Pleural Washings Research Articles

Microbiological confirmation of tuberculous pleurisy with medical thoracoscopy: targeted pleural washing and pleural biopsy.

Due to the pauci-bacillary nature of tuberculous (TB) pleurisy, clinical diagnosis is common, but microbiological confirmation is necessary to determine drug resistance. This study aimed to investigate the diagnostic yield of medical thoracoscopy (MT) for microbiological confirmation of TB pleurisy. Medical records of patients diagnosed as TB pleurisy with microbiological or histologic evidence who underwent MT between May 2015 and July 2023 at Incheon St. Mary's Hospital were retrospectively reviewed. Sensitivities of microbiological results [acid-fast bacilli (AFB) culture or TB-polymerase chain reaction (PCR)] of pre-MT pleural fluid and those of targeted pleural washing fluid and pleural tissues obtained during MT were compared. Difference in sensitivity was verified with McNemar's test. A total of 72 patients were enrolled. With pre-MT pleural fluid, sensitivities of AFB culture and TB PCR were 5.6% (4/72) and 1.4% (1/72), respectively. With targeted pleural washing fluid, sensitivities of AFB culture and TB-PCR were 23.6% (17/72) and 12.5% (9/72), respectively. With pleural tissues, sensitivities of AFB culture and TB-PCR were 18.1% (13/72) and 40.3% (29/72), respectively. MT showed an additional 27.8% [95% confidence interval (95% CI): 14.2-40.1%, P<0.001] of sensitivity gain in AFB culture and 40.3% (95% CI: 25.7-52.5%, P<0.001) of sensitivity gain in TB-PCR. With pleural washing, additional 19.4% (95% CI: 6.8-31.6%, P=0.001) of sensitivity gain in microbiological confirmation was identified, whereas additional 37.5% (95% CI: 22.6-50.2%, P<0.001) of sensitivity gain was identified with pleural biopsy. With MT, 44.4% of additional sensitivity gain in microbiological confirmation of TB pleurisy was identified. This underscores the role of MT in the diagnosis of TB pleurisy.

New Trends in Precision Medicine: A Pilot Study of Pure Light Scattering Analysis as a Useful Tool for Non-Small Cell Lung Cancer (NSCLC) Diagnosis.

Simple SummaryA reliable method for a fast diagnosis of non-small cell lung cancer (NSCLC) would greatly help in improving therapeutic success in personalized medicine approaches. Thus, in the present study, a new idea was proposed: a morphological single-cell analysis approach combined with a microfluidic device for liquid biopsy. The investigation of the NSCLC sample at different culturing times created the possibility of understanding the evolution of different cell types and their morphological changes, making the Circulating Tumour Cells (CTC) predominance against all other cell classes visible.Background: To date, in personalized medicine approaches, single-cell analyses such as circulating tumour cells (CTC) are able to reveal small structural cell modifications, and therefore can retrieve several biophysical cell properties, such as the cell dimension, the dimensional relationship between the nucleus and the cytoplasm and the optical density of cellular sub-compartments. On this basis, we present in this study a new morphological measurement approach for the detection of vital CTC from pleural washing in individual non-small cell lung cancer (NSCLC) patients. Materials and methods: After a diagnosis of pulmonary malignancy, pleural washing was collected from nine NSCLC patients. The collected samples were processed with a density gradient separation process. Light scattering analysis was performed on a single cell. The results of this analysis were used to obtain the cell’s biophysical pattern and, later on, as basis for Machine Learning (ML) on unknown samples. Results: Morphological single-cell analysis followed by ML show a predictive picture for an NSCLC patient, screening that it is possible to distinguish CTC from other cells. Moreover, we find that the proposed measurement approach was fast, reliable, label-free, identifying and count CTC in a biological fluid. Conclusions: Our findings demonstrate that CTC Biophysical Profile by Pure Light Scattering in NSCLC could be used as a promising diagnostic candidate in NSCLC patients.

