Neutrophils are implicated in many inflammatory lung disorders. However, the mechanisms regulating neutrophil migration in acute lung inflammation are incompletely understood. Although, integrin β2 mediates neutrophil migration in lungs in response to many stimuli such as E. coli, integrin involved in <i>S. pneumoniae</i> induced neutrophil migration is not known. Therefore, the role of integrin αvβ3 in neutrophil recruitment was tested. First, it was found that the number of neutrophils expressing the integrin subunits αv and β3 is reduced or remains in lung inflammation induced by E. coli or <i>S. pneumoniae</i>, respectively. Next, the role of integrin αvβ3 using β3 knockout mice (β3-/-) and function blocking antibodies was addressed. Neutrophil recruitment did not vary between wild type and β3-/- mice. Although β3 antibodies reduced neutrophil recruitment, similar effect was observed with isotype antibodies. Therefore, one can conclude that integrin αvβ3 is not critical for neutrophil recruitment in <i>S. pneumoniae</i> induced pneumonia. <p>Apart from integrins, TLR4 also regulate neutrophil migration. Because, the pattern of TLR4 expression at various times of lung inflammation is not known, TLR4 expression during different phases of lung inflammation in a rat model of LPS-induced inflammation was studied. TLR4 expression in the septum increased and decreased at 6h and 12-36h of inflammation, respectively. Since these correlate with the time of increase and decline of neutrophil recruitment, the findings support previously observed requirement for TLR4 in neutrophil recruitment. <p>Neutrophils recruited into the lungs regulate the inflammatory process by controlling subsequent monocyte/macrophage recruitment. The mechanisms involved and the pattern of monocyte/macrophage recruitment in lungs are not completely understood. Therefore, the possible involvement of monocyte chemoattractant protein (MCP)-1, which is a premier chemokine in monocyte/macrophage migration and produced by neutrophils and other cells was tested. This was addressed by quantification of monocytes/macrophages at various times and using neutrophil depletion experiments in LPS-induced lung inflammation in rats. It was found that monocytes/macrophages migrate very early and before neutrophils in addition to their migration in the late phase of acute lung inflammation. Neutrophil depletion abrogated both early as well as the late monocyte/macrophage recruitment without altering the expression of MCP-1. Therefore, possibly other chemokines and not MCP-1 are involved in neutrophil dependent monocyte/macrophage recruitment. <p>To conclude, the experiments further the understanding on acute lung inflammation by ruling-out the involvement of integrin αvβ3 and MCP-1 in β2-independent neutrophil migration and neutrophil dependent monocyte/macrophage recruitment, respectively. Further studies are essential to find the integrins and chemokines operating in the above situations. Equally important will be to understand the functional significance of early recruited monocytes/macrophages in the lung.

