NADPH Oxidase Research Articles

CysLT receptor-mediated NOX2 activation is required for IL-8 production in HMC-1 cells induced by Trichomonas vaginalis-derived secretory products.

Trichomoniasis is caused by a sexually transmitted flagellate protozoan parasite Trichomonas vaginalis. T. vaginalis-derived secretory products (TvSP) contain lipid mediators such as leukotriene B4 (LTB4) and various cysteinyl leukotrienes (CysLTs) which included LTC4, LTD4, and LTE4. However, the signaling mechanisms by which T. vaginalis-induced CysLTs stimulate interleukin (IL)-8 production in human mast cells remain unclear. In this study, we investigated these mechanisms in human mast cells (HMC-1). Stimulation with TvSP resulted in increased intracellular reactive oxygen species (ROS) generation and NADPH oxidase 2 (NOX2) activation compared to unstimulated cells. Pre-treatment with NOX2 inhibitors such as diphenyleneiodonium chloride (DPI) or apocynin significantly reduced ROS production in TvSP-stimulated HMC-1 cells. Additionally, TvSP stimulation increased NOX2 protein expression and the translocation of p47phox from the cytosol to the membrane. Pretreatment of HMC-1 cells with PI3K or PKC inhibitors reduced TvSP-induced p47phox translocation and ROS generation. Furthermore, NOX2 inhibitors or NOX2 siRNA prevented CREB phosphorylation and IL-8 gene expression or protein secretion induced by TvSP. Pretreatment with a CysLTR antagonist significantly inhibited TvSP-induced ROS production, CREB phosphorylation, and IL-8 production. These results indicate that CysLT-mediated activation of NOX2 plays a crucial role in ROS-dependent IL-8 production in human mast cells stimulated by T. vaginalis-secreted CysLTs. These findings enhance our understanding of the inflammatory response in trichomoniasis and may inform the development of targeted therapies to mitigate this response.

Permissive lung neutrophils facilitate tuberculosis immunopathogenesis in male phagocyte NADPH oxidase-deficient mice.

NADPH oxidase 2 (NOX2) is an enzyme responsible for generating reactive oxygen species, primarily found in phagocytes. Chronic Granulomatous Disease (CGD), along with bacterial infections such as Mycobacterium tuberculosis (Mtb), is a representative NOX2-deficient X-linked disease characterized by uncontrolled inflammation. However, the precise roles of host-derived factors that induce infection-mediated hyperinflammation in NOX2-deficient condition remain incompletely understood. To address this, we compared Mtb-induced pathogenesis in Nox2-/- and wild type (WT) mice in a sex-dependent manner. Among age- and sex-matched mice subjected to Mtb infection, male Nox2-/- mice exhibited a notable increase in bacterial burden and lung inflammation. This was characterized by significantly elevated pro-inflammatory cytokines such as G-CSF, TNF-α, IL-1α, IL-1β, and IL-6, excessive neutrophil infiltration, and reduced pulmonary lymphocyte levels as tuberculosis (TB) progressed. Notably, lungs of male Nox2-/- mice were predominantly populated with CD11bintLy6GintCXCR2loCD62Llo immature neutrophils which featured mycobacterial permissiveness. By diminishing total lung neutrophils or reducing immature neutrophils, TB immunopathogenesis was notably abrogated in male Nox2-/- mice. Ultimately, we identified G-CSF as the pivotal trigger that exacerbates the generation of immature permissive neutrophils, leading to TB immunopathogenesis in male Nox2-/- mice. In contrast, neutralizing IL-1α and IL-1β, which are previously known factors responsible for TB pathogenesis in Nox2-/- mice, aggravated TB immunopathogenesis. Our study revealed that G-CSF-driven immature and permissive pulmonary neutrophils are the primary cause of TB immunopathogenesis and lung hyperinflammation in male Nox2-/- mice. This highlights the importance of quantitative and qualitative control of pulmonary neutrophils to alleviate TB progression in a phagocyte oxidase-deficient condition.

Design, Synthesis, and Biological Evaluation of Quinoline (Quinolinone) Derivatives as NADPH Oxidase (NOX) Inhibitors.

