PI3K isoform p110δ role was explored in ubiquitous kinase deleted or conditional PI3K-gene-targeted mice in models of gut inflammation in vivo and PRR-associated events in bone marrow-derived dendritic cells (DCs) in vitro.
The in vivo inactivation of p110δ rendered dysbiosis and a hyper-inflammatory state revealed by 1) DC hyperactivation in the colon lamina propria (cLP) and mesenteric lymph node (mLN), 2) an exaggerated mucosal IFN-γ and IL-17 cytokine production in CD8 T cell in cLP and mLN, and 3) significantly reduced in Treg response (Foxp3 and IL10 producing) T CD4 cells in cLP and mLN. Indeed, we could demonstrate that all the referred results were similarly reproduced in DC-restricted deletion of p110δ kinase.
At molecular level, PRR-mediated activation of DCs defective in p110δ activity shown defect in Akt phosphorylation in parallel with increased stress MAPK signalling and NF-KB activation. These events culminate in increased release of inflammatory mediators and hyperactivation of caspase-1 and inflammasome activation. Besides, DCs defective in p110δ activity shown significative reduction in the production of intra-phagosomal reactive oxygen species (ROS) creating an state of increased acidification inside the vesicle. This acidic state observed in p110δ defective DCs leads to increased antigen processing and degradation impacting negatively in antigen presentation. Still in the mechanisms orchestrated by p110δ, we demonstrated, for the first time, the interaction between p110δ and RAC2 associating the lack of the referred interaction with the defect in PI3K signalling, increased inflammatory mediators and increased inflammasome activation under PRR activation. In addition, we demonstrate that p110δ and RAC2 interaction is important for ROS production vesicle alkalinization that leads to preserve antigen for presentation.
The results of the proposed research have led to the identification of a new role and mechanism of action of a lipid kinase p110δ PI3K isoform in 1. DC-intrinsic NOD2-dependent activation of PI3K signalling pathway, 2. DC-intrinsic anti-inflammatory role in PRR signalling in vitro and in vivo, 3. New PRR-driven regulatory pathways that induce gut tolerance through DC anti-inflammatory properties keeping gut homeostasis, 4. New mechanisms involved in PRR signalling triggered by p110δ PI3K required for proper antigen presentation and gut tolerance. All aspects studied here could be explored to generate anti-/pro-inflammatory DCs in vitro and in vivo.
The results were presented in local events in the Biochemical Pharmacology Department meetings and William Harvey Research Institute seminars and externally at The Francis Crick Institute.
