Severe delayed hypersensitivity reactions to IL-1 and IL-6 inhibitors link to common HLA-DRB1*15 alleles.

OBJECTIVES: Drug reaction with eosinophilia and systemic symptoms (DRESS) is a severe, delayed hypersensitivity reaction (DHR). We observed DRESS to inhibitors of interleukin 1 (IL-1) or IL-6 in a small group of patients with Still's disease with atypical lung disease. We sought to characterise features of patients with Still's disease with DRESS compared with drug-tolerant Still's controls. We analysed human leucocyte antigen (HLA) alleles for association to inhibitor-related DHR, including in a small Kawasaki disease (KD) cohort. METHODS: In a case/control study, we collected a multicentre series of patients with Still's disease with features of inhibitor-related DRESS (n=66) and drug-tolerant Still's controls (n=65). We retrospectively analysed clinical data from all Still's subjects and typed 94/131 for HLA. European Still's-DRESS cases were ancestry matched to International Childhood Arthritis Genetics Consortium paediatric Still's cases (n=550) and compared for HLA allele frequencies. HLA association also was analysed using Still's-DRESS cases (n=64) compared with drug-tolerant Still's controls (n=30). KD subjects (n=19) were similarly studied. RESULTS: Still's-DRESS features included eosinophilia (89%), AST-ALT elevation (75%) and non-evanescent rash (95%; 88% involving face). Macrophage activation syndrome during treatment was frequent in Still's-DRESS (64%) versus drug-tolerant Still's (3%; p=1.2×10-14). We found striking enrichment for HLA-DRB1*15 haplotypes in Still's-DRESS cases versus INCHARGE Still's controls (p=7.5×10-13) and versus self-identified, ancestry-matched Still's controls (p=6.3×10-10). In the KD cohort, DRB1*15:01 was present only in those with suspected anakinra reactions. CONCLUSIONS: DRESS-type reactions occur among patients treated with IL-1/IL-6 inhibitors and strongly associate with common HLA-DRB1*15 haplotypes. Consideration of preprescription HLA typing and vigilance for serious reactions to these drugs are warranted.

miR-155 Contributes to the Immunoregulatory Function of Human Mesenchymal Stem Cells.

Objectives: Mesenchymal stem/stromal cells (MSCs) are widely investigated in regenerative medicine thanks to their immunomodulatory properties. They exert their anti-inflammatory function thanks to the secretion of a number of mediators, including proteins and miRNAs, which can be released in the extracellular environment or in the cargo of extracellular vesicles (EVs). However, the role of miRNAs in the suppressive function of MSCs is controversial. The aim of the study was to identify miRNAs that contribute to the immunomodulatory function of human bone marrow-derived MSCs (BM-MSCs). Methods: Human BM-MSCs were primed by coculture with activated peripheral blood mononuclear cells (aPBMCs). High throughput miRNA transcriptomic analysis was performed using Human MicroRNA TaqMan® Array Cards. The immunosuppressive function of miRNAs was investigated in mixed lymphocyte reactions and the delayed type hypersensitivity (DTH) murine model. Results: Upon priming, 21 out of 377 tested miRNAs were significantly modulated in primed MSCs. We validated the up-regulation of miR-29a, miR-146a, miR-155 and the down-regulation of miR-149, miR-221 and miR-361 in additional samples of primed MSCs. We showed that miR-155 significantly reduced the proliferation of aPBMCs in vitro and inflammation in vivo, using the DTH model. Analysis of miRNA-mRNA interactions revealed miR-221 as a potential target gene that is down-regulated by miR-155 both in primed MSCs and in aPBMCs. Conclusion: Here, we present evidence that miR-155 participates to the immunosuppressive function of human BM-MSCs and down-regulates the expression of miR-221 as a possible inflammatory mediator.

Vps34/PI3KC3 deletion in kidney proximal tubules impairs apical trafficking and blocks autophagic flux, causing a Fanconi-like syndrome and renal insufficiency.