NADPH oxidases (NOXs) are the sole enzyme in the human body that can directly produce reactive oxygen species. Recent studies have shown that NOXs is a very promising target for the treatment of diabetic nephropathy (DN). Here, a series of quinoline(quinolinone) derivatives have been designed based on pharmacophore strategy, synthesized and evaluated. Among them, 19d exhibits potent antiproliferative and NOXs inhibitory activities, and is worthy for further investigation.

Zhuyu Pills regulate p53/SLC7A11 signaling pathway-mediated oxidative damage and ferroptosis to treat atherosclerosis

This article aims to investigate the effect of Zhuyu Pills on atherosclerosis(AS) and decipher the underlying mechanism. The mouse model of AS was established by feeding with a high-fat diet for 12 weeks. The 50 successfully modeled mice with the apolipoprotein E knockout(ApoE~(-/-)) were assigned by the random number table method into 5 groups(n=10): model, low-, medium-, and high-dose(130.54, 261.08, 522.16 mg·kg~(-1), respectively) Zhuyu Pills, and atorvastatin calcium(10.40 mg·kg~(-1)). Ten C57BL/6J mice were selected as the blank group. The blank group and model group were administrated with an equal volume of normal saline, and other groups were administrated with corresponding drugs once a day for 12 weeks. At the end of drug intervention, hematoxylin-eosin(HE) staining was employed to observe the pathological changes of fat in the aorta, liver, and epididymis of mice, and the proportion of aortic plaque area, fat area in epididymis, and nonalcoholic fatty liver disease activity score(NAS) were calculated. Lipid and collagen deposition in the aorta was observed by oil red O staining and Masson staining, respectively, and the proportions of lipid and collagen deposition areas were calculated. The serum levels of superoxide dismutase(SOD), malondialdehyde(MDA), glutathione peroxidase(GSH-Px), and iron ion were measured by colorimetry. The expression of cyclooxygenase 2(COX2), ferritin heavy chain 1(FTH1), cystine/glutamate reverse transporter solute carrier family 7 member 11(SLC7A11), and glutathione peroxidase 4(GPX4) in the aorta was detected by the immunofluorescence assay. The level of tumor protein 53(p53) in the aorta was detected by immunohistochemistry. The protein levels of p53 and SLC7A11 in the aorta were determined by Western blot. The mRNA levels of p53, SLC7A11, GPX4, FTH1, prostaglandin G/H synthase 2(PTGS2), and reduced nicotinamide adenine dinucleotide phosphate oxidase 1(NOX1) in mouse aorta were determined by real-time PCR. The results showed that compared with the blank group, the model group showcased enlarged aortic plaque area, increased collagen fiber deposition, liver lipid deposition, and lipid droplets, and enlarged epididymal adipocytes. In addition, the modeling elevated the levels of iron ion and MDA and lowered the levels of SOD and GSH-Px in the serum, promoted the expression of p53 and COX2, down-regulated the protein and mRNA levels of FTH1, SLC7A11, and GPX4, and up-regulated the mRNA levels of PTGS2 and NOX1 in the aorta. Compared with the model group, low-, medium-, and high-dose Zhuyu Pills and atorvastatin calcium reduced the aortic plaque area, collagen deposition, liver lipid deposition, lipid droplets, and epididymal adipocyte volume, lowered the levels of iron ion and MDA and elevated the levels of SOD and GSH-Px in the serum, inhibited the expression of p53 and COX2, up-regulated the protein and mRNA levels of FTH1, SLC7A11, and GPX4, and down-regulated the mRNA levels of PTGS2 and NOX1 in the aorta. In conclusion, Zhuyu Pills exert definite therapeutic effect on aortic plaque in AS mice by regulating the p53/SLC7A11 signaling pathway to alleviate oxidative damage and inhibit ferroptosis.

The Role of Natural Low Molecular Weight Dicarbonyls in Atherogenesis and Diabetogenesis.

This review summarises the data from long-term experimental studies and literature data on the role of oxidatively modified low-density lipoproteins (LDL) in atherogenesis and diabetogenesis. It was shown that not "oxidized" (lipoperoxide-containing) LDL, but dicarbonyl-modified LDL are atherogenic (actively captured by cultured macrophages with the help of scavenger receptors), and also cause expression of lectin like oxidized low density lipoprotein receptor 1 (LOX-1) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (NOX-1) genes in endotheliocytes, which stimulate apoptosis and endothelial dysfunction. The obtained data allowed us to justify new approaches to pharmacotherapy of atherosclerosis and diabetes mellitus.

