OBJECTIVETo evaluate immunologic mechanisms underlying Aspergillus fumigatus pulmonary infections in immunocompetent Dark Agouti (DA) and Albino Oxford (AO) rats recognized as being susceptible to some inflammatory diseases in different manners.

METHODSLung fungal burden (quantitative colony forming units, CFU, assay), leukocyte infiltration (histology, cell composition) and their function (phagocytosis, oxidative activity, CD11b adhesion molecule expression) and cytokine interferon-γ (IFN-γ) and interleukin-17 and -4 (IL-17 and IL-4) lung content were evaluated following infection (intratracheally, 1x10(7) conidia).

RESULTSSlower reduction of fungal burden was observed in AO rats in comparison with that in DA rats, which was coincided with less intense histologically evident lung cell infiltration and leukocyte recovery as well as lower level of most of the their activities including intracellular myeloperoxidase activity, the capacity of nitroblue tetrazolium salt reduction and CD11b adhesion molecule expression (except for phagocytosis of conidia) in these rats. Differential patterns of changes in proinflammatory cytokine levels (unchanged levels of IFN-γ and transient increase of IL-17 in AO rats vs continuous increase of both cytokines in DA rats) and unchanged levels of IL-4 were observed.

CONCLUSIONGenetically-based differences in the pattern of antifungal lung leukocyte activities and cytokine milieu, associated with differential efficiency of fungal elimination might be useful in the future use of rat models in studies of pulmonary aspergillosis.

Animals ; Aspergillus fumigatus ; immunology ; Cytokines ; metabolism ; Lung ; immunology ; metabolism ; microbiology ; pathology ; Male ; Pulmonary Aspergillosis ; immunology ; Rats

BackgroundVentilator-induced lung injury (VILI) is commonly associated with barrier dysfunction and inflammation reaction. Glutamine could ameliorate VILI, but its role has not been fully elucidated. This study examined the relationship between inflammatory cytokines (interleukin [IL]-6, tumor necrosis factor [TNF]-α, and IL-10) and adherens junctions (E-cadherin, p120-catenin), which were ameliorated by glutamine in VILI, both in vitro and in vivo.

MethodsFor the in vivo study, 30 healthy C57BL/6 mice weighing 25-30 g were randomly divided into five groups with random number table (n = 6 in each group): control (Group C); low tidal volume (Group L); low tidal volume + glutamine (Group L + G); high tidal volume (Group H); and high tidal volume + glutamine (Group H + G). Mice in all groups, except Group C, underwent mechanical ventilation for 4 h. For the in vitro study, mouse lung epithelial 12 (MLE-12) cells pretreated with glutamine underwent cyclic stretching at 20% for 4 h. Cell lysate and lung tissue were obtained to detect the junction proteins, inflammatory cytokines, and lung pathological changes by the Western blotting, cytokine assay, hematoxylin and eosin staining, and immunofluorescence.

ResultsIn vivo, compared with Group C, total cell counts (t = -28.182, P < 0.01), the percentage of neutrophils (t = -28.095, P < 0.01), IL-6 (t = -28.296, P < 0.01), and TNF-α (t = -19.812, P < 0.01) in bronchoalveolar lavage (BAL) fluid, lung injury scores (t = -6.708, P < 0.01), and the wet-to-dry ratio (t = -15.595, P < 0.01) were increased in Group H; IL-10 in BAL fluid (t = 9.093, P < 0.01) and the expression of E-cadherin (t = 10.044, P < 0.01) and p120-catenin (t = 13.218, P < 0.01) were decreased in Group H. Compared with Group H, total cell counts (t = 14.844, P < 0.01), the percentage of neutrophils (t = 18.077, P < 0.01), IL-6 (t = 18.007, P < 0.01), and TNF-α (t = 10.171, P < 0.01) in BAL fluid were decreased in Group H + G; IL-10 in BAL fluid (t = -7.531, P < 0.01) and the expression of E-cadherin (t = -14.814, P < 0.01) and p120-catenin (t = -9.114, P < 0.01) were increased in Group H + G. In vitro, compared with the nonstretching group, the levels of IL-6 (t = -21.111, P < 0.01) and TNF-α (t = -15.270, P < 0.01) were increased in the 20% cyclic stretching group; the levels of IL-10 (t = 5.450, P < 0.01) and the expression of E-cadherin (t = 17.736, P < 0.01) and p120-catenin (t = 16.136, P < 0.01) were decreased in the 20% cyclic stretching group. Compared with the stretching group, the levels of IL-6 (t = 11.818, P < 0.01) and TNF-α (t = 8.631, P < 0.01) decreased in the glutamine group; the levels of IL-10 (t = -3.203, P < 0.05) and the expression of E-cadherin (t = -13.567, P < 0.01) and p120-catenin (t = -10.013, P < 0.01) were increased in the glutamine group.