<p>with biliary cirrhosis are highly susceptible to acute pulmonary dysfunction and suffer from hepato-pulmonary syndrome. The mechanisms of this enhanced susceptibility remain unknown. It is well established that pulmonary intravascular macrophages (PIMs) are present in cattle, horses, goat and sheep and increase susceptibility for lung inflammation. Species such as rat and mouse also recruit PIMs especially in a bile duct ligation model of biliary cirrhosis. The contributions of recruited PIMs to lung inflammation associated with liver dysfunction remain unknown. Therefore, I characterized a bile duct ligation (BDL) model in rats to study role of recruited PIMs in lung inflammation. First, Sprague Dawley rats were subjected to BDL (N=6) or sham surgeries (N=3) and were euthanized at 4 weeks post-surgery. Five rats were used as the controls. Lung tissues were collected and processed for histology, immunohistology, immuno-electron microscopy, enzyme-linked immunosorbant assay (ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR). Light microscopy demonstrated normal lung morphology in sham-operated and control rats but showed septal recruitment of mononuclear cells, which were positive for anti-rat monocytes/macrophage antibody ED-1, in BDL rats (p=0.002). Immuno-electron microscopy confirmed localization of ED-1 in PIMs. BDL rats showed increased lung expression of monocyte chemoattractant protein-1 (MCP-1) protein and mRNA compared to the controls (p=0.017) but not of IL-1â, TNF-á, TGF-â and IL-10. Then, I treated BDL rats (N=5) with gadolinium chloride (GC; 10 mg/Kg body weight intravenous) and found reduced numbers of PIMs (p=0.061) at 48 hours post-treatment along with increased expression of TGF-â and IL-10.</p><p>I challenged control rats (N=5) and BDL rats (N=6) with Escherichia coli lipopolysaccharide (E. coli LPS; 0.1 mg/Kg body weight intravenous). All the BDL rats died within 3 hours of LPS challenge (100% mortality) while the normal LPS-treated rats were euthanized at 6 hours post-treatment. Histology and ED-1 staining showed dramatic increase in the number of septal monocytes/macrophages in BDL+LPS rats compared to normal LPS-treated rats (p=0.000). Staining of lung sections with an LPS antibody localized the LPS in lungs. RT-PCR analyses showed no differences in IL-1â transcript levels between LPS challenged BDL rats and LPS challenged control rats (p=0.746) but ELISA showed increase in IL-1â concentration in LPS challenged BDL rats compared to LPS challenged control rats (p=0.000). TNF-á mRNA (p=0.062) and protein (p=0.000) was increased in BDL+LPS rats compared to the control+LPS rats. Immuno-electron microscopy showed IL-1â and TNF-á in PIMs. BDL rats challenged with LPS showed increased expression of IL-10 mRNA and protein (p=0.000 & 0.002 respectively) in lungs compared to LPS challenged control rats. TGF-â mRNA did not change (p=0.128) but lower protein concentrations (p=0.000) were observed in LPS-treated control rats compared to BDL+LPS. </p><p> To further address the role of PIMs, I treated rats with GC at 6 hours or 48 hours (N=5 each) before LPS challenge. The mortality in the 6 hour group was 20% while all the rats in 48 hour group survived till 6 hours. Histology and ED-1 staining showed decrease in the number of intravascular cells in these groups compared to LPS treated BDL rats (p=0.039 for 6 hour group; p= 0.002 for 48 hour group). There were no differences in IL-1â mRNA in both 6 hour and 48 hour groups compared to the LPS challenged BDL rats (p=0.712 & 0.509 respectively). ELISA showed no decrease in IL-1â concentration in 6 hour GC-treated group but a decrease was noticed at 48 hours compared to LPS challenged BDL rats (p=0.455 & 0.008 respectively). TNF-á mRNA levels were not different between LPS-challenged GC-treated BDL rats and LPS-challenged BDL rats (p=0.499 & 0.297 for 6 hour & 48 hour GC groups respectively). But TNF-á concentration in 48 hour GC group (p=0.001) but not in 6 hour GC group (p=0.572) was lower in comparison to BDL+LPS group. IL-10 mRNA was decreased in both 6 hour and 48 hour GC groups (p=0.038 & 0.000 respectively) compared to LPS challenged BDL rats. ELISA showed decrease in IL-10 concentration in 48 hour GC group (p=0.030) but not in 6 hour GC group (p=0.420). TGF-â mRNA expression was decreased in 48 hour GC group (p=0.000) but not in 6 hour GC group (p=0.182). But GC treatment did not affect TGF-â concentrations. </p><p>The data from these experiments characterize a BDL model to study PIM biology, show PIMs pro-inflammatory potential and their possible role as a therapeutic target in lung inflammation.</p>

