Leukotrienes Produced in Allergic Lung Inflammation Activate Alveolar Macrophages

It has been well-documented that leukotrienes (LTs) are released in allergic lung inflammation and that they participate in the physiopathology of asthma. A role for LTs in innate immunity has recently emerged: Cys-LTs were shown to enhance Fc gamma R-mediated phagocytosis by alveolar macrophages (AMs). Thus, using a rat model of asthma, we evaluated Fc gamma R-mediated phagocytosis and killing of Klebsiella pneumoniae by AMs. The effect of treatment with a cys-LT antagonist (montelukast) on macrophage function was also investigated. Male Wistar rats were immunized twice with OVA/alumen intraperitoneally and challenged with OVA aerosol. After 24 h, the animals were killed, and the AMs were obtained by bronchoalveolar lavage. Macrophages were cultured with IgG-opsonized red blood cells (50: 1) or IgG-opsonized K. pneumoniae (30: 1), and phagocytosis or killing was evaluated. Leukotriene C(4) and nitric oxide were quantified by the EIA and Griess methods, respectively. The results showed that AMs from sensitized and challenged rats presented a markedly increased phagocytic capacity via Fc gamma R (10X compared to controls) and enhanced killing of K. pneumoniae (4X higher than controls). The increased phagocytosis was inhibited 15X and killing 3X by treatment of the rats with montelukast, as compared to the non-treated group. cys-LT addition increased phagocytosis in control AMs but had no effect on macrophages from allergic lungs. Montelukast reduced nitric oxide (39%) and LTC(4) (73%). These results suggest that LTs produced during allergic lung inflammation potentiate the capacity of AMs to phagocytose and kill K. pneumonia via Fc gamma R. Copyright (C) 2010 S. Karger AG, Basel

INSULIN REGULATES CYTOKINES AND INTERCELLULAR ADHESION MOLECULE-1 GENE EXPRESSION THROUGH NUCLEAR FACTOR-kappa B ACTIVATION IN LPS-INDUCED ACUTE LUNG INJURY IN RATS

Diabetic patients have increased susceptibility to infection, which may be related to impaired inflammatory response observed in experimental models of diabetes, and restored by insulin treatment. The goal of this study was to investigate whether insulin regulates transcription of cytokines and intercellular adhesion molecule 1 (ICAM-1) via nuclear factor-kappa B (NF-kappa B) signaling pathway in Escherichia coli LIPS-induced lung inflammation. Diabetic male Wistar rats (alloxan, 42 mg/kg, iv., 10 days) and controls were instilled intratracheally with saline containing LPS (750 mu g/0.4 mL) or saline only. Some diabetic rats were given neutral protamine Hagedorn insulin (4 IU, s.c.) 2 h before LIPS. Analyses performed 6 h after LPS included: (a) lung and mesenteric lymph node IL-1 beta, TNF-alpha, IL-10, and ICAM-1 messenger RNA (mRNA) were quantified by real-time reverse transcriptase-polymerase chain reaction; (b) number of neutrophils in the bronchoalveolar lavage (BAL) fluid, and concentrations of IL-1 beta, TNF-alpha, and IL-10 in the BAL were determined by the enzyme-linked immunosorbent assay; and (c) activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were quantified by Western blot analysis. Relative to controls, diabetic rats exhibited a reduction in lung and mesenteric lymph node IL-1 beta (40%), TNF-alpha (similar to 30%), and IL-10 (similar to 40%) mRNA levels and reduced concentrations of IL-1 beta (52%), TNF-alpha (62%), IL-10 (43%), and neutrophil counts (72%) in the BAL. Activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were almost suppressed in diabetic rats. Treatment of diabetic rats with insulin completely restored mRNA and protein levels of these cytokines and potentiated lung ICAM-1 mRNA levels (30%) and number of neutrophils (72%) in the BAL. Activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were partially restored by insulin treatment. In conclusion, data presented suggest that insulin regulates transcription of proinflammatory (IL-1 beta, TNF-alpha) and anti-inflammatory (IL-10) cytokines, and expression of ICAM-1 via the NF-kappa B signaling pathway.

