p38γ and p38δ modulate innate immune response by regulating MEF2D activation.

Evidence implicating p38γ and p38δ (p38γ/p38δ) in inflammation are mainly based on experiments using Mapk12/Mapk13-deficient (p38γ/δKO) mice, which show low levels of TPL2, the kinase upstream of MKK1-ERK1/2 in myeloid cells. This could obscure p38γ/p38δ roles, since TPL2 is essential for regulating inflammation. Here, we generated a Mapk12D171A/D171A/Mapk13-/- (p38γ/δKIKO) mouse, expressing kinase-inactive p38γ and lacking p38δ. This mouse exhibited normal TPL2 levels, making it an excellent tool to elucidate specific p38γ/p38δ functions. p38γ/δKIKO mice showed a reduced inflammatory response and less susceptibility to lipopolysaccharide (LPS)-induced septic shock and Candida albicans infection than wild-type (WT) mice. Gene expression analyses in LPS-activated wild-type and p38γ/δKIKO macrophages revealed that p38γ/p38δ-regulated numerous genes implicated in innate immune response. Additionally, phospho-proteomic analyses and in vitro kinase assays showed that the transcription factor myocyte enhancer factor-2D (MEF2D) was phosphorylated at Ser444 via p38γ/p38δ. Mutation of MEF2D Ser444 to the non-phosphorylatable residue Ala increased its transcriptional activity and the expression of Nos2 and Il1b mRNA. These results suggest that p38γ/p38δ govern innate immune responses by regulating MEF2D phosphorylation and transcriptional activity.

A Mouse Model of Candidiasis.

The intravenous challenge model of Candida albicans infection in mice is a well-established procedure that mirrors disseminated candidiasis in humans. In this model, in which the fungus is delivered into the bloodstream causing a systemic infection, the kidneys are the primary target organs. Mice develop renal failure and septic shock that recapitulates the progressive sepsis seen in humans during severe clinical cases. This model is used to study inflammation and the host immune response against fungal infection. This chapter describes the intravenous candidiasis infection protocol, detailing different steps from the preparation of the inoculum, injection of Candida, monitoring of animals, collection of tissue from infected mice, sample preparation and analysis of several parameters related to infection and the inflammatory response.

Growth Hormone Reprograms Macrophages toward an Anti-Inflammatory and Reparative Profile in an MAFB-Dependent Manner.

Growth hormone (GH), a pleiotropic hormone secreted by the pituitary gland, regulates immune and inflammatory responses. In this study, we show that GH regulates the phenotypic and functional plasticity of macrophages both in vitro and in vivo. Specifically, GH treatment of GM-CSF-primed monocyte-derived macrophages promotes a significant enrichment of anti-inflammatory genes and dampens the proinflammatory cytokine profile through PI3K-mediated downregulation of activin A and upregulation of MAFB, a critical transcription factor for anti-inflammatory polarization of human macrophages. These in vitro data correlate with improved remission of inflammation and mucosal repair during recovery in the acute dextran sodium sulfate-induced colitis model in GH-overexpressing mice. In this model, in addition to the GH-mediated effects on other immune cells, we observed that macrophages from inflamed gut acquire an anti-inflammatory/reparative profile. Overall, these data indicate that GH reprograms inflammatory macrophages to an anti-inflammatory phenotype and improves resolution during pathologic inflammatory responses.

Myeloid cell deficiency of p38γ/p38δ protects against candidiasis and regulates antifungal immunity.

Candida albicans is a frequent aetiologic agent of sepsis associated with high mortality in immunocompromised patients. Developing new antifungal therapies is a medical need due to the low efficiency and resistance to current antifungal drugs. Here, we show that p38γ and p38δ regulate the innate immune response to C. albicans We describe a new TAK1-TPL2-MKK1-ERK1/2 pathway in macrophages, which is activated by Dectin-1 engagement and positively regulated by p38γ/p38δ. In mice, p38γ/p38δ deficiency protects against C. albicans infection by increasing ROS and iNOS production and thus the antifungal capacity of neutrophils and macrophages, and by decreasing the hyper-inflammation that leads to severe host damage. Leucocyte recruitment to infected kidneys and production of inflammatory mediators are decreased in p38γ/δ-null mice, reducing septic shock. p38γ/p38δ in myeloid cells are critical for this effect. Moreover, pharmacological inhibition of p38γ/p38δ in mice reduces fungal burden, revealing that these p38MAPKs may be therapeutic targets for treating C. albicans infection in humans.