ConclusionsHigh tidal volume mechanical ventilation and 20% cyclic stretching could cause VILI. Glutamine regulates VILI by improving cytokines and increasing the adherens junctions, protein E-cadherin and p120-catenin, to enhance the epithelial barrier function.

Animals ; Cadherins ; metabolism ; Catenins ; metabolism ; Glutamine ; metabolism ; Inflammation ; metabolism ; Interleukin-6 ; metabolism ; Lung ; metabolism ; pathology ; Mice ; Mice, Inbred C57BL ; Ventilator-Induced Lung Injury ; immunology ; metabolism

Adult ; Diagnosis, Differential ; Female ; Histiocytoma, Malignant Fibrous ; immunology ; pathology ; Histiocytosis, Langerhans-Cell ; immunology ; pathology ; Histiocytosis, Sinus ; immunology ; pathology ; Hodgkin Disease ; immunology ; pathology ; Humans ; Ki-1 Antigen ; metabolism ; Lewis X Antigen ; metabolism ; Lung ; immunology ; pathology ; Lung Neoplasms ; immunology ; pathology ; Lymph Nodes ; immunology ; pathology

OBJECTIVETo explore the inhibitory effects of endoplasmic reticulum-retained intrabody on the secretion of type IV collagenase and the invasion of human pulmonary giant cell carcinoma PG cells in vitro.

METHODSTwo expression plasmids were constructed, pcDNA3.1-CP.scFv and pcDNA3.1-ER.scFv encoding cytoplasm-retained and endoplasmic reticulum-retained single chain antibodies against the type IV collagenase, respectively. The intracellular antibody genes were transfected into the human pulmonary giant cell carcinoma PG cells. Western blot was performed to detect the expression of pcDNA3.1-CP.scFv and pcDNA3.1-ER.scFv. Gelatin zymography was performed to detect seretion of type IV collagenase in PG cells and Matrigel assay was employed for determination of the cell invasiveness.

RESULTSBoth of cytoplasm-retained and endoplasmic reticulum-retained introbodies, CP.scFv and ER.scFv, were expressed in PG cells. ER.scFv, significantly inhibited the secretion of type IV collegenase. As shown, matrix metalloproteinase 9 and matrix metalloproteinase 2 were inhibited by 85.7% and by 51.2%, respectively. However, CP.scFv did not show such inhibitory effect. The ER.scFv encoding gene-transfected PG cells were much less invasive than parental or vector control cells, the inhibition rate was 76.3% (P < 0.05), whereas CP.scFv encoding gene-transfected PG cells showed no reduction in invasiveness.

CONCLUSIONThose findings demonstrate that endoplasmic reticulum (ER)-retained intracellular antibody technology may selectively abrogate the activity of type IV collagenase in the protein trafficking and secretory pathway and effectively inhibit tumor cell invasion in vitro. Anti-type IV collagenase intrabody may be further used in cancer gene therapy.

Carcinoma, Giant Cell ; metabolism ; pathology ; Cell Line, Tumor ; Cytoplasm ; immunology ; Endoplasmic Reticulum ; immunology ; Genetic Vectors ; Humans ; Immunoglobulin Variable Region ; metabolism ; physiology ; Lung Neoplasms ; metabolism ; pathology ; Matrix Metalloproteinase 2 ; immunology ; metabolism ; Matrix Metalloproteinase 9 ; immunology ; metabolism ; Neoplasm Invasiveness ; Plasmids ; Transfection

BACKGROUNDCorticosteroid treatment of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) has been one of the most controversial clinical issues in critical care. Although the administration of high-dose corticosteroids does not benefit patients with early septic shock and ARDS, recent clinical trials have indicated that treatment with relatively low-dose corticosteroids (2 to 3 mg/kg/day of methylprednisolone or equivalent) may improve outcome when used for late ARDS or persistent septic shock. The underlying mechanism was not fully clarified. Whether the administration of corticosteroids can arrest neutrophil-driven organ injury once started remains to be elucidated.