Improved renal function after kidney transplantation is associated with heme oxygenase-1 polymorphism

Heme oxygenase-1 (HO-1) has a microsatellite polymorphism based on the number of guanosine-thymidine nucleotide repeats (GT) repeats that regulates expression levels and could have an impact on organ survival post-injury. We correlated HO-1 polymorphism with renal graft function. The HO-1 gene was sequenced (N = 181), and the allelic repeats were divided into subclasses: short repeats (S) (< 27 repeats) and long repeats (L) (>= 27 repeats). A total of 47.5% of the donors carried the S allele. The allograft function was statistically improved six months, two and three yr after transplantation in patients receiving kidneys from donors with an S allele. For the recipients carrying the S allele (50.3%), the allograft function was also better throughout the follow-up, but reached statistical significance only three yr after transplantation (p = 0.04). Considering only those patients who had chronic allograft nephropathy (CAN; 74 of 181), allograft function was also better in donors and in recipients carrying the S allele, two and three yr after transplantation (p = 0.03). Recipients of kidney transplantation from donors carrying the S allele presented better function even in the presence of CAN.

Reduced allergic lung inflammation in rats following formaldehyde exposure: Long-term effects on multiple effector systems

Clinical and experimental evidences show that formaldehyde (FA) exposure has an irritant effect on the upper airways. As being an indoor and outdoor pollutant, FA is known to be a causal factor of occupational asthma. This study aimed to investigate the repercussion of FA exposure on the course of a lung allergic process triggered by an antigen unrelated to FA. For this purpose, male Wistar rats were subjected to FA inhalation for 3 consecutive days (1%, 90-min daily), subsequently sensitized with ovalbumin (OVA)-alum via the intraperitoneal route, and 2 weeks later challenged with aerosolized OVA. The OVA challenge in rats after FA inhalation (FA/OVA group) evoked a low-intensity lung inflammation as indicated by the reduced enumerated number of inflammatory cells in bronchoalveolar lavage as compared to FA-untreated allergic rats (OVA/OVA group). Treatment with FA also reduced the number of bone marrow cells and blood leukocytes in sensitized animals challenged with OVA, which suggests that the effects of FA had not been only localized to the airways. As indicated by passive cutaneous anaphylactic reaction, FA treatment did not impair the anti-OVA IgE synthesis, but reduced the magnitude of OVA challenge-induced mast cell degranulation. Moreover, FA treatment was associated to a diminished lung expression of PECAM-1 (platelet-endothelial cell adhesion molecule 1) in lung endothelial cells after OVA challenge and an exacerbated release of nitrites by BAL-cultured cells. Keeping in mind that rats subjected solely to either FA or OVA challenge were able to significantly increase the cell influx into lung, our study shows that FA inhalation triggers long-lasting effects that affect multiple mediator systems associated to OVA-induced allergic lung such as the reduction of mast cells activation, PECAM-1 expression and exacerbation of NO generation, thereby contributing to the decrease of cell recruitment after the OVA challenge. In conclusion, repeated expositions to air-borne FA may impair the lung cell recruitment after an allergic stimulus, thereby leading to a non-responsive condition against inflammatory stimuli likely those where mast cells are involved. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

High frequency of immature dendritic cells and altered in situ production of interleukin-4 and tumor necrosis factor-alpha in lung cancer

Introduction Antigen-presenting cells, like dendritic cells (DCs) and macrophages, play a significant role in the induction of an immune response and an imbalance in the proportion of macrophages, immature and mature DCs within the tumor could affect significantly the immune response to cancer. DCs and macrophages can differentiate from monocytes, depending on the milieu, where cytokines, like interleukin (IL)-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) induce DC differentiation and tumor necrosis factor (TNF)-alpha induce DC maturation. Thus, the aim of this work was to analyze by immunohistochemistry the presence of DCs (S100+ or CD1a+), macrophages (CD68+), IL-4 and TNF-alpha within the microenvironment of primary lung carcinomas. Results Higher frequencies of both immature DCs and macrophages were detected in the tumor-affected lung, when compared to the non-affected lung. Also, TNF-alpha-positive cells were more frequent, while IL-4-positive cells were less frequent in neoplastic tissues. This decreased frequency of mature DCs within the tumor was further confirmed by the lower frequency of CD14-CD80+ cells in cell suspensions obtained from the same lung tissues analyzed by flow cytometry. Conclusion These data are discussed and interpreted as the result of an environment that does not oppose monocyte differentiation into DCs, but that could impair DC maturation, thus affecting the induction of effective immune responses against the tumor.