Transcription Factor CgSte12 Regulates Pathogenicity by Affecting Appressorium Structural Development in the Anthracnose-Causing Fungus Colletotrichum gloeosporioides.

Colletotrichum gloeosporioides is the causal agent of poplar anthracnose, which induces major economic losses and adversely affects the ecosystem services of poplar forests. The appressorium serves as a penetration structure for many pathogenic fungi, including C. gloeosporioides. The production of mucilage and the formation of penetration pegs are critically important for the appressorium-mediated penetration of host tissues. We previously found that CgPmk1 is a key protein involved in appressorium formation, penetration, and pathogenicity. Although CgSte12, which is a transcription factor that functions downstream of CgPmk1, regulates the formation of penetration pegs, its role in C. gloeosporioides appressorium development and pathogenicity has not been elucidated. Here, we developed C. gloeosporioides CgSTE12 mutants and characterized the molecular and cellular functions of CgSTE12. The results showed that mycelial growth and morphology were not affected in the CgSTE12 knockout mutants, which produced normal melanized appressoria. However, these mutants had less mucilage secreted around the appressoria, impaired appressorial cone formation, and the inability to form penetration pores and pegs, which ultimately led to a significant loss of pathogenicity. Our comparative transcriptome analysis revealed that CgSte12 controls the expression of genes involved in appressorium development and function, including genes encoding cutinases, NADPH oxidase, spermine biosynthesis-related proteins, ceramide biosynthesis-related proteins, fatty acid metabolism-related proteins, and glycerophospholipid metabolism-related proteins. Overall, our findings indicate that CgSte12 is a critical regulator of appressorium development and affects C. gloeosporioides pathogenicity by modulating the structural integrity of appressoria.

Fructooligosaccharides Ameliorate Renal Injury and Dysfunction Through the Modulation of Gut Dysbiosis, Inhibition of Renal Inflammation, Oxidative Stress, Fibrosis, and Improve Organic Anion Transporter 3 Function in an Obese Rat Model.

High-fat diet (HFD) consumption causes obesity and gut dysbiosis which induces kidney injury. It has been reported that prebiotics improve gut dysbiosis and insulin sensitivity and decelerate the progression of kidney disease. This study investigates the impact of fructooligosaccharides (FOS) on renoprotection and the prevention of gut dysbiosis and intestinal barrier injury in obese rats. Wistar rats are treated with HFD for 16 weeks. Then, the HFD fed rats (HF) are given FOS 1gday-1 (HFFOS1), 2gday-1 (HFFOS2), or metformin 30mgkg-1day-1 (HFMET), by intragastric feeding for 8 weeks. Blood, urine, feces, kidney, and intestine are collected to determine the metabolic changes, gut dysbiosis, and the expression of proteins involved in kidney and intestinal injury. FOS can attenuate insulin resistance and hypercholesterolemia concomitant with the inhibition of renal inflammation, oxidative stress, fibrosis, and apoptosis, which are related to the deceleration of the overexpression of renal Toll-like receptor 4 (TLR4) and NADPH oxidase (NOX4). Moreover, FOS shows a greater efficacy than metformin in the reduction of the intestinal injury and loss of tight junction proteins induced by HFD. FOS may be used as a supplement for therapeutic purposes in an obese condition to improve intestinal integrity and prevent renal complications.

MVP enhances FGF21-induced ferroptosis in hepatocellular carcinoma by increasing lipid peroxidation through regulation of NOX4.

