No description available.

Final publication is available at http://www.sciencedirect.com/science/article/pii/S0006291X1530276X / 京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19563号 / 医博第4070号 / 新制||医||1013(附属図書館) / 32599 / 京都大学大学院医学研究科医学専攻 / (主査)教授 河本 宏, 教授 藤田 潤, 教授 髙折 晃史 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Le sepsis est la réaction inflammatoire généralisée secondaire à une infection. C’est une pathologie fréquente et grave qui fait intervenir le système immunitaire. L’action de l’immunité innée se fait par l’activation et le recrutement des monocytes, des cellules mononuclées circulantes qui modulent le phénomène inflammatoire. La mobilisation des monocytes fait intervenir les cytokines chimiotactiques (chimiokines) et leurs récepteurs. Nous nous sommes spécifiquement intéressé dans ce travail au rôle de l’expression monocytaire des récepteurs aux chimiokines CCR2 et CX3CR1 au cours du sepsis. Pour ce faire, nous avons utilisé des modèles murins de sepsis et analysé le rôle d’un polymorphisme génétique de CX3CR1 dans une cohorte de malades atteints de sepsis. Nous avons montré qu’au cours du sepsis, les monocytes présentaient une augmentation de l’adhérence aux parois vasculaire contrôlée par le récepteur CX3CR1. Nous avons également montré que les monocytes inflammatoires jouaient un rôle essentiel dans la régulation du phénomène inflammatoire au cours du sepsis en protégeant le rein des lésions septiques. Cette protection est médiée par l’expression de CX3CR1. L’allèle I249 du gène Cx3cr1, à l’origine d’une augmentation des propriétés adhésives du couple CX3CR1/CX3CL1, est un facteur protecteur dans la survenue d’insuffisance rénale aiguë chez le malade atteint de sepsis. Collectivement, ces travaux confirment un rôle régulateur des monocytes inflammatoires au cours du sepsis et identifient de potentielles nouvelles cibles thérapeutiques. / Sepsis is the generalized inflammatory response secondary to an infection. This is a common and serious condition that involves the immune system. The action of innate immunity in sepsis is mediated by the activation and recruitment of monocytes, which are mononuclear circulating cells, which modulate the inflammatory process. The mobilization of monocytes involves chemotactic cytokines (chemokines) and their receptors. This work was specifically focused on the role of monocyte expression of chemokine receptors CCR2 and CX3CR1 in sepsis. To this end, we used mouse models of sepsis and analyzed the role of a common genetic polymorphism of CX3CR1 in a cohort of patients with sepsis.We have shown that in sepsis, monocytes’ motility was modified with an increase of their adhesion to vascular walls that was controlled in part by CX3CR1. We have also shown that inflammatory monocytes play a key role in the regulation of the inflammatory phenomenon in sepsis and that they protected the kidney from septic lesions via a CX3CR1 mediated adhesion mechanism. The I249 allele of CX3CR1, confering increased adhesive properties to monocytes, is a protective factor regarding the occurrence of acute kidney injury in septic patients. Collectively, these data confirm a a regulatory role for inflammatory monocytes during sepsis and identify potential new therapeutic targets.