Low level laser therapy (LLLT) decreases pulmonary microvascular leakage, neutrophil influx and IL-1 beta levels in airway and lung from rat subjected to LPS-induced inflammation

Background and Objective. Low level laser therapy (LLLT) is a known anti-inflammatory therapy. Herein we studied the effect of LLLT on lung permeability and the IL-1 beta level in LPS-induced pulmonary inflammation. Study Design/Methodology. Rats were divided into 12 groups (n = 7 for each group). Lung permeability was measured by quantifying extravasated albumin concentration in lung homogenate, inflammatory cells influx was determined by myeloperoxidase activity, IL-1P in BAL was determined by ELISA and IL-1P mRNA expression in trachea was evaluated by RT-PCR. The rats were irradiated on the skin over the upper bronchus at the site of tracheotomy after LPS. Results. LLLT attenuated lung permeability. In addition, there was reduced neutrophil influx, myeloperoxidase activity and both IL-1 beta in BAL and IL-1 beta mRNA expression in trachea obtained from animals subjected to LPS-induced inflammation. Conclusion. LLLT reduced the lung permeability by a mechanism in which the IL-1 beta seems to have an important role.

Enhancement of Th1 Lung Immunity Induced by Recombinant Mycobacterium bovis Bacillus Calmette-Guerin Attenuates Airway Allergic Disease

Mycobacterium bovis Bacillus Calmette-Guerin (BCG) has been shown to down-regulate experimental allergic asthma, a finding that reinforced the hygiene hypothesis. We have previously found that recombinant BCG (rBCG) strain that express the genetically detoxified Si subunit of pertussis toxin (rBCG-S1PT) exerts an adjuvant effect that enhances Th1 responses against BCG proteins. Here we investigated the effect of this rBCG-S1PT on the classical ovalbumin-induced mouse model of allergic lung disease. We found that rBCG-S1PT was more effective than wild-type BCG in preventing Th2-mediated allergic immune responses. The inhibition of allergic lung disease was not associated with increased concentration of suppressive cytokines or with an increased number of pulmonary regulatory T cells but was positively correlated with the increase in IFN-gamma-producing T cells and T-bet expression in the lung. In addition, an IL-12-dependent mechanism appeared to be important to the inhibition of lung allergic disease. The inhibition of allergic inflammation was found to be restricted to the lung because when allergen challenge was given by the intraperitoneal route, rBCG-S1PT administration failed to inhibit peritoneal allergic inflammation and type 2 cytokine production. Our work offers a nonclassical interpretation for the hygiene hypothesis indicating that attenuation of lung allergy by rBCG could be due to the enhancement of local lung Th1 immunity induced by rBCG-S1PT. Moreover, it highlights the possible use of rBCG strains as multipurpose immunomodulators by inducing specific immunity against microbial products while protecting against allergic asthma.

Bradykinin inducible receptor is essential to lipopolysaccharide-induced acute lung injury in mice