Ferroptosis is a novel, iron-dependent regulatory cell death mainly caused by an imbalance between the production and degradation of intracellular reactive oxygen species (ROS). Recently, ferroptosis induction has been considered a potential therapeutic approach for hepatocellular carcinoma (HCC). Fibroblast growth factor 21 (FGF21) is a new modulator of ferroptosis; however, the regulatory role of FGF21 in HCC ferroptosis has not been investigated. In this study, we explored the role of FGF21 and its underlying molecular mechanism in the ferroptotic death of HCC cells. We identified Major vault protein (MVP) as a target of FGF21 and revealed that knockdown of MVP inhibited the lipid peroxidation levels of HCC cells by decreasing NADPH oxidase 4 (NOX4, a major source of ROS) transcription, thereby attenuating the effect of FGF21-mediated ferroptosis. On the other hand, MVP overexpression showed the opposite results. Mechanistically, MVP binds to IRF1 and thus interferes with the interaction between IRF1 and the YAP1 promoter, leading to an increase in NOX4 transcription. Importantly, forced expression of IRF1 or downregulation of YAP1 partially reversed the effect of MVP overexpression on HCC ferroptosis. Furthermore, the results in xenograft tumor models suggested that overexpression of MVP can efficiently increase the level of lipid peroxidation invivo. Taken together, these results provide new insights into the regulatory mechanism of ferroptosis in HCC.

Deubiquitinating Enzyme USP19 Regulates Ferroptosis and Mitochondrial Damage in SH-SY5Y Cells by Targeting the NOX4 Protein.

Ferroptosis is extremely relevant to the progression of neurodegenerative pathologies such as Alzheimer's disease (AD). Ubiquitin-specific proteases (USP) can affect the NADPH oxidase family. Our study aimed to elucidate the potential role and molecular basis of a certain USP19 in reducing ferroptosis and mitochondrial injury in AD cells by targeting NOX4 stability. The deubiquitinase USP family gene USP19, which affects the stability of NOX4 protein, was first screened. The cell model of AD was constructed after interfering with SH-SY5Y cells by Aβ1-40, and then SH-SY5Y cells were infected with lentiviral vectors to knock down USP19 and overexpress NOX4, respectively. Finally, the groups were tested for cell viability, changes in cellular mitochondrial membrane potential, lipid reactive oxygen species, intracellular iron metabolism, and NOX4, Mf1, Mf2, and Drp1 protein expression. 5 μmol/L Aβ1-40 intervened in SH-SY5Y cells for 24 h to construct a cell model of AD. Knockdown of USP19 decreased the expression of NOX4 protein, promoted the expression of mitochondrial fusion proteins Mnf1 and Mnf2, and inhibited the expression of the splitting protein Drp1. Furthermore, USP19 knockdown decreased mitochondrial membrane potential, SOD, MDA, intracellular iron content and increased GSH/GSSG ratio in SH-SY5Y cells. Our study revealed that NOX4 protein interacts with USP19 and knockdown of USP19 enhanced ubiquitination to maintain NOX4 protein stability. USP19 attenuates mitochondrial damage in SH-SY5Y cells by targeting NOX4 protein with Aβ1-40.

Altered neutrophil extracellular traps formation among medical residents with sleep deprivation

Resident physicians on long-term night shifts often face sleep deprivation, affecting the immune response, notably neutrophils, vital to innate defense mechanisms. Sleep-deprived residents exhibit altered neutrophil counts and reduced phagocytosis and NADPH oxidase activity, critical to combating infections. Our study focused on neutrophil extracellular traps (NETs), a defense process against pathogens not previously linked to sleep loss. Results revealed that sleep-deprived residents exhibited a 19.8 % reduction in NET formation compared to hospital workers with regular sleep patterns (P < 0.01). Additionally, key NETs proteins, Neutrophil Elastase and Myeloperoxidase, were less active in sleep-deprived individuals (1.53mU; P < 0.01 and 0.95U; P < 0.001 decrease, accordingly). Interestingly, the ability to form NETs resumed to normal levels three months post-residency among pediatric residents. The causal relationship between reduced NETs due to sleep deprivation and the increased susceptibility to infections, as well as its implications for infection severity, is a critical area for further investigation.

Gut-derived wild blueberry phenolic acid metabolites modulate extrinsic cutaneous damage.

