Myeloperoxidase Release Research Articles

Selective Twin NIR‐IIb Ratiometric Luminescence Nanoprobes Enable the Accurate Visualization of MPO‐Mediated Oxidative Stress in Acute Liver Injury

AbstractOxidative stress, orchestrated by myeloperoxidase (MPO), plays crucial roles in the progression of many diseases. Nonetheless, the role of MPO‐mediated oxidative stress in distinct factor‐induced acute liver injuries (ALI) is still up for dispute, mainly due to the lack of probes for in vivo monitoring of MPO releases. Here, a highly selective MPO probe (CSQ) based on the epoxidation biochemical reaction within the MPO‐H2O2‐Cl− system is screened to construct dual near infrared‐IIb (NIR‐IIb) ratiometric (F1550Em, 808Ex/F1550Em, 980Ex) luminescence nanoprobes by integrating CSQ onto down conversion nanoparticles (DCNPs) with and without liver‐targeting moiety (twin NIR‐IIb nanoprobes). Liver‐targeting probes are employed to monitor MPO release, whereas non‐liver‐targeting probes are utilized to assess MPO activity across all cell types. Using twin NIR‐IIb nanoprobes, the MPO‐mediated oxidative stress progressively increased are observed in carbon tetrachloride (CCl4)‐induced ALI over 12 h. In contrast, the MPO‐mediated oxidative stress in acetaminophen (APAP)‐induced ALI initially increased, peaked within 3 h, and then rapidly weakened to normal levels within 12 h. Importantly, the differential release of MPO from neutrophils/Kupfer cells to extracellular fluids in the two types of ALI is revealed. This work reveals significant differences in MPO distribution and the role of MPO‐mediated oxidative stress in CCl4 and APAP‐induced ALI.

In Situ Imaging of Cellular Inflammatory Response to Antibiotic Exposure with a DNAzyme Nanorobot.

Antibiotic-induced inflammation involves the release of myeloperoxidase (MPO), an enzyme whose expression in tissues is associated with the inflammatory pathway. However, existing methods for detecting MPO in cells are limited. In this study, a DNAzyme nanorobot was developed using a scaffold of gold nanoparticles (AuNPs) decorated with functional DNAzyme strands and their fluorophore-labeled substrate strands. The DNAzyme remains inactive due to a self-assembled hairpin structure, with a phosphorothioate (PT) modification inserted into the stem domain. When MPO is present, it triggers a halogenation process that generates hypochlorous acid (HClO). HClO specifically catalyzes the cleavage of the PT-site, releasing free DNAzyme strands to cleave their substrates and generating an increasing fluorescent signal. The detection limit for MPO and its primary product, HClO, were determined to be 0.038 μg/mL and 0.013 μM, respectively. The DNAzyme nanorobot can be readily introduced into cells and function autonomously to differentiate increased MPO/HClO levels caused by antibiotics. This approach was applied to image RAW264.7 cells exposed to four prevalent antibiotics found in the environment (phorbol 12-myristate 13-acetate, erythromycin, penicillin, and tetracycline) as well as antibiotic production wastewater. This nanorobot offers novel strategies for monitoring inflammation to evaluate the health impacts of antibiotic exposure.

Impact of Hypoxia on Neutrophil Degranulation and Inflammatory Response in Alpha-1 Antitrypsin Deficiency Patients.

Alpha-1 antitrypsin deficiency (AATD) is an inflammatory disorder where neutrophils play a key role. Excessive neutrophil activation leads to local hypoxia and tissue damage. Most research on neutrophil function has been conducted under atmospheric conditions (21% O2), which may not represent physiological or pathological conditions. This study aimed to determine the effects of hypoxia on neutrophil degranulation and cytokine production in AATD patients. Neutrophils isolated from 54 AATD patients (31 MZ; 8 SZ; 15 ZZ) and 7 controls (MM) were exposed to hypoxia (1% O2) for 4 h. Neutrophil degranulation was assessed by measuring elastase (NE), myeloperoxidase (MPO), lactoferrin, and matrix metalloproteinase-9 (MMP-9) levels using immunoassay-based methods. Pro-inflammatory (IL-8, IL-1 beta, IL-6, and TNF-alpha) and anti-inflammatory (IL-4 and IL-10) cytokine levels were assessed by a Luminex-based method. Our results indicate a significantly increased release of NE (p = 0.015), MPO (p = 0.042), lactoferrin (p = 0.015), and MMP-9 (p = 0.001) compared to controls. Pro-inflammatory cytokines show a significant rise in IL-8 (p = 0.019), a trend towards increased IL-1 beta (p = 0.3196), no change in IL-6 (p = 0.7329), and reduced TNF-alpha (p = 0.006). Anti-inflammatory cytokines show increased IL-4 (p = 0.057) and decreased IL-10 (p = 0.05703). Increased neutrophil degranulation and inflammatory phenotype are observed in AATD neutrophils under physiological hypoxia.