Clinical Evaluation of Endoscopic Bronchial Occlusion with an Endobronchial Watanabe Spigot for the Management of Intractable Pneumothorax, Pyothorax with Bronchial Fistula, and Postoperative Air Leakage

Objective The present study aimed to evaluate the clinical effectiveness of endoscopic bronchial occlusion (EBO) with endobronchial Watanabe spigots (EWSs) for the management of prolonged pulmonary air leaks, such as intractable pneumothorax, pyothorax with bronchial fistula, and postoperative air leakage. Methods This was a retrospective study. Between April 2005 and March 2018, we recruited 21 patients with intractable pneumothorax (10 cases), pyothorax with bronchial fistula (7 cases), and postsurgical pulmonary fistula (4 cases) in whom appropriate drainage for 2 weeks had been unsuccessful and who were unsuitable for surgery. An EWS was inserted using a flexible bronchoscope via an endotracheal or a tracheostomy tube. Results The mean number of sessions with EWS procedures was 1.94, and the mean number of inserted EWS per patient was 6.5. In addition to EWS procedures, pleural washing and pleural adhesion therapy were performed in all cases with pyothorax, whereas pleural adhesion therapy was performed in three patients with pneumothorax. The successful treatment rate was 85.7%. Reduction of air leakage was observed in 19/21 patients. The mean duration of reduction of air leaks was 4.1 days (median, 1; range, 0-24 days) following EWS procedures. The mean duration from tube insertion to chest tube removal was 43.4 days (median, 29; range, 16-105 days). Complications included spigot migration and infection (aspergillosis); no complications caused significant mortality. Conclusion Performing EBO using an EWS appears to be a reasonable option for the management of intractable pneumothorax, pyothorax with pulmonary fistula, and postoperative air leakage.

Protective effect of Myrsine parvifolia plant extract against the inflammatory process induced by Bothrops jararaca snake venom

Accidents involving snakes from the genus Bothrops sp. constitute the most important cause of snake envenomation in Brazil. The Myrsine genus has been reported to be used in folk medicine against snakebites. In this work, the phytochemical profiles and ability of extracts from Myrsine parvifolia leaves to reduce the inflammatory process (edema, vascular permeability increase and leukocyte migration) induced by Bothrops jararaca venom were investigated in vivo. Chemical compounds were identified by chromatographic and spectroscopy techniques. Total polyphenol, tannin, and flavonoid contents were determined by spectrophotometric methods. Swiss male mice received an oral administration of extracts (100 mg/kg) in different protocols. Paw edema, intraperitoneal vascular permeability and pleurisy models in mice were used to evaluate the antiophidic potential of the extracts. Paw edema was induced by subplantar injection of B. jararaca venom and quantified as the increase in paw volume. Changes in vascular permeability were assessed by measuring the amount of Evans blue dye extravasation. Leukocyte migration was assessed by total and differential counts in the pleural cavity washes. Myricetin, myricetin-3-O-β-arabinopyranoside, quercetin and kaempferol were isolated from the ethyl acetate extract and identified as the primary compounds of the dichloromethane extract. Terpenes and fatty acids were identified in the hexane and dichloromethane extracts. The pretreated group with hydroethanolic and dichloromethane extract reduced total edema (40 and 52%, respectively), vascular permeability increase (32.4 and 32.2%, respectively) and leukocyte influx into the pleural cavity (42 and 39%, respectively), while the group treated with hexane extract showed only reduced edema (37%) induced by B. jararaca venom. The hydroethanolic extract showed better results in all of the tests performed and was also administered by the protocol of post-poisoning, showing maintenance of paw edema reduction and cell migration. These data indicate a potential anti-inflammatory activity of M. parvifolia in poisoning by B. jararaca, especially to reduce local poison effects.

Clinicopathologic analysis of malignant effusions in pediatric patients

The utility of cytologic evaluation of effusion fluids in adults is well established but has been less well documented in the pediatric population. We examined the clinicopathologic characteristics of children with malignant effusions to establish the value of cytologic examination of these specimens. Pleural, pericardial, peritoneal, and intraoperative washing specimens obtained between January 2000 and October 2015 were identified via surgical pathology database search. Relevant clinical and pathologic data was recorded. Statistical analysis of patient groups was performed via two-sided heteroscedastic Student t tests, and Kaplan-Meier analysis was used to compare overall survival between patient groups. 474 effusion/washing specimens obtained from 342 patients were identified: 179 pleural effusions, 220 peritoneal fluids, 28 pericardial effusions, and 47 intraoperative washing specimens. Of these, 13.7% (56 of 474) effusion specimens were positive for malignancy. Among cancer patients with effusions, malignant effusions were seen in 54.5% of patients with rhabdomyosarcoma and 28.8% of patients with hematolymphoid malignancies. Regarding patient outcomes, malignant effusions were associated with statistically significantly shorter overall survival (P = 0.019). When compared with stage IV patients with benign effusions, those with malignant effusions had a shorter 5-year overall survival (P = 0.042); multiple malignant effusions were associated with a dramatically shorter survival than a single malignant effusion. The overall rate of malignant effusions in our patient population was 13.7%. Malignant effusions in children, particularly multiple, portend a poor prognosis. Our observations emphasize the importance of accurate cytologic diagnosis of effusion fluids, and reinforce the value of their cytologic evaluation.

