Multiple lung pathogens such as chemical agents, H5N1 avian flu, or SARS cause high lethality due to acute respiratory distress syndrome. Here we report that Toll-like receptor 4 (TLR4) mutant mice display natural resistance to acid-induced acute lung injury (ALI). We show that TLR4-TRIF-TRAF6 signaling is a key disease pathway that controls the severity of ALI. The oxidized phospholipid (OxPL) OxPAPC was identified to induce lung injury and cytokine production by lung macrophages via TLR4-TRIF. We observed OxPL production in the lungs of humans and animals infected with SARS, Anthrax, or H5N1. Pulmonary challenge with an inactivated H5N1 avian influenza virus rapidly induces ALI and OxPL formation in mice. Loss of TLR4 or TRIF expression protects mice from H5N1-induced ALI. Moreover, deletion of ncf1, which controls ROS production, improves the severity of H5N1-mediated ALI. Our data identify oxidative stress and innate immunity as key lung injury pathways that control the severity of ALI.

Interleukin-6 (IL-6) is a multifunctional cytokine that regulates various aspects of the immune response, acute-phase reaction and haematopoiesis (for reviews see refs 1, 2). In vitro, leukaemia inhibitory factor, oncostatin M, ciliary neurotrophic factor and interleukin-11 display overlapping activities with IL-6. This functional redundancy may be explained by the interactions of specific binding receptors with a common signal-transducing receptor (gp130) (for reviews see refs 3, 4). To elucidate the unique function of IL-6 in vivo, we have disrupted the IL-6 gene by homologous recombination. IL-6-deficient mice develop normally. They fail to control efficiently vaccinia virus and infection with Listeria monocytogenes, a facultative intracellular bacterium. The T-cell-dependent antibody response against vesicular stomatitis virus is impaired. Further, the inflammatory acute-phase response after tissue damage or infection is severely compromised, whereas it is only moderately affected after challenge with lipopolysaccharide. We conclude that IL-6 production induced by injury or infection is an important in vivo SOS signal which coordinates activities of liver cells, macrophages and lymphocytes.

Murine T-helper clones are classified into two distinct subsets (Th1 and Th2) on the basis of their patterns of lymphokine secretion. Th1 clones secrete interleukin-2 (IL-2), tumour necrosis factor-beta (TNF-beta) and interferon-gamma (IFN-gamma), whereas Th2 clones secrete IL-4, IL-5 and IL-10 (ref. 1). These subsets are reciprocally regulated by IL-4, IL-10 and IFN-gamma and differentially promote antibody or delayed-type hypersensitivity responses. To evaluate whether IL-4 is required for mounting Th2 responses, we generated IL-4-mutant mice (IL-4-/-) and assessed the cytokine secretion pattern of T cells both from naive and Nippostrongylus brasiliensis infected mice. CD4+ T cells from naive IL-4-/- mice failed to produce Th2-derived cytokines after in vitro stimulation. The levels of Th2 cytokines IL-5, IL-9 and IL-10 from CD4+ T cells obtained after nematode infection were significantly reduced. The reduced IL-5 production in IL-4-/- mice correlated with reduced helminth-induced eosinophilia, which has been shown to be dependent on IL-5 in vivo. We conclude that IL-4 is required for the generation of the Th2-derived cytokines and that immune responses dependent on these cytokines are impaired.

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer‐reviewed by leading experts in the field, making this an essential research companion.

Tissue-resident macrophages constitute heterogeneous populations with unique functions and distinct gene-expression signatures. While it has been established that they originate mostly from embryonic progenitor cells, the signals that induce a characteristic tissue-specific differentiation program remain unknown. We found that the nuclear receptor PPAR-γ determined the perinatal differentiation and identity of alveolar macrophages (AMs). In contrast, PPAR-γ was dispensable for the development of macrophages located in the peritoneum, liver, brain, heart, kidneys, intestine and fat. Transcriptome analysis of the precursors of AMs from newborn mice showed that PPAR-γ conferred a unique signature, including several transcription factors and genes associated with the differentiation and function of AMs. Expression of PPAR-γ in fetal lung monocytes was dependent on the cytokine GM-CSF. Therefore, GM-CSF has a lung-specific role in the perinatal development of AMs through the induction of PPAR-γ in fetal monocytes.