Anti-inflammatory and antiophidic effects of extract of Hymenaea eriogyne benth and structure-activity relationship prediction of the major markers in silico

Ethnopharmacological relevanceHymenaea eriogyne Benth (Fabaceae) is popularly known as "Jatobá". Despite its use in folk medicine to treat inflammatory disorders, there are no descriptions that show its anti-inflammatory potential.Aim of the study: In this sense, this study aimed to evaluate the anti-inflammatory and antivenom action of bark and leaves extract of H. eriogyne. Materials and methodsThe in vivo anti-inflammatory activity was conducted by carrageenan-induced paw edema and zymosan-induced air pouch models, evaluating the edematogenic effect, leukocyte migration, protein concentration, levels of pro-inflammatory cytokines, malondialdehyde (MDA) and myeloperoxidase (MPO) activity. The antivenom potential was investigated in vitro on the enzymatic action (proteolytic, phospholipase and hyaluronidase) of Bothrops brazili and B. leucurus venom, as well as in vivo on the paw edema model induced by B. leucurus. Furthermore, the influence of its markers (astilbin and rutin) on MPO activity was investigated in silico. For molecular docking, AutodockVina, Biovia Discovery Studio, and Chimera 1.16 software were used. ResultsThe extracts and bark and leaves of H. eriogyne revealed a high anti-inflammatory effect, with a reduction in all inflammatory parameters evaluated. The bark extract showed superior results when compared to the leaf extract, suggesting the influence of the astilbin concentration, higher in the bark, on the anti-inflammatory action. In addition, only the H. eriogyne bark extract was able to reduce MDA, indicating an associated antioxidant effect. Regarding the in vitro antivenom action, the extracts (bark and leaves) revealed the ability to inhibit the proteolytic, phospholipase and hyaluronidase action of both bothropic venom, with a greater effect against B. leucurus venom. In vivo, extracts from the bark and leaves of H. eriogyne (50–200 mg/kg) showed antiedematogenic activity, reducing the release of MPO and pro-inflammatory cytokines, indicating the presence of bioactive components useful in controlling the inflammatory process induced by the venom. In the in silico assays, astilbin and rutin showed reversible interactions of 9 possible positions and orientations towards MPO, with affinities of −9.5 and −10.4 kcal/mol and interactions with Phe407, Gln91, His95 and Arg239, important active pockets of MPO. Rutin demonstrated more effective types of interactions with MPO. ConclusionThis approach reveals for the first time the anti-inflammatory action of H. eriogyne bark and leaf extracts in vivo, as well as its antiophidic potential. Moreover, the distinct effect of pharmacogens as antioxidant agents and distinct effect of astilbin and rutin under MPO sheds light on the different anti-inflammatory mechanisms of bioactive compounds present in H. eriogyne extracts, with high potential for the prospection of new pharmacological agents.

Synovial fibroblasts from children with oligoarticular juvenile idiopathic arthritis induce migration and prolong viability of neutrophils.

Little is known of the processes that trigger neutrophil activation in the joint of patients with oligoarticular juvenile idiopathic arthritis (oJIA), and if synovial fibroblasts (S-Fib) play an important role in the activation. Therefore, we aimed to investigate whether S-Fib derived from oJIA patients drive neutrophil activation. Synovial fluid (SF) was collected from patients with oJIA. S-Fib were isolated from the SF of n = 7 patients through passaging. Subsequently, the S-Fib were primed or not with 20% of pooled SF. Supernatants were used to study migration of neutrophils in a transwell system. Additionally, the influence of S-Fib on neutrophils were studied in co-cultures. Phenotype and viability were assessed by flow cytometry. Neutrophil function was tested through the production of reactive oxygen species (ROS), and supernatants were tested for myeloperoxidase (MPO) release and elastase activity. Supernatants of S-Fib induced neutrophil migration (n = 5, p = 0.0491), which was further pronounced using supernatants from SF-primed S-Fib (p = 0.0063). Additionally, co-culture between SF-primed S-Fib and neutrophils resulted in prolonged viability (n = 5, p = 0.0094), with little effect on activation markers, e.g., CD11b. Conversely, co-culture did not induce functional alterations (n = 4), such as production of ROS (p > 0.1570), release of MPO (p > 0.4934) or elastase activity (p > 0.0904). Finally, supernatant stimulation did not replicate the results of prolonged viability (p = 0.9102), suggesting a role of cell-contact. S-Fib from patients with oJIA induce migration of neutrophils via soluble mediators and, in addition, S-Fib prolong neutrophil viability in a cell-contact dependent manner. These mechanisms could be important for accumulation of neutrophils during arthritis.

