RESUMO
BACKGROUND: Magnetic resonance imaging (MRI) accuracy after neoadjuvant systemic therapy (NST) for breast cancer varies according to hormone receptor (HR), human epidermal growth factor receptor type-2 (HER2) subtype and Ki-67 proliferation index. Whether MRI accuracy varies by genomic signatures is unknown. We examined the accuracy of MRI in the NEONAB trial (Clinicaltrials.gov #: NCT01830244). AIM: To examine the accuracy of MRI to predict pathological response to neoadjuvant therapy for breast cancer in the NEONAB trial. METHODS: Patients with stages II-III breast cancer received sequential epirubicin, cyclophosphamide and nab-paclitaxel and trastuzumab if they were HER2+. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated to assess the utility of preoperative MRI to predict pathological complete response (pCR). Bland-Altman plots were used to assess agreement between MRI and pathological assessment of residual disease. RESULTS: MRI correctly predicted pCR in 64.1% of the cohort. Sensitivity and specificity were 52% and 78%, respectively; PPV 73% and NPV 58%. MRI predicted pCR most accurately in HER2-positive patients; sensitivity 58%, specificity 100%, PPV 100% and NPV 38%. MRI had higher PPV and NPV in tumours with Ki-67 ≥ 15% than tumours with Ki-67 < 15%, 75% versus 50% and 57.5% versus 50%, respectively. In this study, MRI underestimated residual tumour size by 1.65 mm (limits of agreement: 43.07-39.77 mm). CONCLUSIONS: MRI appears more accurate for predicting pCR in HER2+ disease than other subtypes and in cancers with Ki-67 ≥ 15% compared to those with Ki-67 < 15%. Accuracy of MRI in our HR+, RS ≥ 25 cohort is comparable to previous reports of unselected HR+ disease. MRI post-NST should be interpreted in conjunction with HER2 status and Ki-67 index of the primary.
Assuntos
Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/terapia , Imageamento por Ressonância Magnética , Terapia Neoadjuvante , Adulto , Idoso , Albuminas/uso terapêutico , Austrália , Neoplasias da Mama/patologia , Estudos de Coortes , Ciclofosfamida/uso terapêutico , Epirubicina/uso terapêutico , Feminino , Humanos , Antígeno Ki-67/sangue , Pessoa de Meia-Idade , Paclitaxel/uso terapêutico , Receptor ErbB-2/genética , Sensibilidade e Especificidade , Trastuzumab/uso terapêuticoRESUMO
OBJECTIVE: Patients diagnosed with cancer in the Emergency Department (ED) have more advanced disease at diagnosis and poorer outcomes. High rates of initial presentation to ED suggest potential problems with access to care. The aim of this project was to interpret findings in regional/rural Victoria and explore implications for practice. DESIGN: Cross-sectional study linking two independent data sets. SETTING: Regional city of Geelong and surrounding rural areas in south-west Victoria. PARTICIPANTS: All newly diagnosed cancer patients in 2009. MAIN OUTCOME MEASURES: Number of cancer patients diagnosed in the ED. RESULTS: One in five newly diagnosed cancer patients present to ED 6 months prior to cancer diagnosis. One in 10 is diagnosed as a result of their ED visit. Patients presenting to ED were older, more often men and from disadvantaged areas. Symptoms on presentation included chest complaints, bowel obstruction, abdominal pain, anaemia and generalised weakness. Cancer diagnosed in the ED is associated with advanced stage and shorter survival. CONCLUSION: Reasons for presentation to ED would be multifactorial and include complex cases with coexisting symptoms making diagnosis difficult. The general public appear to have a low level of awareness of alternative primary care services or difficulty accessing such information. Some of the changes towards reducing the number of patients presenting to ED will include patient education.
Assuntos
Serviço Hospitalar de Emergência , Neoplasias/diagnóstico , Neoplasias/epidemiologia , Idoso , Bases de Dados Factuais , Feminino , Humanos , Masculino , Vitória/epidemiologiaRESUMO
BACKGROUND AND OBJECTIVE: Natural killer (NK) and natural killer T (NKT)-like cells represent a small but important proportion of effector lymphocytes that we have previously shown to be major sources of pro-inflammatory cytokines and granzymes. We hypothesized that these cells would be increased in the airway in chronic obstructive pulmonary disease (COPD), accompanied by reduced expression of the inhibitory receptor CD94 (Kp43) and increased expression of cytotoxic mediators granzyme B and perforin. METHODS: We measured NK and NKT-like cells and their expression of CD94 in the blood of COPD patients (n = 71; 30 current and 41 ex-smokers), smokers (16) and healthy controls (25), and bronchoalveolar lavage fluid (BALF) from a cohort of subjects (19 controls, 12 smokers, 33 COPD). Activation was assessed by measuring CD69 in blood and the cytotoxic potential of NK cells by measuring granzymes A and B, and using a cytotoxicity assay in blood and BALF. RESULTS: In blood in COPD, there were no significant changes in the proportion of NK or NKT-like cells or expression of granzyme A or NK cytotoxic potential versus controls. There was, however, increased expression of granzyme B and decreased expression of CD94 by both cell types versus controls. The proportion of NK and NKT-like cells were increased in BALF in COPD, associated with increased NK cytotoxicity, increased expression of granzyme B and decreased expression of the inhibitory receptor CD94 by both cell types. CONCLUSIONS: Treatment strategies that target NK and NKT-like cells, their cytotoxicity and production of inflammatory mediators in the airway may improve COPD morbidity.