Malignant pleural effusions because of lung cancer.

The discovery of a pleural effusion in the setting of lung cancer has diagnostic, prognostic, and therapeutic challenges, some of which are addressed in this review. Around 20% of patients with lung cancer have minimal pleural effusions, which are not amenable to a diagnostic thoracentesis. These patients have a poorer overall survival (∼7.5 months) than those without effusions (∼12-18 months), although slightly better than those with proven malignant fluids (∼5.5 months). Tumor genotype techniques are feasible on pleural fluid specimens and clinically helpful in identifying patients who may benefit from targeted therapies. If limited pleural involvement is detected during lung cancer surgery despite the presurgical imaging studies, macroscopic complete resection of the lung tumor is still a treatment option. Cytological examination for cancer cells in pleural cavity washings at the time of thoracotomy (pleural lavage cytology) is recommended to uncover pleural dissemination. Patients with non-small cell lung cancer with visceral pleural invasion might be considered candidates for postsurgical adjuvant therapy. Some predictors of adverse survival in patients with lung cancer include the presence of a minimal pleural effusion, positive pleural lavage cytology, visceral pleural invasion on pathologic examination, and unexpected pleural involvement during surgery.

Gedunin, a natural tetranortriterpenoid, modulates T lymphocyte responses and ameliorates allergic inflammation

T lymphocytes are critical cells involved in allergy. Here, we report that the natural tetranortriterpenoid gedunin impaired allergic responses primarily by modulating T lymphocyte functions. The intraperitoneal (i.p.) administration of gedunin inhibited pleural leukocyte accumulation triggered by intra-pleural (i.pl.) challenge with ovalbumin (OVA) in previously sensitized C57BL/6 mice; this inhibition was primarily due to the impairment of eosinophil and T lymphocyte influx. Likewise, i.pl. pre-treatment with gedunin inhibited eosinophil and T lymphocyte migration into mouse lungs 24h after OVA intra-nasal (i.n.) instillation. Pre-treatment with gedunin diminished the levels of CCL2, CCL3, CCL5, CCL11, Interleukin-5 and leukotriene B4 at the allergic site. In vitro pre-treatment with gedunin failed to inhibit T lymphocyte adhesion and chemotaxis towards pleural washes recovered from OVA-challenged mice, suggesting that gedunin inhibits T lymphocyte migration in vivo via the inhibition of chemotactic mediators in situ. In vivo pre-treatment with gedunin reduced the numbers of CD69+ and CD25+ T lymphocytes in the pleura and CD25+ cells in the thoracic lymph nodes 24h after OVA i.pl. challenge. In accordance, in vitro treatment of T lymphocytes with gedunin inhibited α-CD3 mAb-induced expression of CD69 and CD25, proliferation, Interleukin-2 production and nuclear translocation of NFκB and NFAT. Notably, post-treatment of mice with gedunin reverted OVA-induced lung allergic inflammation by decreasing the T lymphocyte and eosinophil counts and the levels of eosinophilotactic mediators in bronchoalveolar lavage fluid. Our results demonstrate a remarkable anti-allergic effect of gedunin due to its capability to modulate T cell activation and trafficking into the airways.