As the first line of defense, the skin is equipped with various physiological mechanisms positioned to prevent incoming oxidative damage from numerous environmental insults. With persistent exposure to the environment, understanding ways to augment the skin defenses is paramount in protecting from premature aging. In this study, we investigated the ability of five dietary phenolic metabolites, typically found in the bloodstream after wild blueberry consumption, to successfully defend the skin from UV light exposure in a novel ex vivo co-culture model of human skin explants and primary endothelial cells. Skin explants, placed in transwell inserts, were exposed to UV, and subsequently co-cultured with endothelial cells. When the endothelial cells had been pretreated with the bioactive metabolites at physiological concentrations (hippuric acid 3000 nM, isoferulic acid 1000 nM, salicylic acid 130 nM, benzoic acid 900 nM, α-hydroxyhippuric acid 400 nM) cutaneous damage was prevented on the co-cultured with UV-challenged skin explants. Co-culture with non-pretreated endothelial cells did not protect skin explants. Specifically, the pretreatment was able to reduce skin lipid peroxidation (measured as 4-hydroxynonenal protein adducts), and pro-inflammatory enzymes such as cyclooxygenase 2 (COX-2) and NADPH oxidase 4 (NOX-4). Furthermore, pretreatment with the metabolites prevented UV-induced release of inflammatory cytokines such as IL-1β and IL-8 as well as nitric oxides (NO) levels. In addition, the metabolites showed an impressive ability to prevent the loss of cutaneous structural proteins including involucrin and collagen type 1. Of note, endothelial cells cultured with UV exposed skin explants exhibited increased oxidative stress demonstrated by heme oxygenase-1 (HO-1) up-regulation which was significantly prevented in the metabolite treated models. These findings highlight the ability of dietary polyphenolic metabolites to improve cutaneous defenses against extrinsic stressors.

Analysis of Molecular Mechanisms of Chronic Irradiation Effects on Electrical Signals in Wheat Plants

The effect of ionizing radiation (IR) on plants is mainly realized by altering the status of signaling systems and modifying stress signals. Variation potential (VP) is one of the types of electrical signals in plants. IR contributes to an increase in the amplitude of the VP, but the mechanisms of such influence are practically unknown. A possible way to implement changes arising from the action of IR is the regulation of gene expression. In the present work, the changes in the gene expression of participants in the generation and propagation of VP in irradiated plants are investigated. The experiments were performed on 14–15-day-old soft wheat plants (Triticum aestivum L.) grown under chronic irradiation (source 90Sr-90Y) with a dose rate of 31.3 μGy/h. The maximum accumulated dose was about 11.3 mGy. The irradiated plants showed no changes in the expression of calcium (TPC1), anionic (ALMT1 and CLC1), potassium (AKT1) channels, H+-ATPase (HA1), and NADPH oxidase (RBOHs) genes. A decrease in the expression of the SKOR potassium channel gene was revealed. The potassium channel blocker, tetraethylammonium chloride, caused an increase in response amplitude in control plants comparable to the increase in amplitude in the irradiated group. The obtained results indicate that one of the ways IR influences the electrical signals of plants is to inhibit the expression of the potassium channel.

Oxidative Stress and Mitochondria Are Involved in Anaphylaxis and Mast Cell Degranulation: A Systematic Review.

Anaphylaxis, an allergic reaction caused by the massive release of active mediators, can lead to anaphylactic shock (AS), the most severe and potentially life-threatening form of anaphylactic reaction. Nevertheless, understanding of its pathophysiology to support new therapies still needs to be improved. We performed a systematic review, assessing the role and the complex cellular interplay of mitochondria and oxidative stress during anaphylaxis, mast cell metabolism and degranulation. After presenting the main characteristics of anaphylaxis, the oxidant/antioxidant balance and mitochondrial functions, we focused this review on the involvement of mitochondria and oxidative stress in anaphylaxis. Then, we discussed the role of oxidative stress and mitochondria following mast cell stimulation by allergens, leading to degranulation, in order to further elucidate mechanistic pathways. Finally, we considered potential therapeutic interventions implementing these findings for the treatment of anaphylaxis. Experimental studies evaluated mainly cardiomyocyte metabolism during AS. Cardiac dysfunction was associated with left ventricle mitochondrial impairment and lipid peroxidation. Studies evaluating in vitro mast cell degranulation, following Immunoglobulin E (IgE) or non-IgE stimulation, revealed that mitochondrial respiratory complex integrity and membrane potential are crucial for mast cell degranulation. Antigen stimulation raises reactive oxygen species (ROS) production from nicotinamide adenine dinucleotide phosphate (NADPH) oxidases and mitochondria, leading to mast cell degranulation. Moreover, mast cell activation involved mitochondrial morphological changes and mitochondrial translocation to the cell surface near exocytosis sites. Interestingly, antioxidant administration reduced degranulation by lowering ROS levels. Altogether, these results highlight the crucial role of oxidative stress and mitochondria during anaphylaxis and mast cell degranulation. New therapeutics against anaphylaxis should probably target oxidative stress and mitochondria, in order to decrease anaphylaxis-induced systemic and major organ deleterious effects.

