Higher Myeloperoxidase Levels Research Articles

Valor preditivo da mieloperoxidase na identificação de pacientes de alto risco admitidos por dor torácica aguda

Myeloperoxidase (MPO) is a highly expressed enzyme due to leukocyte activation, with multiple atherogenic actions, including LDL cholesterol oxidation, and is related to the instability of atherosclerotic plaque. It is a predictor of adverse events in healthy individuals, patients with heart disease or those undergoing chest pain investigations. To analyze the contribution of MPO to identify patients with acute chest pain, non-ST elevation ECG and at high risk for in-hospital adverse events. Patients presenting acute chest pain and a non-ST elevation ECG, were admitted to the hospital and submitted to serum MPO level measurements and a structured examination protocol. From a cohort of 140 patients, 49 (35%) were diagnosed with acute coronary syndrome, of which 13 patients (9.3%) were diagnosed with non-ST elevation acute myocardial infarction (AMI) (troponin I >1.0 ng/mL). The best MPO cut-off point for AMI was identified as >100 pM using the ROC curve (AUC=0.662; CI 95%=0.532-0.793) revealing elevated sensitivity (92.3%) and negative predictive value (98.1%), however with low specificity (40.2%). In the multivariate analysis, MPO proved to be the only independent variable to diagnose AMI in evolution, with an odds ratio of 8.04 (p=0.048). In patients with acute chest pain and no ST elevation, high MPO levels upon admission to the hospital are an important tool to predict in-hospital adverse events, with an odds ratio of eight for the diagnosis of AMI.

Correlation of New Method for Measuring Plasma Myeloperoxidase with Clinical Behavior in Acute Myeloid Leukemia and Myelodysplastic Syndrome.

We previously reported that levels of myeloperoxidase (MPO) enzyme in plasma of patients with acute myeloid leukemia (AML) may correlate with aggressive disease. However, we found poor reproducibility and significant variability from lot to lot when commercially available kits are used. We developed a cytometric beads assay (CBA) for measuring plasma MPO that shows high reproducibility (CV<10%) with no significant variability with time. More importantly, MPO levels as measured using this CBA assay showed better correlation with clinical behavior in AML and myelodysplastic syndrome (MDS). Based on studying plasma levels of MPO in 98 patients with AML and 50 patients with advanced stage MDS, we found no significant difference in plasma MPO between AML and MDS patients. As expected MPO levels were significantly lower in patients with ALL (N=48) than in AML patient (P=0.04) and MDS (P=0.02). While MPO levels were significantly higher in patients with good cytogenetic abnormalities (P=0.004) in AML patients, MPO levels did not differ between intermediate and poor cytogenetic groups in patients with AML or MDS. Unexpectedly we found a difference between AML and MDS in their correlation with MPO levels and clinical outcome. Patients with advanced stage MDS and high levels of MPO (<300 ng/ml) had significantly longer survival than patients with lower levels (P=0.01). Multivariate analysis showed that this correlation with better survival in this group of MDS patients was independent of cytogenetic grouping. In contrast in AML, higher levels of MPO (>70 ng/ml) correlated with shorter survival (P=0.04). Similarly, patients with acute lymphoblastic leukemia (ALL) and high levels of MPO (>70 ng/ml) had shorter survival (P=0.05). Despite there was a correlation between the commercially available kit for measuring MPO and our CBA method (r=0.71, P<0.0001), the commercial kit did not provide robust data for clinical correlation. This data confirms that confirms that plasma levels of MPO play a role in the biology of the leukemia. Further studies are needed to establish if this role is a reflection of an inflammatory process or a reflection of disease load, although the finding of clinical relevance in ALL suggests that it is more likely reflecting an inflammatory process. [Display omitted]

Myeloperoxidase Is Required for the Combined Apoptotic Effects of Arsenic Trioxide with Ascorbic Acid or Epigallocatechin-3-Gallate in Myeloid Leukemia Cells.