Myeloperoxidase as a Promising Therapeutic Target after Myocardial Infarction.

Coronary artery disease (CAD) and myocardial infarction (MI) remain leading causes of death and disability worldwide. CAD begins with the formation of atherosclerotic plaques within the intimal layer of the coronary arteries, a process driven by persistent arterial inflammation and oxidation. Myeloperoxidase (MPO), a mammalian haem peroxidase enzyme primarily expressed within neutrophils and monocytes, has been increasingly recognised as a key pro-inflammatory and oxidative enzyme promoting the development of vulnerable coronary atherosclerotic plaques that are prone to rupture, and can precipitate a MI. Mounting evidence also implicates a pathogenic role for MPO in the inflammatory process that follows a MI, which is characterised by the rapid infiltration of activated neutrophils into the damaged myocardium and the release of MPO. Excessive and persistent cardiac inflammation impairs normal cardiac healing post-MI, resulting in adverse cardiac outcomes and poorer long-term cardiac function, and eventually heart failure. This review summarises the evidence for MPO as a significant oxidative enzyme contributing to the inappropriate inflammatory responses driving the progression of CAD and poor cardiac healing after a MI. It also details the proposed mechanisms underlying MPO's pathogenic actions and explores MPO as a novel therapeutic target for the treatment of unstable CAD and cardiac damage post-MI.

Exploring the role of myeloperoxidase in the atherosclerotic process in hypoxic mice based on the MAPK signaling pathway

Atherosclerosis (AS) is the common pathophysiological basis of various cardiovascular diseases and the leading cause of death from cardiovascular disease worldwide. When the body is in a hypoxic environment, enhanced oxidative stress and significant accumulation of reactive oxygen species (ROS) in tissue cells exacerbate the inflammatory response, resulting in increased release of myeloperoxidase (MPO), catalyzing the formation of large quantities of hypochlorous acid (HOCl), further oxidative modification of low-density lipoprotein (LDL), and exacerbating the formation and progression of atherosclerotic plaques. The MAPK signaling pathway is important in oxidative stress-mediated promotion of atherogenesis. MPO -/- mice were used in this study to establish a hypoxia model simulating 5000 m altitude and a Western high-fat diet-induced atherosclerosis model for 12 weeks. Exploring the role of MPO in the atherosclerotic process in hypoxic mice by observing the MAPK signaling pathway to provide a therapeutic target for the prevention and treatment of hypoxic atherosclerotic disease in the plateau. We found that hypoxia promotes the formation of atherosclerosis in mice, and the mechanism may be that increased MPO in vivo promotes an inflammatory response, which plays a crucial role in the formation of atherosclerosis. In addition, hypoxia further exacerbates plaque instability by activating the MAPK signaling pathway to upregulate vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP9), which in turn promotes angiogenesis within the plaque. Therefore, a potential target for preventing and treating hypoxic atherosclerotic disease is the inhibition of MPO.

Pioglitazone attenuate level of myeloperoxidases and nitic oxide in psoriatic lesion: a proof-of-concept study in a imiquimod induced psoriasis model in rat.

Psoriasis is a persistent autoimmune inflammatory condition that is primarily affecting the skin. Pioglitazone (PGZ), a peroxisome proliferator activated receptor gamma (PPARγ) agonist, has been reported to have anti-inflammatory effects. However, the role of PGZ in psoriatic disease remains unclear. In this study, we aimed to repurpose the use of the PGZ for the treatment of psoriasis. To investigate its efficacy, we employed an imiquimod (IMQ)-induced rat model. Wistar rats are randomly allocated to four different groups. Group, I served as a negative control, Group II IMQ control, Group III was treated with pioglitazone hydrogel and Group IV received standard drug betamethasone cream. PASI score was monitored on every alternative day and on day 7 animals were sacrificed and histopathology of skin was performed. Level of nitric oxide (NO) and myeloperoxidase (MPO) was also performed using established methods. The results of the experiment revealed that treatment with PGZ significantly (p<0.05) reduced redness, scaling, and skin thickening, surpassing the effectiveness of standard drugs. Our result also indicates that PGZ significantly (p<0.05) inhibits the release of both MPO and NO from the psoriatic lesions. PGZ effectively reduces the severity of psoriasis possibly by inhibiting the accumulation of neutrophil at the psoriatic area which indirectly regulates the release of NO in the affected area. Our study showed we can repurpose the PGZ for the management of psoriasis.