Les monocytes sont des leucocytes dérivés de précurseurs de la moelle osseuse pouvant se différencier en macrophages, cellules dendritiques (CD), ou ostéoclastes (OC). Ils jouent un rôle critique dans la persistance de l'inflammation et la destruction des articulations par le biais de mécanismes encore mal connus. Les monocytes sont composés de deux grandes sous-populations chez la souris, discriminées sur la base du marqueur de surface Ly6C. Il a été suggéré que les OC pouvait être préférentiellement différenciés à partir de la sous-population monocytaire Ly6Chigh, dont l'activation excessive et prolongée est une signature clé de nombreuses pathologies inflammatoires. Parmi les acteurs moléculaires responsables de la régulation de l'expression des gènes, les miARNs jouent un rôle majeur dans de nombreux processus physiologiques, dont la réponse inflammatoire, en régulant finement les programmes génétiques au niveau post-transcriptionnel. Leur implication dans l'ostéoclastogénèse est encore mal connue. Par ailleurs, leur expression est perturbée dans de nombreuses pathologies, dont la polyarthrite rhumatoïde qui implique à la fois un dysfonctionnement de la réponse inflammatoire et de l'homéostasie osseuse. Mon projet de thèse vise à mieux comprendre l'implication des sous-populations monocytaires dans la persistance de l'inflammation et de l'activité des OC au travers de l'étude du rôle de miR-146a dans des conditions physiologiques et inflammatoires. J'ai montré que miR-146a est le miARN le plus différemment exprimé entre monocytes classiques Ly6Chigh et non-classiques Ly6Clow, et que son expression est diminuée dans les Ly6Chigh en conditions arthritiques. J'ai également montré que la perte de miR-146a augmente l'ostéoclastogénèse in vitro et l'érosion osseuse in vivo chez les souris arthritiques. Enfin, la surexpression artificielle de miR-146a dans les monocytes Ly6Chigh inhibe la différenciation osteoclastique et la perte osseuse dans l'arthrite expérimentale chez la souris. Mes résultats suggèrent que miR-146a contrôle l'hétérogénéité fonctionnelle des monocytes et qu'une diminution de son expression dans la sous-population Ly6Chigh serait responsable de l'augmentation de l'osteoclastogénèse et de l'érosion osseuse observées en conditions arthritiques. Pour finir, mes résultats montrent également qu'augmenter l'expression de miR-146a dans les monocytes Ly6Chigh présente un intérêt thérapeutique pour contrecarrer la perte osseuse associée à l'arthrite. / Introduction : Monocytes represent a prototypic cell type when investigating the interplay between immune and skeletal systems as they can give rise to different cell types including dendritic cells, macrophages and osteoclasts (OC), which play key roles in immunity and bone homeostasis. Circulating monocytes consist of at least two main functional subsets, Ly6Chigh and Ly6Clow monocytes. It has been suggested that OC might develop preferentially from the Ly6Chigh monocyte subset, which excessive and prolonged activation is a hallmark of many inflammatory diseases. Among key molecular rheostats of cell fate, micro(mi)RNAs are a class of regulatory RNAs that control basic biological functions and orchestrate inflammatory responses. Few miRNAs have been involved in osteoclastogenesis. The present study aimed at investigating the role of miRNAs in osteoclastogenesis in the context of monocyte subsets, under steady state and inflammatory conditions. Methods & Results : Using genome-wide miRNA expression study we have identified miRNAs and putative targeted pathways that are differentially expressed between Ly6Chigh and LyC6low FACS sorted mouse monocytes, and common to their human counter parts CD14+CD16- and CD14dimCD16+ monocytes, respectively. Among these, miR-146a showed higher expression in Ly6Clow monocytes when compared to Ly6Chigh monocytes. Under inflammatory arthritis conditions, expression of miR-146a in Ly6Chigh monocytes was down regulated as compared to healthy controls. Using mouse deficient for miR-146a, we showed that knockdown of miR-146a increased OC differentiation in vitro. While no bone phenotype was evidenced in miR-146a deficient mice, nor under steady state or ovariectomized conditions, arthritis-induced bone resorption and bone loss were increased in miR-146a knockout mice. Finally, using a liposomal formulation able to delivermiR-146a mimics to Ly6Chigh monocytes upon intravenous injection, we showed that enforced expression of miR-146a led to decreased number of TRAP positive cells within the synovium of arthritic mice, and efficiently reduced bone erosion in inflammatory arthritis. This effect was associated with decreased RelB expression in miR-146a-overexpressing Ly6Chigh osteoclast progenitors. Conclusion : Overall, our results show that specific over-expression of miR-146a in Ly6Chigh monocytes altered OC differentiation and decreased bone erosion in inflammatory arthritis. These data suggest a novel role for miR-146a in controlling osteoclast fate of Ly6Chigh monocyte progenitors and that reduced miR-146a expression in Ly6Chigh monocytes under arthritic conditions contributes to pathogenic bone loss. Finally, delivery of miR-146a mimics to Ly6Chigh monocytes may offer valuable therapeutic potential to interfere with pathological bone loss.

No description available.

In this thesis I examined the role of circulating monocytes and lung macrophages in the pathogenesis of the early fibrotic, progressive fibrotic and resolution phases of pulmonary fibrosis. Pulmonary fibrosis with destruction of lung architecture and consequent respiratory failure and death represents a massive worldwide health burden. Although idiopathic pulmonary fibrosis (IPF) is the archetypal and most common cause of lung fibrosis, numerous respiratory diseases can progress to pulmonary fibrosis, and this usually signifies a worse prognosis. Importantly, the incidence and prevalence of IPF continue to rise and it remains one of the few respiratory conditions for which there are no effective therapies. The mechanisms resulting in pulmonary fibrosis are controversial. Early work in the 1980s and 1990s suggested that lung macrophages were important. However, at the turn of the 21st century there was a shift to a belief that pulmonary fibrosis resulted from aberrant wound healing as a consequence of repetitive epithelial injury from an as yet unknown cause. However, with the ever expanding knowledge of the importance of macrophages in other fibrotic conditions such as the kidney and liver, the potential importance of macrophages in pulmonary fibrosis has become more pertinent. Using an in vivo depletional strategy in several murine models of lung fibrosis, in conjunction with human studies, I sought to characterise the role of circulating monocytes and lung macrophages in the pathogenesis of pulmonary fibrosis. I have established that circulating monocytes and lung macrophages are not critical for the development of early lung fibrosis. In contrast, circulating monocytes and lung macrophages are important during the progressive fibrotic phase of lung fibrosis. Furthermore, my data suggest that the pro-fibrotic alternatively activated macrophages may be the sub-class of macrophages that mediate this fibrogenic effect. In addition and in contrast, I have established that lung macrophages are required for the resolution of fibrosis. This finding is in keeping with important work performed in the field of liver fibrosis. There is an ever increasing literature examining the role of matrix metalloproteinases (MMPs) during tissue fibrosis and repair. My work has suggested that during lung fibrosis there may be compartmental specific functions of MMPs that regulate lung fibrogenesis, although more work is required before this exciting finding can be properly defined.