OBJECTIVETo observe the effects of hydrocortisone (HC, 6 mg/kg) on oxygen free radicals (OFR) released by PMN and pulmonary pathological changes in rat ALI model induced by lipopolysaccharide (LPS), to investigate the possible mechanism through which corticosteroids exert protective effect on ALI.

METHODSA rat model of ALI was induced by peritoneal injection of 2 x 10(12) Escherichia coli/kg. Fifty-six rats were randomly divided into three groups: normal control group, LPS group and HC group (6 mg/kg). Samples were collected 2 h, 4 h and 6 h after giving LPS to LPS and HC group (6 h after giving normal saline in normal control group) to measure the level of OFR released by PMN using chemiluminescence method based on lumino, and to compae of pulmonary pathological changes among the three groups.

RESULTSPathological examination with light microscope in LPS group showed thickened pulmonary interstitia, inflammatory cell infiltration, edema and hemorrhage, which were in accordance with the features of ALI. There were significant differences in the release of OFR by PMN among the three groups (P < 0.01). The level of OFR released by PMN in LPS group was significantly higher than that of the control group, and continued to increase during the observation period (2 - 6 h after LPS). The release of OFR by PMN in HC group was significantly suppressed as compared with LPS group, which was peaked at 4 h after LPS injection (to 98.2%); there were also significant differences in the grades of ALI pathologic changes among the three groups (P < 0.01). The grades of ALI pathologic changes in LPS group were significantly increased when compared with the normal control group (P < 0.05) while significantly decreased in HC group as compared with LPS group (P < 0.05).

CONCLUSIONIt was demonstrated in the LPS induced ALI model that OFR might play an important role in onset of ALI. Intervening with HC (6 mg/kg) treatment could ameliorate the lung injury and exert significant and sustained suppression on the release of OFR by PMN, showing that HC has a protective effect on LPS induced ALI and its theraputic effect occurs possibly through suppression on the release of OFR by PMN.

Acute Lung Injury ; etiology ; immunology ; Animals ; Disease Models, Animal ; Free Radicals ; metabolism ; Glucocorticoids ; pharmacology ; Hydrocortisone ; pharmacology ; Lipopolysaccharides ; adverse effects ; Lung ; immunology ; pathology ; Mice ; Neutrophils ; drug effects ; metabolism

OBJECTIVETo study the clinical and pathological characteristics of pulmonary epithelioid hemangioendothelioma.

METHODSFour cases of pulmonary epithelioid hemangioendothelioma were studied by histopathologic and immunohistochemical examination of lung biopsy specimens.

RESULTSThere were 3 female and 1 male, age 28 to 40 years. Clinically the tumor presented as multiple bilateral small nodules in the lung. Histologically, crown-like clusters of epithelioid tumor cells were obtained which filled in the alveoli locating at the periphery of the tumor nodules, while the central part of the nodules contained myxoid to hyaline matrix. The overall architecture of the lung was still preserved. Additionally, intracytoplasmic vacuoles were seen in tumor cells within which red blood cells were sometimes identified. Tumor cells generally lacked pleomorphism, mitotic activity and necrosis. They were immunohistochemically positive for CD31 and CD34. AE1/AE3 staining was positive in some cases.

CONCLUSIONSPulmonary epithelioid hemangioendothelioma often occurs in a middle-aged woman and represents a distinct clinical pathological entity.

Adult ; Anion Exchange Protein 1, Erythrocyte ; analysis ; Antigens, CD34 ; analysis ; Antiporters ; analysis ; Female ; Hemangioendothelioma, Epithelioid ; immunology ; metabolism ; pathology ; Humans ; Immunohistochemistry ; Lung ; pathology ; Lung Neoplasms ; immunology ; metabolism ; pathology ; Male ; Platelet Endothelial Cell Adhesion Molecule-1 ; analysis

Animals ; Lung ; immunology ; pathology ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; metabolism ; physiopathology ; Spleen ; immunology ; pathology ; fas Receptor ; analysis

OBJECTIVEAllergic asthma is thought to be mediated by CD4+ T lymphocytes producing the Th2-associated cytokines, which play a critical role in the development of the airway hyper-responsiveness and the eosinophilic inflammatory response. The costimulatory pathway CD28/B7 has been shown to play an important role in CD4+ T cell activation in allergic asthma. This study was conducted to evaluate the role of another costimulatory pathway OX40/OX40 ligand (L) in the pathogenesis of allergic asthma in BALB/c mice.