Lipopolysaccharides from gram-negative bacteria are amongst the most common causative agents of acute lung injury, which is characterized by an inflammatory response, with cellular infiltration and the release of mediators/cytokines. There is evidence that bradykinin plays a role in lung inflammation in asthma but in other types of lung inflammation its role is less clear. In the present study we evaluated the role of the bradykinin B(1) receptor in acute lung injury caused by lipopolysaccharide inhalation and the mechanisms behind bradykinin actions participating in the inflammatory response. We found that in C57BI/6 mice, the bradykinin B(1) receptor expression was up-regulated 24 h after lipopolysaccharide inhalation. At this time, the number of cells and protein concentration were significantly increased in the bronchoalveolar lavage fluid and the mice developed airway hyperreactivity to methacholine. In addition, there was an increased expression of tumor necrosis factor-alpha, interleukin-1 beta and interferon-gamma and chemokines (monocytes chemotactic protein-1 and KC) in the bronchoalveolar lavage fluid and in the lung tissue. We then treated the mice with a bradykinin B, receptor antagonist, R-954 (Ac-Orn-[Oic(2), alpha-MePhe(5), D-beta Nal(7), Ile(8)]desArg(9)-bradykinin), 30 min after lipopolysaccharide administration. We observed that this treatment prevented the airway hyperreactivity as well as the increased cellular infiltration and protein content in the bronchoalveolar lavage fluid. Moreover, R-954 inhibited the expression of cytokines/chemokines. These results implicate bradykinin, acting through B(1) receptor, in the development of acute lung injury caused by lipopolysaccharide inhalation. (C) 2010 Elsevier B.V. All rights reserved.

SIGNALING PATHWAYS AND MEDIATORS IN LPS-INDUCED LUNG INFLAMMATION IN DIABETIC RATS: ROLE OF INSULIN

Diabetic patients are more susceptible to infections, and their inflammatory response is impaired. This is restored by insulin treatment. In the present study, we investigated the effect of insulin on LPS-induced signaling pathways and mediators in the lung of diabetic rats. Diabetic male Wistar rats (alloxan, 42 mg/kg i.v., 10 days) and control rats received intratracheal instillation of LPS (750 mu g/0.4 mL) or saline. Some diabetic rats were given neutral protamine Hagedorn insulin (4 IU s.c.) 2 h before LPS. After 6 h, bronchoalveolar lavage was performed for the release of mediators, and lung tissue was homogenized for analysis of LPS-induced signaling pathways. Relative to control rats, diabetic rats exhibited a significant reduction in the LPS-induced phosphorylation of extracellular signal-regulated kinase (64%), p38 (70%), protein kinase B (67%), and protein kinase C alpha (57%) and delta (65%) and in the expression of iNOS (32%) and cyclooxygenase 2 (67%) in the lung homogenates. The bronchoalveolar lavage fluid concentrations of NO (47%) and IL-6 (49%) were also reduced in diabetic rats, whereas the cytokine-induced neutrophil chemoattractant 2 (CINC-2) levels were increased 23%, and CINC-1 was not different from control animals. Treatment of diabetic rats with insulin completely or partially restored all these parameters. In conclusion, data presented show that insulin regulates mitogen-activated protein kinase, phosphatidylinositol 3`-kinase, protein kinase C pathways, expression of the inducible enzymes, cyclooxygenase 2 and iNOS, and levels of IL-6 and CINC-2 in LPS-induced lung inflammation in diabetic rats. These results suggest that the protective effect of insulin in sepsis could be due to modulation of cellular signal transduction factors.

Effect of Age on Outcome of Reduced-Intensity Hematopoietic Cell Transplantation for Older Patients With Acute Myeloid Leukemia in First Complete Remission or With Myelodysplastic Syndrome

Purpose Acute myelogenous leukemia (AML) and myelodysplastic syndrome (MDS) primarily afflict older individuals. Hematopoietic cell transplantation (HCT) is generally not offered because of concerns of excess morbidity and mortality. Reduced-intensity conditioning (RIC) regimens allow increased use of allogeneic HCT for older patients. To define prognostic factors impacting long-term outcomes of RIC regimens in patients older than age 40 years with AML in first complete remission or MDS and to determine the impact of age, we analyzed data from the Center for International Blood and Marrow Transplant Research (CIBMTR). Patients and Methods We reviewed data reported to the CIBMTR (1995 to 2005) on 1,080 patients undergoing RIC HCT. Outcomes analyzed included neutrophil recovery, incidence of acute or chronic graft-versus-host disease (GVHD), nonrelapse mortality (NRM), relapse, disease-free survival (DFS), and overall survival (OS). Results Univariate analyses demonstrated no age group differences in NRM, grade 2 to 4 acute GVHD, chronic GVHD, or relapse. Patients age 40 to 54, 55 to 59, 60 to 64, and >= 65 years had 2-year survival rates as follows: 44% (95% Cl, 37% to 52%), 50% (95% Cl, 41% to 59%), 34% (95% Cl, 25% to 43%), and 36% (95% Cl, 24% to 49%), respectively, for patients with AML (P = .06); and 42% (95% Cl, 35% to 49%), 35% (95% Cl, 27% to 43%), 45% (95% Cl, 36% to 54%), and 38% (95% Cl, 25% to 51%), respectively, for patients with MDS (P = .37). Multivariate analysis revealed no significant impact of age on NRM, relapse, DFS, or OS (all P>.3). Greater HLA disparity adversely affected 2-year NRM, DFS, and OS. Unfavorable cytogenetics adversely impacted relapse, DFS, and OS. Better pre-HCT performance status predicted improved 2-year OS. Conclusion With these similar outcomes observed in older patients, we conclude that older age alone should not be considered a contraindication to HCT.

