affinity to the fucosylated epitope sialyl Lewis x (SLe^x). Increased fucosylation, leading to increased expression of SLe^x on AGP is commonly associated with inflammatory conditions. In the present study, we investigated whether an increased SLe^x expression would affect the Ca²⁺‐mobilizing effect of AGP. AGP with elevated fucose content isolated from patients with untreated chronic joint inflammation showed a decreased [Ca²⁺]_i modulatory effect on neutrophils compared to normally fucosylated AGP. Furthermore a hyperfucosylated AGP form produced by in vitro fucosylation, that consequently had an elevated expression of SLe^x, could not elicit a [Ca²⁺]_i increase in neutrophils. The role of the carbohydrate portion of AGP in modulating neutrophil responses was further strengthened by showing that synthetic glycoconjugates carrying oligosaccharides with terminal α2–3 or α2–6 linked sialic acid were able to mimic the Ca²⁺‐mobilizing effect of AGP whereas a synthetic glycoconjugate carrying SLe^x was not. Based on these data, we conclude that increased fucosylation can alter the ability of AGP to induce neutrophil signalling and further supports an important role of the oligosaccharide chains of AGP in the modulation of leukocyte functions during an inflammatory process.     

Dexamethasone inhibits apoptosis of human neutrophils induced by reactive oxygen species

Neutrophils are completely differentiated cells that die in tissues a few days after they migrate from the vascular compartment as a consequence of a rigouous apoptotic program. Many of the mediators produced during an inflammatory response delay neutrophil apoptosis allowing a more efficient removal of microorganisms but also favoring the tissue damage by reactive oxygen species (ROS) and lysosomal proteins released by neutrophils. Glucocorticoids delay the apoptosis of neutrophils but the mechanisms are not completely understood. To investigate the inhibition of glucocorticoids on neutrophil apoptosis we have used the glucose/glucose oxidase (G/GO) system as a constant source of hydrogen peroxide. When neutrophils are incubated in the presence of the G/GO system, a significant acceleration of their apoptotic response is observed. Preincubation with 10^–6 M, 10^–7 M, 10^–8 M or 10^–9 M of dexamethasone, negatively modulated the spontaneous and G/GO induced apoptosis of neutrophils. Then the G/GO system is a useful model to simulate the oxidative stress of neutrophils, and that the effect of DXM on neutrophil apoptosis depends, at least in part, on blocking the proapoptotic effect of ROS.     

NADPH oxidases and ROS signaling in the gastrointestinal tract

Reactive oxygen species (ROS), initially categorized as toxic by-products of aerobic metabolism, have often been called a double-edged sword. ROS are considered indispensable when host defense and redox signaling is concerned and a threat in inflammatory or degenerative diseases. This generalization does not take in account the diversity of oxygen metabolites being generated, their physicochemical characteristics and their production by distinct enzymes in space and time. NOX/DUOX NADPH oxidases are the only enzymes solely dedicated to ROS production and the prime ROS producer for intracellular and intercellular communication due to their widespread expression and intricate regulation. Here we discuss new insights of how NADPH oxidases act via ROS as multifaceted regulators of the intestinal barrier in homeostasis, infectious disease and intestinal inflammation. A closer look at monogenic VEOIBD and commensals as ROS source supports the view of H₂O₂ as key beneficial messenger in the barrier ecosystem.     

Rapid effects of hypoxia on H+ homeostasis in articular chondrocytes

Articular chondrocytes experience low oxygen (O₂) levels compared with many other tissues, and values fall further in disease states. Chondrocyte intracellular pH (pH_i) is a powerful modulator of matrix synthesis and is principally regulated by Na⁺-H⁺ exchange (NHE). In equine chondrocytes, NHE is inhibited when cells are incubated for 3 h at low O₂, leading to intracellular acidosis. O₂-dependent changes in reactive oxygen species (ROS) levels appear to underlie this effect. The present study examines whether hypoxia can influence chondrocyte NHE activity and pH_i over shorter timescales using the pH-sensitive fluoroprobe BCECF in cells isolated not only from equine cartilage but also from bovine tissue. O₂ levels in initially oxygenated solutions gassed with N₂ fell to approximately 1% within 2 h. A progressive fall in pH_i and acid extrusion capacity was observed, with statistically significant effects (P?<?0.05) apparent within 3 h. For equine and bovine cell populations subjected to step change in O₂ by resuspension in hypoxic (1%) solutions, a decline in acid extrusion and pH_i was observed within 10 min and continued throughout the recording period. This effect represented inhibition of the NHE-mediated fraction of acid extrusion. Cells subjected to hypoxic solutions supplemented with CoCl₂ (100 μM) or antimycin A (100 µM) to raise levels of ROS did not acidify. The conserved nature and rapidity of the response to hypoxia has considerable implications for chondrocyte homeostasis and potentially for the maintenance of cartilage integrity.     