Suppressive effect of resveratrol, catechin and their conformationally constrained analogs on neutrophil extracellular trap formation by HL-60-derived neutrophils.

Neutrophil extracellular trap (NET) formation is a unique self-defense mechanism of neutrophils; however, it is also involved in many diseases, including atherosclerosis. Resveratrol and catechin are antioxidants with anti-atherosclerotic properties. Here, we examined the effects of resveratrol, catechin, and other related compounds on NET formation. HL-60-derived neutrophils were pretreated with resveratrol and other compounds before stimulation with phorbol-myristate acetate (PMA). DNA and myeloperoxidase released from neutrophils were determined. Resveratrol suppressed the DNA release from neutrophils in a dose-dependent manner. NET formation was enhanced by 1-palmitoyl-2-oxovaleroyl phosphatidylcholine (POVPC), a truncated form of oxidized phospholipid, and resveratrol suppressed NET formation induced by POVPC and PMA. Furthermore, we designed several analogs of resveratrol or catechin whose conformation was restricted by the inhibition of the free rotation of aromatic rings. The conformationally constrained analogs were more effective at inhibiting NET formation; however, their inhibitory function decreased when compound was a large, hydrophobic analog. The most potent compounds, planar catechin and resveratrol, suppressed myeloperoxidase release from activated neutrophils. In addition, these compounds suppressed DNA release from neutrophils stimulated with calcium ionophore. These results suggest that resveratrol, catechin and their analogs exert anti-NET effects, and that constraining the geometry of these compounds enhanced their inhibitory effects.

Quercetina inibe a liberação das armadilhas extracelulares dos neutrófilos e a atividade tóxica delas sobre células A549

Abstract Neutrophil extracellular traps (NETs) were first reported as a microbicidal strategy for activated neutrophils. Through an immunologic response against several stimuli, neutrophils release their DNA together with proteins from granules, nucleus, and cytoplasm (e.g., elastase and myeloperoxidase). To date, NETs have been implicated in tissue damage during intense inflammatory processes, mainly when their release is dependent on oxygen radical generation. Flavonoids are antioxidant and anti-inflammatory agents; of these, quercetin is commonly found in our daily diet. Therefore, quercetin could exert some protective activity against tissue damage induced by NETs. In our in vitro assays, quercetin reduced NETs, myeloperoxidase (MPO), and elastase release from neutrophils stimulated with phorbol 12-myristate 13-acetate (PMA). The activity of these enzymes also decreased in the presence of quercetin. Quercetin also reduced the cytotoxic effect of NETs on alveolar cells (A549 cell line). Further, in silico assays indicated favorable interactions between quercetin and NET proteins (MPO and elastase). Overall, our results demonstrate that quercetin decreases deleterious cellular effects of NETs by reducing their release from activated neutrophils, and diminishing the enzymatic activity of MPO and elastase, possibly through direct interaction.

Intrahepatic neutrophil accumulation and extracellular trap formation are associated with posthepatectomy liver failure.

Posthepatectomy liver failure (PHLF) represents a life-threatening complication with limited therapeutic options. Neutrophils play a critical and dynamic role during regeneratory processes, but their role in human liver regeneration is incompletely understood, especially as underlying liver disease, detectable in the majority of patients, critically affects hepatic regeneration. Here we explored intrahepatic neutrophil accumulation and neutrophil extracellular traps (NETs) in patients with PHLF and validated the functional relevance of NETs in a murine partial hepatectomy (PHx) model. We investigated the influx of neutrophils, macrophages, eosinophils, and mast cells and the presence of their respective extracellular traps in liver biopsies of 35 patients undergoing hepatectomy (10 patients with PHLF) before and after the initiation of liver regeneration by fluorescence microscopy. In addition, NET formation and neutrophil activation were confirmed by plasma analysis of 99 patients (24 patients with PHLF) before and up to 5 days after surgery. Furthermore, we inhibited NETs via DNase I in a murine PHx model of mice with metabolically induced liver disease. We detected rapid intrahepatic neutrophil accumulation, elevated levels of myeloperoxidase release, and NET formation in regenerating human livers, with a significantly higher increase of infiltrating neutrophils and NETs in patients with PHLF. Circulating markers of neutrophil activation, including elastase, myeloperoxidase, and citrullinated histone H3, correlated with markers of liver injury. In a murine PHx model, we showed that the inhibition of NET accelerated hepatocyte proliferation and liver regeneration. Patients with PHLF showed accelerated intrahepatic neutrophil infiltration and NET formation, which were associated with liver damage. Further, we identified postsurgical myeloperoxidase levels as predictive markers for adverse outcomes and observed that blocking NETs in a murine PHx model accelerated tissue regeneration.