Arsenic trioxide (As2O3) is a drug used world-wide that selectively causes the death of acute promyelocytic leukemia (APL) cells by novel mechanisms and induces complete clinical remission in 90% of patients without significant toxicity. The outstanding success of As2O3 therapy in relapsed APL patients has not been replicated by an equivalent success in other types of acute myeloid leukemia (AML). We have studied As2O3 mechanism(s) of action and identified agents to use in combination to improve the use of As2O3 as a treatment for other types of AML. As2O3 produces higher levels of H2O2 and apoptosis in APL NB4 cells than in other AML cells at therapeutic concentrations of 1-2 uM. As2O3 does not induce apoptosis in HL-60 cells but is synergistic with ascorbic acid (AA) or epigallocatechin-3-gallate (EGCG). Both AA and EGCG produce H2O2 which is augmented by the addition of As2O3. Apoptosis induction by As2O3 in combination with AA or EGCG is inhibited by catalase and the antioxidant N-acetylcysteine. Myeloperoxidase (MPO), a major neutrophil enzyme, augments H2O2-induced apoptosis by converting it into more potent reactive oxygen species. HL-60, NB4, SKNO-1 and PLB985 cells which express high level of MPO, but not U937 and K562 cells without expression of MPO, are responsive to As2O3 plus AA or EGCG-induced apoptosis. HP-100 cells, a subclone of HL-60 cells without MPO expression, are resistant to both combination treatments. MPO stable transfection sensitizes K562 cells to apoptosis following treatment with As2O3 and AA or EGCG. NADPH oxidase is an enzyme complex which generates O2− and H2O2 in neutrophils. In APL NB4 cells, As2O3 induces expression of NADPH oxidase members and is thought to participate in H2O2 production required for As2O3 response. X-CGD cells, a subclone of PLB-985 with targeted disruption of the gp91phox gene (Zhen et al, PNAS 90:9832, 1993) and loss of NADPH oxidase activity but with high levels of MPO activity, are responsive to treatment with As2O3 and AA or EGCG. These results suggest that AA and EGCG enhance As2O3-induced apoptosis through a synergistic production of H2O2 which is independent NADPH oxidase activation in non-APL AML cells. MPO augments the apoptotic effect of H2O2 produced by As2O3 plus AA or EGCG and it could be used as a marker to predict the sensitivity of AML cells to both combination treatments.

The Nitric Oxide Donor LA-419 [S-(6-Nitro-oxi-hexahydro-furo[3,2-b]furan-3-1-il)thioacetate] Prevents Intestinal Dysmotility, Bacterial Translocation, and Inflammation in a Rat Model of Enteritis

Indomethacin induces a chronic model of inflammatory bowel disease (IBD) characterized by spontaneous relapses of inflammation, bacterial translocation, and long-lasting motor disturbances derived from cyclical up-regulated inducible nitric-oxide synthase (iNOS) and sustained down-regulated neuronal NOS (nNOS). The aims of this study were to evaluate whether LA-419 [S-(6-nitro-oxi-hexahydro-furo[3,2-b]furan-3-1-il)thioacetate], a NO-donor drug, could re-establish the normal expression of NOS and, hence, prevent the development of intestinal dysmotility, bacterial translocation, and relapses of inflammation associated to this model. Enteritis was induced in rats by administration of indomethacin with and without treatment with a novel NO-donor: LA-419 (0.5 mg/ml in the drinking water). Inflammatory reaction was evaluated by measuring blood leukocytes, serum tumor necrosis factor, and tissue myeloperoxidase. Intestinal motor activity was evaluated using strain-gauges. Ileal expression of iNOS and nNOS mRNA was determined by reverse transcription-polymerase chain reaction. Bacterial translocation was evaluated in cultures from mesenteric lymph nodes. The indomethacin-induced acute inflammatory reaction was associated with a rise in blood leukocytes and tumor necrosis factor. In the chronic stage, blood leukocyte monitoring allowed the selection of animals in active and inactive phases. Active phase was associated with iNOS up-regulation, high myeloperoxidase levels, hypomotility, and bacterial translocation. In contrast, inactive phase was associated with hypermotility and absence of bacterial translocation. LA-419 treatment restored nitric-oxide synthase isoenzyme expression and prevented the oscillation of both inflammatory and motor parameters that could be cyclically observed in inflamed rats. LA-419 also prevented intestinal dysmotility, bacterial translocation, and relapses of intestinal inflammation. LA-419 might be a novel therapeutic approach to prevent acute inflammatory relapses in patients with IBD.