Assuntos
Células Matadoras Naturais/patologia , Subfamília D de Receptores Semelhantes a Lectina de Células NK/metabolismo , Células T Matadoras Naturais/patologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Sistema Respiratório/metabolismo , Sistema Respiratório/patologia , Adulto , Idoso , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Lavagem Broncoalveolar , Estudos de Casos e Controles , Granzimas/metabolismo , Humanos , Células Matadoras Naturais/metabolismo , Lectinas Tipo C/metabolismo , Pessoa de Meia-Idade , Células T Matadoras Naturais/metabolismo , Perforina/metabolismo , Fumar/metabolismoRESUMO
Although the importance of the macrophage complement receptor immunoglobulin (CRIg) in the phagocytosis of complement opsonized bacteria and in inflammation has been established, the regulation of CRIg expression remains undefined. Because cellular activation during inflammation leads to the release of arachidonate, a stimulator of leukocyte function, we sought to determine whether arachidonate regulates CRIg expression. Adding arachidonate to maturing human macrophages and to prematured CRIg(+) macrophages caused a significant decrease in the expression of cell-surface CRIg and CRIg mRNA. This effect was independent of the metabolism of arachidonate via the cyclooxygenase and lipoxygenase pathways, because it was not inhibited by the nonsteroidal anti-inflammatory drugs indomethacin and nordihydroguaiaretic acid. Studies with specific pharmacological inhibitors of arachidonate-mediated signaling pathways showed that protein kinase C was involved. Administration of dexamethasone to macrophages caused an increase in CRIg expression. Studies with proinflammatory and immunosuppressive cytokines showed that IL-10 increased, but interferon-γ, IL-4, and transforming growth factor-ß1 decreased CRIg expression on macrophages. This down- and up-regulation of CRIg expression was reflected in a decrease and increase, respectively, in the phagocytosis of complement opsonized Candida albicans. These data suggest that a unique inflammatory mediator network regulates CRIg expression and point to a mechanism by which arachidonate and dexamethasone have reciprocal effects on inflammation.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Ácido Araquidônico/farmacologia , Dexametasona/farmacologia , Macrófagos/efeitos dos fármacos , Fagocitose/efeitos dos fármacos , Receptores de Complemento 3b/metabolismo , Candida albicans/imunologia , Células Cultivadas , Citocinas/metabolismo , Regulação para Baixo , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Lipoxigenase/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Fagocitose/imunologia , Prostaglandina-Endoperóxido Sintases/metabolismo , RNA Mensageiro/metabolismoRESUMO
Defective efferocytosis may perpetuate inflammation in smokers with or without chronic obstructive pulmonary disease (COPD). Macrophages may phenotypically polarize to classically activated M1 (proinflammatory; regulation of antigen presentation) or alternatively activated M2 (poor antigen presentation; improved efferocytosis) markers. In bronchoalveolar lavage (BAL)-derived macrophages from control subjects and smoker/ex-smoker COPD subjects, we investigated M1 markers (antigen-presenting major histocompatibility complex [MHC] Classes I and II), complement receptors (CRs), the high-affinity Fc receptor involved with immunoglobulin binding for phagocytosis (Fc-gamma receptor, FcγR1), M2 markers (dendritic cell-specific intercellular adhesion molecule-grabbing nonintegrin [DC-SIGN] and arginase), and macrophage function (efferocytosis and proinflammatory cytokine production in response to LPS). The availability of glutathione (GSH) in BAL was assessed, because GSH is essential for both M1 function and efferocytosis. We used a murine model to investigate macrophage phenotype/function further in response to cigarette smoke. In lung tissue (disaggregated) and BAL, we investigated CRs, the available GSH, arginase, and efferocytosis. We further investigated the therapeutic effects of an oral administration of a GSH precursor, cysteine l-2-oxothiazolidine-4-carboxylic acid (procysteine). Significantly decreased efferocytosis, available GSH, and M1 antigen-presenting molecules were evident in both COPD groups, with increased DC-SIGN and production of proinflammatory cytokines. Increased CR-3 was evident in the current-smoker COPD group. In smoke-exposed mice, we found decreased efferocytosis (BAL and tissue) and available GSH, and increased arginase, CR-3, and CR-4. Treatment with procysteine significantly increased GSH, efferocytosis (BAL: control group, 26.2%; smoke-exposed group, 17.66%; procysteine + smoke-exposed group, 27.8%; tissue: control group, 35.9%; smoke-exposed group, 21.6%; procysteine + smoke-exposed group, 34.5%), and decreased CR-4 in lung tissue. Macrophages in COPD are of a mixed phenotype and function. The increased efferocytosis and availability of GSH in response to procysteine indicates that this treatment may be useful as adjunct therapy for improving macrophage function in COPD and in susceptible smokers.
