Redox-dependent plasticity of oxMIF facilitates its interaction with CD74 and therapeutic antibodies.

MIF is a ubiquitous protein involved in proinflammatory processes, which undergoes an oxidation-driven conformational change to oxidized (ox)MIF. We demonstrate that hypochlorous acid, produced by neutrophil-released myeloperoxidase (MPO) under inflammatory conditions, effectively oxidizes MIF into the oxMIF isoform, which is specifically recognized by the anti-oxMIF therapeutic antibody, ON104. NMR investigation of MIF oxidized by the MPO system revealed increased flexibility throughout the MIF structure, including at several catalytic and allosteric sites. Mass spectrometry of MPO-oxMIF revealed methionines as the primary site of oxidation, whereas Pro2 and Tyr99/100 remained almost unmodified. ELISA, SPR and cell-based assays demonstrated that structural changes caused by MPO-driven oxidation promoted binding of oxMIF to its receptor, CD74, which does not occur with native MIF. These data reveal the environment and modifications that facilitate interactions between MIF and its pro-inflammatory receptor, and a route for therapeutic intervention targeting the oxMIF isoform.

Integrating network pharmacology and experimental evaluation to explore the complementary therapeutic effect and mechanism of melatonin in periodontitis.

Objective: To explore the potential targets for melatonin in the treatment of periodontitis through network pharmacologic analysis and experimental validation via in vivo animal models and in vitro cellular experiments. Materials and methods: In this study, we first screened melatonin targets from Pharm Mapper for putative targets, Drug Bank, and TCMSP databases for known targets. Then, disease database was searched and screened for differential expressed genes associated with periodontitis. The intersection of disease and melatonin-related genes yielded potential target genes of melatonin treatment for periodontitis. These target genes were further investigated by protein-protein interaction network and GO/KEGG enrichment analysis. In addition, the interactions between melatonin and key target genes were interrogated by molecular docking simulations. Then, we performed animal studies to validate the therapeutic effect of melatonin by injecting melatonin into the peritoneal cavity of ligation-induced periodontitis (LIP) mice. The effects of melatonin on the predicted target proteins were also analyzed using Western blot and immunofluorescence techniques. Finally, we constructed an in vitro cellular model and validated the direct effect of melatonin on the predicted targets by using qPCR. Results: We identified 8 potential target genes by network pharmacology analysis. Enrichment analysis suggests that melatonin may treat periodontitis by inhibiting the expression of three potential targets (MPO, MMP8, and MMP9). Molecular docking results showed that melatonin could effectively bind to MMP8 and MMP9. Subsequently, melatonin was further validated in a mouse LIP model to inhibit the expression of MPO, MMP8, and MMP9 in the periodontal tissue. Finally, we verified the direct effect of melatonin on the mRNA expression of MPO, MMP8, and MMP9 in an in vitro cellular model. Conclusions: Through a combination of network pharmacology and experimental validation, this study provides a more comprehensive understanding of the mechanism of melatonin to treat periodontitis. Our study suggests that MPO, MMP8, and MMP9 as key target genes of melatonin to treat periodontitis. These findings present a more comprehensive basis for further investigation into the mechanisms of pharmacological treatment of periodontitis by melatonin.

Croton stipulaceus Kunth, a native Mexican medicinal plant with antioxidant and anti-inflammatory activities / Croton stipulaceus Kunth, planta medicinal mexicana con actividad antioxidante y antiinflamatoria

Leaves of Croton stipulaceuswere extracted (EHex, ECHCl3and EEtOH extracts) to assesstheir antioxidant potential, anti-inflammatory activity in murine models and acute toxicity. EEtOH showed the highest effect in DPPH (37.80% inhibition), FRAP (1065.00 ± 55.30 µmolFe2+) and total polyphenols (231.24 ± 9.05 meq AG/gM). EHex was the most active, ~ 50% inhibition of TPA-induced ear edema; while EEtOH (dose of 2 mg/ear) showed the highest inhibition in the chronic model (97% inhibition), and inhibited MPO activity (48%). In carrageenan-induced edema, ECHCl3(dose 500 mg/kg) was the most active. None of the extracts showed acute toxicity (LD50) at 2 g/kg (p.o.). This work is the first report that supports the traditional use of C. stipulaceusas an anti-inflammatory.