GC/MS and 2D NMR-based approach to evaluate the chemical profile of hydroalcoholic extract from Agaricus blazei Murill and its anti-inflammatory effect on human neutrophils

Ethnopharmacological relevanceAgaricus blazei Murill (AbM) is one of the main mushrooms used for medicinal purposes. The use of AbM in the preparation of teas is widespread mainly in Asian countries, while in Brazil it is used as a functional food to combat inflammatory diseases and cancer. Aim of the studyThe main focus of this study was the characterization of the chemical profile of the hydroalcoholic extract of Agaricus blazei Murill (AbE), as well as the evaluation of its cytotoxic and anti-inflammatory potential using human neutrophils. Materials and methodsThe extract was prepared by dynamic maceration using a mixture of ethanol and water (70/30, v v−1) as solvent. The chemical profile characterization was carried out by 2D NMR and GC-MS techniques. The cytotoxicity of AbE was evaluated through studies of hemolytic potential, cell viability and membrane integrity. The anti-inflammatory activity was analyzed by a PMA-induced neutrophil degranulation assay. ResultsChemical analysis of AbE revealed the presence of 28 metabolites in its composition, with mannitol as the major compound. AbE at 1–200 μg mL−1 and mannitol at 4–160 μg mL−1, showed low hemolytic and cytotoxic potential against human red blood cells and neutrophils. Furthermore, both were able to significantly reduce the release of myeloperoxidase. ConclusionsThese results indicate that AbE is a promising natural product to be incorporated into pharmaceutical dosage forms intended for the adjuvant treatment of inflammatory diseases.

Interactions between Integrin α9β1 and VCAM-1 Promote Neutrophil Hyperactivation and Deep Vein Thrombosis in a Murine Model of Lung Cancer