Assuntos
Macrófagos Alveolares/citologia , Ácido Pirrolidonocarboxílico/farmacologia , Fumar/efeitos adversos , Tiazolidinas/farmacologia , Animais , Apoptose , Broncoscopia/métodos , Citometria de Fluxo/métodos , Humanos , Inflamação , Interleucina-12/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Macrófagos Alveolares/efeitos dos fármacos , Camundongos , Fagocitose , Fenótipo , Doença Pulmonar Obstrutiva Crônica , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Despite recent reviews of best practice for the treatment of Australian venomous bites and stings, there is controversy about some aspects of care, particularly the use of antivenom. Our aim was to understand current attitudes and practice in the management of suspected snake envenoming. A single-stage, cross-sectional survey of Australian emergency care physicians who had treated snake envenomation in the previous 36 months was conducted. Hospital pharmacists were also invited to complete a survey about antivenom availability, usage, and wastage in Australian hospitals. The survey was available between 5 March and 16 June 2019. A total of 121 snake envenoming cases were reported, and more than a third (44.6%) of patients were not treated with antivenom. For those treated with antivenom (n = 67), 29 patients (43%) received more than one ampoule. Nearly a quarter of respondents (21%) identified that antivenom availability was, or could be, a barrier to manage snake envenoming, while cost was identified as the least important factor. Adverse reactions following antivenom use were described in 11.9% of cases (n = 8). The majority of patients with suspected envenoming did not receive antivenom. We noted variation in dosage, sources of information, beliefs, and approaches to the care of the envenomed patient.
Assuntos
Mordeduras de Serpentes/epidemiologia , Venenos de Serpentes , Animais , Antivenenos/administração & dosagem , Atitude , Austrália/epidemiologia , Estudos Transversais , Venenos Elapídicos , Elapidae , Humanos , Mordeduras de Serpentes/terapia , Inquéritos e QuestionáriosRESUMO
Behavioural assessment units (BAU) have been established in emergency departments (EDs) to provide short-term observation, treatment, and care to people experiencing acute behavioural disturbance. A prospective observational study was conducted in a cohort of adult patients admitted to one BAU located within an ED (July-December 2017) to compare clinical characteristics, treatment outcomes, and use of restrictive interventions for those who received a specialist mental health (MH) assessment with those who did not. Of the 457 patients, 61.5% received a specialist MH assessment. This group had a lower acuity (Australasian Triage Score 10.4%; CI 0.2-2.0% vs 13.6%; CI 9.3-19.5%); more arrived with police (28.8%; CI 23.8-34.3 vs 5.1%; CI 2.7-9.4%); and were subjected to restrictive interventions while in the BAU. Security responses for unarmed threat (code grey) were higher (10.9%; CI 7.8-15.0% vs 4.4%; CI 2.3-8.5%), as was the use of chemical restraint (4.2%; CI 2.4-7.2 vs 0.0% CI 0.0 - 2.1%). Those requiring specialist MH assessment had a longer length of stay (12.7 vs 5.2 hours). Further development of the BAU model of care must include targeted, evidence-based strategies to minimize the use of restrictive interventions and ensure timely access to acute mental health services.
Assuntos
Serviços de Saúde Mental , Saúde Mental , Adulto , Serviço Hospitalar de Emergência , Hospitalização , Humanos , Tempo de Internação , TriagemRESUMO
In chronic obstructive pulmonary disease (COPD) apoptotic bronchial epithelial cells are increased, and their phagocytosis by alveolar macrophages (AM) is decreased alongside bacterial phagocytosis. Epithelial cellular lipids, including those exposed on uncleared apoptotic bodies, can become oxidized, and may be recognized and presented as non-self by antigen presenting cells. CD1b is a lipid-presenting protein, previously only described in dendritic cells. We investigated whether CD1b is upregulated in COPD AM, and whether lipid oxidation products are found in the airways of cigarette smoke (CS) exposed mice. We also characterise CD1b for the first time in a range of macrophages and assess CD1b expression and phagocytic function in response to oxidised lipid. Bronchoalveolar lavage and exhaled breath condensate were collected from never-smoker, current-smoker, and COPD patients and AM CD1b expression and airway 8-isoprostane levels assessed. Malondialdehyde was measured in CS-exposed mouse airways by confocal/immunofluorescence. Oxidation of lipids produced from CS-exposed 16HBE14o- (HBE) bronchial epithelial cells was assessed by spectrophotometry and changes in lipid classes assessed by mass spectrometry. 16HBE cell toxicity was measured by flow cytometry as was phagocytosis, CD1b expression, HLA class I/II, and mannose receptor (MR) in monocyte derived macrophages (MDM). AM CD1b was significantly increased in COPD smokers (4.5 fold), COPD ex-smokers (4.3 fold), and smokers (3.9 fold), and AM CD1b significantly correlated with disease severity (FEV1) and smoking pack years. Airway 8-isoprostane also increased in smokers and COPD smokers and ex-smokers. Malondialdehyde was significantly increased in the bronchial epithelium of CS-exposed mice (MFI of 18.18 vs 23.50 for control). Oxidised lipid was produced from CS-exposed bronchial epithelial cells (9.8-fold of control) and showed a different overall lipid makeup to that of control total cellular lipid. This oxidised epithelial lipid significantly upregulated MDM CD1b, caused bronchial epithelial cell toxicity, and reduced MDM phagocytic capacity and MR in a dose dependent manner. Increased levels of oxidised lipids in the airways of COPD patients may be responsible for reduced phagocytosis and may become a self-antigen to be presented by CD1b on macrophages to perpetuate disease progression despite smoking cessation.