Hepatocellular carcinoma (HCC), the fastest rising cancer in the United States and increasing in Europe, often occurs with nonalcoholic steatohepatitis (NASH). Mechanisms underlying NASH and NASH-induced HCC are largely unknown. We developed a mouse model recapitulating key features of human metabolic syndrome, NASH, and HCC by long-term feeding of a choline-deficient high-fat diet. This induced activated intrahepatic CD8(+) T cells, NKT cells, and inflammatory cytokines, similar to NASH patients. CD8(+) T cells and NKT cells but not myeloid cells promote NASH and HCC through interactions with hepatocytes. NKT cells primarily cause steatosis via secreted LIGHT, while CD8(+) and NKT cells cooperatively induce liver damage. Hepatocellular LTβR and canonical NF-κB signaling facilitate NASH-to-HCC transition, demonstrating that distinct molecular mechanisms determine NASH and HCC development.

Do endocannabinoids (eCBs) participate in long-term synaptic plasticity in the brain? Using pharmacological approaches and genetically altered mice, we show that stimulation of prelimbic cortex afferents at naturally occurring frequencies causes a longterm depression of nucleus accumbens glutamatergic synapses mediated by eCB release and presynaptic CB1 receptors. Translation of glutamate synaptic transmission into eCB retrograde signaling involved metabotropic glutamate receptors and postsynaptic intracellular Ca 2؉ stores. These findings unveil the role of the eCB system in activity-dependent long-term synaptic plasticity and identify a mechanism by which marijuana can alter synaptic functions in the endogenous brain reward system. E xogenous cannabinoids, such as the active component of Cannabis sativa L, (Ϫ)-transdelta9-tetrahydrocannabinol, as well as endocannabinoids (eCBs; anandamide and 2-arachidonoyl-glycerol) share the same target in the central nervous system: a Gi͞Go-coupled receptor named CB1 (1, 2). Activation of CB1 receptors inhibits both inhibitory and excitatory synaptic transmission in the hippocampus (3, 4), substantia nigra pars compacta (5), the cerebellum (6), the prefrontal cortex (7), and the nucleus accumbens (NAc) (8, 9). Both eCBs and CB1 have been involved in a short-lasting form of synaptic regulation: the ''depolarization-induced suppression'' of both inhibitory (10-15) and excitatory transmission (16). However, the involvement of eCB in long-lasting activity-dependent synaptic plasticity remained to be documented. The mesocorticolimbic dopaminergic system, and particularly the NAc, is essential to the reinforcing properties of addictive drugs (17, 18). Cannabinoids activate mesolimbic dopamine neurons (19) and increase NAc dopamine levels (20), similarly to other drugs of abuse. Our finding that CB1 are localized at the excitatory afferents to the NAc where exogenous cannabimimetics inhibit glutamatergic synaptic transmission (9) raised two questions: how does synaptic activity lead to the production of eCBs in the NAc, and what are the physiological correlates of eCB release on synaptic transmission? Methods Slice Preparation and Electrophysiology. Whole-cell patch-clamp and extracellular field recordings were made from medium spiny neurons in parasagittal slices of mouse NAc. These methods have been described in detail previously (9). In brief, mice (male C57BL6, 4-6 weeks) were anesthetized with isoflurane and decapitated. The brain was sliced (300-400 m) in the parasagittal plane by using a vibratome and maintained in physiological saline at 4°C. Slices containing the NAc were stored at least 1 h at room temperature before being placed in the recording chamber and superfused (2 ml͞min) with artificial cerebrospinal fluid that contained (in mM): 126 NaCl, 2.5 KCl, 1.2 MgCl 2 , 2.4 CaCl 2, 18 NaHCO 3 , 1.2 NaH 2 PO 4 , and 11 glucose, and was equilibrated with 95% O 2 ͞5% CO 2 . All experiments were done at room temperature. The superfusion medium contained picrotoxin (100 M) to...

Although inflammation is an essential component of the protective response to fungi, its dysregulation may significantly worsen fungal diseases. We found here that the IL‐23/IL‐17 developmental pathway acted as a negative regulator of the Th1‐mediated immune resistance to fungi and played an inflammatory role previously attributed to uncontrolled Th1 cell responses. Both inflammation and infection were exacerbated by a heightened Th17 response against Candida albicans and Aspergillus fumigatus, two major human fungal pathogens. IL‐23 acted as a molecular connection between uncontrolled fungal growth and inflammation, being produced by dendritic cells in response to a high fungal burden and counter‐regulating IL‐12p70 production. Both IL‐23 and IL‐17 subverted the inflammatory program of neutrophils, which resulted in severe tissue inflammatory pathology associated with infection. Our data are the first demonstrating that the IL‐23/IL‐17 pathway promotes inflammation and susceptibility in an infectious disease model. As IL‐23‐driven inflammation promotes infection and impairs antifungal resistance, modulation of the inflammatory response represents a potential strategy to stimulate protective immune responses to fungi.See accompanying commentary: http://dx.doi.org/10.1002/eji.200737804