Peroxide Release Research Articles

Cytosolic DNA‒Mediated STING-Dependent Inflammation Contributes to the Progression of Psoriasis

Psoriasis is a chronic inflammatory skin disease characterized by an active dynamic interplay between immune cells and keratinocytes (KCs). STING is a universal receptor that recognizes cytosolic DNA and triggers innate immune activation. This study aims to elucidate the role of STING in the inflammation in psoriasis. STING deficiency alleviated psoriatic symptoms and inflammation in mouse models of psoriasis. Stimulation of macrophages with double-stranded DNA induced STING-dependent release of TNF-α and hydrogen peroxide invitro. Furthermore, incubation of KCs with TNF-α or hydrogen peroxide increased oxidative DNA damage, induced nuclear DNA release into the cytosol, and inhibited double-stranded DNA‒induced degradation of STING protein. More importantly, transfection of KCs with double-stranded DNA synergized with TNF-α or hydrogen peroxide to induce STING-dependent activation of NF-κB and subsequent expression of Il1b, Ccl20, and Cxcl10. Finally, exposure to 5,6-dimethylxanthenone-4-acetic acid (a STING agonist) aggravated psoriatic symptoms and inflammation in wild-type mice but not in STING-deficient mice. Collectively, STING functioned as a self-DNA sensor in macrophages and KCs of psoriatic skin. Cytosolic DNA-induced activation of STING in immune cells and KCs acted synergistically and contributed to the inflammation in psoriasis.

Subtropical Freshwater Cyanobacterial Blooms as Hydrogen Peroxide Hot Spots

In aquatic environments, chemical and biological processes represent major sources of reactive oxygen species (ROS) such as hydrogen peroxide. Diurnal patterns of hydrogen peroxide in surface water may suggest photochemical processes affect ROS dynamics. We hypothesized that thin surface hydrogen peroxide maxima could be detected through millimeter-scale depth profiling. These maxima were previously overlooked because they cannot be detected using standard surface water sampling techniques. Combined field and laboratory experiments demonstrated that cyanobacterial blooms could create extremely high concentrations of hydrogen peroxide in a thin layer of surface water (5–60 mm). Hydrogen peroxide concentrations were higher within surface scums of Microcystis aeruginosa than in areas without scum. The cyanobacterial scum increased the temperature of the ambient water, and hydrogen peroxide concentrations were higher at locations exposed to sunlight that at locations in the shade. We found that scum samples directly collected from the field could release hydrogen peroxide to the surrounding water when the biomass was exposed by the light, while 20 laboratory cultures tested did not. The production of hydrogen peroxide from cyanobacteria may depend on the physiological condition of cells. Overall, we identified thin surface water layers during subtropical freshwater cyanobacterial blooms as hydrogen peroxide hot spots.

Development of bifunctional vaginal suppositories by joint use terconazole and probiotic for treatment and prophylaxis of vulvovaginal candidiasis

Objective The objective of this article is the creation and investigation of bifunctional suppositories that are endowed with antifungal and probiotic properties intended for the treatment of vulvovaginal candidiasis (VVC) and colonization of the vaginal cavity by powerful lactobacilli. Materials and methods Freeze-dried Lactobacillus delbrueckii MH10 was used as a probiotic, which is an active producer of H2O2 isolated from a healthy woman’s vagina. Hydrophilic PEG 4000/400, amphiphilic Suppocire AP, and lipophilic Novata ABPH bases were used for the preparation of vaginal suppositories and each included 80 mg terconazole and 25 mg (∼109 CFU) lactobacilli. The release kinetic of active agents from suppositories was studied using the British Pharmacopeia’s basket dissolution method, and the physicochemical properties were studied using known methods. Bioadhesion of suppositories was evaluated by flow rate from polyacrylamide substrate inclined at 60°. Antagonism of lactobacillus against fungi was studied using the disk diffusion method and joint cultivation in simulated vaginal fluid. Results The release rate of active ingredients from the suppositories was PEG 4000/400 > Suppocire AP > Novata ABPH in decreasing order. Suppositories made of Suppocire AP possessed higher adhesion ability to artificial mucosa and longer shelf life. It was revealed that during joint cultivation L. delbrueckii MH10 kills more than 90% of C. albicans population. Conclusions Due to its physicochemical, biopharmaceutical, and bioadhesive properties, the base Suppocire AP is preferable for manufacturing vaginal bifunctional suppositories. Lactobacilli kill C. albicans by coaggregating and direct releasing hydrogen peroxide onto target cells.