Assuntos
Antígenos CD1/imunologia , Metabolismo dos Lipídeos , Macrófagos Alveolares/imunologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Líquido da Lavagem Broncoalveolar/citologia , Feminino , Citometria de Fluxo , Volume Expiratório Forçado , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Masculino , Malondialdeído/metabolismo , Espectrometria de Massas , Camundongos , Microscopia de Fluorescência , Pessoa de Meia-Idade , Oxirredução , Doença Pulmonar Obstrutiva Crônica/imunologia , Fumar/efeitos adversos , Poluição por Fumaça de Tabaco/efeitos adversos , Capacidade Vital , Adulto JovemRESUMO
The compound 4-hydroxynonenal (4-HNE) is the major aldehyde formed during lipid peroxidation of omega-6-polyunsaturated fatty acids and has been suggested to regulate inflammatory responses because it inhibits tumor necrosis factor (TNF) mRNA production in the human monocytic cell line THP-1. Here we demonstrate that 4-HNE inhibits TNF and interleukin-1beta production in human monocytes in response to lipopolysaccharide. The main action of 4-HNE occurred at the pretranscriptional level; there was no effect on TNF mRNA production or stability when 4-HNE was added after stimulation. The mechanism of action of 4-HNE appears to be downstream of lipopolysaccharide-receptor binding. In the human monocytic MonoMac 6 cell line, 4-HNE caused selective inhibition of the activity of the mitogen-activated protein kinases p38 and ERK1/ERK2, but not JNK. However, in monocytes, the activities of all three kinases were inhibited, suggesting that the effects of 4-HNE were exerted at points upstream of ERK1/ERK2 and JNK as the levels of the phosphorylated kinases were reduced. In contrast, p38 phosphorylation was not inhibited, suggesting that 4-HNE affects kinase activity. 4-HNE also inhibited nuclear factor-kappaB activation in monocytes. In view of the roles of p38, ERK1/ERK2, JNK, and nuclear factor-kappaB in inflammation, the data suggest that 4-HNE, at nontoxic concentrations, has anti-inflammatory properties, most likely through an effect on these signaling molecules, and could lead to the development of novel treatments for inflammatory diseases.
Assuntos
Aldeídos/farmacologia , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/enzimologia , Linhagem Celular , Ativação Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Ácidos Graxos Ômega-6/metabolismo , Humanos , Proteínas I-kappa B/metabolismo , Interleucina-1beta/biossíntese , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Oxirredução/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estabilidade de RNA/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: This study evaluated the feasibility of achieving high response rates in stage II or III breast cancer by tailoring neoadjuvant therapy using clinical and histopathological features and the Oncotype DX Breast Recurrence Score. Genomic determinants of response and resistance were also explored. PATIENTS AND OUTCOME MEASURES: Fifty-one patients were enrolled. The primary cohort comprised 40 patients: 15 human epidermal growth factor receptor type 2 (HER2)-amplified; 15 triple-negative (TNBC); and ten hormone receptor (HR)-positive, HER2-non-amplified tumours; with recurrence scores ≥25. Patients were treated with epirubicin and cyclophosphamide, followed by nab-paclitaxel, with the addition of trastuzumab if HER2-amplified. The primary endpoint was pathological complete response (pCR) in the breast. Pre- and post-treatment tumour samples underwent variant burden, gene and gene pathway, mutational signature profile and clonal evolution analyses. RESULTS: The pCR rates were: overall 55% (n = 22), HER2-amplified 80% (n = 12), triple-negative 46% (n = 7) and HR-positive, HER2-non-amplified 30% (n = 3). Grade 3 or 4 adverse events included febrile neutropenia (8%), neutropenia (18%), sensory neuropathy (5%), deranged transaminases (5%), fatigue (2%), diarrhoea (2%), and pneumothorax (2%). Molecular analyses demonstrated strong similarities between residual disease and matched primary tumour. ATM signalling pathway alterations and the presence of a COSMIC Signature 3 implied the majority of tumours contained some form of homologous repair deficiency. ATM pathway alterations were identified in the subset of TNBC patients who did not achieve pCR; Signature 3 was present in both pCR and non-pCR subgroups. Clonal evolution analyses demonstrated both persistence and emergence of chemoresistant clones. CONCLUSIONS: This treatment regime resulted in a high rate of pCR, demonstrating that tailored neoadjuvant therapy using a genomic recurrence score is feasible and warrants further investigation. Molecular analysis revealed few commonalities between patients. For TNBC future clinical gains will require precision medicine, potentially using DNA sequencing to identify specific targets for individuals with resistant disease. TRIAL REGISTRATION: Clinicaltrials.gov NCT01830244.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Terapia Neoadjuvante , Receptor ErbB-2 , Neoplasias de Mama Triplo Negativas , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Ciclofosfamida/administração & dosagem , Ciclofosfamida/efeitos adversos , Epirubicina/administração & dosagem , Epirubicina/efeitos adversos , Feminino , Seguimentos , Humanos , Pessoa de Meia-Idade , Nanopartículas/uso terapêutico , Paclitaxel/administração & dosagem , Paclitaxel/efeitos adversos , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Albumina Sérica Humana/administração & dosagem , Albumina Sérica Humana/efeitos adversos , Trastuzumab/administração & dosagem , Trastuzumab/efeitos adversos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismoRESUMO
Despite anti-TNF therapy advancements for inflammatory diseases such as rheumatoid arthritis, the burden of diseases remains high. An 11-mer TNF peptide, TNF70-80, is known to stimulate selective functional responses compared to the parent TNF molecule. Here, we show that TNF70-80 binds to the TNF receptor, activating p38 MAP kinase through TNF receptor-associated factor 2. Using truncated TNFR mutants, we identify the sequence in TNFRI which enables p38 activation by TNF70-80. Peptides with this TNFRI sequence, such as TNFRI206-211 bind to TNF and inhibit TNF-induced p38 activation, respiratory burst, cytokine production and adhesion receptor expression but not F-Met-Leu-Phe-induced respiratory burst in neutrophils. TNFRI206-211 does not prevent TNF binding to TNFRI or TNF-induced stimulation of ERK, JNK and NF-κB. TNFRI206-211 inhibits bacterial lipopolysaccharide-induced peritonitis, carrageenan-induced and antigen-induced paw inflammation, and respiratory syncytial virus-induced lung inflammation in mice. Our findings suggest a way of targeting TNF-p38 pathway to treat chronic inflammatory disorders.
Assuntos
Anti-Inflamatórios/farmacologia , Artrite Experimental/tratamento farmacológico , Fragmentos de Peptídeos/farmacologia , Peritonite/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Pneumonia/tratamento farmacológico , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/metabolismo , Artrite Experimental/genética , Artrite Experimental/imunologia , Artrite Experimental/patologia , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Humanos , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Fragmentos de Peptídeos/biossíntese , Fragmentos de Peptídeos/genética , Peritonite/genética , Peritonite/imunologia , Peritonite/patologia , Complexo Glicoproteico GPIb-IX de Plaquetas/genética , Complexo Glicoproteico GPIb-IX de Plaquetas/imunologia , Pneumonia/genética , Pneumonia/imunologia , Pneumonia/patologia , Pneumonia Viral/genética , Pneumonia Viral/imunologia , Pneumonia Viral/patologia , Ligação Proteica , Receptores do Fator de Necrose Tumoral/antagonistas & inibidores , Receptores do Fator de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/imunologia , Explosão Respiratória/efeitos dos fármacos , Vírus Sinciciais Respiratórios/imunologia , Vírus Sinciciais Respiratórios/patogenicidade , Transdução de Sinais , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Our previous studies have shown that nutritional zinc restriction exacerbates airway inflammation accompanied by an increase in caspase-3 activation and an accumulation of apoptotic epithelial cells in the bronchioles of the mice. Normally, apoptotic cells are rapidly cleared by macrophage efferocytosis, limiting any secondary necrosis and inflammation. We therefore hypothesized that zinc deficiency is not only pro-apoptotic but also impairs macrophage efferocytosis. Impaired efferocytic clearance of apoptotic epithelial cells by alveolar macrophages occurs in chronic obstructive pulmonary disease (COPD), cigarette-smoking and other lung inflammatory diseases. We now show that zinc is a factor in impaired macrophage efferocytosis in COPD. Concentrations of zinc were significantly reduced in the supernatant of bronchoalveolar lavage fluid of patients with COPD who were current smokers, compared to healthy controls, smokers or COPD patients not actively smoking. Lavage zinc was positively correlated with AM efferocytosis and there was decreased efferocytosis in macrophages depleted of Zn in vitro by treatment with the membrane-permeable zinc chelator TPEN. Organ and cell Zn homeostasis are mediated by two families of membrane ZIP and ZnT proteins. Macrophages of mice null for ZIP1 had significantly lower intracellular zinc and efferocytosis capability, suggesting ZIP1 may play an important role. We investigated further using the human THP-1 derived macrophage cell line, with and without zinc chelation by TPEN to mimic zinc deficiency. There was no change in ZIP1 mRNA levels by TPEN but a significant 3-fold increase in expression of another influx transporter ZIP2, consistent with a role for ZIP2 in maintaining macrophage Zn levels. Both ZIP1 and ZIP2 proteins were localized to the plasma membrane and cytoplasm in normal human lung alveolar macrophages. We propose that zinc homeostasis in macrophages involves the coordinated action of ZIP1 and ZIP2 transporters responding differently to zinc deficiency signals and that these play important roles in macrophage efferocytosis.