De las hojas de Croton stipulaceusse obtuvieron diferentes extractos (EHex, ECHCl3y EEtOH) evaluando el potencial antioxidante y la actividad antiinflamatoria en modelos murinos y la toxicidad aguda. El EEtOH mostró mayor efecto en DPPH (37.80% inhibición), FRAP (1065.00 ± 55.30 µmolFe2+) y polifenolestotales (231.24 ± 9.05 meq AG/gM). El EHex fue el más activo, cercano al 50% de inhibición del edema auricular inducido con TPA; mientras que el EEtOH (dosis de 2 mg/oreja) mostró la mayor inhibición en el modelo crónico (97% inhibición), e inhibió la actividad de la MPO (48%). En el edema inducido con carragenina, el ECHCl3(dosis 500 mg/kg) fue el más activo. Ninguno de los extractos mostró una toxicidad aguda (DL50) mayor a 2 g/kg (p.o). Este trabajo es el primer reporte que sustenta el uso tradicional de C. stipulaceuscomo antiinflamatorio.

Active myeloperoxidase: a promising biomarker to differentiate "acute" and "low-grade" peri-prosthetic joint infections from aseptic failures.

Introduction: The accurate distinction between periprosthetic joint infections (PJI) and aseptic failures (AF) is of paramount importance due to differences in treatment. However, this could be challenging by using the current criteria. Various synovial fluid biomarkers are being assessed to improve the diagnostic accuracy. Myeloperoxidase (MPO), an enzyme contained in the granules of neutrophils, may be a promising biomarker for PJI. Methods: Synovial fluids of 99 patients (n = 65 PJI according to EBJIS criteria; n = 34 AF) were collected in two specialized orthopedic centers. PJI were divided into acute (n = 33) and low-grade (n = 32) according to previously published classification. An activity assay specific for active MPO was performed in each sample. Ability of MPO to correctly discriminate patients with PJI from AF was determined by ROC analysis. The best discriminating cut-off value was determined by calculating the J Youden index. For all analyses, a P value < 0.05 was considered statistically significant. Results: Active MPO was higher in PJI than AF (P < 0.0001). The ROC analysis revealed a significant area under the curve (AUC: 0.86; 95% CI: 0.78-0.93, P < 0.0001). A cut-off value of 561.9 U/mL, with good sensitivity (0.69) and specificity (0.88), discriminated between AF and PJI (accuracy 75.76%, 95% CI: 66.11-83.81%, positive likelihood ratio 5.88, 95% CI: 2.31-14.98 and negative likelihood ratio 0.35, 95%CI: 0.24-0.51). No difference in MPO levels was found between acute and chronic low-grade PJI. Conclusion: The proposed assay appears to be a reliable and affordable tool for detecting the active MPO in synovial fluid, with promising characteristics of sensitivity and specificity in discriminating both acute and low-grade PJI from AF. Further studies are needed to confirm MPO diagnostic cut-off values and validate their use in the routine clinical practice.

Association between metabolic syndrome and salivary MMP-8, myeloperoxidase in periodontitis.

OBJECTIVE: This study investigated the effect of metabolic syndrome (MetS) on periodontal clinical parameters and salivary biomarkers' matrix metalloproteinase-8 (MMP-8) and myeloperoxidase (MPO) in patients with periodontitis. METHODS: A total of 120 participants aged 25-55 were categorized into three groups: MetS with periodontitis (n = 40); systemically healthy with periodontitis (n = 40); and systemically and periodontally healthy controls (n = 40). Data collected included systemic parameters like waist circumference (WC), blood pressure (BP), high- and low-density lipoproteins, triglycerides (TG), fasting blood sugar (FBS), and glycated hemoglobin (HbA1c). Periodontal parameters estimated included bleeding on probing score (BoP), full-mouth plaque score (FMPS), periodontal probing depth (PPD), clinical attachment loss (CAL), and the number of missing teeth. Unstimulated whole saliva was analyzed via ELISA for active MMP-8 (aMMP-8), total MMP-8 (tMMP-8), and MPO. RESULTS: Participants with MetS and periodontitis exhibited significantly higher periodontal parameters, salivary aMMP-8, and MPO (26.26 vs. 24.1 ng/mL and 13.53 vs. 11.55 ng/mL compared to systemically healthy periodontitis patients) (all p < 0.01). Positive correlations occurred between aMMP-8 and WC, TG, and FBS (p < 0.01), and between MPO and WC, BP, and TG (p < 0.01). CONCLUSIONS: The positive associations between these biomarkers and metabolic parameters indicate their potential utility for monitoring cardiovascular and glycemic risk in patients with periodontal disease.