Kidney proximal tubular cells (PTCs) are highly specialized for ultrafiltrate reabsorption and serve as paradigm of apical epithelial differentiation. Vps34/PI3-kinase type III (PI3KC3) regulates endosomal dynamics, macroautophagy and lysosomal function. However, its in vivo role in PTCs has not been evaluated. Conditional deletion of Vps34/PI3KC3 in PTCs by Pax8-Cre resulted in early (P7) PTC dysfunction, manifested by Fanconi-like syndrome, followed by kidney failure (P14) and death. By confocal microscopy, Vps34∆/∆ PTCs showed preserved apico-basal specification (brush border, NHERF-1 versus Na+/K+-ATPase, ankyrin-G) but basal redistribution of late-endosomes/lysosomes (LAMP-1) and mis-localization to lysosomes of apical recycling endocytic receptors (megalin, cubilin) and apical non-recycling solute carriers (NaPi-IIa, SGLT-2). Defective endocytosis was confirmed by Texas-red-ovalbumin tracing and reduced albumin content. Disruption of Rab-11 and perinuclear galectin-3 compartments suggested mechanistic clues for defective receptor recycling and apical biosynthetic trafficking. p62-dependent autophagy was triggered yet abortive (p62 co-localization with LC3 but not LAMP-1) and PTCs became vacuolated. Impaired lysosomal positioning and blocked autophagy are known causes of cell stress. Thus, early trafficking defects show that Vps34 is a key in vivo component of molecular machineries governing apical vesicular trafficking, thus absorptive function in PTCs. Functional defects underline the essential role of Vps34 for PTC homeostasis and kidney survival.

Unique microRNAs appear at different times during the course of a delayed-type hypersensitivity reaction in human skin.

Diphencyprone (DPCP) is a hapten that induces delayed-type hypersensitivity (DTH) reactions. MicroRNAs (miRNAs) are short non-coding RNAs that negatively regulate gene expression and have been implicated in various inflammatory skin diseases, but their role in DTH reactions is not well understood. We generated global miRNA expression profiles (using next-generation sequencing) of DPCP reactions in skin of seven healthy volunteers at 3, 14 and 120 days after challenge. Compared to placebo-treated sites, DPCP-challenged skin at 3 days (peak inflammation) had 127 miRNAs significantly deregulated. At 14 days (during resolution of inflammation), 43 miRNAs were deregulated and, at 120 days (when inflammation had completely resolved), six miRNAs were upregulated. While some miRNAs have been observed in psoriasis or atopic dermatitis, most of the deregulated miRNAs have not yet been studied in the context of skin biology or immunology. Across the three time points studied, many but not all miRNAs were uniquely expressed. As various miRNAs may influence T cell activation, this may indicate that the miRNAs exclusively expressed at different time points function to promote or resolve skin inflammation, and therefore, may inform on the paradoxical ability of DPCP to treat both autoimmune conditions (alopecia areata) and conditions of ineffective immunity (melanoma).

Defective lymphoid organogenesis underlies the immune deficiency caused by a heterozygous S32I mutation in IκBα.

Patients with ectodermal dysplasia with immunodeficiency (ED-ID) caused by mutations in the inhibitor of NF-κB α (IκBα) are susceptible to severe recurrent infections, despite normal T and B cell numbers and intact in vitro lymphocyte function. Moreover, the outcome of hematopoietic stem cell transplantation (HSCT) in these patients is poor despite good engraftment. Mice heterozygous for the IκBα S32I mutation found in patients exhibited typical features of ED-ID. Strikingly, the mice lacked lymph nodes, Peyer's patches, splenic marginal zones, and follicular dendritic cells and failed to develop contact hypersensitivity (CHS) or form germinal centers (GCs), all features not previously recognized in patients and typical of defective noncanonical NF-κB signaling. Lymphotoxin ß receptor (LTßR)-driven induction of chemokines and adhesion molecules mediated by both canonical and noncanonical NF-κB pathways was impaired, and levels of p100 were markedly diminished in the mutant. IκBα mutant → Rag2(-/-), but not WT→IκBα mutant, bone marrow chimeras formed proper lymphoid organs and developed CHS and GCs. Defective architectural cell function explains the immunodeficiency and poor outcome of HSCT in patients with IκBα deficiency and suggests that correction of this niche is critical for reconstituting their immune function.

Tuberculin skin test negativity is under tight genetic control of chromosomal region 11p14-15 in settings with different tuberculosis endemicities.

A substantial proportion of subjects exposed to a contagious tuberculosis case display lack of tuberculin skin test (TST) reactivity. We previously mapped a major locus (TST1) controlling lack of TST reactivity in families from an area in South Africa where tuberculosis is hyperendemic. Here, we conducted a household tuberculosis contact study in a French area where the endemicity of tuberculosis is low. A genome-wide analysis of TST negativity identified a significant linkage signal (P < 3 × 10(-5)) in close vicinity of TST1. Combined analysis of the 2 samples increased evidence of linkage (P = 2.4 × 10(-6)), further implicating genetic factors located on 11p14-15. This region overlaps the TNF1 locus controlling mycobacteria-driven tumor necrosis factor α production.