Assuntos
Proteínas de Transporte/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Fagocitose/imunologia , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Zinco/metabolismo , Animais , Líquido da Lavagem Broncoalveolar , Proteínas de Transporte/genética , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Linhagem Celular , Citosol/metabolismo , Modelos Animais de Doenças , Etilenodiaminas/farmacologia , Feminino , Expressão Gênica , Humanos , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/metabolismo , Camundongos , Camundongos Knockout , Doença Pulmonar Obstrutiva Crônica/genéticaRESUMO
In chronic obstructive pulmonary disease (COPD/emphysema) we have shown a reduced ability of lung and alveolar (AM) macrophages to phagocytose apoptotic cells (defective 'efferocytosis'), associated with evidence of secondary cellular necrosis and a resultant inflammatory response in the airway. It is unknown whether this defect is present in cancer (no COPD) and if so, whether this results from soluble mediators produced by cancer cells. We investigated efferocytosis in AM (26 controls, 15 healthy smokers, 37 COPD, 20 COPD+ non small cell lung cancer (NSCLC) and 8 patients with NSCLC without COPD) and tumor and tumor-free lung tissue macrophages (21 NSCLC with/13 without COPD). To investigate the effects of soluble mediators produced by lung cancer cells we then treated AM or U937 macrophages with cancer cell line supernatant and assessed their efferocytosis ability. We qualitatively identified Arachidonic Acid (AA) metabolites in cancer cells by LC-ESI-MSMS, and assessed the effects of COX inhibition (using indomethacin) on efferocytosis. Decreased efferocytosis was noted in all cancer/COPD groups in all compartments. Conditioned media from cancer cell cultures decreased the efferocytosis ability of both AM and U937 macrophages with the most pronounced effects occurring with supernatant from SCLC (an aggressive lung cancer type). AA metabolites identified in cancer cells included PGE2. The inhibitory effect of PGE2 on efferocytosis, and the involvement of the COX-2 pathway were shown. Efferocytosis is decreased in COPD/emphysema and lung cancer; the latter at least partially a result of inhibition by soluble mediators produced by cancer cells that include PGE2.
Assuntos
Dinoprostona/biossíntese , Neoplasias Pulmonares/complicações , Pulmão/metabolismo , Pulmão/patologia , Macrófagos Alveolares/patologia , Doença Pulmonar Obstrutiva Crônica/complicações , Enfisema Pulmonar/complicações , Adulto , Idoso , Líquido da Lavagem Broncoalveolar , Linhagem Celular Tumoral , Ciclo-Oxigenase 2/metabolismo , Demografia , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Fagocitose , Doença Pulmonar Obstrutiva Crônica/patologia , Enfisema Pulmonar/patologia , Frações Subcelulares/metabolismoRESUMO
We have previously shown that the defective ability of alveolar macrophages (AM) to phagocytose apoptotic cells ('efferocytosis') in chronic obstructive pulmonary disease/emphysema (COPD) could be therapeutically improved using the C-type lectin, mannose binding lectin (MBL), although the exact mechanisms underlying this effect are unknown. An S-type lectin, galectin-3, is also known to regulate macrophage phenotype and function, via interaction with its receptor CD98. We hypothesized that defective expression of galectin/CD98 would be associated with defective efferocytosis in COPD and that mechanisms would include effects on cytoskeletal remodeling and macrophage phenotype and glutathione (GSH) availability. Galectin-3 was measured by ELISA in BAL from controls, smokers and current/ex-smokers with COPD. CD98 was measured on AM using flow cytometry. We assessed the effects of galectin-3 on efferocytosis, CD98, GSH, actin polymerisation, rac activation, and the involvement of PI3K (using ß-actin probing and wortmannin inhibition) in vitro using human AM and/or MH-S macrophage cell line. Significant decreases in BAL galectin-3 and AM CD98 were observed in BAL from both current- and ex-smoker COPD subjects vs controls. Galectin 3 increased efferocytosis via an increase in active GTP bound Rac1. This was confirmed with ß-actin probing and the role of PI3K was confirmed using wortmannin inhibition. The increased efferocytosis was associated with increases in available glutathione and expression of CD98. We provide evidence for a role of airway lectins in the failed efferocytosis in COPD, supporting their further investigation as potential macrophage-targeted therapies.