Made available in DSpace on 2014-06-11T19:30:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-02-12Bitstream added on 2014-06-13T20:21:27Z : No. of bitstreams: 1 takahagi_en_dr_botib.pdf: 614550 bytes, checksum: a7740b5af41df8726b1c4a6469ca54e5 (MD5) / Receptores Toll-like RECONHECER componentes distintos da FUNGOS para iniciar a resposta imune inata. Examinamos se Paracoccidioides brasiliensis OU SEUS gp43 antígenos imunodominantes pode modular IN VITRO E TLR2 TLR4 expressão e produção TRIGGER citocinas por monócitos humanos. Monócitos de indivíduos saudáveis ​​foram incubadas com gp43 LYPOPOLYSACCHARIDE, (LPS) ou mortas pelo calor FORMAS leveduriforme do P. brasiliensis em uma proporção de 50 monócitos por célula fúngica (Pb18) a 37 º C por 4h e 18h. A EXPRESSÃO DE TLR2 E TLR4 SOBRE SUPERFÍCIE monócitos, e TNF-alfa, IL-10 e IL-12p40 PRODUÇÃO foram determinados por citometria de fluxo e ELISA respectivamente. Os resultados mostraram que a estimulação de monócitos COM OU LPS Pb18 PROMOVIDO UP-regulação da expressão TLR2 E TLR4 SOBRE SUPERFÍCIE EM RELAÇÃO monócitos às células não-estimulados EM AMBAS AS 4H e 18h de cultura, e aumentaram os níveis de TNF-alfa, IL- 10 e IL-12p40 PRINCIPALMENTE EM 18H DA CULTURA. POR OUTRO LADO, E EXPRESSÃO TLR4 BAIXA ALTA TLR2 são evocados por gp43 AT 4H DA CULTURA, associadas a altos níveis de TNF-alfa. No entanto, após 18H UMA MUDANÇA DE ALTA E BAIXA TLR2 TLR4 EXPRESSÕES foi seguido por elevados níveis de IL-10 e IL-12p40. Estes resultados sugerem que a gp43 pode induzir um desequilíbrio entre RESPOSTAS pró e anti-inflamatório em FUNGOS-monócitos interações, um efeito modulatório na CAMINHO TLR. AS TNF-alfa pode estar envolvida na patogênese da paracoccidioidomicose, O EFEITO regulatórios induzidos por gp43, VIA upregulation de TLR2 EXPRESSÃO E IL-10 produção, pode ser importante para proteger contra a lesão de tecido que é descrito neste micose. / Toll-like receptors recognize distinct components of fungi to initiate the innate immune íesponse. We examined whether Paracoccidioides brasiliensis ar its immunodominant antigen gp43 can modulate in vitro TLR2 and TLR4 expression and trigger cytokine production by human monocytes. Monocytes from healthy individuaIs were incubated with gp43, IypopoIysaccharide (LPS) or heat-killed yeast forms of P. brasiliensis in a ratio of 50 monocytes per funga! cell (pb 18) at 37°C for 4h and I8h. The expression of TLR2 and TLR4 on monocyte surface, and TNF-alpha, IL-Io and IL-I2p4o production were determined by flow cytometry and ELISA respectiveIy. The results showed that monocyte stimuIation with LPS ar Pb18 promoted up-reguIation ofthe TLR2 and TLR4 expression on monocyte surface in reIation to the non-stimuIated cells at both 4h and I8h ofculture, and induced higher IeveIs ofTNF-aIpha, IL-Io and IL-12p4o mainly at I8h of culture. On the other hand, high TLR4 and low TLR2 expression were elicited by gp43 at 4h of cuIture, associated with higher Ievels of TNF-aIpha. However, after 18h a change to high TLR2 and !ow TLR4 expressions was followed by e1evated levels of IL-Io and IL-12p4o. These results suggest that gp43 might induce an imbalance between pro- and anti-inflammatory responses in fungal-monocyte interactions by a modulatory effect on TLR pathway. As TNF-alpha may be invoIved in the pathogenesis of paracoccidioidomycosis, the regulatory effect induced by gp43, via upregulation of TLR2 expression and IL-Io production, can be important to protect against tissue injury which is described in this mycosis.

Abstract not available

Thèse de doctorat : Ingénierie Cellulaire et Tissulaire : Nancy 1 : 2008. / Titre provenant de l'écran-titre.

Thesis (Ph. D.)--University of Washington, 2001. / Vita. Includes bibliographical references (leaves 232-255).