Very Slowly Progressive Microscopic Polyangiitis: Comparative Analysis with Rapidly Progressive Forms.

Microscopic polyangiitis (MPA) is predominantly characterized by rapidly progressive glomerulonephritis (RPGN) associated with myeloperoxidase anti-neutrophil cytoplasmic antibodies (MPO-ANCA). Nonetheless, up to 30% of cases of ANCA-associated vasculitis (AAV) may exhibit a more indolent progression toward renal failure, an aspect less frequently discussed and understood in medical literature. This study seeks to clarify the clinical and pathological distinctions between the slowly and rapidly progressive forms of MPA, thereby enhancing understanding of their distinct pathogeneses and treatment responses. We conducted a comparative analysis of two patients diagnosed with MPA under the 2022 American College of Rheumatology/the European Alliance of Associations for Rheumatology (ACR/EULAR) classification. Evaluations included laboratory tests such as serum creatinine levels, serology for MPO-ANCA, and renal biopsies. Patient 1 exhibited a mere 1.07% decrease in estimated glomerular filtration rate (eGFR) over 6 months, significantly below the RPGN threshold, and demonstrated sclerotic glomerular pathology without active inflammation. This patient also showed lower levels of MPO-ANCA, Birmingham Vasculitis Activity Score (BVAS), and C-reactive protein. Conversely, Patient 2 experienced an 89.9% reduction in eGFR over the same timeframe, accompanied by acute systemic inflammation. The comparative clinical analysis of these cases illuminates clear differences in disease activity. Slowly progressive MPA is marked by lesser disease activity that fosters chronic inflammation, leading to a more gradual decline in renal function. Early diagnosis, facilitated by initial measurements of MPO-ANCA, can enhance disease management and improve patient outcomes.

Regulatory T cells effectively downregulate the autoimmune anti-MPO response and ameliorate anti-MPO induced glomerulonephritis in mice.

Regulation of autoreactive cells is key for both prevention and amelioration of autoimmune disease. A better understanding of the key cell population(s) responsible for downregulation of autoreactive cells would provide necessary foundational insight for cellular-based therapies in autoimmune disease. Utilizing a mouse model of anti-myeloperoxidase (MPO) glomerulonephritis, we sought to understand which immune cells contribute to downregulation of the anti-MPO autoimmune response. MPO-/- mice were immunized with whole MPO to induce an anti-MPO response. Anti-MPO splenocytes were then transferred into recipient mice (Rag2-/- mice or WT mice). Anti-MPO titers were followed over time. After anti-MPO splenocyte transfer, WT mice are able to downregulate the anti-MPO response while anti-MPO titers persist in Rag2-/- recipients. Reconstitution with WT splenocytes into Rag2-/- recipients prior to anti-MPO splenocyte transfer enabled mice to downregulate the anti-MPO immune response. Therefore, wildtype splenocytes contain a cellular population that is capable of downregulating the autoimmune response. Through splenocyte transfer, antibody depletion experiments, and purified cell population transfers, we confirmed that the regulatory T cell (Treg) population is responsible for the downregulation of the anti-MPO autoimmune response. Further investigation revealed that functional Tregs from WT mice are capable of downregulating anti-MPO antibody production and ameliorate anti-MPO induced glomerulonephritis. These data underscore the importance of functional Tregs for control of autoimmune responses and prevention of end-organ damage due to autoimmunity.