Background: Venous thromboembolism (VTE) is highly prevalent in patients with lung cancer and is associated with increased morbidity and mortality. Ectopic secretion of granulocyte colony-stimulating factor (G-CSF), a critical regulator of neutrophil trafficking, is commonly observed in lung cancer. In recent years, compelling evidence has emerged implicating neutrophils and neutrophil extracellular traps (NETosis) in cancer-associated thrombosis. Integrin α9β1 is highly expressed on neutrophils, upregulated upon neutrophil activation, promotes NETosis, and is known to stabilize neutrophil adhesion to activated endothelium via multiple ligands including vascular cell adhesion molecule 1 (VCAM-1). Interactions between integrin α9 and VCAM-1 have been reported to reduce neutrophils apoptosis. Given the relevance of integrin α9 in neutrophil migration and apoptosis, granulopoiesis, and thromboinflammation, here we evaluated the mechanistic role of neutrophil integrin α9β1 in the pathophysiology of cancer-associated thrombosis. Methods: Lung cancer was induced by the intravenous administration of 3X10 5 Lewis lung carcinoma cells. Deep vein thrombosis (DVT) was induced in neutrophil-specific α9 -/- (α9 fl/flMrp8Cre +/-) and littermate controls (α9 fl/flMrp8Cre -/-) by partial ligation of the inferior vena cava (IVC). Plasma levels of the thrombin-antithrombin (TAT) complex, fibrinogen, G-CSF, elastase, and myeloperoxidase (MPO) were assessed by ELISA. To evaluate the potential mechanisms by which neutrophil integrin α9-VCAM-1 interactions promote DVT, we performed unbiased RNA sequencing from 1) control neutrophils stimulated with and without VCAM-1 and, 2) neutrophils of α9 fl/flMrp8Cre +/- and α9 fl/flMrp8Cre -/- mice stimulated with VCAM-1. Computational modeling, docking, virtual screening, and the cell-free assay were used to identify small-molecule inhibitors of integrin α9-VCAM-1 interaction. Results: Mice with lung cancer exhibited significantly increased levels of TAT, G-CSF, blood neutrophil/lymphocyte ratio, cell-free DNA, elastase, and MPO, suggesting a hypercoagulable and inflammatory state. Tumor-bearing and non-tumor-bearing neutrophil-specific α9 -/- mice exhibited reduced DVT severity, as evident by reduced IVC thrombus weight and thrombus length compared to littermate controls, while thrombosis incidence was comparable between the groups. Reduced DVT severity in neutrophil-specific α9 -/- mice was also associated with decreased neutrophil content and citrullinated histone H3 (CitH3) expression in IVC thrombi. Integrin α9 deficient neutrophils incubated with VCAM-1 exhibited reduced release of elastase and MPO, decreased endothelial cell apoptosis, and reduced adhesion to the activated endothelium at venous shear rate compared to control neutrophils incubated with VCAM-1. Bioinformatic analysis of RNA sequencing data from VCAM-1 stimulated neutrophils from α9 fl/flMrp8Cre +/- and control mice showed distinct transcriptional profiles. Hierarchical clustering of gene expression revealed gene clusters that were highly downregulated in stimulated neutrophils from α9 fl/flMrp8Cre +/- mice. Gene ontology enrichment revealed that highly downregulated genes corresponded to biological processes involved in the inflammatory response, cytokine production, NF-κB and ERK signaling pathways, granulocyte migration, and chemotaxis. Virtual screening and cell-free assay identified macitentan as a potent inhibitor of integrin α9-VCAM-1 interactions. In vitro treatment with macitentan reduced neutrophil adhesion to VCAM-1 and also to activated endothelial cells at venous shear rate. In vivo, macitentan treatment at a dose of 5 mg/kg exhibited reduced DVT severity, decreased plasma levels of elastase and MPO, and reduced neutrophil and CitH3 levels in IVC thrombi. Conclusion: Using genetic and pharmacological approaches in combination with transcriptomic and pharmacological studies, we described a previously unknown pathway involving integrin α9 and VCAM-1 in promoting cancer-associated thrombosis. We found that integrin α9-VCAM1 axis is critical for neutrophil hyperactivation, and its pharmacological inhibition significantly reduces DVT severity. The results of this study suggest that neutrophil integrin α9 is a key factor regulating neutrophil hyperactivation and cancer-associated thrombosis.

Effects of photobiomodulation in the experimental acetic acid-induced colitis: comparison between male and female.

Ulcerative colitis (UC) is an important chronic and multifactorial disease, which alters the colon mucosal with a significant impact on life quality affecting both men and women. The difference between genders causes changes in the inflammatory processes, modulating the development of several diseases. The available drugs to treat UC exhibit limited outcomes and side effects; thus, new therapies are needed. Photobiomodulation (PBM) emerges as potential treatment by modulating the inflammatory process without side effects and low costs. The aim of this study was to evaluate the effects of PBM in acetic acid-induced UC comparing the responses between male and females. For this purpose, male and female Wistar rats (36) were submitted to induction of UC by rectal administration of 10% acetic acid (colitis group) and treated or not with PBM (colitis-PBM group) (LED, 660nm, 100 mW, 150s) in three points: right side and left of the ventral surface and in the external anal region. Non-manipulated rats were used as control (basal group). We investigated the disease activity index (DAI score), myeloperoxidase enzyme activity (MPO) and release of cytokines in the intestine homogenates, and histological analysis. PBM reduces DAI score, MPO activity, and mast cell degranulation while increased mucous production in both females and males. Moreover, PBM reduced histopathological score as well as the levels of IL-6 and IL-4 in the bowel only in males. We also showed reduced levels of IL-1beta and TNF-alpha after PBM in both males and females, while the levels of IL-10 and IFN-gamma were increased. In conclusion, despite our study has shown some differences between males and females, PBM attenuated the biomarkers of UC in both genders constituting a potential combined treatment that is non-invasive and low cost.

Multi-Oxidant Environment as a Suicidal Inhibitor of Myeloperoxidase.