Plasma Concentrations of Myeloperoxidase Predict Mortality After Myocardial Infarction

This study investigated relationships between plasma myeloperoxidase (MPO), protein oxidation markers, and clinical outcome retrospectively in patients after acute myocardial infarction (MI). Reactive oxidants are implicated in cardiovascular disease, and elevated plasma MPO is reported to predict adverse outcome in acute coronary syndromes. Detailed demographic information, radionuclide ventriculography, neurohormone measurements, and clinical history were obtained for 512 acute MI patients at hospital admission. Plasma levels of MPO and protein carbonyls were measured in patients and 156 heart-healthy control subjects. 3-chlorotyrosine was measured in selected patients. Patient mortality was followed for 5 years. Plasma MPO and protein carbonyl concentrations were higher in MI patients 24 h to 96 h after admission than in control subjects (medians: MPO 55 ng/ml vs. 39 ng/ml, and protein carbonyls 48 pmol/mg vs. 17 pmol/mg protein, p < 0.001 for each). Both markers were significantly correlated with each other and with cardiovascular hormone levels. Chlorotyrosine was not elevated in patients with high MPO or carbonyl levels. Above-median levels of MPO but not protein carbonyls were independently predictive of mortality (odds ratio 1.8, 95% confidence interval 1.0 to 3.0, p = 0.034). Patients with above-median MPO levels in combination with above-median plasma amino-terminal pro-brain natriuretic peptide (NT-proBNP) or below-median left ventricular ejection fraction (LVEF) had significantly greater mortality compared with other patients. Myeloperoxidase and protein carbonyl levels are elevated in plasma after acute MI, apparently via independent mechanisms. High MPO is a risk factor for long-term mortality and adds prognostic value to LVEF and plasma NT-proBNP measurements.

Functional Polymorphism of the Myeloperoxidase Gene in Hypertensive Nephrosclerosis Dialysis Patients

Myeloperoxidase (MPO) may play an important role not only in host defense reactions but also in local inflammations, especially in atherosclerotic diseases such as hypertensive nephrosclerosis (HN). Paradoxically, MPO-deficient mice have been reported to show increased atherosclerosis compared with wild mice, although higher MPO levels are thought to exacerbate atherosclerotic disease. To clarify the genetic role of MPO in HN, we examined the function and distribution of the -463G/A polymorphism located in the promoter region of the MPO gene with ex vivo flow cytometry analysis and a study in end-stage renal disease patients, respectively. This polymorphism has been reported to have a functional significance in vitro, with the A allele being associated with lower MPO expression. In the present study, we also found significantly higher reactive oxygen species (ROS) production with peripheral neutrophils isolated from subjects with the GG genotype compared with those from subjects with other genotypes by flow cytometry assay with 2-[6-(4'-amino) phenoxy-3H-xanthen-3-on-9-yl] benzoic acid (APF), which shows higher sensitivity with hypochlorite (OCl(-)). Genotyping the -463G/A polymorphism in HN, chronic glomerulonephritis (CGN) and diabetic nephropathy (DM) patients who were under hemodialysis treatment demonstrated that the GG genotype was more frequent in the HN group than in the CGN and DM groups. However, the distribution of the GG genotype in the HN group was similar to that in healthy individuals. Although the -463G/A polymorphism is associated with ROS production, careful interpretation may be required to conclude that the -463G/A polymorphism can serve as a useful marker of atherosclerosis and cardiovascular events in dialysis patients.

Upregulation of myeloperoxidase in patients with opticospinal multiple sclerosis: Positive correlation with disease severity

To clarify the role of myeloperoxidase (MPO) in multiple sclerosis (MS), we measured serum MPO levels in 86 Japanese patients with relapsing remitting MS, 47 with opticospinal MS (OSMS) and 39 with conventional MS (CMS), and 85 healthy subjects by sandwich enzyme immunoassays and analyzed relationships with clinical features. We found a significant increase in serum MPO in OSMS patients at relapse and remission, and in CMS patients at remission compared with controls. By logistic regression analysis, the clinical variable associated with high level of MPO at remission in OSMS patients (higher than the mean ± 2 S.D. of healthy controls) was only Kurtzke's Expanded Disability Status Scale (EDSS) score in blood sampling ( p = 0.0245); that is, a greater EDSS scores in the high MPO group, whereas in CMS none were associated. The results of our study suggest that MPO levels in remission are related with severe tissue destruction in OSMS.