NOX4 and its association with myeloperoxidase and osteopontin in regulating endochondral ossification.

IMPORTANCE: Endochondral ossification plays an important role in skeletal development. Recent studies have suggested a link between increased intracellular reactive oxygen species (ROS) and skeletal disorders. Moreover, previous studies have revealed that increasing the levels of myeloperoxidase (MPO) and osteopontin (OPN) while inhibiting NADPH oxidase 4 (NOX4) can enhance bone growth. This investigation provides further evidence by showing a direct link between NOX4 and MPO, OPN in bone function. OBJECTIVE: This study investigates NOX4, an enzyme producing hydrogen peroxide, in endochondral ossification and bone remodeling. NOX4's role in osteoblast formation and osteogenic signaling pathways is explored. METHODS: Using NOX4-deficient (NOX4-/-) and ovariectomized (OVX) mice, we identify NOX4's potential mediators in bone maturation. RESULTS: NOX4-/- mice displayed significant differences in bone mass and structure. Compared to the normal Control and OVX groups. Hematoxylin and eosin staining showed NOX4-/- mice had the highest trabecular bone volume, while OVX had the lowest. Proteomic analysis revealed significantly elevated MPO and OPN levels in bone marrow-derived cells in NOX4-/- mice. Immunohistochemistry confirmed increased MPO, OPN, and collagen II (COLII) near the epiphyseal plate. Collagen and chondrogenesis analysis supported enhanced bone development in NOX4-/- mice. CONCLUSIONS AND RELEVANCE: Our results emphasize NOX4's significance in bone morphology, mesenchymal stem cell proteomics, immunohistochemistry, collagen levels, and chondrogenesis. NOX4 deficiency enhances bone development and endochondral ossification, potentially through increased MPO, OPN, and COLII expression. These findings suggest therapeutic implications for skeletal disorders.

Staphylococcal peroxidase inhibitor (SPIN): Investigation of the inhibitory N-terminal domain via a stabilizing disulfide insertion.

Staphylococcus aureus secretes an array of small proteins that inhibit key enzyme-catalyzed reactions necessary for proper function of the human innate immune system. Among these, the Staphylococcal Peroxidase Inhibitor, SPIN, blocks the activity of myeloperoxidase (MPO) and thereby disrupts the HOCl-generating system of neutrophils. Previous studies on S. aureus SPIN have shown that it relies on a C-terminal α-helical bundle domain to mediate initial binding to MPO, but requires a disordered N-terminal region to fold into a ß-hairpin conformation to inhibit MPO activity. To further investigate the structure/function relationship of SPIN, we introduced two cysteine residues into its N-terminal region to trap SPIN in its MPO-bound conformation and characterized the modified protein, which we refer to here as SPIN-CYS. Although control experiments confirmed the presence of the disulfide bond in SPIN-CYS, solution structure determination revealed that the N-terminal region of SPIN-CYS adopted a physically constrained series of lariat-like structures rather than a well-defined ß-hairpin. Nevertheless, SPIN-CYS exhibited a gain in inhibitory potency against human MPO when compared to wild-type SPIN. This gain of function persisted even in the presence of deleterious mutations within the C-terminal α-helical bundle domain. Surface plasmon resonance studies showed that the gain in potency arose through an increase in apparent affinity of SPIN-CYS for MPO, which was driven primarily by an increased association rate with MPO when compared to wild-type SPIN. Together, this work provides new information on the coupled binding and folding events required to manifest biological activity of this unusual MPO inhibitor.

The mitochondrial TSPO ligand Atriol mitigates metabolic-associated steatohepatitis by downregulating CXCL1.