Tissue inflammation drives the infiltration of innate immune cells that generate reactive species to kill bacteria and recruit adaptive immune cells. Neutrophil activation fosters the release of myeloperoxidase (MPO) enzyme, a heme-containing protein generating hypochlorous acid (HOCl) from hydrogen peroxide (H2O2) and chloride ions. MPO-dependent oxidant formation initiates bioactive oxidation and chlorination products and induces oxidative post-translational modifications (oxPTMs) on proteins and lipid oxidation. Besides HOCl and H2O2, further reactive species such as singlet oxygen and nitric oxide are generated in inflammation, leading to modified proteins, potentially resulting in their altered bioactivity. So far, knowledge about multiple free radical-induced modifications of MPO and its effects on HOCl generation is lacking. To mimic this multi-oxidant microenvironment, human MPO was exposed to several reactive species produced simultaneously via argon plasma operated at body temperature. Several molecular gas admixes were used to modify the reactive species type profiles generated. MPO was investigated by studying its oxPTMs, changes in protein structure, and enzymatic activity. MPO activity was significantly reduced after treatment with all five tested plasma gas conditions. Dynamic light scattering and CD-spectroscopy revealed altered MPO protein morphology indicative of oligomerization. Using mass spectrometry, various oxPTMs, such as +1O, +2O, and +3O, were determined on methionine and cysteine (Cys), and -1H-1N+1O was detected in asparagine (Asp). The modification types identified differed between argon-oxygen and argon-nitrogen plasmas. However, all plasma gas conditions led to the deamidation of Asp and oxidation of Cys residues, suggesting an inactivation of MPO due to oxPTM-mediated conformational changes.

Glucosinolate-rich broccoli sprouts protect against oxidative stress and improve adaptations to intense exercise training

Oxidative stress plays a vital role for the adaptive responses to physical training. However, excessive oxidative stress can precipitate cellular damage, necessitating protective mechanisms to mitigate this effect. Glucosinolates, found predominantly in cruciferous vegetables, can be converted into isothiocyanates, known for their antioxidative properties. These compounds activate crucial antioxidant defence pathways and support mitochondrial function and protein integrity under oxidative stress, in both Nrf2-dependent and independent manners. We here administered glucosinolate-rich broccoli sprouts (GRS), in a randomized double-blinded cross-over fashion to 9 healthy subjects in combination with daily intense exercise training for 7 days. We found that exercise in combination with GRS significantly decreased the levels of carbonylated proteins in skeletal muscle and the release of myeloperoxidase into blood. Moreover, it lowered lactate accumulation during submaximal exercise, and attenuated the severe nocturnal hypoglycaemic episodes seen during the placebo condition. Furthermore, GRS in combination with exercise improved physical performance, which was unchanged in the placebo condition.

Myeloperoxidase is a critical mediator of anthracycline-induced cardiomyopathy

Cardiotoxicity is a major complication of anthracycline therapy that negatively impacts prognosis. Effective pharmacotherapies for prevention of anthracycline-induced cardiomyopathy (AICM) are currently lacking. Increased plasma levels of the neutrophil-derived enzyme myeloperoxidase (MPO) predict occurrence of AICM in humans. We hypothesized that MPO release causally contributes to AICM. Mice intravenously injected with the anthracycline doxorubicin (DOX) exhibited higher neutrophil counts and MPO levels in the circulation and cardiac tissue compared to saline (NaCl)-treated controls. Neutrophil-like HL-60 cells exhibited increased MPO release upon exposition to DOX. DOX induced extensive nitrosative stress in cardiac tissue alongside with increased carbonylation of sarcomeric proteins in wildtype but not in Mpo−/− mice. Accordingly, co-treatment of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with DOX and MPO aggravated loss of hiPSC-CM-contractility compared to DOX treatment alone. DOX-treated animals exhibited pronounced cardiac apoptosis and inflammation, which was attenuated in MPO-deficient animals. Finally, genetic MPO deficiency and pharmacological MPO inhibition protected mice from the development of AICM. The anticancer efficacy of DOX was unaffected by MPO deficiency. Herein we identify MPO as a critical mediator of AICM. We demonstrate that DOX induces cardiac neutrophil infiltration and release of MPO, which directly impairs cardiac contractility through promoting oxidation of sarcomeric proteins, cardiac inflammation and cardiomyocyte apoptosis. MPO thus emerges as a promising pharmacological target for prevention of AICM.

Functional immunophenotyping of blood neutrophils identifies novel endotypes of viral response in preschool children with recurrent wheezing