Acute Inflammatory and Neuropathic Pain in Lewis and Fischer Rats

Inbred, histocompatible Lewis and Fischer 344 rats (LEW and FIS) have been used to identify an inverse relationship between hypothalamic-pituitary-adrenal (HPA) axis activity and susceptibility to autoimmune and chronic inflammatory disorders, with LEW showing blunted HPA axis activity and increased susceptibility toward the development of autoimmunity and chronic inflammation, and FIS showing the opposite relationship. In the present study, LEW and FIS were used to determine the relationship between HPA axis function and acute inflammatory pain (carrageenan-induced hindpaw inflammation) and neuropathic pain (partial sciatic nerve ligation; PSNL). The results showed that carrageenan-induced thermal and mechanical allodynia and hyperalgesia were greater in FIS than in LEW. Similarly, FIS showed more carrageenan-induced hindpaw swelling and higher levels of myeloperoxidase (a measure of neutrophils) in the carrageenan-inflamed hindpaw. After PSNL, LEW showed a profound mechanical allodynia and hyperalgesia, whereas mechanical sensitivity in FIS was unaltered. However, FIS, but not LEW, developed thermal allodynia and hyperalgesia after PSNL. These results provide strong evidence for a positive relationship between HPA axis activity and acute inflammatory pain. The results also support a relationship between HPA axis activity and neuropathic pain, but the relationship is complex and may depend on the pain assay.

Serum Clara Cell Protein as an Indicator of Pulmonary Impairment in Occupational Exposure at Aluminum Foundry

Although some of the exposures in aluminum (Al) smelting have been well characterized, and respiratory disorders in aluminum production workers are well known, the relationship between internal aluminum loads and appropriate lung biomarkers have not been elucidated. The aim of our work was to carry out a comprehensive investigation in workers employed in the Aluminum Foundry Casting Department with special reference to currently existing hygiene standards, known as threshold limit values (TLV) based on aluminum effects on the respiratory system. The measurement of serum anti-inflammatory Clara cell protein (CC16) was employed as a peripheral marker of the lung epithelium function. A group of 50 casting smelters, 5 locksmiths, 11 sawyers and auxiliary workers exposed to dust containing 14% of aluminum, and a group of 42 controls were examined. Respiratory function tests were performed and forced volume capacity (FVC), forced expiratory volume in 1 s (FEV1), forced expiratory volume in the first percent (FEV1%), forced expiratory flows in 50% VC (FEV50), and markers of foundry workers' exposure and body burden, Al concentration in the breathing zone, blood and urine, biomarkers of the effects of exposure, concentration of CC16 and hyaluronic acid (HA) in serum were determined in all examined workers. Additional measurements comprised determinations of serum iron (Fe) levels, myeloperoxidase (MPO), eosinophil cationic protein (ECP), immunoglobulin E (IgE), glutathione S-transferase (GST), and superoxide dismutase (SOD) activity in erythrocytes. The group of casting smelters was characterized by the highest levels of aluminum in urine (Al-U) (43.7 microg L(-1)), high levels of MPO, ECP and IgE, high SOD activity, low CC16 levels, and low activity of GST. Lower Al-U excretion was observed in locksmiths (35.2 microg L(-1)) and sawyers (21.7 microg L(-1)). Serum CC16 proved to be the most sensitive biomarker, showing high inverse relationship with serum Al (Al-S) concentrations in casting smelters (p = 0.006). The study showed that in conditions of occupational exposure, dusts containing Al2O3 < 1 mg m(-3) cause changes in the respiratory system and biomarkers in serum, especially in CC16, connected with altered functioning of this system. Changes in concentrations of the examined biomarkers and also in respiratory parameters of the study subjects were observed when Al-U concentration was > 40 microg L (-1).