Use of non-steroidal anti-inflammatory drugs in the treatment of bacterial respiratory tract infections of lambs

Respiratory tract infections, which are common in sheep farming, involve interactions between the host's immune system, pathogenic microorganisms such as viruses, bacteria, parasites and environmental conditions. Respiratory diseases can affect sheep of any age and breed. However, acute bacterial and viral pneumonia is most common in those aged three to twelve months due to decreased maternal antibody titers. Although lower respiratory tract infections initially started as a condition caused by a single agent, many cases progress to a complex disease involving both bacterial and viral components, making the diagnosis of the disease complicated by the emergence of secondary infections. Although the inflammatory reaction that occurs in respiratory infections is part of the normal healing process, an extreme reaction can impair healing and severely damage the airways. It is optimal to limit the inflammatory reaction by removing the harmful agent, isolating the damaged tissue, and controlling this tissue. In cases of death due to acute pneumonia, antibiotic drugs are used to prevent further deaths and to treat the herd. It has been found that NSAIDs reduce the release of toxic peroxide which causes the proliferation of fibroblasts and the deposition of collagen in inflammatory processes associated with the respiratory tract. It had been reported that clinical findings improve more rapidly when NSAIDs (non-steroidal anti-inflammatory drugs) were used together with antibiotics in respiratory system infections. In this review, the importance of using NSAIDs in the respiratory tract infections of sheep has been mentioned.

Confined Electrochemiluminescence at Microtube Electrode Ensembles for Local Sensing of Single Cells†

Main observation and conclusionChemical analysis of single cells plays essential roles in both fundamental research and clinical applications. In this work, the electrochemiluminescence (ECL) generated at vertically aligned gold microtube electrode ensembles (MEEs) was applied for the local chemical sensing of single cells. Thanks to the structure of MEEs and the surface‐confined nature of ECL generation, the ECL is confined in single microtubes and the local variations of hydrogen peroxide concentration can be revealed by the changes of ECL signals at single MEEs. With this methodology, we studied the hydrogen peroxide efflux from single living cells, showing the subcellular heterogeneity in hydrogen peroxide release. With sufficient spatial resolution for the parallel single cell sensing as well as advantages including easy preparation and high throughput, the MEEs provide a promising platform for subcellular electroanalysis.

Mixtures of natural antimicrobials can reduce Campylobacter jejuni, Salmonella enterica and Clostridium perfringens infections and cellular inflammatory response in MDCK cells

BackgroundThe classification of natural antimicrobials as potential antibiotic replacements is still hampered by the absence of clear biological mechanisms behind their mode of action. This study investigated the mechanisms underlying the anti-bacterial effect of a mixture of natural antimicrobials (maltodextrin, citric acid, sodium citrate, malic acid, citrus extract and olive extract) against Campylobacter jejuni RC039, Salmonella enterica SE 10/72 and Clostridium perfringens ATCC® 13124 invasion of Madin–Darby Canine Kidney cells (MDCK).ResultsMinimum sub-inhibitory concentrations were determined for Campylobacter jejuni (0.25%), Salmonella enterica (0.50%) and Clostridium perfringens (0.50%) required for the in vitro infection assays with MDCK cells. The antimicrobial mixture significantly reduced the virulence of all three pathogens towards MDCK cells and restored the integrity of cellular tight junctions through increased transepithelial resistance (TEER) and higher expression levels of ZO-1 (zonula occludens 1) and occludin. This study also identified the ERK (external regulated kinase) signalling pathway as a key mechanism in blocking the pro-inflammatory cytokine production (IL-1β, IL-6, IL-8, TNF-α) in infected cells. The reduction in hydrogen peroxide (H2O2) production and release by infected MDCK cells, in the presence of the antimicrobial mixture, was also associated with less tetrathionate formed by oxidation of thiosulphate (p < 0.0001).ConclusionThe present study describes for the first time that mixtures of natural antimicrobials can prevent the formation of substrates used by bacterial pathogens to grow and survive in anaerobic environments (e.g. tetrathionate). Moreover, we provide further insights into pathogen invasion mechanisms through restoration of cellular structures and describe their ability to block the ERK–MAPK kinase pathway responsible for inflammatory cytokine release

Mutant U2AF1-induced differential alternative splicing causes an oxidative stress in bone marrow stromal cells.

