Introduction: NKT cells are categorised as borderline between NK and T cells, sharing phenotypic and functional characteristics of both cells, demonstrating their capacity to contritube to both pro- or anti-inflammatory processes. However, the role of these cells among lung transplant recipients remains largely unknown. The aim of this study was to determine the role of NKT cells following lung transplantation. Methods: NKT cells were quantified and characterised according to markers of: activation (CD107a, CD161, NKG2D) and immunomodulation (CD200 and CD200R) in peripheral blood and BALs. NKT cell numbers and phenotypes were correlated to clinical variables: immunosuppression, acute rejection, acute infections (viral, bacterial and fungal), bronchiolitis obliterans syndrome (BOS grade), lung function, and demographic variables. Interactions between NKT cells and the transplanted lung were linked by determining the relative expression of immunomodulatory ligand CD200 in lung biopsies. In vitro models were employed to determine the role of NKT cells to acute lung injury, either alone or in combination with cells of the mononuclear phagocyte system (MPS). Results: Higher numbers of immunomodulatory NKT cells (CD200+ and CD200R+) were found as lung function decreased. Data from peripheral blood indicates that recipients whose donors or themselves had been exposed to CMV infection demonstrated increased numbers of NKT cells. Patients with active EBV infections demonstrated higher NKT cell numbers expressing CD200 and CD200R. Data from BALs, indicates that patients with active fungal infections present higher immunomodulatory (CD200R) NKT cells and lower cytotoxicity marker (CD107a). In peripheral blood, lung recipients demonstrated higher NKT cell numbers compared to healthy volunteers. However, the lower relative mean expression of functional markers in the lung transplant group suggests that cells are less active. In vitro cultures with immunosuppressants demonstrated that cell cycle inhibitors (MMF and AZA) and corticosteroids (Prednisolone) are likely to inhibit NKT cell proliferation, while calcineurin inhibitors (Cyclosporine A and Tacrolimus) decrease the relative mean expression of activation markers. Clinical observations indicate that higher doses of Azathioprine may correlate with increased NKT cell numbers and the relative expression of CD200 and CD200R. However, under these conditions the relative expression of activation marker NKG2D decreases. In vitro data from the acute injury model indicates that NKT cells are capable to migrate into the injured lung and become activated following transmigration which is facilitated by the presence of monocytes. We also observed the interaction of NKT cells with endothelial cells, monocytes and macrophages. Also, the relative mean expression of CD200 and CD200R increased at the capillary layer, regardless of injury while upregulation of activation markers (CD107a, CD161 and NKG2D) was found at the capillary layer, following injury. In contrast, the alveolar layer demonstrated a decrease in both activation and immunomodulatory markers, following acute injury. Conclusions: Despite immunosuppression, NKT cells remain present in peripheral blood and BAL following lung transplantation. NKT cell proliferation is likely to be reduced by effect of cell cycle inhibitors, while calcineurin inhibitors exert an immunomodulatory effect. Our data indicates that NKT cells can participate in inflammatory and immunomodulatory events at the alveolar bilayer. Their capacity to infiltrate the lungs was assisted by cells of the mononuclear phagocyte system (MPS), which play an important role in antigen presentation and modulation of acute injury. Further research is needed to elucidate the signals and mechanisms occurring between NKT and MPS interactions and the outcomes these populations drive in acute lung injury.