MMP-9 expression is associated with leukocytic but not endothelial markers in brain arteriovenous malformations

Brain arteriovenous malformations (BAVM) have high matrix metalloproteinase-9 (MMP-9) expression, the source of which is unclear. We hypothesized MMP-9 production might be due to inflammation in BAVM. Compared to control brain tissues (n = 5), BAVM tissue (n = 139) had a higher expression (by ELISA) of myeloperoxidase (MPO) (193 +/- 189 vs. 6 +/- 3, ng/mg, P < .001), MMP-9 (28 +/- 32 vs. 0.7 +/- 0.6, ng/mg, P < .001), and IL-6 (102 +/- 218 vs. 0.1 +/- 0.1, pg/mg, P < .001), but not eNOS (114 +/- 87 vs. 65 +/- 9, pg/mg, P = .09). MMP-9 expression in BAVM highly correlated with myeloperoxidase (R2 = .76, P < .001), as well as with IL-6 (R2 = .32, P < .001). In contrast, MMP-9 in BAVM poorly correlated with the endothelial marker, eNOS (R2 = .03, P = .05), and CD31 (R2 = .004, P = .57). Compared to non-embolized patients (n = 46), patients with pre-operative embolization (n = 93) had higher levels of myeloperoxidase (236 +/- 205 vs. 106 +/- 108, ng/mg, P < .001) and MMP-9 (33 +/- 35 vs. 16 +/- 20, ng/mg, P < .001), however the correlation between MMP-9 and myeloperoxidase was equally strong for both groups (R2 = .69, n = 93, P < .001, for both). MMP-9 expression correlated with the lipocalin-MMP-9 complex, suggesting neutrophils as the MMP-9 source. MPO co-localized with majority of MMP-9 signal by immunohistochemistry. Our data suggest that inflammation is a prominent feature of BAVM lesional phenotype, and neutrophils appear to be a major source of MMP-9 in these lesions.

Oncogenic TLS-ERG Fusion Protein Promotes Differentiation of Myeloid Progenitors into Myeloblasts but Blocks Their Terminal Differentiation.

In a subset of acute myelogenous leukemia (AML) and chronic myelogenous leukemia (CML) in blast crisis, the TLS (translocation liposarcoma) gene is fused to the ERG (ets-related gene) gene through a recurrent t(16;21) translocation. This chromosomal translocation results in the generation of the TLS-ERG fusion protein. More recently, the same TLS-ERG fusion protein has also been identified in Ewing's sarcoma in children. To investigate the oncogenic mechanism of TLS-ERG, we stably expressed TLS-ERG or its mutant TLS-ERGΔets in mouse L-G myeloid progenitor cells through retroviral transduction. L-G cells normally proliferate in the presence of IL-3 and undergo terminal differentiation into hypersegmented neutrophils when treated with G-CSF. In contrast, expression of TLS-ERG in L-G cells led to spontaneous differentiation into immature myeloblasts but G-CSF treatment of these cells failed to induce terminal differentiation. L-G myeloblasts harboring TLS-ERG were characterized by high levels of myeloperoxidase (MPO), a phenomenon also observed in immature myeloblasts isolated from human leukemia patients with the t(16;21) translocation. As deletion of the ets DNA-binding domain from TLS-ERG abolished its ability to promote spontaneous myeloblast differentiation or the ability to up-regulate the MPO gene, we further examined how TLS-ERG fusion protein might affect the functions of other regulators such as PU.1, an ets transcription factor known to play a critical role in the differentiation of myeloid progenitors. Though both proteins have similar DNA-binding property, TLS-ERG differs from PU.1 through binding directly to RNA polymerase II as well as through escaping repression by histone modification enzymes. Together, these findings suggest that the TLS-ERG fusion protein can cause blockage of myeloid differentiation by interfering with the gene expression program normally controlled by key regulators.