BACKGROUND AND AIMS: The mitochondrial translocator protein (TSPO, 18 kDa) is pivotal in binding cholesterol and facilitating its transfer from the outer to the inner mitochondrial membrane. Atriol is a TSPO ligand disrupting cholesterol binding by targeting the cholesterol-recognition amino acid consensus domain. Prior research has shown that TSPO deficiency improved metabolic-associated steatohepatitis (MASH). We hypothesized that Atriol may have the potential to alleviate MASH. METHODS AND RESULTS: In vitro cell culture studies revealed that Atriol treatment effectively mitigated MASH by restoring mitochondrial function, inhibiting the NF-κB signaling pathway, and reducing hepatic stellate cell (HSC) activation. SD male rats were fed a GAN diet for 10 months to induce MASH. During the final two weeks of feeding, rats received intraperitoneal Atriol administration daily. Atriol treatment significantly ameliorated MASH by reducing lipid accumulation, diminishing hepatic lobular inflammation and fibrosis, decreasing cell death, and inhibiting excessive bile acid synthesis. Moreover, Atriol restored mitochondrial function in primary hepatocytes isolated from MASH rats. In search of the mechanism(s) governing these effects, we found that Atriol downregulated the proinflammatory chemokine CXCL1 through the NF-κB signaling pathway or via myeloperoxidase (MPO) in HSCs and Kupffer cells. Additionally, in vitro, studies further suggested that CXCL1 treatment induced dysfunctional mitochondria, inflammation, HSCs activation, and macrophage migration, whereas Atriol countered these effects. Finally, the mitigating effects of Atriol on MASH were reproduced by pharmacological inhibition of NF-κB or MPO and neutralization of CXCL1. CONCLUSION: Atriol ameliorates MASH both in vitro and in vivo, demonstrating its potential therapeutic benefits in managing MASH.

The Therapeutic Potential of Neutrophil Extracellular Traps and NLRP3 Inflammasomes in Mycoplasma pneumoniae Pneumonia.

INTRODUCTION: Mycoplasma pneumoniae (MP) is the most common pathogen of community-acquired pneumonia in children. However, the role of neutrophil extracellular traps (NETs) in the pathogenesis of MP is unclear. METHODS: Both the level of NETs were detected between the 60 MP pneumonia patients and 20 healthy controls, whose the clinical characteristics were compared. Additionally, NETs formation induced by community-acquired respiratory distress syndrome (CARDS) toxin was also analyzed through transcriptome sequencing. RESULTS: The levels of cell-free DNA, Cit-H3, and MPO-DNA complexes were significantly increased in the patients with MP pneumonia. Importantly, both cell-free DNA and LDH were higher in hospitalized patients with severity than those without severity. In addition, CARDS toxin induced the NETs formation in vitro and in vivo. Transcriptomics GO and KEGG pathway analysis indicate that NOD like receptor signaling pathway and Toll-like receptor signaling pathway are significantly enriched. Finally, we found that DNase I significantly attenuated the higher levels of Cit-H3, and up-regulation of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) by down-regulating the expression of NLRP3 and Caspase1(p20) in the lung tissues. DISCUSSION: These results indicate that inhibiting excessive activation of NLRP3 inflammasomes, and NETs formation may alleviate MP pneumonia.

Identification of biomarkers in multiple myeloma: A comprehensive study combining microarray analysis and Mendelian randomization.

Despite remarkable advancements in the treatment of multiple myeloma (MM), relapse remains a challenge. However, the mechanisms underlying this disease remain unclear. This study aimed to identify potential biomarkers that could open new avenues for MM treatment. Microarray data and clinical characteristics of patients with MM were obtained from the Gene Expression Omnibus database. Differential expression analysis and protein-protein interaction (PPI) network construction were used to identify hub genes associated with MM. Predictive performance was further assessed using receiver operating characteristic curves and nomogram construction. Functional enrichment analysis was conducted to investigate possible mechanisms. Mendelian randomization (MR) was used to evaluate the causal relationship between the crucial gene and MM risk. Topological analysis of the PPI network revealed five hub genes associated with MM, with myeloperoxidase (MPO) being the key gene owing to its highest degree and area under the curve values. MPO showed significant differences between patients with MM and controls across all datasets. Functional enrichment analysis revealed a strong association between MPO and immune-related pathways in MM. MR analysis confirmed a causal relationship between MPO and the risk of MM. By integrating microarray analysis and MR, we successfully identified and validated MPO as a promising biomarker for MM that is potentially implicated in MM pathogenesis and progression through immune-related pathways.