Granzyme B releases vascular endothelial growth factor from extracellular matrix and induces vascular permeability.

The formation of unstable, leaky neovessels underlies the pathogenesis of many chronic inflammatory diseases. Granzyme B (GZMB) is an immune-derived serine protease that accumulates in the extracellular matrix (ECM) during chronic inflammation and is capable of cleaving fibronectin (FN). Vascular endothelial growth factor (VEGF) is a potent vascular permeabilizing agent that is sequestered in the ECM through its interaction with FN. As GZMB levels are elevated in chronic inflammatory diseases that are associated with increased vascular permeability, the role of GZMB in the regulation of VEGF bioavailability and vascular permeability were assessed. GZMB was added to either VEGF bound to FN or VEGF bound to endothelial cell (EC)-derived ECM. Supernatants containing released VEGF were assessed to determine VEGF activity by treating EC and evaluating VEGF receptor-2 (VEGFR2) phosphorylation. GZMB released VEGF from both FN and from EC-derived matrix, whereas GZMB inhibition prevented FN cleavage and VEGF release. GZMB-mediated VEGF release resulted in significant phosphorylation of VEGFR2. The role of GZMB-mediated VEGF release in altering vascular permeability was also assessed in vivo using Miles/Evans blue permeability assay. GZMB induced a significant VEGF-dependent increase in vascular permeability in vivo that was reduced in the presence of an anti-VEGF-neutralizing antibody. Inflammatory-mediated vascular leakage was also assessed in GZMB-KO mice using a delayed-type hypersensitivity model. GZMB-KO mice exhibited reduced microvascular leakage compared with C57\B6 controls. GZMB increases vascular permeability in part through the proteolytic release of ECM-sequestered VEGF, leading to VEGFR2 activation and increased vascular permeability in vivo. These findings present a novel role for GZMB as a modulator of vascular response during chronic inflammation.

Characterization of T cell receptors of Th1 cells infiltrating inflamed skin of a novel murine model of palladium-induced metal allergy.

Metal allergy is categorized as a delayed-type hypersensitivity reaction, and is characterized by the recruitment of lymphocytes into sites of allergic inflammation. Because of the unavailability of suitable animal models for metal allergy, the role of T cells in the pathogenesis of metal allergy has not been explored. Thus, we developed a novel mouse model for metal allergy associated with infiltration of T cells by multiple injections of palladium (Pd) plus lipopolysaccharide into the footpad. Using this model, we characterized footpad-infiltrating T cells in terms of phenotypic markers, T cell receptor (TCR) repertoires and cytokine expression. CD3+ CD4+ T cells accumulated in the allergic footpads 7 days after Pd challenge. The expression levels of CD25, interleukin-2, interferon-γ and tumor necrosis factor, but not interleukin-4 and interleukin-5, increased in the footpads after challenge, suggesting CD4+ T helper 1 (Th1) cells locally expanded in response to Pd. Infiltrated T cells in the footpads frequently expressed AV18-1 and BV8-2 T cell receptor (TCR) chains compared with T cells in the lymph nodes and exhibited oligoclonality. T-cell clones identified from Pd-allergic mouse footpads shared identical CDR3 sequences containing AV18-1 and BV8-2. These results suggest that TCR AV18-1 and BV8-2 play dominant and critical parts in the antigen specificity of Pd-specific Th1 cells.

PI3Kγ kinase activity is required for optimal T-cell activation and differentiation.