Statins downregulate myeloperoxidase gene expression in macrophages

Statins, inhibitors of HMG-CoA reductase, have pleiotropic benefits independent of cholesterol levels, including anti-oxidant and anti-inflammatory effects. Here, we investigate the effect of statins on myeloperoxidase (MPO) expression. MPO, expressed in foam cell macrophages, was recently shown to oxidize the ApoA-1 component of HDL, impairing ABCA-1 mediated cholesterol efflux. High levels of serum MPO correlate with increased risk of CAD events. Findings here show that statins strongly inhibit MPO mRNA expression in human and murine monocyte-macrophages. Suppression was reversed by downstream intermediates of HMG-CoA reductase, mevalonate, and geranylgeranylpyrophosphate, but not farnesylpyrophosphate. An inhibitor of geranylgeranyltransferase, GGTI-286, mimics the effects of statins, indicating geranylgeranylation is key to MPO expression. Reduction of MPO mRNA levels was observed in vivo in leukocytes from statin-fed mice, correlating with reductions in MPO protein and enzyme activity. These findings suggest that the pleiotropic protections afforded by statins may be due in part to suppression of MPO expression.

Characterization of a rat model to study acute neuroinflammation on histopathological, biochemical and functional outcomes

Neuroinflammation is one of the events occurring after acute brain injuries. The aim of the present report was to characterize a rat model to study acute neuroinflammation on the histopathological, biochemical and functional outcomes. Lipopolysaccharide (LPS), known as a strong immunostimulant, was directly injected into the hippocampus. The spatiotemporal evolution of inducible NOS (iNOS) and cell death was studied from 6 h to 7 days. A perfect time course correlation was observed between iNOS immunoreactivity and iNOS activity showing an acute, expansive and transient iNOS induction in the hippocampus with a peak at 24 h. It was associated with a marked increase in NO metabolite (NO x ) levels, and a high level of myeloperoxidase (MPO) activity. This inflammation precedes a massive cellular loss including at least neurons and astrocytes, and a drop of constitutive NOS activity, restrictive to the ipsilateral hippocampus from 48 h after LPS injection. Moreover, sensorimotor function impairment occurred from 24 h to 7 days with a maximum at 24 h post-LPS injection. Therefore, we characterized an in vivo model of acute neuroinflammation and neurodegeneration, in relation with a neurological deficit, which may be a powerful tool for mechanistic studies and for further evaluation of the potential neuroprotective agents.

Clinical Relevance of Circulating Myeloperoxidase (MPO) in Acute Myeloid Leukemia (AML) and Myelodysplastic Syndrome (MDS).

MPO, a hemoprotein expressed in polymorphonuclear neutrophils and their precursors, is an important component of the antimicrobial activity of phagocytes and is associated with inflammatory response. Cellular expression levels of MPO, assessed by cytochemical staining, are used extensively for the diagnosis and subtyping of various leukemias. However, the association of circulating (plasma or serum) MPO levels with hematologic values and clinical outcomes of AML and MDS has not been reported. We investigated whether MPO, presumably released from leukemic cells, can be detected in the plasma of patients with AML and MDS. We further assessed the association of plasma MPO levels with hematologic characteristics and outcomes. 144 patients with AML, 28 patients with MDS, and 100 control subjects were included in the analysis. Patients with AML (median 19.3, range 1.0–9514.7 ng/mL) and MDS (median 13.6, range 1.0–3021.9 ng/mL) had significantly higher plasma MPO levels than control subjects (median, 4.9; range, 3.5–20.6 ng/mL). Because MPO levels and overall survival did not differ significantly between AML and MDS patients, the 2 groups were considered together in subsequent analyses. MPO levels were inversely correlated with overall survival (Cox regression analysis, P<0.001). In the multivariate model, the inverse correlation between MPO levels and survival was independent of cytogenetics, performance status, anticident hematologic disease (AHD), age, and diagnosis (AML vs MDS). Response to therapy in AML and MDS patients was not associated with plasma MPO levels. However, responders with high MPO levels had shorter remission duration (CRD), although this difference did not reach statistical significance (P = 0.07). As expected, MPO levels correlated with the level of differentiation of leukemic cells (FAB subclassification): patients with M0 and M6 had lower MPO levels than those with M2, M3, or M4 (P = 0.0001). MPO levels also correlated positively with white cell count, beta2-microglobulin, monocyte, LDH, BUN, and creatinine levels. These data indicate that plasma MPO levels are elevated in AML and MDS and may be useful as a prognostic factor. Our findings also raise the question of how the proinflammatory properties of MPO might affect the clinical course of these diseases.