Inhibition of the neutrophil NADPH oxidase by adenosine is associated with increased movement of flavocytochrome b between subcellular fractions

Adenosine is a potent inhibitor of reactive oxygen species (ROS) production by the NADPH oxidase in fMLF-stimulated neutrophils. Although much is known about the pharamacology and signal transduction of this effect, it is not known how adenosine affects assembly and localization of the NADPH oxidase components within the neutrophil. We report here that adenosine pretreatment of fMLF-stimulated neutrophils results in decreased plasma membrane/secretory granule content of the flavocytochrome b components (p22^phox and gp91^phox) of the NADPH oxidase, which correlates with inhibition of ROS production. Adenosine treatment did not affect upregulation of secretory and specific granule surface markers, confirming that degranulation was not impaired by adenosine. However, adenosine treatment did result in increased movement of cell-surface flavocytochrome b to heavy granule fractions in fMLF-stimulated neutrophils. These data suggest that adenosine-mediated effects on neutrophil ROS production are due, in part to endocytosis and/or redistribution of flavocytochrome b between various subcellular compartments.     

Both decorin-binding proteins A and B are critical for the overall virulence of Borrelia burgdorferi

Both decorin-binding proteins (DbpA and DbpB) of the Lyme disease spirochete Borrelia burgdorferi bind decorin and glycosaminoglycans, two important building blocks of proteoglycans that are abundantly found in the extracellular matrix (ECM) and connective tissues as well as on cell surfaces of mammals. As an extracellular pathogen, B. burgdorferi resides primarily in the ECM and connective tissues and between host cells during mammalian infection. The interactions of B. burgdorferi with these host ligands mediated by DbpA and DbpB potentially influence various aspects of infection. Here, we show that both DbpA and DbpB are critical for the overall virulence of B. burgdorferi in the murine host. Disruption of the dbpBA locus led to nearly a 10⁴-fold increase in the 50% infectious dose (ID₅₀). Complementation of the mutant with either dbpA or dbpB reduced the ID₅₀ from over 10⁴ to roughly 10³ organisms. Deletion of the dbpBA locus affected colonization in all tissues of infected mice. The lack of dbpA alone precluded the pathogen from colonizing the heart tissue, and B. burgdorferi deficient for DbpB was recovered only from 42% of the heart specimens of infected mice. Although B. burgdorferi lacking either dbpA or dbpB was consistently grown from joint specimens of almost all infected mice, it generated bacterial loads significantly lower than the control. The deficiency in either DbpA or DbpB did not reduce the bacterial load in skin, but lack of both significantly did. Taken together, the study results indicate that neither DbpA nor DbpB is essential for mammalian infection but that both are critical for the overall virulence of B. burgdorferi.     

Effects of vitamin D-induced supernatant of placental explants from preeclamptic women on oxidative stress and nitric oxide bioavailability in human umbilical vein endothelial cells

The study evaluated the effect of the supernatant of placental explants from preeclamptic (PE) and normotensive (NT) pregnant women after tissue treatment with or without vitamin D (VD) on oxidative stress and nitric oxide (NO) bioavailability in human umbilical vein endothelial cells (HUVEC). Placental explants were prepared from eight NT and eight PE women, and supernatants were obtained after incubation with or without hydrogen peroxide (H₂O₂) and/or VD. HUVEC were cultured for 24 h with supernatants, and the following parameters were analyzed in HUVEC cultures: NO, nitrate (NO₃ ^-), and nitrite (NO₂ ^-) levels, lipid peroxidation, and intracellular reactive oxygen species (ROS). Results showed that the production of NO₃ ^-, NO₂ ^-, malondialdehyde (MDA), and ROS were significantly higher in HUVEC treated with explant supernatant from PE compared to NT pregnant women, while the supernatant of PE explants treated with VD led to a decrease in these parameters. A significantly high production of NO was detected in HUVEC cultured with control supernatant of NT group, and in cultures treated with supernatant of PE explants treated with VD. Taken together, these results demonstrated that cultures of placental explants from PE women with VD treatment generated a supernatant that decreased oxidative stress and increased the bioavailability of NO in endothelial cells.     

Design of high-performance anti-adhesion agent using injectable gel with an anti-oxidative stress function

Postsurgical tissue adhesion formation caused by inflammation and oxidative stress is one of the serious issues because it induces severe clinical disorders. In this study, we designed redox injectable gel (RIG) which covalently possesses nitroxide radicals as a reactive oxygen species (ROS) scavenger for high performance anti-adhesion agent. The redox flower micelles exhibiting gelation under physiological conditions were prepared by a polyion complex (PIC) between polyamine-PEG-polyamine triblock copolymer possessing nitroxide radicals as a side chain of polyamine segments and poly(acrylic acid). RIG showed prolonged local retention in the abdominal cavity of the mice, which was monitored by in vivo imaging system (IVIS). Compared with a commercial anti-adhesion agent (Seprafilm^®, Genzyme, Cambridge, MA), RIG dramatically inhibited the formation of tissue adhesions via a combination of physical separation and biological elimination of generated ROS in talc-induced adhesion model mice. Treatment with RIG suppressed inflammatory cytokines and neutrophil invasion, suppressing the increase in peritoneal membrane thickness. It is also emphasized that RIG suppressed the increase of white blood cells level, indicating that the present RIG treatment effectively prevents diffusion of local inflammation to entire body. These findings indicate that RIG has a great potential as a high performance anti-adhesion agent.