Phosphatidylinositol-3-kinase gamma (PI3Kγ) is a leukocyte-specific lipid kinase with signaling function downstream of G protein-coupled receptors to regulate cell trafficking, but its role in T cells remains unclear. To investigate the requirement of PI3Kγ kinase activity in T-cell function, we studied T cells from PI3Kγ kinase-dead knock-in (PI3Kγ(KD/KD)) mice expressing the kinase-inactive PI3Kγ protein. We show that CD4(+) and CD8(+) T cells from PI3Kγ(KD/KD) mice exhibit impaired TCR/CD28-mediated activation that could not be rescued by exogenous IL-2. The defects in proliferation and cytokine production were also evident in naïve and memory T cells. Analysis of signaling events in activated PI3Kγ(KD/KD) T cells revealed a reduction in phosphorylation of protein kinase B (AKT) and ERK1/2, a decrease in lipid raft formation, and a delay in cell cycle progression. Furthermore, PI3Kγ(KD/KD) CD4(+) T cells displayed compromised differentiation toward Th1, Th2, Th17, and induced Treg cells. PI3Kγ(KD/KD) mice also exhibited an impaired response to immunization and a reduced delayed-type hypersensitivity to Ag challenge. These findings indicate that PI3Kγ kinase activity is required for optimal T-cell activation and differentiation, as well as for mounting an efficient T cell-mediated immune response. The results suggest that PI3Kγ kinase inhibitors could be beneficial in reducing the undesirable immune response in autoimmune diseases.

Accumulation of invariant NKT cells into inflamed skin in a novel murine model of nickel allergy.

Nickel (Ni) can cause delayed-type hypersensitivity reactions, which are thought to be mediated by the accumulation of T cells into inflamed skin. Accumulated T cells at the developmental stages in metal allergy are poorly characterized because a suitable animal model has not been established. To investigate the accumulated T cells in allergic inflamed skin, we generated a novel murine model of Ni-induced allergy. The murine model of Ni allergy was induced by two sensitizations of Ni plus lipopolysaccharide solution into the groin followed by three challenges with Ni solution into the footpad. Here we show that a specific TCR repertoire bearing Vα14Jα18, called natural killer (NK) T cells, was expanded monoclonally in BALB/c or C57BL/6 mice. Accumulation of NKT cells was characterized as CD4(+) or CD4(-)CD8(-) T cells. These results suggested that NKT cells are major pathogenic T cells at the elicitation phase of Ni allergy.

Decorin potentiates interferon-γ activity in a model of allergic inflammation.

The proteoglycan decorin modulates leukocyte recruitment during delayed-type hypersensitivity responses. Decorin-deficient (Dcn(-/-)) mice show reduced edema formation during the first 24 h with a concurrent attenuated recruitment of CD8(+) leukocytes in the inflamed Dcn(-/-) ears. The aim of this study was to elucidate the molecular pathways affected by the loss of decorin. In vivo, reduced numbers of CD8(+) cells in Dcn(-/-) ears correlated with a reduced interferon-γ (Ifn-γ) and CXCL-10 expression. In vitro, Dcn(-/-) lymphocytes displayed an increased adhesion to brain microvascular (bEnd.3) endothelial cells. Decorin treatment of bEnd.3 increased Icam1 and down-regulated Vcam1 expression after TNF-α stimulation. However, Dcn(-/-) and wild-type lymphocytes produced IFN-γ after activation with CD3ε. Upon incubation with decorin, endothelial cells and fibroblasts responded differently to IFN-γ and TNF-α; CCL2 in bEnd.3 cells was more prominently up-regulated by TNF-α compared with IFN-γ. Notably, both factors were more potent in the presence of decorin. Compared with TNF-α, IFN-γ treatment induced significantly more CXCL-10, and both factors increased synthesis of CXCL-10 in the presence of decorin. The response to IFN-γ was similar in Dcn(-/-) and wild-type fibroblasts, an additional source of CXCL-10. However, addition of decorin yielded significantly more CXCL-10. Notably, decorin increased the stability of IFN-γ in vitro and potentiated IFN-γ-induced activation of STAT-1. Furthermore, only dermatan sulfate influenced IFN-γ signaling by significantly increasing CXCL-10 expression in contrast to decorin protein core alone. Our data demonstrate that decorin modulates delayed-type hypersensitivity responses by augmenting the induction of downstream effector cytokines of IFN-γ and TNF-α, thereby influencing the recruitment of CD8(+) lymphocytes into the inflamed tissue.

Divergent effects of endogenous and exogenous glucocorticoid-induced leucine zipper in animal models of inflammation and arthritis.