Autofluorescence spectroscopy in liver transplantation: Preliminary results from a pilot clinical study

The evaluation of graft function at various stages after transplantation is relevant, particularly at the moment of organ harvest, when a decision must be made whether to use the organ. Autofluorescence spectroscopy is noninvasive technique to monitor the metabolic condition of a liver graft throughout its course, from an initial evaluation in the donor, through cold ischemia transportation, to reperfusion and reoxygenation in the recipient. Preliminary results are presented in six liver transplantations spanning the periods from liver harvest to implant. The laser-induced fluorescence spectrum at 532-mn excitation was investigated before cold perfusion (autofluorescence), during cold ischemia, at the back table procedure, as well as 5 and 60 minutes after reperfusion. The results showed that the fluorescence analysis was sensitive to changes during the transplantation procedure. Fluorescence spectroscopy potentially provides a real-time, noninvasive technique to monitor liver graft function. The information could potentially be valuable for surgical decisions and transplant success.

Early Changes in Gene Expression Induced by Tobacco Smoke: Evidence for the Importance of Estrogen within Lung Tissue

Lung cancer is the leading cause of cancer deaths in the United States, surpassing breast cancer as the primary cause of cancer-related mortality in women. The goal of the present study was to identify early molecular changes in the lung induced by exposure to tobacco smoke and thus identify potential targets for chemoprevention. Female A/J mice were exposed to either tobacco smoke or HEPA-filtered air via a whole-body exposure chamber (6 h/d, 5 d/wk for 3, 8, and 20 weeks). Gene expression profiles of lung tissue from control and smoke-exposed animals were established using a 15K cDNA microarray. Cytochrome P450 1b1, a phase I enzyme involved in both the metabolism of xenobiotics and the 4-hydroxylation of 17 beta-estradiol (E(2)), was modulated to the greatest extent following smoke exposure. A panel of 10 genes were found to be differentially expressed in control and smoke-exposed lung tissues at 3, 8, and 20 weeks (P < 0.001). The interaction network of these differentially expressed genes revealed new pathways modulated by short-term smoke exposure, including estrogen metabolism. In addition, E(2) was detected within murine lung tissue by gas chromatography-coupled mass spectrometry and immunohistochemistry. Identification of the early molecular events that contribute to lung tumor formation is anticipated to lead to the development of promising targeted chemopreventive therapies. In conclusion, the presence of E2 within lung tissue when combined with the modulation of cytochrome P450 1b1 and other estrogen metabolism genes by tobacco smoke provides novel insight into a possible role for estrogens in lung cancer. Cancer Prev Res; 3(6); 707-17. (C) 2010 AACR.

Base Excision Repair Activities Differ in Human Lung Cancer Cells and Corresponding Normal Controls