Diagnosis of Chronic Granulomatous Disease: Strengths and Challenges in the Genomic Era.

Chronic granulomatous disease (CGD) is a group of rare primary inborn errors of immunity characterised by a defect in the phagocyte respiratory burst, which leads to severe and life-threatening infective and inflammatory complications. Despite recent advances in our understanding of the genetic and molecular pathophysiology of X-linked and autosomal recessive CGD, and growth in the availability of functional and genetic testing, there remain significant barriers to early and accurate diagnosis. In the current review, we provide an up-to-date summary of CGD pathophysiology, underpinning current methods of diagnostic testing for CGD and closely related disorders. We present an overview of the benefits of early diagnosis and when to suspect and test for CGD. We discuss current and historical methods for functional testing of NADPH oxidase activity, as well as assays for measuring protein expression of NADPH oxidase subunits. Lastly, we focus on genetic and genomic methods employed to diagnose CGD, including gene-targeted panels, comprehensive genomic testing and ancillary methods. Throughout, we highlight general limitations of testing, and caveats specific to interpretation of results in the context of CGD and related disorders, and provide an outlook for newborn screening and the future.

Furanodienone induces apoptosis via regulating the PRDX1/MAPKs/p53/caspases signaling axis through NOX4-derived mitochondrial ROS in colorectal cancer cells

Furanodienone, a biologically active constituent of sesquiterpenes isolated from Rhizome Curcumae, has been reported to induce apoptosis in human colorectal cancer (CRC) cells by promoting the generation of reactive oxygen species (ROS). However, the source of ROS and how it manipulates apoptosis in CRC cells remains to be elucidated. Herein, we assessed the potential role of the well-known sources of intracellular ROS—mitochondrial electron transport chain and the nicotinamide adenine dinucleotide phosphate oxidases (NOXs), on furanodienone-induced cell death. The results indicated that furanodienone substantially increased the levels of mitochondrial ROS, which were subsequently eliminated by the general NOX inhibitor. Specifically, the nuclear factor kappa-B (NF-κB) translocation triggered a significant rise in the expression of NOX4, an isoform of the NOXs family, upon furanodienone treatment. Nevertheless, the specific NOX4 inhibitor GLX351322 attenuated cell apoptosis and mitochondrial ROS production. As a result, ROS burst induced by furanodienone suppressed the expression of peroxiredoxin1 (PRDX1), a redox signaling protein overexpressed in CRC cells, through a nuclear factor-erythroid-2-related factor 2 (Nrf2)-dependent pathway, thus amplifying the mitogen-activated protein kinases (MAPKs)/p53-mediated apoptotic signaling by increasing the p-p38, p-JNK levels, as well as the cleaved caspases −3, −8 and −9. In vivo experiments further confirmed the anti-proliferative impact of PRDX1 following furanodienone treatment. In summary, the study demonstrated that furanodienone-induced apoptosis in CRC cells is initiated by mitochondrial ROS derived from NOX4, which targeted the PRDX1 and activated the downstream MAPKs/p53-mediated caspase-dependent signaling pathway. Our findings may provide novel insights into the development of adjuvant drugs for CRC treatment and therapeutic applications.

Synthetic thymic hexapeptide in the correction of alterations of antibacterial immune defense and normalization of the profile of proinflammatory cytokines in immunocompromized Children with local unlimited acute peritonitis