OBJECTIVE: Glucocorticoid-induced leucine zipper (GILZ) has effects on inflammatory pathways that suggest it to be a key inhibitory regulator of the immune system, and its expression is exquisitely sensitive to induction by glucocorticoids. We undertook this study to test our hypothesis that GILZ deficiency would exacerbate experimental immune-mediated inflammation and impair the effects of glucocorticoids on inflammation and, correspondingly, that exogenous GILZ would inhibit these events. METHODS: GILZ(-/-) mice were generated using the Cre/loxP system, and responses were studied in delayed-type hypersensitivity (DTH), antigen-induced arthritis (AIA), K/BxN serum-transfer arthritis, and lipopolysaccharide (LPS)-induced cytokinemia. Therapeutic expression of GILZ via administration of recombinant adeno-associated virus expressing the GILZ gene (GILZ-rAAV) was compared to the effects of glucocorticoid in collagen-induced arthritis (CIA). RESULTS: Increased T cell proliferation and DTH were observed in GILZ(-/-) mice, but neither AIA nor K/BxN serum-transfer arthritis was affected, and GILZ deficiency did not affect LPS-induced cytokinemia. Deletion of GILZ did not impair the effects of exogenous glucocorticoids on CIA or cytokinemia. In contrast, overexpression of GILZ in joints significantly inhibited CIA, with an effect similar to that of dexamethasone. CONCLUSION: Despite effects on T cell activation, GILZ deficiency had no effect on effector pathways of arthritis and was unexpectedly redundant with effects of glucocorticoids. These findings do not support the hypothesis that GILZ is central to the actions of glucocorticoids, but the efficacy of exogenous GILZ in CIA suggests that further evaluation of GILZ in inflammatory disease is required.

Mesenchymal stem cells: a double-edged sword in regulating immune responses.

Mesenchymal stem cells (MSCs) have been employed successfully to treat various immune disorders in animal models and clinical settings. Our previous studies have shown that MSCs can become highly immunosuppressive upon stimulation by inflammatory cytokines, an effect exerted through the concerted action of chemokines and nitric oxide (NO). Here, we show that MSCs can also enhance immune responses. This immune-promoting effect occurred when proinflammatory cytokines were inadequate to elicit sufficient NO production. When inducible nitric oxide synthase (iNOS) production was inhibited or genetically ablated, MSCs strongly enhance T-cell proliferation in vitro and the delayed-type hypersensitivity response in vivo. Furthermore, iNOS(-/-) MSCs significantly inhibited melanoma growth. It is likely that in the absence of NO, chemokines act to promote immune responses. Indeed, in CCR5(-/-)CXCR3(-/-) mice, the immune-promoting effect of iNOS(-/-) MSCs is greatly diminished. Thus, NO acts as a switch in MSC-mediated immunomodulation. More importantly, the dual effect on immune reactions was also observed in human MSCs, in which indoleamine 2,3-dioxygenase (IDO) acts as a switch. This study provides novel information about the pathophysiological roles of MSCs.

Dissecting the causes of atopic dermatitis in children: less foods, more mites.

Atopic dermatitis (AD) is a common, chronic or chronically relapsing, multifactorial skin disease that mainly occurs in children but affects also adults. AD usually begins early in life and often concerns people with a personal or family history of asthma and allergic rhinitis. AD is characterized by eczematous changes in the epidermis and originates from a late, T-cell mediated reaction associated to the formation and production of memory T-cell of TH2 type, occurrence of homing receptor at skin level and cutaneous lymphocyte-associated (CLA) antigens. Extrinsic or allergic AD, but not intrinsic AD, shows high total serum IgE levels and the presence of specific IgE for environmental and food allergens. A pivotal role in the pathogenesis of AD is played by filaggrin, a protein contained in the granular layer of the epidermis regulating the aggregation of keratin filaments. Mutation in the filaggrin gene causes decreased barrier function of the corny layers of the epidermis. This favours the enter through the skin of environmental allergens, especially the house dust mite, that further facilitates such entering by the proteolytic activity of its major allergen Der p 1. In fact, recent advances suggest that the dust mite, more than foods, is the major cause of allergic AD. As far as the causal diagnosis of AD is concerned, there is notable evidence supporting the capacity of the atopy patch test (APT) to reproduce the pathophysiologic events of AD. This makes APT a valuable diagnostic tool for AD.

Cystatin C influences the autoimmune but not inflammatory response to cartilage type II collagen leading to chronic arthritis development.