Prognostic Value of Myeloperoxidase in Patients With Chest Pain

Brennan, Marie-Luise; Penn, Marc S.; Van Lente, Frederick; Nambi, Vijay; Shishenbor, Mehdi H.; Aviles, Ronnier J.; Goormastic, Marlene; Pepoy, Michael L.; McErlean, Ellen S.; Topol, Eric J.; Nissen, Steven E.; Hazen, Stanley L.

Natural killer cells participate in bacterial clearance during septic peritonitis through interactions with macrophages.

Natural killer (NK) cells have a well-established role in host defense against viral infections and malignancies. However, their function in bacterial infection and sepsis is poorly defined. We hypothesized that NK cells, as a major producer of interferon-gamma during sepsis, would be important in host defense against bacterial infections. Cecal ligation and puncture (CLP) was performed on Swiss Webster mice depleted of NK cells by pretreatment with anti-asialo GM1 and control mice given immunoglobulin G (IgG) antibody. NK cell-depleted mice had significantly higher anaerobic bacterial counts in the liver and peritoneal lavage fluid, as well as higher aerobic counts in the liver and blood 4 h after CLP. Macrophage phagocytosis, nitric oxide production, and interleukin (IL)-6 levels at 4 h were also decreased in mice depleted of NK cells compared with controls. Greater neutrophil influx into the peritoneum, indicated by higher myeloperoxidase levels, was also seen in NK cell-depleted mice. At 8 and 18 h after CLP, bacterial counts were similar between groups, and overall survival rates were not significantly different. Peritoneal IL-12 levels significantly increased by 18 h in normal mice, but not in NK cell-depleted animals. Our data suggest that NK cells participate in the early local and systemic eradication of bacteria and regulation of IL-12 during polymicrobial sepsis. These effects are likely due to their interactions with macrophages.

Serum Eosinophil Cationic Protein, Myeloperoxidase, Tryptase, Eotaxin and Th2-Like Cytokines in Dermatitis herpetiformis

Background: Dermatitis herpetiformis (DH) is a subepidermal bullous disease characterized by a neutrophilic and eosinophilic infiltrate, together with activated lymphocytes that show preferential polarization towards the Th2-like phenotype. To date, the role of neither soluble mediators, such as cytokines, nor serum levels of myeloperoxidase (MPO), eosinophil cationic protein (ECP) and tryptase (TRY) have been studied in the peripheral blood of DH. In this study we investigated 40 DH subjects for possible alterations in MPO, ECP, TRY, eotaxin, interleukin (IL) 4 and IL-5 concentrations to evaluate the role of neutrophils, eosinophils and mast cells in the induction of skin lesions. Sera from 20 healthy subjects were tested as controls. Methods: Eotaxin, IL-4 and IL-5 serum contents were determined by a solid-phase enzyme-linked immunosorbent assay. A double antibody radioimmunoassay was used to measure serum levels of MPO, ECP and TRY. Results: Normal levels of eotaxin and IL-4 were found in all DH subjects; increased IL-5 levels were found only in 1 subject. The MPO levels were significantly higher in DH subjects than in healthy controls (p < 0.01). There was also a significant difference between the serum ECP levels of DH and healthy controls (p < 0.05). TRY was not revealed in the serum of DH subjects. Conclusions: The determination of Th2-like cytokines and TRY serum levels is not important in quantifying the involvement of activated lymphocytes and mast cells in DH; however, the finding of high MPO and ECP serum levels seems to confirm that these products are good markers of the presence of activated neutrophils and eosinophils.

Neutrophil-associated activation markers in healthy smokers relates to a fall in DLCOand to emphysematous changes on high resolution CT