Inhalation of chemical substances can cause irritation to airways and in high doses acute airway injury. When mice are exposed to the alkylating nitrogen mustard analogue melphalan they develop an acute airway inflammation with a rapid influx of neutrophils to the lungs. The acute phase is followed by long-term respiratory complications characterized by bronchitis, lung fibrosis, and airway hyperreactivity.      In this thesis, a mouse model for chemical airway inflammation was established and the effects on the lungs in a time span from 6 hours up to 3 months were investigated in order to study both acute effects and possible chronic injury. We find that treatment with corticosteroids, e.g. dexamethasone, effectively blocks the inflammatory reaction in several ways: Neutrophil influx to the lungs is diminished, the expression of the proinflammatory cytokines interleukin (IL) -6 and IL-1b is decreased and edema formation as well as development of lung fibrosis is mitigated. In acute airway inflammation we show that the antioxidant vitamin E can be used as a possible complement to corticosteroids but not as a replacement since it causes insufficient downregulation of the inflammatory response. We show the importance of the T lymphocytes as they play a prominent role in the pathogenesis of long-term lung injuries caused by melphalan. Especially the minor gd T cell subset is of major importance orchestrating a number of responses including the acute cytokine and neutrophil response and late-phase lung fibrosis. In order to find the critical time for dexamethasone treatment, mice were exposed to melphalan, treated with dexamethasone at specific time points and lung physiology and airway reactivity was measured in anaesthetized, tracheostomized mice using a small animal ventilator. From these results we conclude that an early treatment, i.e. within one hour after exposure, with dexamethasone is needed to prevent chronic lung injury.  This thesis was undertaken with the main goal to better understand the pathogenesis of melphalan-induced airway inflammation. We believe that our findings have shed new light in this area of research and hope that this increased knowledge may be of future clinical use.

Mechanically ventilated patients are at risk of developing Ventilator-Associated Lung Injury (VALI). Improved ventilation strategies by lung-protective settings may cause hypercapnia. This study investigated whether attenuation of VALI is attributed to protective ventilation with low tidal volume (VT) or hypercapnia. Lung injury was induced in rats by instillation of 1.25M HCl. Ten rats each were ventilated for 4 hours with: Conventional Normocapnia (highVT), Lung-Protective Ventilation (VT¬ 8mL/Kg), Injurious Normocapnia (highVT, added dead space), Conventional Hypercapnia (highVT, inhaled CO2), Protective Hypercapnia (VT 8mL/Kg, inhaled CO2) and Permissive Hypercapnia (VT 8mL/Kg, hypoventilation). Lung-Protective Ventilation reduced pulmonary edema compared to Conventional and Injurious Normocapnia. Therapeutic hypercapnia reduced alveolar damage and inflammation by reducing IL-6 and MCP-1 in the lung, and IL-1? and TNF-? systemically. Therapeutic hypercapnia may be more effective in attenuating some of the biomarkers of VALI and protecting the lung than protective ventilation alone.

No description available.

Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / A pancreatite aguda (PA) Ã considerada uma situaÃÃo de emergÃncia abdominal, na forma grave da doenÃa os pacientes desenvolvem acentuada resposta inflamatÃria sistÃmica e SÃndrome de DisfunÃÃo de MÃltiplos ÃrgÃos (SDMO). Um terÃo das mortes relacionadas com PA acontecem antes da admissÃo hospitalar, e a maior parte dos casos estÃo relacionados com lesÃo pulmonar aguda (LPA) e sÃndrome do desconforto respiratÃrio agudo (SDRA). Objetivos: Avaliar as alteraÃÃes inflamatÃrias e funcionais do pulmÃo no curso da pancreatite aguda experimental induzida