Preschool children with recurrent wheezing are heterogeneous, with differing responses to respiratory viral infections. Although neutrophils are crucial for host defense, their function has not been studied in this population. We performed functional immunophenotyping on isolated blood neutrophils from 52 preschool children with recurrent wheezing (aeroallergen sensitization, n= 16; no sensitization, n= 36). Blood neutrophils were purified and cultured overnight with polyinosinic:polycytidylic acid [poly(I:C)] as a viral analog stimulus. Neutrophils underwent next-generation sequencing with Reactome pathway analysis and were analyzed for cytokine secretion, apoptosis, myeloperoxidase, and extracellular DNA release. CD14+ monocytes were also exposed to neutrophil culture supernatant and analyzed for markers of M1 and M2 activation. A total of 495 genes, related largely to the innate immune system and neutrophil degranulation, were differently expressed in children with versus without aeroallergen sensitization. Functional experiments identified more neutrophil degranulation and extracellular trap formation (ie, more myeloperoxidase and extracellular DNA) and less neutrophil proinflammatory cytokine secretion in children with aeroallergen sensitization. Neutrophils also shifted CD14+ monocytes to a more anti-inflammatory (ie, M2) phenotype in sensitized children and a more proinflammatory (ie, M1) phenotype in nonsensitized children. Although both groups experienced viral exacerbations, annualized exacerbation rates prompting unscheduled health care were also higher in children without aeroallergen sensitization after enrollment. Systemic neutrophil responses to viral infection differ by allergic phenotype and may be less effective in preschool children without allergic inflammation. Further studies of neutrophil function are needed in this population, which often has less favorable therapeutic responses to inhaled corticosteroids and other therapies directed at type 2-high inflammation.

Reduction of neutrophil extracellular traps accelerates inflammatory resolution and increases bone formation on titanium implants.

Neutrophils are the most abundant immune cells in the blood and the first cells to be recruited to the biomaterial implantation site. Neutrophils are fundamental in recruiting mononuclear leukocytes to mount an immune response at the injury site. Neutrophils exert significant pro-inflammatory effects through the release of cytokines and chemokines, degranulation and release of myeloperoxidase (MPO) and neutrophil elastase (NE), and the production of large DNA-based networks called neutrophil extracellular traps (NETs). Neutrophils are initially recruited and activated by cytokines and pathogen- and damage-associated molecular patterns, but little is known about how the physicochemical composition of the biomaterial affects their activation. This study aimed to understand how ablating neutrophil mediators (MPO, NE, NETs) affected macrophage phenotype in vitro and osseointegration in vivo. We discovered that NET formation is a crucial mediator of pro-inflammatory macrophage activation, and inhibition of NET formation significantly suppresses macrophage pro-inflammatory phenotype. Furthermore, reducing NET formation accelerated the inflammatory phase of healing and produced greater bone formation around the implanted biomaterial, suggesting that NETs are essential regulators of biomaterial integration. Our findings emphasize the importance of the neutrophil response to implanted biomaterials and highlight innate immune cells' regulation and amplification signaling during the initiation and resolution of the inflammatory phase of biomaterial integration. STATEMENT OF SIGNIFICANCE: Neutrophils are the most abundant immune cells in blood and are the first to be recruited to the injury/implantation site where they exert significant pro-inflammatory effects. This study aimed to understand how ablating neutrophil mediators affected macrophage phenotype in vitro and bone apposition in vivo. We found that NET formation is a crucial mediator of pro-inflammatory macrophage activation. Reducing NET formation accelerated the inflammatory phase of healing and produced greater appositional bone formation around the implanted biomaterial, suggesting that NETs are essential regulators of biomaterial integration.

Human Whole Blood Interactions with Craniomaxillofacial Reconstruction Materials: Exploring In Vitro the Role of Blood Cascades and Leukocytes in Early Healing Events.

The present study investigated early interactions between three alloplastic materials (calcium phosphate (CaP), titanium alloy (Ti), and polyetheretherketone (PEEK) with human whole blood using an established in vitro slide chamber model. After 60 min of contact with blood, coagulation (thrombin-antithrombin complexes, TAT) was initiated on all test materials (Ti > PEEK > CaP), with a significant increase only for Ti. All materials showed increased contact activation, with the KK-AT complex significantly increasing for CaP (p < 0.001), Ti (p < 0.01), and PEEK (p < 0.01) while only CaP demonstrated a notable rise in KK-C1INH production (p < 0.01). The complement system had significant activation across all materials, with CaP (p < 0.0001, p < 0.0001) generating the most pronounced levels of C3a and sC5b-9, followed by Ti (p < 0.001, p < 0.001) and lastly, PEEK (p < 0.001, p < 0.01). This activation correlated with leukocyte stimulation, particularly myeloperoxidase release. Consequently, the complement system may assume a more significant role in the early stages post implantation in response to CaP materials than previously recognized. Activation of the complement system and the inevitable activation of leukocytes might provide a more favorable environment for tissue remodeling and repair than has been traditionally acknowledged. While these findings are limited to the early blood response, complement and leukocyte activation suggest improved healing outcomes, which may impact long-term clinical outcomes.