Plasma levels of bactericidal/permeability-increasing protein correlate with systemic inflammation in acute coronary syndrome.

Background: Neutrophils play important roles in atherosclerosis and atherothrombosis. Bactericidal/permeability-increasing protein (BPI) is mainly expressed in the granules of human neutrophils in response to inflammatory stress. This observational, cross-sectional study investigated the plasma level of BPI in patients with acute coronary syndrome (ACS) and its correlation with blood neutrophil counts and circulating inflammatory biomarkers. Methods: A total of 367 patients who had acute chest pain and who were admitted to our hospital for coronary angiography (CAG) and/or percutaneous coronary intervention (PCI) from May 1, 2020 to August 31, 2020 were recruited. Among them, 256 had a cardiac troponin value above the 99th percentile upper reference limit and were diagnosed with ACS. The remaining patients (n = 111) were classified as non-ACS. The TIMI and GRACE scores were calculated at admission. The Gensini score based on CAG was used to determine atherosclerotic burden. Plasma levels of interleukin (IL)-1ß, myeloperoxidase-DNA (MPO-DNA), high sensitivity C-reactive protein (hs-CRP), S100A8/A9, and BPI were measured using enzyme-linked immunosorbent assays. Correlations of plasma BPI levels with examination scores and levels of circulating inflammatory biomarkers were explored. Receiver operating characteristic (ROC) curve analysis was used to determine the diagnostic efficacy of BPI for ACS and myocardial infarction. Results: Patients in the ACS group showed significantly higher plasma BPI levels compared to the non-ACS group (46.42 ± 16.61 vs. 16.23 ± 6.19 ng/mL, p < 0.05). Plasma levels of IL-1ß, MPO-DNA, hs-CRP, and S100A8/A9 in the ACS group were also significantly higher than those in the non-ACS group (all p < 0.05). In addition, plasma BPI levels were positively correlated with the TIMI, GRACE, and Gensini scores (r = 0.176, p = 0.003; r = 0.320, p < 0.001; r = 0.263, p < 0.001, respectively) in patients with ACS. Plasma BPI levels were also positively correlated with blood neutrophil counts (r = 0.266, p < 0.001) and levels of circulating inflammatory biomarkers (IL-1ß, r = 0.512; MPO-DNA, r = 0.452; hs-CRP, r = 0.554; S100A8/A9, r = 0.434; all p < 0.001) in patients with ACS. ROC curve analysis revealed that the diagnostic efficacy of BPI for ACS was not inferior to that of IL-1ß, MPO-DNA, hs-CRP, S100A8/A9, or blood neutrophil counts. ROC analysis also showed that the diagnostic efficacy of BPI for myocardial infarction was not inferior to that of creatine kinase (CK)-MB or cardiac troponin I. Conclusion: BPI is associated with systemic inflammation in ACS and may be involved in the process of atherosclerosis and atherothrombosis. The potential of BPI as a prognostic and diagnostic biomarker for ACS should be investigated in clinical settings.

Modification of extracellular matrix proteins by oxidants and electrophiles.

The extracellular matrix (ECM) is critical to biological architecture and determines cellular properties, function and activity. In many situations it is highly abundant, with collagens and elastin being some of the most abundant proteins in mammals. The ECM comprises of multiple different protein species and sugar polymers, with both different isoforms and post-translational modifications (PTMs) providing a large variety of microenvironments that play a key role in determining tissue structure and health. A number of the PTMs (e.g. cross-links) present in the ECM are critical to integrity and function, whereas others are deleterious to both ECM structure and associated cells. Modifications induced by reactive oxidants and electrophiles have been reported to accumulate in some ECM with increasing age. This accumulation can be exacerbated by disease, and in particular those associated with acute or chronic inflammation, obesity and diabetes. This is likely to be due to higher fluxes of modifying agents in these conditions. In this focused review, the role and effects of oxidants and other electrophiles on ECM are discussed, with a particular focus on the artery wall and atherosclerotic cardiovascular disease. Modifications generated on ECM components are reviewed, together with the effects of these species on cellular properties including adhesion, proliferation, migration, viability, metabolic activity, gene expression and phenotype. Increasing data indicates that ECM modifications are both prevalent in human and mammalian tissues and play an important role in disease development and progression.

