Differential roles of PI3-Kinase, MAPKs and NF-κB on the manipulation of dendritic cell T_h1/T_h2 cytokine/chemokine polarizing profile

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• 作者：Bruno Miguel Neves ; Maria Teresa Cruz ; Vera Francisco ; Cá ; rmen Garcia-Rodriguez ; Ricardo Silvestre ; Anabela Cordeiro-da-Silva ; Augusto M. Dinis ; Maria Teresa Batista ; Carlos B. Duarte ; Maria Celes
• 关键词：Dendritic cells ; T-cell polarization ; Signal transduction pathways ; Cytokines ; Chemokines
• 刊名：Molecular Immunology
• 出版年：2009
• 出版时间：August 2009
• 年：2009
• 卷：46
• 期：13
• 页码：2481-2492
• 全文大小：1696 K

文摘

Dendritic cells (DC) are professional antigen-presenting cells with a unique capacity to initiate and modulate immune responses by their ability to prime naïve T-cells. Upon stimuli, DC experience several morphologic, phenotypic and functional changes in a process referred to as maturation. This process is crucial to the biological functions of DC since their maturation status confer them the ability to polarize distinct T-cell subsets. In this work we explored the relevance of PI3-Kinase, Mitogen-Activated Protein Kinases (MAPKs) and NF-kappaB on cytokines/chemokines and co-stimulatory molecules expression. As experimental model, we used a fetal skin-derived dendritic cell line (FSDC) induced to mature by treatment with lipopolysaccharide (LPS). Morphology and ultrastructure were analyzed by confocal and electron microscopies, respectively. Levels of phosphorylated proteins were evaluated by Western blot, production of cytokines/chemokines was analyzed by protein arrays and the expression of surface molecules was evaluated by flow cytometry. The effect of specific inhibitors of the studied signaling pathways on the transcription of cytokines/chemokines and co-stimulatory molecules was accessed by Quantitative Real-Time RT-PCR. The results showed that LPS induces significant morphological and ultrastructural changes in FSDC. Western blot analysis revealed that LPS challenge promotes an early and transient activation of NF-κB, ERK1/2, p38 MAPK, along with a more sustained PI3 kinase/AKT activation. The co-stimulatory CD40, CD80, CD86 and antigen-presenting MHC class I and II molecules were increased and among secreted molecules, interleukin IL-6, CCL5, G-CSF, CCL2, CXCL2 were strongly up-regulated. Using a pharmacological approach we observed that LPS-induced increase of these molecules was differentially regulated by the distinct signaling pathways. Moreover, the polarizing T_h2 cytokines/chemokines induced by LPS in FSDC were found to be positively regulated by NF-κB and ERK and negatively modulated by p38 MAPK. Altogether these results suggest that the use of pharmacological inhibitors to manipulate DC maturation, namely the polarizing T_h1/T_h2 cytokine/chemokine profile, may be useful in the development of more specific immunotherapeutic protocols.