TLR4 | EPFL Graph Search

Le Toll Like Receptor 4, ou TLR 4, parfois également nommé CD284 (Cluster de différenciation 284), est un récepteur membranaire, membre de la famille des récepteurs de type toll (TLR), qui appartient à la famille des récepteurs de reconnaissance de formes (PRR pour Pattern Recognition Receptor). Son poids moléculaire est d'environ 95 kDa et son gène est le TLR4 situé sur le chromosome 9 humain. Il est retrouvé sur les macrophages ainsi que sur certains adipocytes (pré-adipocytes et adipocytes matures) et sur certaines microglies du système nerveux central. Les différents TLR présentent différents modèles d'expression. Ce récepteur est exprimé le plus abondamment dans le placenta et dans la sous-population myélo-monocytaire des leucocytes. Les TLR sont hautement conservés de la Drosophile à l'homme et partagent des similitudes structurelles et fonctionnelles. Ils reconnaissent les modèles moléculaires associés aux agents pathogènes (PAMP pour Pathogen-Associated Molecular Pattern) qui sont exprimés sur des agents infectieux et interviennent dans la production de cytokines nécessaires au développement d'une immunité efficace. Son activation conduit à une voie de signalisation intracellulaire NF-κB et à la production de cytokines inflammatoires, responsables de l'activation du système immunitaire inné. Il est bien connu pour la reconnaissance du lipopolysaccharide (LPS), aussi nommé endotoxine, un composant présent dans de nombreuses bactéries à Gram négatif (par exemple Neisseria spp.) et certaines bactéries à Gram positif. Ses ligands comprennent également plusieurs protéines virales, un polysaccharide et diverses protéines endogènes telles que les lipoprotéines de basse densité, les bêta-défensines et la protéine de choc thermique. Plusieurs variantes de transcription de ce gène ont été trouvées, mais le potentiel de codage de protéines de la plupart d'entre elles est incertain.

Engineering the Extra Domain A of fibronectin as a vaccine adjuvant and study of its role in tissue regeneration

Ziad Julier

The extracellular matrix (ECM) protein fibronectin (FN) is a remarkably multifaceted molecule, including numerous fibronectin type III (FNIII) repeats carrying out different functions, which despite being extensively studied for over thirty years still presents certain functions that remain incompletely elucidated. The alternatively spliced FN variant containing the extra domain A (FNIII EDA), located between FNIII 11 and FNIII 12, is expressed in sites of injury, chronic inflammation, and solid tumors. Although its function is not well understood, FNIII EDA is known to agonize Toll-like receptor 4 (TLR4) and appears to be involved in the progression of different skin diseases while studies also suggest that FNIII EDA is involved in tissue repair. The aim of this thesis is thus to better understand some of the immunological and regenerative functions of FNIII EDA and to translate these findings to develop new vaccination strategies based on endogenous proteins. One important task in vaccinology is to develop delivery strategies by which to present antigen, along TLR agonists, so as to induce a desirable adaptive immune response. The use of TLR agonists is associated with sometimes severe constraints and concerns about safety, causing inflammatory responses. Thus, the development of safe and effective vaccines with fewer side effects is a strong need and requires the development of new adjuvants with a correct balance between a strong activation of the immune system and a low toxicity for the patient. Here, by producing various FN fragments containing FNIII EDA, we show that the immunological activity of FNIII EDA depends on its local intramolecular context within the FN chain. N-terminal extension of the isolated FNIII EDA with its neighboring FNIII repeats enhanced its activity in agonizing TLR4, while C-terminal extension abrogated it. In addition, we reveal that an elastase 2 cleavage site is present between FNIII EDA and FNIII 12. Moreover, as FNIII EDA is naturally present within the ECM, we incorporated FNIII EDA bound in a fibrin matrix to mimic the in vivo situation and create an adjuvant-rich microenvironment. Such matrices were shown to induce functional cytotoxic CD8+ T cell responses in two murine cancer models. Furthermore, combinations of specific TLR-agonists have the ability to synergize to induce immune responses that are greater than the sum of their individual effect. Hence, we demonstrate that combination of the TLR9 agonist CpG with FNIII EDA synergizes to induce an efficient immune response while keeping the dose of the toxic CpG at a minimum. Here, the combination of both adjuvants induced a potent immune response sufficient to slow down tumor growth in a murine tumor model as well as to break immune tolerance to eradicate circulating hepatitis B virus (HBV) in a transgenic HBV mouse model. Finally in the last part of this thesis we revealed that FNIII EDA is a growth factor binding domain of FN, with a strong affinity for the platelet-derived growth factor-AA (PDGF-AA). We also demonstrated that FNIII EDA and PDGF-AA have together the ability to enhance fibroblasts proliferation and differentiation into myofibroblasts. In addition, we explored the effects of fibrin matrices functionalized with variants of FNIII EDA on wound closure.

EPFL2016

Engineering the Extra Domain A of fibronectin as a vaccine adjuvant and study of its role in tissue regeneration

Ziad Julier

EPFL2016