Assuntos
Lectinas/metabolismo , Macrófagos Alveolares/imunologia , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Enfisema Pulmonar/imunologia , Enfisema Pulmonar/metabolismo , Actinas/metabolismo , Adulto , Idoso , Androstadienos/farmacologia , Animais , Arginase/metabolismo , Líquido da Lavagem Broncoalveolar/imunologia , Feminino , Proteína-1 Reguladora de Fusão/metabolismo , Galectina 3/metabolismo , Galectina 3/farmacologia , Glutationa/metabolismo , Humanos , Lectinas/farmacologia , Macrófagos Alveolares/efeitos dos fármacos , Macrófagos Alveolares/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Fosfatidilinositol 3-Quinases/metabolismo , Fumar , Wortmanina , Proteínas rac1 de Ligação ao GTP/metabolismoRESUMO
Cytotoxic CD8(+) T-cells mount immune responses to cancer via cytotoxic pathways including granzyme B. Cancer cells are also known to develop immune evasion mechanisms. We hypothesised that lung cancer cells would over-express the granzyme B-inhibitor, proteinase inhibitor-9 (PI-9) and down-regulate granzyme B expression by neighbouring CD8(+) T-cells. We investigated PI-9 expression in lung cancer cell lines, and primary lung cancer cells obtained at curative lung resection from cancer patients with/without chronic obstructive pulmonary disease (COPD). Granzyme B and PI-9 expression was also determined in CD8(+) T-cells from the cancer and non-cancer areas of resected lung tissue and from bronchoalveolar lavage (BAL). We then evaluated the effects of conditioned media from lung cancer cell lines on granzyme B expression and the cytotoxic activity of CD8(+) T-cells. PI-9 was highly expressed in lung cancer cell lines. Increased PI-9 expression was also observed in primary cancer cells vs. epithelial cells from non-cancer tissue or bronchial brushing-derived normal primary large airway epithelial cells. Expression significantly correlated with cancer stage. Significantly reduced granzyme B was noted in CD8(+) T-cells from cancer vs. non-cancer tissue. Granzyme B production by CD8(+) T-cells was reduced in the presence of conditioned media from lung cancer cell lines. Our data suggest that lung cancer cells utilise their increased PI-9 expression to protect from granzyme B-mediated cytotoxicity as an immune evasion mechanism, a function that increases with lung cancer stage.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Granzimas/metabolismo , Neoplasias Pulmonares/metabolismo , Serpinas/metabolismo , Evasão Tumoral , Idoso , Líquido da Lavagem Broncoalveolar , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Carcinoma Pulmonar de Células não Pequenas/complicações , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Casos e Controles , Extratos Celulares/farmacologia , Linhagem Celular Tumoral , Meios de Cultivo Condicionados , Citotoxicidade Imunológica , Feminino , Humanos , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/complicações , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Doença Pulmonar Obstrutiva Crônica/complicações , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologiaRESUMO
Apoptosis of bronchial epithelial cells and the phagocytic clearance of these cells by alveolar macrophages (a process termed efferocytosis) are integral processes leading to repair of airway epithelial injury. Efferocytosis allows for the removal of apoptotic material with minimal inflammation and prevents the development of secondary necrosis and ongoing inflammation. Defective efferocytosis and the increased presence of apoptotic cells have been identified in the airways of subjects with chronic obstructive pulmonary disease (COPD). There are three major potential causes for this accumulation of apoptotic cells: (i) increased apoptosis per se as a result of an increase in apoptotic mediators, (ii) defects in the recognition of apoptotic cells by AM and (iii) failure to clear the unwanted cells by the process of efferocytosis. The implications of these processes in COPD and novel treatment strategies aimed at improving clearance of apoptotic cells form the focus of the present review.
Assuntos
Apoptose , Fagocitose , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Animais , Sistemas de Liberação de Medicamentos , Humanos , Inflamação/fisiopatologia , NecroseRESUMO
The role of JNK in neutrophil chemotaxis and killing of microbial pathogens remains unclear. Using a recently described cell-permeable peptide inhibitor of the JNK pathway, based on the JBD of JIP-1, coupled to the protein transduction domain of HIV-TAT (TAT-JIP), in association with control peptides, we demonstrate that the JNK pathway plays a major role in regulating human neutrophil chemotaxis and killing of microbial pathogens. Serum-opsonized Staphylococcus aureus elicited JNK activation and c-jun phosphorylation. The activation of the JNK pathway and bactericidal activity were inhibited by the TAT-JIP peptide. The stimulation of oxygen radical generation by S. aureus was dependent on the JNK signaling pathway, as was the phagocytosis of serum-opsonized bacteria. Chemotaxis to activated serum complement but not random migration was inhibited by the TAT-JIP peptide. The findings demonstrate a major role for the JNK signaling pathway in neutrophil-mediated defense against microbial pathogens.