CCL25 induces α4β7integrin-dependent migration of IL-17+γδ T lymphocytes during an allergic reaction

Herein, we provide evidence that during allergic inflammation, CCL25 induces the selective migration of IL-17(+) γδ T cells mediated by α(4) β(7) integrin. Intrapleural injection of CCL25 into ovalbumin (OVA)-immunized C57BL/6 mice triggered the accumulation of γδ T lymphocytes expressing CCR9 (CCL25 receptor) and α(4) β(7) integrin in the pleura, but failed to attract αβ T lymphocytes. CCL25 attracted CCR6(+) γδ T cells producing IL-17 (but not IFN-γ or IL-4). OVA challenge triggered increased production of CCL25 followed by the accumulation of CCR9(+) , α(4) β(7) (+) , and CCR6(+) /IL-17(+) γδ T cells into the pleural cavities of OVA-immunized mice, which was inhibited by the in vivo neutralization of CCL25. The in vivo blockade of α(4) β(7) integrin also inhibited the migration of IL-17(+) γδ T lymphocytes (but not of αβ T lymphocytes) into mouse pleura after OVA challenge, suggesting that the CCL25/α(4) β(7) integrin pathway is selective for γδ T cells. In addition, α(4) β(7) integrin blockade impaired the in vitro transmigration of γδ T cells across endothelium (which expresses α(4) β(7) ligands VCAM-1 and MadCAM-1), which was induced by CCL25 and by cell-free pleural washes recovered from OVA-challenged mice. Our results reveal that during an allergic reaction, CCL25 drives IL-17(+) γδ T-cell mobilization to inflamed tissue via α(4) β(7) integrin and modulates IL-17 levels.

Cytological examination of pleural cavity lavage accompanied by the study of gene promoter hypermethylation of p16 and O6-methylguanine-DNA-methyltransferase genes in diagnostics of non-small cell lung cancer metastatic changes into pleura.

Aim of the studyMetastases of non-small cell lung cancer (NSCLC) into pleura disqualify a patient from surgery and present a bad prognostic index. The aim of the study was to find out whether washing out the pleural cavity in such cases and examining obtained washings for presence of cancer cells will help to detect early NSCLC metastases into pleura, and also whether negative results of the cytology determine whether hypermethylation of these genes will increase the sensitivity of this examination.Material and methodsThe study consisted of the examination of 76 patients, including 59 operated on for NSCLC and 17 operated on for other reasons. Pleural washing fluid collected during the surgery was subjected to cytological examination as well as examined to determine the presence of promoter region hypermethylation of p16 and MGMT genes.ResultsPositive cytological results of pleural lavage were confirmed in 4 persons (7%) with NSCLC. The presence of promoter region hypermethylation of one or both examined genes was found in 3 patients (18%) in the control group and in 47 (80%) in the study group. Sex, occupational exposure, smoking cigarettes, and NSCLC histological type did not have an influence on the presence of cancer cells or hypermethylation in the pleural lavage fluid. Positive cytology results were more frequent at the T4 stage of NSCLC. Hypermethylation was more frequent in the research group (p < 0.01). Cancer cells and hypermethylation did not occur more frequently in pleural lavage fluid of patients with metastases into pleura.ConclusionsThe cytological examination and promoter region hypermethylation assessment of the p16 gene and MGMT gene in pleural lavage cells do not allow one to detect early metastasis of NSCLC into pleura.

Leukotriene B4 mediates γδ T lymphocyte migration in response to diverse stimuli

Herein, we investigated the involvement of the 5-LO-derived lipid mediator LTB(4) in gammadelta T cell migration. When injected into the i.pl. space of C57BL/6 mice, LTB(4) triggered gammadelta T lymphocyte mobilization in vivo, a phenomenon also observed in in vitro chemotaxis assays. The i.pl. injection of Escherichia coli endotoxin (LPS) triggered increased levels of LTB(4) in pleural cavities. The in vivo inhibition of LTB(4) biosynthesis by the 5-LO inhibitor zileuton or the FLAP inhibitor MK886 attenuated LPS-induced gammadelta T cell accumulation into pleural cavities. Accordingly, 5-LO KO mice failed to recruit gammadelta T cells into the inflammatory site after i.pl. LPS. Antagonists of the high-affinity LTB(4) receptor BLT1, CP105,696, and LY292476 also attenuated LPS-induced gammadelta T cell accumulation in pleural cavities as well as in vitro chemotaxis toward pleural washes obtained from LPS-simulated mice. LTB(4)/BLT1 also accounted for gammadelta T cell migration induced by i.pl. administration of Mycobacterium bovis BCG or antigen in sensitized mice. BLT1 was expressed on naïve, resident as well as LPS-recruited gammadelta T cells. Isolated gammadelta T cells were found to undergo F-actin cytoskeleton reorganization when incubated with LTB(4) in vitro, confirming that gammadelta T lymphocytes can respond directly to LTB(4). In addition to its direct effect on gammadelta T cells, LTB(4) triggered their accumulation indirectly, via modulation of CCL2 production in mouse pleural cavities. These data show that gammadelta T cell migration into the pleural cavity of mice during diverse inflammatory responses is dependent on LTB(4)/BLT1.