The study of dysregulatory disorders of the immune system underlying the immunopathogenesis of severe purulent-inflammatory diseases (PIDs) is important for development of new therapeutic tactics of restoring antibacterial defense. Acute peritonitis (AP) is a severe PID of the abdominal cavity, the course of which are dependent on the treatment, cytokine balance and adequate functioning of the immunity. Objective: to evaluate the modulating effects on the immune system and the levels of proinflammatory cytokines of the synthetic thymic hexapeptide, active substance of Imunofan, included in the complex postoperative treatment (CPOT) of immunocompromised children with local AP. Clinical and immunological examination of 20 immunocompromised children aged 5–12 years with local AP was carried out before (study group 1, SG1) and after (study group 1a, SG1a) CPOT including synthetic thymic hexapeptide (Arginyl-alpha-AspartylLysyl-Valyl-Tyrosyl-Arginine, НР), alongside 20 conditionally healthy children (comparison group, SG). The content of T and B lymphocytes, natural killer cells (NK), levels of serum pro-inflammatory cytokines IL-1β, IL-6, TNFα, IL-8, IL-18, phagocytic and microbicidal activity of neutrophils (NG) were assessed. In SG1, before treatment, a decrease in the number of T lymphocytes, T helpers, CTL-lymphocytes, NK and an increase in the level of B lymphocytes was revealed. Defects in the effector functions of NG were determined: impaired bacterial antigen killing and decreased NADPH oxidase activity. It was established that in case of AP in immunocompromised children, the cytokine profile is characterized by overproduction of studied proinflammatory and neutrophil-associated cytokines. After complex treatment including immunomodulatory therapy, there was a restoration of the content of T lymphocytes, T helper cells, TCTL lymphocytes, an increase in the number of NK and decrease in the level of B lymphocytes. In addition, regression of the levels of inflammatory, including neutrophil-associated, cytokines and emergence of effector functions of NG due to restoration of NADPH oxidases activity, was noted. Thus, the restoration of immunological parameters in AP leads to earlier regression of the purulent-inflammatory process in the abdominal cavity and to the absence of postoperative complications. The clinical and immunological effects of the immunomodulatory therapy program with inclusion of the drug based on HP determines the feasibility of its use in the postoperative period in immunocompromised children with local AP.

Signaling by Neutrophil G Protein-Coupled Receptors that Regulate the Release of Superoxide Anions.

In human peripheral blood, the neutrophil granulocytes (neutrophils) are the most-abundant white blood cells. These professional phagocytes are rapidly recruited from the bloodstream to inflamed tissues by chemotactic factors that signal danger. Neutrophils, which express many receptors that are members of the large family of G protein-coupled receptors (GPCRs), are critical for the elimination of pathogens and inflammatory insults, as well as for the resolution of inflammation leading to tissue repair. Danger-signaling molecular patterns such as the N-formylated peptides that are formed during bacterial and mitochondrial protein synthesis and recognized by formyl peptide receptors (FPRs) and free fatty acids recognized by free fatty acid receptors (FFARs) regulate neutrophil functions. Short peptides and short chain fatty acids activate FPR1 and FFA2R, respectively, while longer peptides and fatty acids activate FPR2 and GPR84, respectively. The activation profiles of these receptors include the release of reactive oxygen species (ROS) generated by the NADPH oxidase. Activation of the oxidase and the production of ROS are processes that are regulated by proinflammatory mediators, including TNFα and GM-CSF. The receptors have signaling and functional similarities, although there are also important differences, not only between the two closely related neutrophil FPRs, but also between the FPRs and the FFARs. In neutrophils, these receptors never walk alone, and additional mechanistic insights into the regulation of the GPCRs and the novel regulatory mechanisms underlying the activation of NADPH oxidase advance our understanding of the role of receptor transactivation in the regulation of inflammatory reactions.

GGPPS Negatively Regulates the Formation of Neutrophil Extracellular Traps in Lipopolysaccharide-Induced Acute Lung Injury.