Assuntos
Quimiotaxia de Leucócito/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Neutrófilos/imunologia , Infecções Estafilocócicas/imunologia , Western Blotting , Quimiotaxia de Leucócito/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Células HL-60 , Humanos , Imunoprecipitação , Neutrófilos/efeitos dos fármacos , Neutrófilos/enzimologia , Fagocitose/imunologia , Staphylococcus aureus/imunologiaRESUMO
INTRODUCTION: While consumption of omega-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) has been recommended for those at risk of inflammatory disease such as rheumatoid arthritis, the mechanism of their anti-inflammatory effect remains to be clearly defined, particularly in relation to the dose and type of n-3 LCPUFA. The objective of this study was to determine whether varying the levels of n-3 LCPUFA in erythrocyte membrane lipids, following dietary supplementation, is associated with altered numbers and function of circulating leukocytes conducive to protection against inflammation. METHODS: In a double-blind and placebo-controlled study, 44 healthy subjects aged 23 to 63 years consumed either standard or n-3 LCPUFA-enriched versions of typical processed foods, the latter allowing a target daily consumption of 1 gram n-3 LCPUFA. After six months, peripheral blood leukocyte and subpopulation proportions and numbers were assessed by flow cytometry. Leukocytes were also examined for lymphoproliferation and cytokine production, neutrophil chemotaxis, chemokinesis, bactericidal, adherence and iodination activity. Erythrocytes were analyzed for fatty-acid content. RESULTS: Erythrocyte n-3 LCPUFA levels were higher and absolute leukocyte and lymphocyte numbers were lower in subjects consuming n-3 enriched foods than in controls. There were no changes in the number of neutrophils, monocytes, T cells (CD3+), T-cell subsets (CD4+, CD8+) and B cells (CD19+). However, natural killer (NK) (CD3-CD16+CD56+) cell numbers were lower in n-3 supplemented subjects than in controls and were inversely related to the amount of eicosapentaenoic acid or docosahexaenoic acid in erythrocytes. No significant correlations were found with respect to lymphocyte lymphoproliferation and production of IFN-gamma and IL-2, but lymphotoxin production was higher with greater n-3 LCPUFA membrane content. Similarly, neutrophil chemotaxis, chemokinesis, bactericidal activity and adherence did not vary with changes in erythrocyte n-3 LCPUFA levels, but the iodination reaction was reduced with higher n-3 LCPUFA content. CONCLUSION: The data show that regular long-term consumption of n-3 enriched foods leads to lower numbers of NK cells and neutrophil iodination activity but higher lymphotoxin production by lymphocytes. These changes are consistent with decreased inflammatory reaction and tissue damage seen in patients with inflammatory disorders receiving n-3 LCPUFA supplementation.
Assuntos
Gorduras Insaturadas na Dieta/administração & dosagem , Ingestão de Alimentos/imunologia , Ácidos Graxos Ômega-3/administração & dosagem , Alimentos Fortificados , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Adulto , Idoso , Método Duplo-Cego , Humanos , Terapia de Imunossupressão , Células Matadoras Naturais/citologia , Contagem de Linfócitos/métodos , Pessoa de Meia-IdadeRESUMO
The role of the extracellular signal-regulated kinase (ERK) 1 and ERK2 in the neutrophil chemotactic response remains to be identified since a previously used specific inhibitor of MEK1 and MEK2, PD98059, that was used to provide evidence for a role of ERK1 and ERK2 in regulating chemotaxis, has recently been reported to also inhibit MEK5. This issue is made more critical by our present finding that human neutrophils express mitogen-activated protein (MAP) kinase/ERK kinase (MEK)5 and ERK5 (Big MAP kinase), and that their activities were stimulated by the bacterial tripeptide, formyl methionyl-leucyl-phenylalanine (fMLP). Dose response studies demonstrated a bell-shaped profile of fMLP-stimulated MEK5 and ERK5 activation, but this was left-shifted when compared with the profile of fMLP-stimulated chemotaxis. Kinetics studies demonstrated increases in kinase activity within 2 min, peaking at 3-5 min, and MEK5 activation was more persistent than that of ERK5. There were some similarities as well as differences in the pattern of activation between fMLP-stimulated ERK1 and ERK2, and MEK5-ERK5 activation. The up-regulation of MEK5-ERK5 activities was dependent on phosphatidylinositol 3-kinase. Studies with the recently described specific MEK inhibitor, PD184352, at concentrations that inhibited ERK1 and ERK2 but not ERK5 activity demonstrate that the ERK1 and ERK2 modules were involved in regulating fMLP-stimulated chemotaxis and chemokinesis. Our data suggest that the MEK5-ERK5 module is likely to regulate neutrophil responses at very low chemoattractant concentrations whereas at higher concentrations, a shift to the ERK1/ERK2 and p38 modules is apparent.