Neutrophil extracellular trap-associated protein in cerebrospinal fluid for prognosis evaluation of adult bacterial meningitis: a retrospective case-control study.

BACKGROUND AND OBJECTIVES: Central nervous system infections, typified by bacterial meningitis, stand as pivotal emergencies recurrently confronted by neurologists. Timely and precise diagnosis constitutes the cornerstone for efficacious intervention. The present study endeavors to scrutinize the influence of inflammatory protein levels associated with neutrophils in cerebrospinal fluid on the prognosis of central nervous system infectious maladies. METHODS: This retrospective case series study was undertaken at the Neurology Department of the Second Hospital of Shandong University, encompassing patients diagnosed with infectious encephalitis as confirmed by PCR testing and other diagnostic modalities spanning from January 2018 to January 2024. The quantification of MPO and pertinent inflammatory proteins within patients' cerebrospinal fluid was accomplished through the utilization of ELISA. RESULTS: We enlisted 25 patients diagnosed with bacterial meningitis, ascertained through PCR testing, and stratified them into two groups: those with favorable prognoses (n = 25) and those with unfavorable prognoses (n = 25). Following assessments for normality and variance, notable disparities in CSF-MPO concentrations emerged between the prognostic categories of bacterial meningitis patients (P < 0.0001). Additionally, scrutiny of demographic data in both favorable and unfavorable prognosis groups unveiled distinctions in CSF-IL-1ß, CSF-IL-6, CSF-IL-8, CSF-IL-18, CSF-TNF-α levels, with correlation analyses revealing robust associations with MPO. ROC curve analyses delineated that when CSF-MPO ≥ 16.57 ng/mL, there exists an 83% likelihood of an adverse prognosis for bacterial meningitis. Similarly, when CSF-IL-1ß, CSF-IL-6, CSF-IL-8, CSF-IL-18, and CSF-TNF-α levels attain 3.83pg/mL, 123.92pg/mL, 4230.62pg/mL, 35.55pg/mL, and 35.19pg/mL, respectively, there exists an 83% probability of an unfavorable prognosis for bacterial meningitis. CONCLUSION: The detection of neutrophil extracellular traps MPO and associated inflammatory protein levels in cerebrospinal fluid samples holds promise in prognosticating bacterial meningitis, thereby assuming paramount significance in the prognostic evaluation of patients afflicted with this condition.

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.

Myeloperoxidase induces monocyte migration and activation after acute myocardial infarction.

Introduction: Myocardial infarction (MI) is a significant contributor to morbidity and mortality worldwide. Many individuals who survive the acute event continue to experience heart failure (HF), with inflammatory and healing processes post-MI playing a pivotal role. Polymorphonuclear neutrophils (PMN) and monocytes infiltrate the infarcted area, where PMN release high amounts of the heme enzyme myeloperoxidase (MPO). MPO has numerous inflammatory properties and MPO plasma levels are correlated with prognosis and severity of MI. While studies have focused on MPO inhibition and controlling PMN infiltration into the infarcted tissue, less is known on MPO's role in monocyte function. Methods and results: Here, we combined human data with mouse and cell studies to examine the role of MPO on monocyte activation and migration. We revealed a correlation between plasma MPO levels and monocyte activation in a patient study. Using a mouse model of MI, we demonstrated that MPO deficiency led to an increase in splenic monocytes and a decrease in cardiac monocytes compared to wildtype mice (WT). In vitro studies further showed that MPO induces monocyte migration, with upregulation of the chemokine receptor CCR2 and upregulation of inflammatory pathways identified as underlying mechanisms. Conclusion: Taken together, we identify MPO as a pro-inflammatory mediator of splenic monocyte recruitment and activation post-MI and provide mechanistic insight for novel therapeutic strategies after ischemic injury.

Myricetin reduces neutrophil extracellular trap release in a rat model of rheumatoid arthritis, which is associated with a decrease in disease severity.