por ceruleÃna. MÃtodos: PA foi induzida em Ratos Wistar, machos pensando 100-150g, pela administraÃÃo de 4 doses de ceruleÃna (20Âg/kg) com intervalo de uma hora e os grupos controle receberam apenas soluÃÃo salina. ApÃs 24 horas, os animais foram sedados, analgesiados e traqueostomizados e anÃlise da funÃÃo pulmonar foi realizada atravÃs da espirometria, onde foram avaliados Fluxo, Volume Corrente (VC), FrequÃncia RespiratÃria (FR) e Volume Minuto (VM), e da mecÃnica pulmonar onde foram observados ElastÃncia DinÃmica (Edin), ComplacÃncia DinÃmica (Cdin), PressÃo de Pico, ResistÃncia (Res). Lavado bronco-alveolar (LBA) foi realizado para contagem total e diferencial de cÃlulas. Amostra de sangue arterial foi colhida para avaliaÃÃo dos parÃmetros gasomÃtricos. Em seguida os animais foram sacrificados e nÃveis sÃricos de amilase, lipase, EPO, TNF-α, GRO-KC, MIP-1, VEGF, IL-1β, IL-2, IL-6, IL-10, IL-12, IL-17, IL-18 e de malondialdeÃdo (MDA) foram medidos. Atividade de mieloperoxidase (MPO) e avaliaÃÃo histolÃgica de pÃncreas e pulmÃo foram determinadas. AlÃm disso, amostras de sangue venoso foram colhidas para avaliaÃÃo de translocaÃÃo bacteriana. Resultados: NÃveis sÃricos de amilase, lipase, citocinas, MDA e atividade de MPO pancreÃtica e pulmonar estavam aumentados nos animais com PA; houve danos ao tecido pancreÃtico e pulmonar, revelados na histologia, nos animais que receberam ceruleÃna, quando comparados ao grupo controle. O LBA dos animais tratados com ceruleÃna demonstrou maior quantidade de cÃlulas, sendo predominantemente macrÃfagos. Gasometria arterial nÃo apresentou diferenÃa significativa entre os grupos. Fluxo, VC e VM se mostraram diminuÃdos nos animais com PA; FR permaneceu inalterada. Edin e PressÃo de Pico estavam maiores e Cdin estava menor nos animais com PA e nÃo houve alteraÃÃes na Res. Estudo da bacteremia foi negativo em ambos grupos. ConclusÃo: CeruleÃna induz PA em ratos com elevaÃÃo dos nÃveis de amilase e lipase pancreÃtica, com alteraÃÃes histopatolÃgicas no pÃncreas e no pulmÃo dependente do infiltrado neutrofÃlico, radicais livres e citocinas inflamatÃrias. PA induz alteraÃÃes espiromÃtricas e na mecÃnica pulmonar que nÃo sÃo dependentes de processo infeccioso. / Acute pancreatitis (AP) is considered an emergency abdominal, the severe form of the disease patients develop intense systemic inflammatory response and Multiple Organ Dysfunction Syndrome (MODS). About one-third of all deaths from acute pancreatitis has been reported to occur prior to admission to hospital, and in most cases, is associated with acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Objectives: To assess the inflammatory and functional lung alterations in the course of AP induced by cerulein. Methods: Male Wistar rats (100-150g) were treated four times with one hour interval, intraperitoneally with cerulein (20 μg / kg, suspended in saline) or saline. Twenty-four hours after the first injection of cerulein, the animals were anesthetized, tracheostomized and placed in a spirometer for small animals and with following parameters evaluated: Flow, Volume(VC), Respiratory Frequency(RF) and Minute Volume(MV), and lung mechanics were observed where Dynamic Elastance (Edyn), Dynamic Compliance (Cdyn), Peak Pressure, Resistance (Raw). Bronchoalveolar lavage (BAL) was performed to count and differential cell. Arterial blood sample was drawn for assessment of pulmonary gas exchange parameters. Then the animals were sacrificed and serum amylase, lipase, EPO, TNF-α, GRO-KC, MIP-1, VEGF, IL-1β, IL-2, IL-6, IL-10, IL-12, IL-17, IL-18 and malondialdehyde (MDA) were measured. Myeloperoxidase activity (MPO) and histological evaluation of pancreas and lung were determined. In addition, venous blood samples were collected for evaluation of bacterial translocation. Results: Serum levels of amylase, lipase, cytokines, MDA and MPO activity of pancreatic and lung were increased in animals with PA, there was damage