Smoking is a risk factor for developing chronic obstructive pulmonary disease (COPD), but there are no good indicators for early identification of subjects who will develop symptomatic COPD. The aim of this study was to investigate inflammatory mechanisms related to changes in lung function and emphysematous changes on high resolution computed tomography (HRCT) in ‘healthy’ smokers. Subjects were 60-year-old men from a population study. Bronchoscopy was performed in 30 smokers and 18 who had never smoked. Blood tests, lung function measurements and HRCT were carried out in 58 and 34 subjects, respectively.In comparison with never-smokers, smokers had higher levels of myeloperoxidase (MPO), human neutrophil lipocalin (HNL), eosinophil cationic protein (ECP) and lysozyme in blood, higher levels of MPO, interleukin-8 (IL-8) and HNL in bronchial lavage (BL), and of IL-8, HNL and interleukin-1 β (IL-1 β) in bronchoalveolar lavage (BAL). Smokers also had lower levels of Clara cell protein 16 (CC-16) in blood. HNL in BL and BAL showed strong correlations to other inflammatory markers (MPO, IL-8, IL-1 β). The variations in MPO in BL were explained by variations in HNL (R2=0·69), while these variations in BAL were explained by variations in HNL and IL-1β (R2=0·76). DLCOwas the lung function variable most closely related to MPO and IL-8 in BL and BAL and to IL-1 β in BAL. In a multiple regression analysis, MPO, IL-1 β, IL-8 and CC-16 in BL and MPO in BAL contributed to the explanation of variations in DLCOto 41% and 22%, respectively, independent of smoking habits. In smokers with emphysematous lesions on HRCT, HNL in BAL correlated to emphysema score (rs=0·71).We conclude that ‘healthy’ smoking men with a near normal FEV1show signs of inflammation in the lower airways that are related to a decrease in DLCOand to emphysematous lesions on HRCT. This inflammation seems to be the result of both monocyte/macrophage and neutrophil activation.

Serum ECP and MPO are increased during exacerbations of chronic bronchitis with airway obstruction

Many studies have demonstrated that, in asthma, serum levels of eosinophil cationic protein (ECP) are related to the activity and severity of the disease and can be used to evaluate the response to steroid treatment. During exacerbations of chronic bronchitis, airway inflammation shows some features of asthmatic inflammatory processes, with recruitment of eosinophils and recovery of significant amounts of ECP in bronchial lavage fluid (BAL). Involvement of neutrophils, with high levels of myeloperoxidase (MPO), is, on the contrary, typical of this latter disease, and not shared with asthma. In spite of the information collected with BAL and bronchial biopsy studies, few data still exist on serum levels of these proteins in chronic bronchitis. The objective of this study was to assess if serum levels of ECP and MPO are specifically increased in exacerbations of chronic bronchitis, as compared to other non-asthmatic acute respiratory disturbances. Serum ECP, MPO and immunoglobulin E (IgE) levels were measured in 17 non-atopic patients with exacerbation of chronic bronchitis with airway obstruction (COPD) and in 11 control subjects seeking emergency medical treatment for unrelated acute respiratory problems. Spirometry was performed in patients able to give the necessary collaboration. All the subjects of this study were recruited from the emergency department. Both ECP and MPO were significantly increased in serum from patients with exacerbated COPD (22.2 +/- 4.1 vs 9.5 +/- 1.4 mcg/L and 853 +/- 168 vs 375 +/- 41 mcg/L) and a strong correlation existed between these two variables (r = 0.782). A further control group was made of 11 patients with stable COPD. These subjects had levels of both ECP (13.1 +/- 2.7 mcg/L) and MPO (469 +/- 71) significantly lower than patients with exacerbated disease and higher than those without COPD. We conclude that serum ECP and MPO are increased during the exacerbations of COPD. These observations can give a basis for further studies aimed to evaluate the utility of these two proteins as markers of activity and severity of COPD.

Sequential development of pulmonary hemorrhage with MPO-ANCA complicating anti–glomerular basement membrane antibody-mediated glomerulonephritis

We report a case of rapidly progressive glomerulonephritis caused by anti–glomerular basement membrane (anti-GBM) antibodies that progressed to end-stage renal disease in a 67-year-old woman with diabetes. Intensive combined immunosuppressive therapy with methylprednisolone bolus, oral prednisone, and cyclophosphamide led to negativity of anti-GBM antibodies but was not able to restore renal function. After 28 months of hemodialysis, the patient suddenly presented with pulmonary hemorrhage. In this setting, high levels of myeloperoxidase (MPO)-antineutrophil cytoplasmic antibody (ANCA) and negative anti-GBM antibodies were found. Therapy with oral prednisone and cyclophosphamide led to resolution of pulmonary hemorrhage and negativity of MPO-ANCA.