A case of advanced ovarian cancer upstaged on the bases of pleural washing cytology

We encountered a case of ovarian cancer that had positive pleural washing cytology diagnosed by thoracoscopy. A 65-year-old woman was referred to our hospital with a sensation of abdominal fullness. A whole-body examination revealed a solid tumor of the ovary and accumulation of ascites. Neither pleural effusion nor lung metastasis was found at the time. Laparotomy showed extensive dissemination and we classified the stage as IIIc. As the diaphragm was thickened with dissemination, we tried to strip the diaphragm and used thoracoscopy to identify the thoracic area. No tumor or pleural effusion was detected. We performed pleural biopsy and pleural washing cytology. Finally, malignant cells were found in the pleural washing cytology. Pathological findings showed serous papillary adenocarcinoma of the ovary in this stage IV case. We suggest in this report that thoracoscopy is a useful method for making the correct surgical staging.

Videothoracoscopy in Pleural Empyema Following Methicillin-ResistantStaphylococcus aureus(MRSA) Lung Infection

Our study shows the different therapeutic procedures in 64 patients with pleural effusion due to MRSA pneumonia. The thoracostomy tube associated with pleural washing was decisive in 10 simple effusion patients. Video-assisted thoracic surgery allowed a complete resolution of the disease in 22 complex parapneumonic effusion patients. In 20 of 32 patients with frank pus in the pleural cavity, the videothoracoscopic insufflation of carbon dioxide (CO2) before thoracotomy facilitated the dissection of the lung tissue. In 12 patients, this approach was not applied because of cardiac insufficiency. Videothoracoscopy and decortication after thoracotomy ensured the recovery of functions.

Pilot Study on Transdiaphragmatic Thoracoscopic-Assisted Pleural Biopsy and Intrathoracic Washing Cytology for Stage IIIc Ovarian Cancer With Diaphragmatic Metastases

The significance of investigations of thoracic cavities as well as pleural biopsy and intrathoracic washing cytology through transdiaphragmatic thoracoscopy for stage IIIc ovarian cancer with diaphragmatic metastases was assessed as a prospective pilot study. Eligibility criteria were established to include patients with stage IIIc ovarian cancer in whom pleural effusions were not detected preoperationally, but prominent diaphragmatic metastases were observed when the abdomen was opened and those who submitted consent. Transdiaphragmatic thoracoscopy was performed after diaphragm stripping. Then, biopsy of the lesion suspected to be disseminated or the plural membrane of the thoracic opening was performed followed by washing cytology using physiological saline. Ten subjects were enrolled. Disseminated lesions were observed on plural membranes, and positive results were obtained in biopsy and washing cytology in 3 subjects. In addition, positive results were seen with biopsy alone in 2 subjects and with washing cytology alone in 2 subjects. Hence, a total of 7 subjects (70.0%) were up-staged to the level of stage IV. Postoperational complications were not observed in any of these cases. It was suggested that stage IIIc ovarian cancer with prominent diaphragmatic metastasis may advance to the level of stage IV from a clinical point of view even if carcinomatous pleural effusions are not detected pre-operationally. Therefore, it is thought that this operational method is useful in the management of progressive ovarian cancer.

Mechanisms of T-Lymphocyte Accumulation during Experimental Pleural Infection Induced byMycobacterium bovisBCG

Tuberculous pleurisy is a frequent extrapulmonary manifestation characterized by accumulation of fluid and inflammatory cells in the pleural space. Here, we investigated the mechanisms of T-lymphocyte accumulation in the pleural space by using a murine model of pleurisy induced by Mycobacterium bovis BCG. Intrathoracic (i.t.) injection of BCG (4.5 x 10(5) bacteria/cavity) induced accumulation of T lymphocytes in the pleural cavities of C57BL/6 mice. We observed the presence of CFU in pleural washes conducted 1, 2, 3, 7, and 15 days after pleurisy induction. Pretreatment with fucoidan inhibited T-lymphocyte accumulation at 1 day, but not at 15 days, after BCG-induced pleurisy. Accordingly, adoptive transfer of fluorescein isothiocyanate-labeled blood mononuclear cells to infected mice showed that T lymphocytes migrated into the pleural cavity 1 day (but not 15 days) after BCG injection. Cell-free pleural wash fluids recovered from mice 1 day after BCG i.t. stimulation (day 1 BCG-PW), but not day 7 or day 15 BCG-PW, induced in vitro T-cell transmigration, which was dependent on L-, P-, and E-selectins. In contrast, day 7 BCG-PW (but not day 1 BCG-PW) induced in vitro T-lymphocyte proliferation via interleukin-2 (IL-2) and gamma interferon (IFN-gamma). Accordingly, in vivo IL-2 or IFN-gamma neutralization abolished T-lymphocyte accumulation 7 days after pleurisy induction. Our results demonstrate that pleural infection induced by BCG leads to T-lymphocyte accumulation in two waves. The acute phase depends on selectin-mediated migration, while the second wave of T-lymphocyte accumulation seems to depend on a local proliferation induced by cytokines produced in situ.

Involvement of CC chemokines in T lymphocyte trafficking during allergic inflammation: the role of CCL2/CCR2 pathway

In the present study, we show that the intra-thoracic injection of ovalbumin (OVA, 12.5 microg per cavity) into C57BL/10 mice induced a significant increase in gammadelta T lymphocyte numbers in the pleural cavity, blood and thoracic lymph node of challenged mice. Such increase was significant within 12 h, peaked within 48 h and returned to basal counts within 120 h. Levels of CC chemokine ligand (CCL)-2/monocyte chemotactic protein-1, CCL5/regulated upon activation, normal T cell expressed and secreted, CCL3/macrophage inflammatory protein-1 alpha and CCL25/thymus-expressed chemokine were above control values in pleural washes recovered 24 h after OVA challenge (OPW) and were likely produced by pleural macrophages and mesothelial cells. Antigenic challenge also induced an up-regulation in CC chemokine receptor (CCR)-2, CCR5 and CCR9 on gammadelta T cells from pleural cavities, blood and lymph nodes, suggesting that cells found in mice pleural cavity migrate from secondary lymphoid organs into the inflammatory site via blood stream. The in vitro neutralization of CCL2 (but not of CCL3, CCL5 or CCL25) abrogated OPW-induced gammadelta T lymphocyte transmigration. Confirming such results, the in vivo administration of alpha-CCL2 mAb inhibited gammadelta T lymphocyte accumulation in the pleural cavity of challenged mice, whereas the blockade of CCL3, CCL5 or CCL25 showed no effect on gammadelta T cell mobilization. In addition, OVA challenge failed to induce gammadelta T lymphocyte accumulation in the pleural cavity of C57BL/6 CCR2 knockout mice, which also showed decreased numbers of these cells in blood and lymph nodes when compared with wild-type mice. Overall, such results demonstrate that CCR2/CCL2 pathway is crucial for gammadelta T lymphocyte mobilization during the allergic response.

P1-004: Role of cytology accompanied by the study of the occurrence of p16INK4a and O6-methylguanine-DNA transferase (MGMT) promoter methylation in the cells of the pleural cavity washings fluid in early diagnostics of non-microcellular lung caner (NSCLC) metastatic changes into pleura

P1-004: Role of cytology accompanied by the study of the occurrence of p16INK4a and O6-methylguanine-DNA transferase (MGMT) promoter methylation in the cells of the pleural cavity washings fluid in early diagnostics of non-microcellular lung caner (NSCLC) metastatic changes into pleura

Inhibition of allergen-induced eosinophil recruitment by natural tetranortriterpenoids is mediated by the suppression of IL-5, CCL11/eotaxin and NFκB activation