to pancreatic tissue and lung histology revealed, in animals that received cerulein compared to the control group. There was an increase in the number of BAL cells, predominantly macrophages. Arterial blood gas analysis showed no significant difference between groups. Flow, and MV proved lower in animals with PA; FR remained unchanged. Edyn and pressure peak were larger and Cdyn was lower in animals with PA and no changes in Res. There was no translocation in any groups. Conclusion: Cerulein induced AP in rats with elevated serum amylase and pancreatic lipase, with histopathological changes in the pancreas and lung dependent neutrophilic infiltrate, free radicals and inflammatory cytokines. PA induces spirometric and lung mechanics alterations that are not dependent on bacterial translocation

Orientadores: Ilma Aparecida Paschoal, Mônica Corso Pereira, Lilian Tereza Lavras Costallat / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-19T08:01:04Z (GMT). No. of bitstreams: 1 Leite_MarivoneArruda_M.pdf: 553765 bytes, checksum: 523c010d8a4b64d307b61b14f6c9b8d4 (MD5) Previous issue date: 2011 / Resumo: Objetivo: avaliar os pacientes com LES sem sintomas respiratórios significativos, por meio do teste de caminhada de 6 minutos (TC6). Casuística e Métodos: foram selecionados 45 pacientes com LES estáveis. Os pacientes foram avaliados quanto à dispneia (MRC), foram submetidos a testes de função pulmonar (espirometria e manovacuometria) e realizaram TC6 (protocolo da ATS/ERS). Dois parâmetros de TC6 (distância e dessaturação) foram comparados com as outras variáveis estudadas. Resultados: Dos 45 pacientes, 42 eram mulheres com idade média de 39±11.4 anos; a média do tempo de doença foi 121±93.1 meses; valor da média do MRC 2±0; a média da CVF foi 85.9±34.2%; a média do VEF1 67.5±21.6%; média PiMáx 82±58.4%; média PeMáx 78±37.3%; média FC em repouso 75±12.8 bpm; FR em repouso 19±5.3bpm; média da distância caminhada no TC6 foi 478±82m; média SpO2 em repouso foi 98±0.8%; média da queda da SpO2 foi de 4±6 pontos. Quando a população de estudo foi dividida de acordo com o valor da distância caminhada, a FC antes do TC6 foi significantemente menor no grupo que caminhou '> ou =' 400m quando comparados com o grupo que caminhou < 400m (p=0.0043), assim como o valor da Escala de Borg (p=0.0036); de acordo com a presença de dessaturação, a FC ao final do teste foi significantemente maior no grupo que dessaturou (p=0.0170), PeMáx (p=0.0282) e a distância caminhada no TC6 (p=0.0291) foi significativamente menor, enquanto que a PiMáx mostrou uma tendência para também ser menor (p=0.0504). CVF<limite inferior de normal era significativamente associado com o grupo que dessaturou (p=0.0274). Conclusão: comparada com a distância caminhada no TC6, a dessaturação foi melhor parâmetro para identificar os pacientes com comprometimento nos índices dos testes de função respiratória / Abstract: Objective: to evaluate SLE patients without overt respiratory symptoms by means of six-minute walk test (6MWT). Casuistic and Methods: 45 stable SLE patients were enrolled. Patients were evaluated for dyspnoea (MRC), underwent pulmonary function tests (spirometry and manovacuometry ) and performed the 6MWT protocol (ATS / ERS). Two parameters of the 6MWT (distance anddesaturation) were compared with the other variables. Results: Of the 45 patients, 42 were women with mean age 39±11.4 years; mean duration of disease was 121±93.1 months; mean value of MRC was 2±0; mean FVC 85.9±34.2%; mean FEV1 was 67.5±21.6%; mean MIP was 82±58.4%; mean MEP was 78±37.3%; mean heart rate at rest was 75±12.8 bpm; mean respiratory rate at rest was 19±5.3bpm; mean 6MWD was 478±82m; mean SpO2 at rest was 98±0.8%; mean fall in SpO2 was 4±6 points. When the study population was divided according to the cut-off value of 400m of walk distance heart rate immediately before the test was significant smaller in those who walked less than 400m (p=0.0043) as was the value