METHODSAn allergic asthma model in BALB/c mice was established. Thirty-six BALB/c mice were randomly divided into three groups with 12 in each. Mice in treatment group (group B) were treated with neutralizing anti-OX40L monoclonal antibody (mAb, 300 microg per mouse) during the sensitization period. Mice in two control groups, asthma model group (group A) and IgG antibody group (group C) were treated with normal saline (NS) and control IgG respectively instead of anti-OX40L mAb. Bronchoalveolar lavage fluid (BALF) was collected from the mice of each group for counting the total number of white blood cells (including neutrophil granulocyte, lymphocyte, monocyte and eosinophil granulocyte) and the proportions of these cells. The levels of IL-4 and INF-gamma in BALF were measured by ELISA. Lungs were removed for morphological examination after HE and PAS staining, and expression of OX40 in lungs was evaluated by immunohistochemical method.

RESULTS(1) The count of total number of white blood cells in BALF (x10(6)/ml) was lower in group B than that of group A and group C (26.6 +/- 4.6 vs. 36.8 +/- 5.2 and 34.3 +/- 6.9, respectively), the difference between the treatment group (group B) and two control groups (groups A and C) was significant; The proportions of eosinophils and lymphocytes in the BALF (%) were lower in group B than those in group A and group C (eosinophils 15.1 +/- 2.6 vs. 20.0 +/- 4.1 and 19.9 +/- 3.9, respectively; lymphocytes 7.0 +/- 0.9 vs. 8.9 +/- 1.6 and 8.6 +/- 1.8, respectively), the difference between the treatment group and two control groups was significant. (2) The IL-4 level in BALF (pg/ml) was lower in group B than that in group A and group C (672 +/- 58 vs. 809.57 +/- 106.00 and 784 +/- 58, respectively), but the INF-gamma levels in BALF (pg/ml) were higher than those in group A and group C (0.86 +/- 0.09 vs. 0.69 +/- 0.15 and 0.67 +/- 0.13 respectively), and all the differences were statistically significant. (3) The expression of OX40 in the lungs of mice in group B were at a lower level than that of group A and group C, and the morphological changes of asthma were ameliorated in the mice of the treatment group. The signs of mice in treatment group were obviously ameliorated as compared to the two control groups.

CONCLUSIONBlocking the costimulatory pathway by administering the neutralizing anti-OX40L mAb during the sensitization period of allergic asthma model could balance the Th1/Th2 responses, inhibit lung inflammation and ameliorate the signs of mice model of asthma.

Animals ; Antibodies, Monoclonal ; administration & dosage ; immunology ; pharmacology ; Antigens, Differentiation ; immunology ; metabolism ; Asthma ; immunology ; metabolism ; pathology ; therapy ; Bronchoalveolar Lavage Fluid ; cytology ; immunology ; Disease Models, Animal ; Enzyme-Linked Immunosorbent Assay ; Eosinophils ; immunology ; Female ; Immunoglobulin G ; administration & dosage ; immunology ; pharmacology ; Immunohistochemistry ; Interferon-gamma ; analysis ; immunology ; Interleukin-4 ; analysis ; immunology ; Leukocyte Count ; Leukocytes ; immunology ; Lung ; drug effects ; immunology ; pathology ; Lymphocytes ; immunology ; Membrane Glycoproteins ; immunology ; Mice ; Mice, Inbred BALB C ; Ovalbumin ; toxicity ; Tumor Necrosis Factors ; immunology

OBJECTIVETo investigate whether dendritic cells pulsed with whole tumor lysates (WTL) could in vitro elicit antitumor T cell responses in patients with non-small-cell lung cancer (NSCLC).

METHODSMonocyte-derived immature DCs (imDCs) generated in the presence of human recombinant granulocyte-macrophage colony stimulating factor and interleukin-4 from peripheral blood mononuclear cell of NSCLC patients, and then were induced to mature by pulsing autologous WTL (DCs/WTL) or by the addition of TNF-alpha(TNF/DCs). FACS and MLR assay were used to monitor their phenotypic changes and capacity to stimulate allogeneic and autologous T cell proliferation. DCs/WTL activated with TNF-alpha (* DCs/WTL) were cocultured in vitro with autologous T cells for eliciting antitumor CTLs. T cell mediated antitumor responses were measured by IFN-gamma enzyme-linked immunospot (ELISPOT) assay for WTL-specific IFN-gamma releasing T cells and by lactate dehydrogenase release (LDH) assay for lysis of autologous tumor cells, respectively.