Alternative splicing (AS) is a critical regulatory process of gene expression. In bone marrow microenvironment, AS plays a critical role in mesenchymal stem cells fate determination by forming distinct isoforms of important regulators. As a spliceosome factor, U2AF1 is essential for the catalysis of pre-mRNA splicing, and its mutation can cause differential AS events. In the present study, by forced expression of mutant U2AF1 (U2AF1S34F) in the mouse bone marrow stroma OP9 cells, we determine AS changes in U2AF1S34F transduced OP9 cells and investigate their role in stroma cell biological functions. We find that abundant differential RNA splicing events are induced by U2AF1S34F in OP9 cells. U2AF1S34F causes increased generation of hydrogen peroxide, promotes production of cytokines and chemokines. U2AF1S34F transduced OP9 cells also exhibit dysfunction of mitochondria. RNA-seq data, gene ontology (GO), and gene set enrichment analysis reveal that differentially expressed genes downregulated in response to U2AF1S34F are enriched in peroxisome component and function. U2AF1S34F can also cause release of hydrogen peroxide from OP9 cells. Furthermore, we investigate the influence of U2AF1S34F-induced oxidative stress in stromal cells on hematopoietic cells. When co-culturing mouse bone marrow mononuclear cells with OP9 cells, the U2AF1S34F expressing OP9 cells induce phosphorylation of histone H2AX in hematopoietic cells. Collectively, our results reveal that mutant U2AF1-induced differential AS events cause oxidative stress in bone marrow stromal cells and can further lead to DNA damage and genomic instability in hematopoietic cells.

Active Site Hydrogen Bonding Induced in Cytochrome P450cam by Effector Putidaredoxin.

Cytochrome P450s are diverse and powerful catalysts that can activate molecular oxygen to oxidize a wide variety of substrates. Catalysis relies on effective uptake of two electrons and two protons. For cytochrome P450cam, an archetypal member of the superfamily, the second electron must be supplied by the redox partner putidaredoxin (Pdx). Pdx also plays an effector role beyond electron transfer, but after decades the mechanism remains under investigation. We applied infrared spectroscopy to heme-ligated CN- to examine the influence of Pdx binding. The results indicate that Pdx induces the population of a conformation wherein the CN- ligand forms a strong hydrogen bond to a solvent water molecule, experimentally corroborating the formation of a proposed proton delivery network. Further, characterization of T252A P450cam implicates the side chain of Thr252 in regulating the population equilibrium of hydrogen-bonded states within the P450cam/Pdx complex, which could underlie its role in directing activated oxygen toward product formation and preventing reaction uncoupling through peroxide release.

ON THE KINETICS OF ANODIC PROCESSES AT OXIDATION OF AQUEOUS SOLUTIONS OF DIMETHYL SULFOXIDE

Dimethyl sulfoxide is a feedstock for a large number of organic substances syntheses. Nowadays research is considerably focused on the production of general products of dimethyl sulfoxide oxidation – dimethyl sulfone and methane sulfonic acid. Dimethyl sulfone is well–known as a food supplement for the treating and strengthening of human joints and ligaments. dimethyl sulfone is basically synthesized by oxidation of dimethyl sulfoxide in hot 30 % hydrogen peroxide in glacial acetic acid. Synthesis is accompanied by significant losses of hydrogen peroxide, the target product has to be significantly purified. It becomes possible to control the synthesis of pure dimethyl sulfone and methane sulfonic acid when using the electrochemical method of oxidation of dimethyl sulfoxide in its aqueous solution with chemically resistant anode and high overvoltage of oxygen reaction Controlled synthesis is relevant because sulfur tends to change the oxidation rate. Study of kinetics of anodic processes at platinum electrode was performed in the dimethyl sulfoxide concentration range about 1.0…4.0 mol∙dm–3. Current raise was observed at potentials that are more positive than 1.3…1.4 V. This potential range corresponds to oxygen release. Dissolved sulfuric acid (0.2 mol∙dm–3) was added in order to inhibit the oxygen release and achieve the potential for the formation of peroxide radicals in aqueous solutions of dimethyl sulfoxide. It is known that sulfate ions are adsorbed on the surface of the platinum anode, displacing molecules of protonated water. This allows to shift the potentials and increase of the electrolysis current in 0.2 mol∙dm–3 H2SO4 to 1.7…1.9 V. It indicates the processes of formation of peroxide radicals on the surface of the platinum anode. Further shift of the anode potential into more positive area than 2.00…2.05 V leads to a rapid increase in current density. At such potentials, dimethyl sulfoxide and dimethyl sulfone are oxidized to methane sulfonic acid with a parallel oxygen and hydrogen peroxide release. Current–voltage study has shown that the oxidation of dimethyl sulfoxide in aqueous solutions runs through the formation of dimethyl sulfone. When conducting electrochemical synthesis with control of the anode potential, it is possible to produce dimethyl sulfone without further oxidation to methane sulfonic acid. The addition of 0.2 mol∙dm–3 H2SO4 to aqueous dimethyl sulfoxide solutions inhibits oxygen release and intensifies oxidation of dipole dimethyl sulfoxide molecules adsorbed on the platinum surface. The influence of adsorption processes on the kinetics of anode processes at the platinum anode in aqueous solutions of dimethyl sulfoxide at high anode potentials has been studied.

DUOX2 variants associate with preclinical disturbances in microbiota-immune homeostasis and increased inflammatory bowel disease risk.

A primordial gut-epithelial innate defense response is the release of hydrogen peroxide by dual NADPH oxidase (DUOX). In inflammatory bowel disease (IBD), a condition characterized by an imbalanced gut microbiota-immune homeostasis, DUOX2 isoenzyme is the highest induced gene. Performing multiomic analyses using 2872 human participants of a wellness program, we detected a substantial burden of rare protein-altering DUOX2 gene variants of unknown physiologic significance. We identified a significant association between these rare loss-of-function variants and increased plasma levels of interleukin-17C, which is induced also in mucosal biopsies of patients with IBD. DUOX2-deficient mice replicated increased IL-17C induction in the intestine, with outlier high Il17c expression linked to the mucosal expansion of specific Proteobacteria pathobionts. Integrated microbiota/host gene expression analyses in patients with IBD corroborated IL-17C as a marker for epithelial activation by gram-negative bacteria. Finally, the impact of DUOX2 variants on IL-17C induction provided a rationale for variant stratification in case control studies that substantiated DUOX2 as an IBD risk gene. Thus, our study identifies an association of deleterious DUOX2 variants with a preclinical hallmark of disturbed microbiota-immune homeostasis that appears to precede the manifestation of IBD.

Role of AMPK in Determining the Mediator of Flow Induced Dilation in the Human Microvasculature

An important measure of microvascular endothelial function is flow-induced dilation (FID), the ability of an arteriole to dilate in response to increased flow. In healthy adults, FID is predominantly mediated by endothelial-derived nitric oxide (NO), a vasoactive compound with anti-inflammatory properties. In microvessels from patients with coronary artery disease (CAD), FID occurs via formation and release of hydrogen peroxide (H2O2), a pro-inflammatory vasoactive mediator. We have previously shown that exogenous adiponectin restores NO-mediated FID in CAD arterioles, however the mechanism remains unknown. Because AMP kinase (AMPK), a downstream target of adiponectin receptors, increases cellular NO, we hypothesized that inhibition of AMPK would promote H2O2 as the mediator of FID in arterioles from healthy, nonCAD subjects. Human microvessels (80-200µm) were dissected from discarded surgical adipose specimens, cannulated onto micropipettes and pressurized. Arterioles were pre-constricted with endothelin-1 and changes in vessel wall diameter were measured during graded increases in flow using videomicroscopy. FID was observed in healthy arterioles treated with the AMPK inhibitor, Compound C (1µM, 16hrs) in the presence of the NO scavenger c-PTIO (100µM) (77.9 % max dilation ± 6, n=3), as well as with the H2O2 degradation enzyme PEG-Catalase (500U), (62.4% max dilation ±15, n=3). Interestingly, FID was suppressed only in the presence of both c-PTIO and PEG-Catalase (42.1% max dilation ±16, n=3) compared to controls (69.4% max dilation ±6, n=3). Together, these data may suggest that NO and H2O2 compensate for loss of one another during inhibition of AMPK. Future studies will investigate whether adiponectin-induced restoration of NO requires activation of AMPK as thorough understanding of this pathway during FID is critical for understanding the mechanism by which adiponectin improves microvascular endothelial function during disease.

Glucose oxidase: Applications, sources, and recombinant production

Glucose oxidase is a subset of oxidoreductase enzymes that catalyzes the transfer of electrons from an oxidant to a reductant. Glucose oxidases use oxygen as an external electron acceptor that releases hydrogen peroxide (H2 O2 ). Glucose oxidase has many applications in commercial processes, including improving the color and taste, increasing the persistence of food materials, removing the glucose from the dried egg, and eliminating the oxygen from different juices and beverages. Moreover, glucose oxidase, along with catalase, is used in glucose testing kits (especially in biosensors) to detect and measure the presence of glucose in industrial and biological solutions (e.g., blood and urine specimens). Hence, glucose oxidase is a valuable enzyme in the industry and medical diagnostics. Therefore, evaluating the structure and function of glucose oxidase is crucial for modifying as well as improving its catalytic properties. Finding different sources of glucose oxidase is an effective way to find the type of enzyme with the desired catalysis. Besides, the recombinant production of glucose oxidase is the best approach to produce sufficient amounts of glucose oxidase for various uses. Accordingly, the study of various aspects of glucose oxidase in biotechnology and bioprocessing is crucial.

Antimicrobial properties of Lactobacillus cell‐free supernatants against multidrug‐resistant urogenital pathogens

The healthy vaginal microbiota is dominated by Lactobacillus spp., which provide an important critical line of defense against pathogens, as well as giving beneficial effects to the host. We characterized L. gasseri 1A‐TV, L. fermentum 18A‐TV, and L. crispatus 35A‐TV, from the vaginal microbiota of healthy premenopausal women, for their potential probiotic activities. The antimicrobial effects of the 3 strains and their combination against clinical urogenital bacteria were evaluated together with the activities of their metabolites produced by cell‐free supernatants (CFSs). Their beneficial properties in terms of ability to interfere with vaginal pathogens (co‐aggregation, adhesion to HeLa cells, biofilm formation) and antimicrobial activity mediated by CFSs were assessed against multidrug urogenital pathogens (S. agalactiae, E. coli, KPC‐producing K. pneumoniae, S. aureus, E. faecium VRE, E. faecalis, P. aeruginosa, P. mirabilis, P. vulgaris, C. albicans, C. glabrata). The Lactobacilli tested exhibited an extraordinary ability to interfere and co‐aggregate with urogenital pathogens, except for Candida spp., as well as to adhere to HeLa cells and to produce biofilm in the Lactobacillus combination. Lactobacillus CFSs and their combination revealed a strong bactericidal effect on the multidrug resistant indicator strains tested, except for E. faecium and E. faecalis. The antimicrobial activity was maintained after heat treatment but decreased after enzymatic treatment. All Lactobacilli showed lactic dehydrogenase activity and production of D‐ and L‐lactic acid isomers on Lactobacillus CFSs, while only 1A‐TV and 35A‐TV released hydrogen peroxide and carried helveticin J and acidocin A bacteriocins. These results suggest that they can be employed as a new vaginal probiotic formulation and bio‐therapeutic preparation against urogenital infections. Further, in vivo studies are needed to evaluate human health benefits in clinical situations.

Quantitative measurements of free and immobilized RgDAAO Michaelis-Menten constant using an electrochemical assay reveal the impact of covalent cross-linking on substrate specificity.

Challenges facing enzyme-based electrochemical sensors include substrate specificity, batch to batch reproducibility, and lack of quantitative metrics related to the effect of enzyme immobilization. We present a quick, simple, and general approach for measuring the effect of immobilization and cross-linking on enzyme activity and substrate specificity. The method can be generalized for electrochemical biosensors using an enzyme that releases hydrogen peroxide during its catalytic cycle. Using as proof of concept RgDAAO-based electrochemical biosensors, we found that the Michaelis-Menten constant (Km) decreases post immobilization, hinting at alterations in the enzyme kinetic properties and thus substrate specificity. We confirm the decrease in Km electrochemically by characterizing the substrate specificity of the immobilized RgDAAO using chronoamperometry. Our results demonstrate that enzyme immobilization affects enzyme substrate specificity and this must be carefully evaluated during biosensor development.