of Borg scale(p=0.0036); according to the presence of desaturation '> or =' 4, heart rate at the end of the test was significant higher in those who desaturate (p=0.0170), MEP (p=0.0282) and 6MWD (p=0.0291) were significantly smaller, MIP showed a tendency towards being smaller(p=0.0504). FVC < inferior limit of normal was significantly associated with the group that desaturates (p=0.0274). Conclusion: compared with 6MWD, desaturation was better suited to find the patients with the most compromised indexes in respiratory function tests / Mestrado / Clinica Medica / Mestre em Clinica Medica

京都大学 / 新制・課程博士 / 博士(医学) / 甲第23071号 / 医博第4698号 / 新制||医||1049(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 伊達 洋至, 教授 YOUSSEFIAN Shohab, 教授 平井 豊博 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Acute lung injury (ALI) is a critical manifestation of sepsis/septic shock. Heat shock protein A12B (HSPA12B), an endothelial cell-expressed heat shock protein, shows a negative regulation of lipopolysaccharide (LPS)-induced inflammation in myocardium and endothelial cells. However, it is unclear whether HSPA12B exerts protective effects against ALI during sepsis/septic shock. In this study, we treated HSPA12B transgenic mice (Tg) and wild type littermates (WT) with LPS for 6 h to induce endotoxemia. LPS treatment significantly caused pulmonary injuries as evidenced by microarchitecture destruction, vascular leakage and neutrophil recruitment in lungs of WT mice. However, the LPS-induced pulmonary injuries were significantly attenuated in Tg mice. Moreover, the LPS-induced activation of extracellular signal-regulated kinases (ERKs) and upregulation of intercellular adhesion molecule-1 (ICAM-1) and Cyclooxygenase-2 (Cox-2) were inhibited in Tg lungs compared with that in WT mice. Additionally, Tg lungs showed a significant lower level of vascular endothelial growth factor (VEGF) compared with WT mice. Our results demonstrate a pulmonary protective effect of HSPA12B against endotoxin challenge, which indicates management of HSPA12B expression could serve as a potential therapeutic target for ALI during sepsis/septic shock.

We have recently reported that antioxidant liposomes can be used as antidotes for mustard gas induced lung injury in guinea pigs. The maximum protection was achieved with a liposome composed of tocopherols (α, γ, δ) and N-acetylcysteine (NAC) when administered after 5 min of exposure of 2-chloroethyl ethyl sulfide (CEES), a half sulfur mustard gas. We also reported an association of mustard gas-induced lung injury with an activation of MAPK/AP-1 signaling pathway and cell proliferation. The objective of the present study was to investigate whether CEES-induced MAPKs/AP-1 signaling pathway is influenced by antioxidant liposome therapy. A single dose (200 μl) of the antioxidant liposome was administered intratracheally after 5 min of exposure of CEES (0.5 mg/kg). The animals were sacrificed after 1 h and 30 days of CEES exposure. Although the liposome treatment did not have any significant effect on the activation of the MAPKs family (ERK1/2, p38 and JNK1/2), it significantly counteracted the CEES-induced activation of AP-1 transcription factors and corresponding increase in the protein levels of Fos, ATF and Jun family members. The liposome treatment significantly blocked the CEES-induced increase in the protein levels of cyclin D1, a cell cycle protein and PCNA, a cell differentiation marker. Furthermore, it protected lung against CEES-induced inflammation and infiltration of neutrophils, eosinophils and erythrocytes in the alveolar space. This suggests that the protective effect of antioxidant liposome against CEES-induced lung damage is mediated via control of AP-1 signaling.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20965号 / 医博第4311号 / 新制||医||1026(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 伊達 洋至, 教授 平井 豊博, 教授 江藤 浩之 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM