Rajkumar Noubade

The induction of antibody production by IL-6 is indirectly mediated by IL-21 produced by CD4+ T cells

Interleukin (IL) 6 is a proinflammtory cytokine produced by antigen-presenting cells and nonhematopoietic cells in response to external stimuli. It was initially identified as a B cell growth factor and inducer of plasma cell differentiation in vitro and plays an important role in antibody production and class switching in vivo. However, it is not clear whether IL-6 directly affects B cells or acts through other mechanisms. We show that IL-6 is sufficient and necessary to induce IL-21 production by naive and memory CD4+ T cells upon T cell receptor stimulation. IL-21 production by CD4+ T cells is required for IL-6 to promote B cell antibody production in vitro. Moreover, administration of IL-6 with inactive influenza virus enhances virus-specific antibody production, and importantly, this effect is dependent on IL-21. Thus, IL-6 promotes antibody production by promoting the B cell helper capabilities of CD4+ T cells through increased IL-21 production. IL-6 could therefore be a potential coadjuvant to enhance humoral immunity.

A Crohn/'s disease variant in Atg16l1 enhances its degradation by caspase 3

Crohn’s disease is a debilitating inflammatory bowel disease (IBD) that can involve the entire digestive tract. A single-nucleotide polymorphism (SNP) encoding a missense variant in the autophagy gene ATG16L1 (rs2241880, Thr300Ala) is strongly associated with the incidence of Crohn’s disease. Numerous studies have demonstrated the effect of ATG16L1 deletion or deficiency; however, the molecular consequences of the Thr300Ala (T300A) variant remains unknown. Here we show that amino acids 296–299 constitute a caspase cleavage motif in ATG16L1 and that the T300A variant (T316A in mice) significantly increases ATG16L1 sensitization to caspase-3-mediated processing. We observed that death-receptor activation or starvation-induced metabolic stress in human and murine macrophages increased degradation of the T300A or T316A variants of ATG16L1, respectively, resulting in diminished autophagy. Knock-in mice harbouring the T316A variant showed defective clearance of the ileal pathogen Yersinia enterocolitica and an elevated inflammatory cytokine response. In turn, deletion of the caspase-3-encoding gene, Casp3, or elimination of the caspase cleavage site by site-directed mutagenesis rescued starvation-induced autophagy and pathogen clearance, respectively. These findings demonstrate that caspase 3 activation in the presence of a common risk allele leads to accelerated degradation of ATG16L1, placing cellular stress, apoptotic stimuli and impaired autophagy in a unified pathway that predisposes to Crohn’s disease.

Activation of p38 MAPK in CD4 T cells controls IL-17 production and autoimmune encephalomyelitis

Although several transcription factors have been shown to be critical for the induction and maintenance of IL-17 expression by CD4 Th cells, less is known about the role of nontranscriptional mechanisms. Here we show that the p38 MAPK signaling pathway is essential for in vitro and in vivo IL-17 production by regulating IL-17 synthesis in CD4 T cells through the activation of the eukaryotic translation initiation factor 4E/MAPK-interacting kinase (eIF-4E/MNK) pathway. We also show that p38 MAPK activation is required for the development and progression of both chronic and relapsing-remitting forms of experimental allergic encephalomyelitis (EAE), the principal autoimmune model of multiple sclerosis. Furthermore, we show that regulation of p38 MAPK activity specifically in T cells is sufficient to modulate EAE severity. Thus, mechanisms other than the regulation of gene expression also contribute to Th17 cell effector functions and, potentially, to the pathogenesis of other Th17 cell-mediated diseases.

NRROS negatively regulates reactive oxygen species during host defence and autoimmunity.

Reactive oxygen species (ROS) produced by phagocytes are essential for host defence against bacterial and fungal infections. Individuals with defective ROS production machinery develop chronic granulomatous disease. Conversely, excessive ROS can cause collateral tissue damage during inflammatory processes and therefore needs to be tightly regulated. Here we describe a protein, we termed negative regulator of ROS (NRROS), which limits ROS generation by phagocytes during inflammatory responses. NRROS expression in phagocytes can be repressed by inflammatory signals. NRROS-deficient phagocytes produce increased ROS upon inflammatory challenges, and mice lacking NRROS in their phagocytes show enhanced bactericidal activity against Escherichia coli and Listeria monocytogenes. Conversely, these mice develop severe experimental autoimmune encephalomyelitis owing to oxidative tissue damage in the central nervous system. Mechanistically, NRROS is localized to the endoplasmic reticulum, where it directly interacts with nascent NOX2 (also known as gp91phox and encoded by Cybb) monomer, one of the membrane-bound subunits of the NADPH oxidase complex, and facilitates the degradation of NOX2 through the endoplasmic-reticulum-associated degradation pathway. Thus, NRROS provides a hitherto undefined mechanism for regulating ROS prodution—one that enables phagocytes to produce higher amounts of ROS, if required to control invading pathogens, while minimizing unwanted collateral tissue damage.

Central histamine H3 receptor signaling negatively regulates susceptibility to autoimmune inflammatory disease of the CNS.

Histamine (HA), a biogenic amine with a broad spectrum of activities in both physiological and pathological settings, plays a key regulatory role in experimental allergic encephalomyelitis, the autoimmune model of multiple sclerosis. HA exerts its effect through four G protein-coupled receptors designated HA receptor H1, H2, H3, and H4. We report here that, compared with wild-type animals, mice with a disrupted HA H3 receptor (H3RKO), the expression of which is normally confined to cells of the nervous system, develop more severe disease and neuroinflammation. We show that this effect is associated with dysregulation of blood–brain barrier permeability and increased expression of MIP-2, IP-10, and CXCR3 by peripheral T cells. Our data suggest that pharmacological targeting of the H3R may be useful in preventing the development and formation of new lesions in multiple sclerosis, thereby significantly limiting the progression of the disease.

Histamine receptor H1 is required for TCR-mediated p38 MAPK activation and optimal IFN-γ production in mice

Histamine receptor H1 (H1R) is a susceptibility gene in both experimental autoimmune encephalomyelitis (EAE) and experimental autoimmune orchitis (EAO), 2 classical T cell-mediated models of organ-specific autoimmune disease. Here we showed that expression of H1R in naive CD4+ T cells was required for maximal IFN-gamma production but was dispensable for proliferation. Moreover, H1R signaling at the time of TCR ligation was required for activation of p38 MAPK, a known regulator of IFN-gamma expression. Importantly, selective reexpression of H1R in CD4+ T cells fully complemented both the IFN-gamma production and the EAE susceptibility of H1R-deficient mice. These data suggest that the presence of H1R in CD4+ T cells and its interaction with histamine regulates early TCR signals that lead to Th1 differentiation and autoimmune disease.

Cutting Edge: The Y Chromosome Controls the Age-Dependent Experimental Allergic Encephalomyelitis Sexual Dimorphism in SJL/J Mice

Multiple sclerosis is a sexually dimorphic, demyelinating disease of the CNS, and experimental allergic encephalomyelitis (EAE) is its principal autoimmune model. Young male SJL/J mice are relatively resistant to EAE whereas older males and SJL/J females of any age are susceptible. By comparing a wide age range of proteolipid protein peptide 139–151 immunized mice, we found that female disease severity remains constant with age. In contrast, EAE disease severity increases with age in SJL/J males, with young males having significantly less severe disease and older males having significantly more disease than equivalently aged females. To determine whether the Y chromosome contributes to this sexual dimorphism, EAE was induced in consomic SJL/J mice carrying a B10.S Y chromosome (SJL.YB10.S). EAE was significantly more severe in young male SJL.YB10.S mice compared with young male SJL/J mice. These studies show that a Y chromosome-linked polymorphism controls the age-dependent EAE sexual dimorphism observed in SJL/J mice.