The purpose of this study was to compare power output and ventilatory measurements between the steep ramp test (SR) and both the 30-second Wingate anaerobic (WAT) and standard cardiopulmonary exercise tests (CPET) in chronic obstructive pulmonary disease (COPD). 11 patients (7 males and 4 females) underwent spirometry, a CPET, WAT and SR test. Repeated measures ANOVA was used to compare the differences between the peak work rate of the CPET (CPET_peak), SR (SR_peak), and the average power of the WAT (W_avg). The W_avg was higher than the SR_peak, which was higher than the CPET (231.2 ± 113.4, 156.8 ± 67.9, 65.9 ± 35.9, p>0.05 respectively). There were no differences found between the tests at end-exercise for inspiratory reserve volume (IRV), ventilation (V_E), and end-expiratory lung volume (EELV). Tidal volume (V_T) was also compared between the tests as a percentage of the inspiratory capacity (IC) remaining at end-exercise and no differences were found. The similarity between the ventilatory measures indicates a similar level of constraint, despite the large difference in work rates achieved, in all 3 tests. This shows that a standard CPET underestimates leg power in COPD patients, and the WAT and SR may be better indicators of leg muscle power and anaerobic type exercise.

Despite extensive research, the mechanisms behind cardiovascular effects of subchronic environmental tobacco smoke (ETS) remain unclear, but may be related to ETS-induced inflammation and oxidative stress. Additionally, the protective role of resveratrol (RES), a natural antioxidant available in red grapes, is controversial. We hypothesized that the polycyclic aromatic hydrocarbon (PAH) component of ETS is responsible for causing adverse cardiovascular effects. We also hypothesized that the administration of RES is protective against the adverse cardiovascular effects of ETS. In order to address these hypotheses, male juvenile pigs (4-weeks old) were exposed to ETS or ambient air for 28 consecutive days (1 hr/day) and effects compared to 7 days of i.v. injection of the PAH, benzo-a-pyrene (BAP; 5 mg/kg daily). In another experiment, pigs were sham-exposed or ETS-exposed, with or without oral RES treatment (5mg/kg daily). In all experiments, endothelial and left ventricular function were assessed by flow mediated dilation (FMD), and echocardiography, respectively, while blood pressure was evaluated by oscillometry. At the termination of each experiment, serum nitrotyrosine, total nitrate/nitrite (NOx) and C-reactive protein (CRP) were measured as well as hepatic and pulmonary ethoxyresorufin-o-deethylase (EROD) activity to indicate cytochrome P450 1A1 (CYP1A1) expression. Finally, the correlation between pulmonary inflammation and adverse cardiovascular effects was investigated by measuring total and differential white blood cell (WBC) count as well as leukocyte elastase activity in bronchoalveolar lavage fluid at the termination of each experiment. ETS exposure, but not BAP treatment, resulted in a significant impairment of FMD (P<0.0001) and increased left ventricular end diastolic volume (P=0.0032). Cotreatment with RES failed to restore the ETS induced impairment of FMD (P>0.05). However, a trend pointing to an increase in ejection fraction (EF) was noted (P=0.072). ETS, BAP and RES treatments failed to have any effect on blood pressure (P>0.05). BAP injection caused a significant increase in serum nitrotyrosine (P=0.0146) and CRP (P=0.012), but not serum NOx levels (P>0.05). In contrast, ETS exposure resulted in a significant increase in CRP serum levels (P=0.0092), a trend pointing to increased serum nitrotyrosine (P=0.105), and no change in serum NOx levels (P>0.05). The increased nitrotyrosine and CRP with ETS exposure was not reversed by RES administration (P>0.05). ETS exposure increased EROD activity in the lung (P=0.0093), but not the liver (P=0.12). In contrast, BAP treatment had the opposite effect (lung EROD: P=0.621, liver EROD: P=0.01), while RES administration had no effect (P>0.05). ETS exposure (P=0.0139), but not BAP treatment (P=0.723), resulted in increased WBC count in BAL fluid which was not affected by RES administration (P>0.05). These results show that ETS exposure causes lung inflammation, systemic inflammation, oxidative stress-mediated inactivation of nitric oxide and impaired endothelial function. In contrast, BAP failed to alter endothelial function, downstream of the lung, despite systemic inflammation and increased oxidative stress. Furthermore, RES failed to restore endothelial function, or decrease systemic inflammation and oxidative stress. Taken together, these results suggest either that pulmonary inflammatory responses or pulmonary increases in CYP1A1 activity may be more important links to endothelial dysfunction than systemic inflammation and nitric oxide bioactivity. The beneficial effects of RES by itself are manifested only at the cardiac level by improving the ejection fraction, but the work in this thesis failed to detect any ability of RES to ameliorate ETS cardiovascular effects.

In recent decade, lung cancers had the highest incidence and mortality rate among all cancers in Taiwan. Among lung cancers, adenocarcinoma was the most frequent type. The chemotherapy was still the main choice in treating lung cancer by the mechanism of destroying DNA, but the response rate kept low. The function of DNA repair makes cancer cells resistant to chemotherapy. Therefore, this study focused on the effect of cancer cell growth by silencing Mre11. The first part of this study was to make a tissue microarray consisting of adenocarcinoma from 57 patients. Immunohistochemistry staining for Mre11 was done. The correlation of Mre11 expression and clinical variables with survival was analyzed. The second part was tried to knockdown Mre11 in A549 cell by shRNA. Another A549 cell line containing empty vector was selected as control group. These cell lines were then ready for XTT method, soft agar colony formation assay, flow cytometry and nude mice assay. In the clinical data, the absence of lymph node and distant site metastasis were good prognosis factor for longer survival. Although the high expression on Mre11 had longer survival, this variable was not a true independent factor. On XTT method and soft agar colony formation assay, the A549 cells with Mre11 knockdown had a slower proliferation and fewer colony numbers, respectively. The cell cycle demonstrated an elevated G0/G1 and S phase and depressed G2/M phase in A549 cells with Mre11 knockdown. The tumor arising from A549 cells with Mre11 knockdown in the nude mice also had a smaller size. Based on the above study, inhibition of Mre11 may result in a reduction of tumor growth and provide another choice to treat lung cancer.

The Reelin gene located on the human chromosome region 7q22, encodes an extracellular matrix glycoprotein, a ligand for ApoER2 and low-density lipoprotein receptors (LDL) Receptor, is required for mediating the correct positioning of neurons during embryonic brain development1. In the current study, first we applied RT-PCR and immunohistochemistry analysis (IHC) analysis on tissue microarrays (TMA) to verify the Reelin expression patterns in a variety of adult tissues, suggesting additional roles for Reelin in stabling the cyto-architecture and controlling the remodeling of many organs during development. Second, we report the Reelin expression status in tumorigenesis. We discover that the loss of Reelin expression is associated with multiple types of cancers, including more than 80% of both breast and colorectal cancers. Interestingly, our study also found suspension small cell lung cancer (SCLC) cell lines that grow as large aggregates retained high Reelin expression, whereas attached non small cell lung cancer cultures do not. That may imply the Reelin expression may be also associated with cell culture morphology and growth characteristics in the in vitro culture system for lung cancers. Our results here also demonstrated that epigenetic silencing of Reelin expression by DNA hypermethylation in tumors directly correlates with loss of Reelin expression in many cancers. Reelinmethylation was reversed and expression restored by treating tumor cell lines with the demethylating agent 5-aza-2-deoxycytidine. In conclusion, from the molecular basis of Reelingene inactivation in human cancer here, we propose that the Reelinvariation in more than 80% of breast and colorectal cancers makes it a significant novel tumor marker.

Tyrosine kinases regulate fundamental signal pathways in cells including cell proliferation, motility, and differentiation. The kinase activity is tightly controlled in normal cells but is usually excessive activated in cancers. Several tyrosine kinase inhibitors are used in cancer therapies nowadays. Our novel tyrosine kinase inhibitor, 1J-309, is a multiple kinase inhibitor that targets several receptors including vascular endothelial growth factor receptors (VEGFRs). We find 1J-309 dramatically reduces cell proliferation of VEGFR3+/VEGF-C+ A549 human lung cancer cells by decreasing the expression of CDK1 and cyclin B1 following growth arrest at G2/M phase. After long term drug treatment, 1J-309 causes cell death. Moreover, 1J-309 represses CDK1 expression at early stage but it does not change CDK1 RNA expression and protein stability. Additionally, 1J-309 significantly decreases the migration ability of A549 cells. 1J-309 also reduces gelatin-related invasion potency. The AKT and p38 MAPK activity are significantly repressed by 1J-309 and it dramatically drives the expression of tumor suppressor, p53, at low-dose treatment. Our results demonstrate that 1J-309 significantly attenuates cell proliferation by inducing G2/M growth arrest, reduces the invasion and migration potency, and promotes a dramatic increase of p53 in A549 cells.