Oxidative damage to DNA is thought to play a role in carcinogenesis by causing Mutations, and indeed accumulation of oxidized DNA bases has been observed in samples obtained from tumors but not from surrounding tissue within the same patient. Base excision repair (BER) is the main pathway for the repair of oxidized modifications both in nuclear and mitochondrial, DNA. In order to ascertain whether diminished BER capacity might account for increased levels of oxidative DNA damage in cancer cells, the activities of BER enzymes in three different lung cancer cell lines and their non-cancerous counterparts were measured using oligonucleotide substrates with single DNA lesions to assess specific BER enzymes. The activities of four BER enzymes, OGG1, NTH1, UDG and APE1, were compared in mitochondrial and nuclear extracts. For each specific lesion, the repair activities were similar among the three cell lines used. However, the specific activities and cancer versus control comparison differed significantly between the nuclear and mitochondrial compartments. OGG1 activity, as measured by 8-oxodA incision, was upregulated in cancer cell mitochondria but down-regulated in the nucleus when compared to control cells. Similarly, NTH1 activity was also up-regulated in mitochondrial extracts from cancer cells but did not change significantly in the nucleus. Together, these results support the idea that alterations in BER capacity are associated with carcinogenesis.

Cytotoxic and genotoxic effects of tambjamine D, an alkaloid isolated from the nudibranch Tambja eliora, on Chinese hamster lung fibroblasts

Marine organisms have been shown to be potential sources of bioactive compounds with pharmaceutical applications. Previous chemical investigation of the nudibranch Tambja eliora led to the isolation of the alkaloid tambjamine D. Tambjamines have been isolated from marine sources and belong to the family of 4-methoxypyrrolic-derived natural products, which display promising immunosuppressive and cytotoxic properties. Their ability to intercalate DNA and their pro-oxidant activity may be related to some of the biological effects of the 4-methoxypyrrolic alkaloids. The aim of the present investigation was to determine the cytotoxic, pro-oxidant and genotoxic properties of tambjamine D in V79 Chinese hamster lung fibroblast cells. Tambjamine D displayed a potent cytotoxic effect in V79 cells (IC50 1.2 mu g/mL) evaluated by the MTT assay. Based on the MTT result, V79 cells were treated with different concentrations of tambjamine D (0.6. 1.2. 2.4 and 4.8 mu g/mL). After 24 h, tambjamine D reduced the number of viable cells in a concentration-dependent way at all concentrations tested. assessed by the trypan blue dye exclusion test. The hemolytic assay showed that the cytotoxic activity of tambjamine D was not related to membrane disruption (EC50 > 100 mu g/mL). Tambjamine D increased the number of apoptotic cells in a concentration-dependent manner at all concentrations tested according to acridine orange/ethidium bromide staining, showing that the alkaloid cytotoxic effect was related to the induction of apoptosis. MTT reduction was stimulated by tambjamine D, which may indicate the generation of reactive oxygen species. Accordingly, treatment of cells with tambjamine D increased nitrite/nitrate at all concentrations and TBARS production starting at the concentration corresponding to the IC50. Tambjamine D, also, induced DNA strand breaks and increased the micronucleus cell frequency as evaluated by comet and micronucleus tests, respectively, at all concentrations evaluated. showing a genotoxic risk induced by tambjamine D. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Push or pull? : The relationship between quality of care and use of complementary and alternative medicine among patients with lung cancer in Sweden

Little is known about relationships between quality of care (QoC) and use of complementary and alternative medicine (CAM) among patients with lung cancer (LC). Purpose: This study examines CAM-use among patients with LC in Sweden, associations between QoC and CAM-use among these patients, and reported aspects of LC-care perceived as particularly positive and negative by patients, as well as suggestions for improving QoC. Methods: Survey data from 94 patient members of the Swedish LC patient organization about CAM-use and QoC as measured by the instrument “Quality from the patient’s perspective” were analyzed. Results: Fifty (53%) LC-patients used CAM, with 40 of the CAM-users reporting that CAM helped them. The most common CAMs used were dietary supplements and natural remedies, followed by prayer. Significantly more patients reported using prayer and meditation for cure than was the case for other types of CAM used. Less than half the CAM-users reported having spoken with staff from the biomedical health care system about their CAM-use. Patients provided numerous suggestions for improving LC-care in a variety of areas, aiming at a more effective and cohesive care trajectory. No differences in QoC were found between CAM-users and non-CAM-users, but differences in CAM-use i.e. type of CAM, reasons for using CAM, and CAM-provider consulted could be associated with different experiences of care. Conclusions: It is important to recognize that CAM-users are not a homogeneous group but might seek different types of CAMs and CAM-providers in different situations depending on experiences of care.