The present study reports the anti-allergic activity of a group of six different tetranortriterpenoids (TNTP) isolated from the seeds of Carapa guianensis Aublet: 6a-acetoxygedunin, 7-deacetoxy-7-oxogedunin, andirobin, methyl angolensate, 6a-acetoxyepoxyazadiradione and gedunin. Oral pretreatment with TNTP significantly inhibited total leukocyte and eosinophil accumulation in C57BL/10 mice pleural cavities 24 h after the intrathoracic (i.t.) injection of ovalbumin (OVA), but had no effect on CD4, CD8 or γδ T lymphocyte accumulation. Pleural washes recovered from 6 h OVA-stimulated mice (OPW) pretreated with TNTP failed to induce shape change in eosinophil in vitro, indicating the inhibition of eosinophilotactic chemokines by TNTP. In accordance with such results, ELISA assays showed decreased levels of CCL11/eotaxin and IL-5 in OPW recovered from TNTP pretreated mice within 6 h. TNTP oral pretreatment inhibited nuclear factor-κB (NFκB) translocation into the nucleus in pleural leukocytes recovered from previously sensitized mice after antigenic challenge. In addition, the incubation of splenocytes recovered from previously sensitized mice with TNTP also inhibited NFκB activation after OVA stimulation. Taken together, these results indicate that the inhibition of allergic eosinophilia by TNTP is correlated with the inhibition of CCL11/eotaxin and IL-5 generation through NFκB signaling pathway impairment in mice.

In vitro and in vivo responses of murine granulocytes to human complement-derived, haemolytically inactive C5b67 (iC5b67).

Haemolytically inactive C5b67 (iC5b67), which was made from purified human components and decayed to a haemolytically inactive form, was evaluated as an agonist for murine leucocytes both in vitro and in vivo. In an in vitro assay, iC5b67 stimulated chemotaxis for both neutrophils purified from mouse bone marrow and splenic eosinophils of IL-5 transgenic mice. The stimulation was dose-dependent, with high dose inhibition. As with human neutrophils, iC5b67 also failed to up-regulate CR3 (CD11b/CD18) expression and to stimulate superoxide generation in murine bone marrow neutrophils, in vitro. In vivo, iC5b67 elicited an inflammatory response in a mouse model of pleuritis. A marked infiltration of neutrophils, which peaked at 4 h, was followed by an infiltration of eosinophils and mononuclear leucocytes. This inflammatory response was dose- and time-dependent. However, the protein concentration in the pleural wash fluid did not increase, indicating that iC5b67 did not induce a capillary leak. Although the infiltration of neutrophils could not be reproduced by pure C7 or human serum albumin (HSA), C5b6 did induce an influx of neutrophils. We were able to document the existence of C7, both antigenically and functionally, in pleural washes of normal mice, making it likely that the activity of C5b6 resulted from the in situ formation of C5b67 and iC5b67. The mouse model of pleuritis promises to be a useful in vivo system in which to evaluate the pro- and anti-inflammatory effects of iC5b67 that have been noted in vitro.

LPS induces eosinophil migration via CCR3 signaling through a mechanism independent of RANTES and Eotaxin.

Mounting evidence suggests that lipopolysaccharide (LPS) modulates bronchoconstriction and eosinophil function in asthma. We have investigated the role of different chemokines in the eosinophil influx to the pleural cavity after LPS stimulation. Expression of mRNA for eotaxin, regulated on activation, normal T cells expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, MIP-2, and monocyte chemotactic protein (MCP)-1 was increased in cells recovered from the mouse pleural cavity 6 h after LPS administration. Eotaxin and RANTES, but not MIP-1alpha, protein levels were also increased in cell-free pleural washes recovered 6 h after LPS stimulation (LPW). Antimurine eotaxin and antimurine RANTES antibodies (Abs) failed to inhibit LPS-induced eosinophil influx into mouse pleural cavity in vivo. Pertussis toxin inhibited LPW-induced eosinophil shape change in vitro, suggesting the involvement of G protein-coupled receptors in LPW signaling. Blockade of CCR3 receptors diminished eosinophil shape change induced by LPW fractions in vitro and LPS-induced eosinophil accumulation in vivo. To investigate further contribution of CC chemokines, we administered a 35-kD CC chemokine neutralizing protein (vCKBP) in vivo. vCKBP inhibited the eosinophil accumulation induced by eotaxin and ovalbumin, but did not block that induced by LPS or LPW. Our data suggest that LPS-induced eosinophil accumulation depends on G protein-coupled CCR3 receptor activation, through a mechanism independent of eotaxin, RANTES, or other vCKBP-inhibitable CC chemokines.

Usefullness of carcinoembryonic antigen levels in the pleural washings as a predictor of survival after surgical resection for stage I non-small cell lung cancer

Usefullness of carcinoembryonic antigen levels in the pleural washings as a predictor of survival after surgical resection for stage I non-small cell lung cancer