Several computer-aided diagnostic (CAD) methods for solitary pulmonary nodules (SPNs) have been proposed, which can be divided into two major categories: (1) the morphometric CT method, and (2) the perfusion CT method. The first goal of this work is to introduce a neural network-based CAD method of lung nodule diagnosis by combining morphometry and perfusion characteristics by perfusion CT. The proposed approach has the following distinctive features. Firstly, this work develops a very efficient semi-automatic procedure to segment entire nodules. Secondly, reliable nodule classification can be achieved by using only two time-point perfusion CT feature measures (precontrast and 90 s). This greatly reduces the amount of radiation exposure to patients and the data processing time. As demonstrated in previous work, classification tuberculomas from malignancies has been considered to be a challenging task. However the diagnosis accuracy for tuberculomas reaches 92.9% by applying the proposed CAD method. Another goal of this work is, by investigating the relative merits of 2D and 3D methods, to develop a two-stage approach that combines the simplicity of 2D and the accuracy of 3D methods. Experimental results show statistically significant differences between the diagnostic accuracy of 2D and 3D methods. The results also show that with a very minor drop in diagnostic performance the two-stage approach can significantly reduce the number of nodules needed to be processed by the 3D method and thus alleviates the computational demand.

Das Krankheitsbild des schweren akuten Lungenversagens wurde erstmals in den 60er Jahren des 20. Jahrhunderts beschrieben. Initial als relativ seltene Erkrankung angese¬hen, zeigen neuere Untersuchungen eine sehr viel größere Inzidenz. Die Letalität ist mit ca. 40% nach wie vor sehr hoch und die Therapie des ARDS stellt eine komplexe inten¬sivmedizinische Herausforderung dar. Die Patienten profitieren daher von einer frühzei¬tigen Verlegung in ein Behandlungszent¬rum. Die vorliegende Arbeit untersucht in einer retrospektiven Kohortenstudie die Transporte von Patienten mit schwerem Lungenver¬sagen hinsichtlich Sicherheit, auftretender Komplikatio¬nen und den Auswirkungen des Transportes auf die Morbidität und Letalität. Über einen Zeit¬raum von 10 Jahren wurden insgesamt 187 Patiententransporte in die Untersuchung eingeschlos¬sen. Im Untersu¬chungszeitraum nahm die Zahl der luftgebundenen Transporte deut¬lich zu. Damit ver¬bunden war eine erhebliche Reduktion der Transportzeiten. Es fand sich ein signifikan¬ter Zu¬sammenhang sowohl zwischen der Transportdauer als auch der Transportdistanz und der Häu¬figkeit von Komplikationen. Die auffälligste Veränderung im Transportver¬lauf war eine signifi¬kante Erhöhung des PEEP und eine damit verbundene Verbes¬se¬rung der Oxygenie¬rung. Durch Wahl des optimalen PEEP-Niveaus und den Einsatz von NO gelang es häufig den Gasaustausch zu verbessern, so dass der Einsatz der mobi¬len ECMO-Einheit auf wenige Aus¬nahmefälle be¬schränkt blieb. Die vorliegende Unter¬suchung zeigt, dass Transporte von Patienten mit ARDS auch über große Entfer¬nungen sicher durchgeführt werden können und die deutlich höheren Überlebensraten, die in spezialisierten ARDS-Behandlungszentren erreicht werden, rechtfertigen die po¬tentiel¬len Risiken und Gefahren, die mit einem Transport dieser kritisch kranken Pati¬enten verbunden sind.

Thesis (M. Phil.)--University of Hong Kong, 2009. / Includes bibliographical references (leaves 164-173). Also available online.