RESULTSWhen monocytes-derived imDCs from the patients with NSCLC (n = 10) were pulsed with autologous WTL for a day at 30 microg total protein of WTL per 10(6) DCs/ml, this led to up-regulation of CD1a, CD83 and CD86 as well as HLA-DR, and also led to marked stimulation of allogeneic T cell proliferating activity, which was comparable to that of TNF/DCs. However, their capacity of stimulating autologous T cell proliferation in vitro was significantly more potent than those of TNF/DCs (P < 0.05). The numbers of WTL-specific IFN-gamma releasing T cells in 1/3 cultures after one week exposure to * DCs/WTL was increased significantly compared with those pulsing with TNF/DCs plus IL-2 or IL-2 alone (P = 0.05). T cells derived by priming of non-adherent PBMCs with * DCs/WTL after 14 days in vitro stimulation were significantly more responsive to autologous tumor cells compared with LAK (n = 3, P < 0.05), but its cytotoxicity against K562 cells was also comparable to LAK cells.

CONCLUSIONMonocyte-derived DCs from NSCLC patients could serve as functional APC. The * DCs/WTL may effectively elicit T cell-mediated antitumor response in vitro and enhance NK killing activity.

Antigens, CD1 ; metabolism ; Carcinoma, Non-Small-Cell Lung ; immunology ; Cell Culture Techniques ; Cytotoxicity, Immunologic ; Dendritic Cells ; immunology ; HLA-DR Antigens ; metabolism ; Humans ; Interferon-gamma ; secretion ; K562 Cells ; Killer Cells, Lymphokine-Activated ; immunology ; Leukocytes, Mononuclear ; immunology ; pathology ; Lung Neoplasms ; immunology ; Lymphocyte Culture Test, Mixed ; T-Lymphocytes, Cytotoxic ; immunology ; pathology ; Tumor Necrosis Factor-alpha ; pharmacology

OBJECTIVETo investigate the effect and mechanism of B16F10-ESAT-6-gpi/IL-21 tumor cell vaccine on pulmonary metastasis in mouse model of melanoma.

METHODSTwelve 8-week old female C57BL/6 mice were used in this study. The mice were injected with wild-type B16F10 cells through tail vein after immunization with B16F10-ESAT-6-gpi/IL-21 tumor cell vaccine, and the pulmonary metastasis was observed. The CD4(+) and CD8(+) T cells were isolated by magnetic activated cell sorting, and then used for the detection of CFSE/7-AAD cytotoxicity by flow cytometry. Serum from the mice immunized with tumor-cell vaccine was used to detect IFN-γ expression by ELISA. The expression of TGF-β2, ZEB1, E-cadherin, and N-cadherin of tumor tissues was detected by RT-PCR and immunofluorescence, respectively.

RESULTSThe mice vaccinated with B16F10-ESAT-6-gpi/IL-21 had significantly fewer nodules in the lung and lower lung weight [(285.8 ± 19.01) mg vs. (406.3 ± 27.12) mg], with lower levels of TGF-β2, ZEB1 and N-cadherin proteins but higher level of E-cadherin protein within the tumor tissue, as compared with the control mice. Meanwhile, the immunized mice had significantly increased CD8(+) T cell killing activity [(42.62 ± 3.465)% vs. (22.29 ± 1.804)%] and IFN-γ expression level [(55.200 ± 7.173) pg/ml vs. (6.435 ± 1.339) pg/ml] over the control mice.

CONCLUSIONSThe B16F10-ESAT-6-gpi/IL-21 vaccine can inhibit the metastasis of melanoma in the lung in vaccinated melanoma-bearing mice. This inhibitory effect is associated with CD8(+) T cell immune response and a higher level of IFN-γ, which may influence on the mesenchymal-epithelial transition of tumor cells.

Animals ; CD8-Positive T-Lymphocytes ; immunology ; Cadherins ; metabolism ; Cancer Vaccines ; immunology ; Cell Line, Tumor ; Epithelial-Mesenchymal Transition ; Female ; Homeodomain Proteins ; metabolism ; Humans ; Interferon-gamma ; metabolism ; Interleukins ; immunology ; Lung ; pathology ; Lung Neoplasms ; metabolism ; secondary ; Melanoma ; metabolism ; pathology ; Mice ; Mice, Inbred C57BL ; Neoplasm Transplantation ; Organ Size ; Transcription Factors ; metabolism ; Transforming Growth Factor beta2 ; metabolism ; Zinc Finger E-box-Binding Homeobox 1