Synthesis, Cytotoxicity and Anti-Proliferative Activity Against AGS Cells of New 3(2H)-Pyridazinone Derivatives Endowed with a Piperazinyl Linker.

Novel twenty-three 3(2H)-pyridazinone derivatives were designed and synthesized based on the chemical requirements related to the anti-proliferative effects previously demonstrated within this scaffold. The introduction of a piperazinyl linker between the pyridazinone nucleus and the additional (un)substituted phenyl group led to some compounds endowed with a limited cytotoxicity against human gingival fibroblasts (HGFs) and good anti-proliferative effects against gastric adenocarcinoma cells (AGS) as evaluated by MTT and LDH assays, using doxorubicin as a positive control. Successive analyses revealed that the two most promising representative compounds (12 and 22) could exert their effects by inducing oxidative stress as demonstrated by the hydrogen peroxide release and the morphological changes (cell blebbing) revealed by light microscopy analysis after the haematoxylin-eosin staining. Moreover, to further assess the apoptotic process induced by compounds 12 and 22, Bax expression was measured by flow cytometry. These findings enlarged our knowledge of the structural requirements in this scaffold to display valuable biological effects against cancerous cell lines.

Development of Controlled-Release Carbamide Peroxide Loaded Nanoemulgel for Tooth Bleaching: In Vitro and Ex Vivo Studies.

Burst release of carbamide peroxide (CP) from traditional hydrogels causes severe inflammation to periodontal tissues. The present study explores the development of a novel CP nanoemulgel (CP-NG), an oil-in-water nanoemulsion-based gel in which CP was loaded with a view to controlling CP release. CP solid dispersions were prepared, using white soft paraffin or polyvinylpyrrolidone-white soft paraffin mixture as a carrier, prior to formulating nanoemulsions. It was found that carrier type and the ratio of CP to carrier affected drug crystallinity. Nanoemulsions formulated from the optimized CP solid dispersions were used to prepare CP-NG. It was found that the ratio of drug to carrier in CP solid dispersions affected the particle size and zeta potential of the nanoemulsions as well as drug release behavior and tooth bleaching efficacy of CP-NG. Drug release from CP-NG followed a first-order kinetic reaction and the release mechanism was an anomalous transport. Drug release rate decreased with an increase in solid dispersion carriers. CP-NG obtained from the solid dispersion with a 1:1 ratio of CP to the polymer mixture is suitable for sustaining drug release with high tooth bleaching efficacy and without reduction of enamel microhardness. The developed CP-NG is a promising potential tooth bleaching formulation.

Application of Hydrogen Peroxide Sensitive Fluorescen Nanoparticles in Atherosclerosis.

ROS plays an important role in the formation of atherosclerotic plaque, especially hydrogen peroxide, which can stimulate macrophages to express pro-inflammatory cytokines and chemokines to enhance the inflammatory activity of macrophages. Therefore, the development of drugs with sensitive release of hydrogen peroxide is safe and effective for the treatment of atherosclerosis The side chain hydroxyl groups of pHEMA were grafted with oxalylsimvastatin (SIM), fluoropolyethylene glycol (fpeg) and macrophage target molecule ISO-1 to prepare hydrogen peroxide sensitive fluorescent drug loaded nanomicelles (phema-simfpeg-iso-1). By using hydrogen peroxide sensitive and controlled-release drug, it can target macrophage, at the same time, improve the characteristics of traditional polyethylene glycol only as hydrophilic chain, and synthesize polyethylene glycol with fluorescence function, so that the polymer can have probe ability without modifying fluorescence substance, which is suitable for the diagnosis and treatment of atherosclerosis. The results show that the water-soluble nanoparticles show good biocompatibility and peroxide Hydrogen sensitivity and fluorescence ability provide new materials for the development of a nano system for the diagnosis and treatment of atherosclerosis.