Histamine H4 Receptor Optimizes T Regulatory Cell Frequency and Facilitates Anti-Inflammatory Responses within the Central Nervous System

Histamine is a biogenic amine that mediates multiple physiological processes, including immunomodulatory effects in allergic and inflammatory reactions, and also plays a key regulatory role in experimental allergic encephalomyelitis, the autoimmune model of multiple sclerosis. The pleiotropic effects of histamine are mediated by four G protein-coupled receptors, as follows: Hrh1/H1R, Hrh2/H2R, Hrh3/H3R, and Hrh4/H4R. H4R expression is primarily restricted to hematopoietic cells, and its role in autoimmune inflammatory demyelinating disease of the CNS has not been studied. In this study, we show that, compared with wild-type mice, animals with a disrupted Hrh4 (H4RKO) develop more severe myelin oligodendrocyte glycoprotein (MOG)35\x{2013}55-induced experimental allergic encephalomyelitis. Mechanistically, we also show that H4R plays a role in determining the frequency of T regulatory (TR) cells in secondary lymphoid tissues, and regulates TR cell chemotaxis and suppressor activity. Moreover, the lack of H4R leads to an impairment of an anti-inflammatory response due to fewer TR cells in the CNS during the acute phase of the disease and an increase in the proportion of Th17 cells.

Deubiquitinase DUBA is a post-translational brake on interleukin-17 production in T cells

T-helper type 17 (TH17) cells that produce the cytokines interleukin-17A (IL-17A) and IL-17F are implicated in the pathogenesis of several autoimmune diseases. The differentiation of TH17 cells is regulated by transcription factors such as RORγt, but post-translational mechanisms preventing the rampant production of pro-inflammatory IL-17A have received less attention. Here we show that the deubiquitylating enzyme DUBA is a negative regulator of IL-17A production in T cells. Mice with DUBA-deficient T cells developed exacerbated inflammation in the small intestine after challenge with anti-CD3 antibodies. DUBA interacted with the ubiquitin ligase UBR5, which suppressed DUBA abundance in naive T cells. DUBA accumulated in activated T cells and stabilized UBR5, which then ubiquitylated RORγt in response to TGF-β signalling. Our data identify DUBA as a cell-intrinsic suppressor of IL-17 production.

Pertussis toxin induces angiogenesis in brain microvascular endothelial cells.

Pertussis toxin (PTX) is an ancillary adjuvant used to elicit experimental allergic encephalomyelitis (EAE), the principal autoimmune model of multiple sclerosis. One mechanism whereby PTX potentiates EAE is to increase blood–brain barrier (BBB) permeability. To elucidate further the mechanism of action of PTX on the BBB, we investigated the genomic and proteomic responses of isolated mouse brain endothelial cells (MBEC) following intoxication. Among ∼14,000 mouse genes tracked by cDNA microarray, 34 showed altered expression in response to PTX. More than one‐third of these genes have roles in angiogenesis. Accordingly, we show that intoxication of MBEC induces tube formation in vitro and angiogenesis in vivo. The global effect of PTX on signaling protein levels and phosphorylation in MBEC was investigated by using Kinex antibody microarrays. In total, 113 of 372 pan‐specific and 58 of 258 phospho‐site‐specific antibodies revealed changes ≥25% following intoxication. Increased STAT1 Tyr‐701 and Ser‐727 phosphorylation; reduced phosphorylation of the activating phospho‐sites in Erk1, Erk2, and MAPKAPK2; and decreased phosphorylation of arrestin β1 Ser‐412 and Hsp27 Ser‐82 were confirmed by Kinetworks multi‐immunoblotting. The importance of signal transduction pathways on PTX‐induced MBEC tube formation was evaluated pharmacologically. Inhibition of phospholipase C, MEK1, and p38 MAP kinase had little effect, whereas inhibition of cAMP‐dependent protein kinase, protein kinase C, and phosphatidylinositol 3‐kinase partially blocked tube formation. Taken together, these findings are consistent with the concept that PTX may lead to increased BBB permeability by altering endothelial plasticity and angiogenesis.