Rheumatoid arthritis (RA) is a chronic disease characterized by joint inflammation and severe disability. However, there is a lack of safe and effective drugs for treating RA. In our previous study, we discovered that myricetin (MC) and celecoxib have a synergistic effect in the treatment of RA. We conducted in vitro and in vivo experiments to further investigate the effects and mechanisms of action of MC. Our findings demonstrated that MC treatment effectively reduced the release of neutrophil extracellular traps (NETs) and alleviated the inflammatory response in RA. Mechanistic studies showed that MC prevents the entry of PADI4 and MPO into the cell nucleus, thereby protecting DNA from decondensation. In a rat arthritis model, MC improved histological changes in ankle joints and suppressed NET-related signaling factors. In conclusion, MC protects the ankle joints against arthritis by inhibiting MPO and PADI4, thereby reducing NET release. The pharmacological mechanism of MC in RA involves the inhibition of NET release.

Association of the immediate perioperative dynamics of circulating DNA levels and neutrophil extracellular traps formation in cancer patients.

Objectives: Elevated circulating DNA (cirDNA) concentrations were found to be associated with trauma or tissue damage which suggests involvement of inflammation or cell death in post-operative cirDNA release. We carried out the first prospective, multicenter study of the dynamics of cirDNA and neutrophil extracellular trap (NETs) markers during the perioperative period from 24 h before surgery up to 72 h after curative surgery in cancer patients. Methods: We examined the plasma levels of two NETs protein markers [myeloperoxidase (MPO) and neutrophil elastase (NE)], as well as levels of cirDNA of nuclear (cir-nDNA) and mitochondrial (cir-mtDNA) origin in 29 colon, prostate, and breast cancer patients and in 114 healthy individuals (HI). Results: The synergistic analytical information provided by these markers revealed that: (i) NETs formation contributes to post-surgery conditions; (ii) post-surgery cir-nDNA levels were highly associated with NE and MPO in colon cancer [r = 0.60 (P < 0.001) and r = 0.53 (P < 0.01), respectively], but not in prostate and breast cancer; (iii) each tumor type shows a specific pattern of cir-nDNA and NETs marker dynamics, but overall the pre- and post-surgery median values of cir-nDNA, NE, and MPO were significantly higher in cancer patients than in HI. Conclusion: Taken as a whole, our work reveals the association of NETs formation with the elevated cir-nDNA release during a cancer patient's perioperative period, depending on surgical procedure or cancer type. By contrast, cir-mtDNA is poorly associated with NETs formation in the studied perioperative period, which would appear to indicate a different mechanism of release or suggest mitochondrial dysfunction.

Staphylococcal peroxidase inhibitor (SPIN): Residue-level investigation of the helical bundle domain.

Myeloperoxidase is a critical component of the antibacterial arsenal of neutrophils, whereby it consumes H2O2 as an oxidant to convert halogen and pseudohalogen anions into cytotoxic hypohalous acids. Following phagocytosis by neutrophils, the human pathogen Staphylococcus aureus secretes a potent myeloperoxidase inhibitory protein, called SPIN, as part of its immune evasion repertoire. The matured S. aureus SPIN polypeptide consists of only 73 residues yet contains two functional domains: whereas the 60 residue C-terminal helical bundle domain is responsible for MPO binding, the 13 residue N-terminal domain is required to inhibit MPO. Previous studies have informed understanding of the SPIN N-terminal domain, but comparatively little is known about the helical domain insofar as the contribution of individual residues is concerned. To address this limitation, we carried out a residue-level structure/function investigation on the helical bundle domain of S. aureus SPIN. Using sequence conservation and existing structures of SPIN bound to human MPO as a guide, we selected residues L49, E50, H51, E52, Y55, and Y75 for interrogation by site-directed mutagenesis. We found that loss of L49 or E52 reduced SPIN activity by roughly an order of magnitude, but that loss of Y55 or H51 caused progressively greater loss of inhibitory potency. Direct binding studies by SPR showed that loss of inhibitory potency in these SPIN mutants resulted from a diminished initial interaction between the inhibitor and MPO. Together, our studies provide new insights into the structure/function relationships of SPIN and identify positions Y55 and H51 as critical determinants of SPIN function.