Oxygen-release microspheres capable of releasing oxygen in response to environmental oxygen level to improve stem cell survival and tissue regeneration in ischemic hindlimbs

Stem cell transplantation has been extensively explored to promote ischemic limb vascularization and skeletal muscle regeneration. Yet the therapeutic efficacy is low due to limited cell survival under low oxygen environment of the ischemic limbs. Therefore, continuously oxygenating the transplanted cells has potential to increase their survival. During tissue regeneration, the number of blood vessels are gradually increased, leading to the elevation of tissue oxygen content. Accordingly, less exogenous oxygen is needed for the transplanted cells. Excessive oxygen may induce reactive oxygen species (ROS) formation, causing cell apoptosis. Thus, it is attractive to develop oxygen-release biomaterials that are responsive to the environmental oxygen level. Herein, we developed oxygen-release microspheres whose oxygen release was controlled by oxygen-responsive shell. The shell hydrophilicity and degradation rate decreased as the environmental oxygen level increased, leading to slower oxygen release. The microspheres were capable of directly releasing molecular oxygen, which are safer than those oxygen-release biomaterials that release hydrogen peroxide and rely on its decomposition to form oxygen. The released oxygen significantly enhanced mesenchymal stem cell (MSC) survival without inducing ROS production under hypoxic condition. Co-delivery of MSCs and microspheres to the mouse ischemic limbs ameliorated MSC survival, proliferation and paracrine effects under ischemic conditions. It also significantly accelerated angiogenesis, blood flow restoration, and skeletal muscle regeneration without provoking tissue inflammation. The above results demonstrate that the developed microspheres have potential to augment cell survival in ischemic tissues, and promote ischemic tissue regeneration in a safer and more efficient manner.

VIP modulates human macrophages phenotype via FPRL1 via activation of RhoA-GTPase and PLC pathways.

This study is aimed at uncovering the signaling pathways activated by vasoactive intestinal peptide in human macrophages MATERIALS: Human peripheral blood mononuclear cell-derived macrophages were used for the in vitro investigation of the VIP-activated signaling pathways. Time-course and dose-response experiments and siRNA were used in human macrophages co-challenged with various concentrations of VIP and different MAPK pharmacologic inhibitors to investigate signaling pathways activated by VIP. Flow analysis was performed to assess the levels of CD11b, CD35 and CD66. Luminescence spectrometry was used to measure the levels of the released hydrogen peroxide and the intracellular calcium levels in the media. Macrophages incubated with VIP showed increased phospho-AKT and phospho-ERK1/2 levels in a GTP-RhoA-GTPase-dependent manner. Similarly, VIP increased intracellular release of H2O2 and calcium via PLC and GTP-RhoA-GTPase, in addition to inducing the expression of CD11b, CD35, CD66 and MMP9. Furthermore, VIP activated P38 MAPK through the cAMP/PKA pathway but was independent of both PLC and RhoA signaling. The above-mentioned VIP effects were mediated via activation of the FPRL1 receptor. VIP/FPRL1/VPAC/GTP-RhoA-GTPase signaling modulated macrophages phenotype through activation of multiple signaling pathways including ERK1/2, AKT, P38, ROS, cAMP and calcium.

Aqueous Decomposition Behavior of Solid Peroxides: Effect of pH and Buffer Composition on Oxygen and Hydrogen Peroxide Formation

The ability of solid peroxides to provide sustained release of both oxygen and hydrogen peroxide makes them potentially suitable for oxygen release or antibacterial applications. Most recent reports using solid peroxides to augment oxygen levels do so by compounding solid peroxide powders in polymers to retard the aqueous decomposition. Compounds with peroxidase activity may be added to reduce hydrogen peroxide toxicity. Peroxides are rarely pure and are mixed with oxide and themselves decompose to form hydroxides in water. Therefore, even if buffering strategies are used, locally the pH at the surface of aqueously immersed peroxide particles is inevitably alkaline. Since pH affects the decomposition of peroxides and hydrogen peroxide stability, this study compared for the first-time the aqueous decomposition products of hydrogen and inorganic peroxides that are in use or have been used for medical applications of have been evaluated preclinically; calcium peroxide (CaO2), magnesium peroxide (MgO2), zinc peroxide (ZnO2), sodium percarbonate (Na2CO3.1.5H2O2) and hydrogen peroxide (H2O2). Since plasma can be approximated to be carbonate buffered phosphate solution, we maintained pH using carbonate and phosphate buffers and compared results with citrate buffers. For a given peroxide compound, we identified not only a strong effect of pH but also of buffer composition on the extent to which oxygen and hydrogen peroxide formation occurred. The influence of buffer composition was not previously appreciated, thereby establishing in vitro parameters for better design of intentional release of specific decomposition species.