Acute respiratory distress syndrome (ARDS) and acute lung injury (ALI) are life-threatening diseases. Neutrophil extracellular traps (NETs) play a key role in lung damage. Geranylgeranyl diphosphate synthase (GGPPS) is associated with the development of inflammatory diseases. We aimed to explore the role of GGPPS in NETs formation in ARDS/ALI. First, lung pathological changes in lipopolysaccharide (LPS)-induced ALI mice after myeloid-specific GGPPS deletion were evaluated. The level of NETs formation was analyzed by immunofluorescence, PicoGreen assay and Western blotting. Next, we determined the role of GGPPS in NETs formation and underlying mechanisms using immunofluorescence, flow cytometry, DCFH-DA, and SYTOX GREEN staining in vitro. Finally, the correlation between GGPPS expression incirculating neutrophils and dsDNA levels in plasma was evaluated. Myeloid-specific GGPPS deletion mice showed increased NETs deposition in lung tissue and aggravated histopathological damage of lung tissue. In vitro, GGPPS deficiency in neutrophils resulted in increased NETs formation by Phorbol-12-myristate-13-acetate (PMA), which was reversed by Geranylgeranyl diphosphate (GGPP). In addition, inhibitors blocking protein kinase C (PKC) and NADPH-oxidase (NOX) decreased NETs formation induced by GGPPS deletion. Importantly, GGPPS expression in circulating neutrophils was decreased in ARDS patients compared with the healthy control, and the level of dsDNA in plasma of ARDS patients was negatively correlated with the GGPPS expression. Taken together, GGPPS deficiency in neutrophils aggravates LPS-induced lung injury by promoting NETs formation via PKC/NOX signaling. Thus, neutrophil GGPPS could be a key therapeutic target for ARDS.

Anti-inflammatory and antioxidant mechanisms of coniferaldehyde in lipopolysaccharide-induced neuroinflammation: Involvement of AMPK/Nrf2 and TAK1/MAPK/NF-κB signaling pathways

Microglia are primarily involved in inflammatory reactions and oxidative stress in the brain; as such reducing microglial activation has been proposed as a potential therapeutic strategy for neurodegenerative disorders. Herein, we investigated the anti-inflammatory and antioxidant activities of coniferaldehyde (CFA), a naturally occurring cinnamaldehyde derivative, on activated microglia to evaluate its therapeutic potential. CFA inhibited the production of nitric oxide (NO) and proinflammatory cytokines, such as tumor necrosis factor-α, interleukin (IL)-1β, and IL-6, in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. CFA also inhibited intracellular reactive oxygen species levels and oxidative stress markers such as 4-HNE and 8-OHdG. Detailed mechanistic studies showed that CFA exerted anti-inflammatory effects by inhibiting TAK1-mediated MAP kinase/NF-κB activation and upregulating AMPK signaling pathways. In addition, CFA exerted antioxidant effects by inhibiting the NADPH oxidase subunits and by increasing the expression of antioxidant enzymes such as HO-1, NQO1, and catalase by upregulating Nrf2 signaling. Finally, we confirmed the effects of CFA on the brains of the LPS-injected mice. CFA inhibited microglial activation and the expression of proinflammatory markers and increased Nrf2-driven antioxidant enzymes. Furthermore, CFA inhibited the production of 4-HNE and 8-OHdG in the brains of LPS-injected mice. As a result, CFA's significant anti-inflammatory and antioxidant properties may have therapeutic applications in neuroinflammatory disorders related with oxidative stress and microglial activation.

NADPH oxidase 5: Where are we now and which way to proceed?

Since the incorporation of mitochondria in early eukaryotes cells struggle to keep the deleterious effects of reactive oxygen species (ROS), mainly originating from the respiratory chain, at bay. Evolutionary adaptation to ROS burden went so far that by acting as messenger and effector molecules, ROS became important in maintaining homeostasis. The evolutionary success of this phenomenon is underscored by the arising of professional ROS-generating enzymes, namely the family of NADPH oxidases (NOXes). NOXes, by shaping ROS levels at different subcellular locations and in extracellular space, are involved in such fundamental functions as proliferation, differentiation, apoptosis, host defense, fertilization, and hormone biosynthesis. NOX5, being a calcium-regulated professional ROS source exerts its function at the crossroad of these two fundamental but potentially deleterious intracellular signaling pathways (i.e. Ca2+ and ROS). The expression of NOX5 in the adult human body under unchallenged conditions is restricted to very few sites, among which the two major tissue groups are genital organs (mainly testis) and immune tissues (mainly spleen). In cases of increased cellular proliferation and protein synthesis (e.g., diverse tumors, cultured primary cells, or sites of tissue damage) the expression and activity of NOX5 is often upregulated in various tissues. This and the evolutionary conserved nature of NOX5 would imply a very fundamental role for this enzyme, but intriguingly the genomes of rodents essentially lack the NOX5 gene. The latter fact had been a major obstacle in determining the physiological roles of NOX5 in normal tissues until the very recent generation of a NOX5-deficient rabbit model.