INTRODUCTION: Collagen-induced arthritis (CIA) is a mouse model for rheumatoid arthritis (RA) and is induced after immunization with type II collagen (CII). CIA, like RA, is an autoimmune disease leading to destruction of cartilage and joints, and both the priming and inflammatory phases have been suggested to be dependent on proteases. In particular, the cysteine proteases have been proposed to be detrimental to the arthritic process and even immunomodulatory. A natural inhibitor of cysteine proteases is cystatin C. METHODS: Cystatin C-deficient, sufficient and heterozygous mice were tested for onset, incidence and severity of CIA. The effect of cystatin C-deficiency was further dissected by testing the inflammatory effector phase of CIA; that is, collagen antibody-induced arthritis model and priming phase, that is, T cell response both in vivo and in vitro. In addition, in order to determine the importance of T cells and antigen-presenting cells (APCs), these cell populations were separated and in vitro T cell responses determined in a mixed co-culture system. Finally, flow cytometry was used in order to further characterize cell populations in cystatin C-deficient mice. RESULTS: Here, we show that mice lacking cystatin C, develop arthritis at a higher incidence and an earlier onset than wild-type controls. Interestingly, when the inflammatory phase of CIA was examined independently from immune priming then cystatin C-deficiency did not enhance the arthritis profile. However, in line with the enhanced CIA, there was an increased T cell and B cell response as delayed-type hypersensitivity reaction and anti-CII antibody titers were elevated in the cystatin C-deficient mice after immunization. In addition, the ex vivo naïve APCs from cystatin C-deficient mice had a greater capacity to stimulate T cells. Interestingly, dendritic cells had a more activated phenotype in naïve cystatin C-deficient mice. CONCLUSIONS: The lack of cystatin C enhances CIA and primarily affects in vivo priming of the immune system. Although the mechanism of this is still unknown, we show evidence for a more activated APC compartment, which would elevate the autoimmune response towards CII, thus resulting in an enhanced development of chronic arthritis.

Endogenous collagen peptide activation of CD1d-restricted NKT cells ameliorates tissue-specific inflammation in mice.

NKT cells in the mouse recognize antigen in the context of the MHC class I-like molecule CD1d and play an important role in peripheral tolerance and protection against autoimmune and other diseases. NKT cells are usually activated by CD1d-presented lipid antigens. However, peptide recognition in the context of CD1 has also been documented, although no self-peptide ligands have been reported to date. Here, we have identified an endogenous peptide that is presented by CD1d to activate mouse NKT cells. This peptide, the immunodominant epitope from mouse collagen type II (mCII707-721), was not associated with either MHC class I or II. Activation of CD1d-restricted mCII707-721-specific NKT cells was induced via TCR signaling and classical costimulation. In addition, mCII707-721-specific NKT cells induced T cell death through Fas/FasL, in an IL-17A-independent fashion. Moreover, mCII707-721-specific NKT cells suppressed a range of in vivo inflammatory conditions, including delayed-type hypersensitivity, antigen-induced airway inflammation, collagen-induced arthritis, and EAE, which were all ameliorated by mCII707-721 vaccination. The findings presented here offer new insight into the intrinsic roles of NKT cells in health and disease. Given the results, endogenous collagen peptide activators of NKT cells may offer promise as novel therapeutics in tissue-specific autoimmune and inflammatory diseases.

Altered immune response in mice deficient for the G protein-coupled receptor GPR34.

The X-chromosomal GPR34 gene encodes an orphan G(i) protein-coupled receptor that is highly conserved among vertebrates. To evaluate the physiological relevance of GPR34, we generated a GPR34-deficient mouse line. GPR34-deficient mice were vital, reproduced normally, and showed no gross abnormalities in anatomical, histological, laboratory chemistry, or behavioral investigations under standard housing. Because GPR34 is highly expressed in mononuclear cells of the immune system, mice were specifically tested for altered functions of these cell types. Following immunization with methylated BSA, the number of granulocytes and macrophages in spleens was significantly lower in GPR34-deficient mice as in wild-type mice. GPR34-deficient mice showed significantly increased paw swelling in the delayed type hypersensitivity test and higher pathogen burden in extrapulmonary tissues after pulmonary infection with Cryptococcus neoformans compared with wild-type mice. The findings in delayed type hypersensitivity and infection tests were accompanied by significantly different basal and stimulated TNF-α, GM-CSF, and IFN-γ levels in GPR34-deficient animals. Our data point toward a functional role of GPR34 in the cellular response to immunological challenges.

[Pharmacogenetics]. / Pharmakogenetik.

Allergic reactions to drugs belong to the group of B-reactions, which are characterized not only by the pharmacologic profile of the drug but also by individual risk factors. Pharmacogenetic approaches facilitate identifying these risk factors. The most important example for Caucasian patients is NIDR (non-immediate drug reaction) to abacavir. Chinese patients possess risk factors for NIDR to allopurinol and anticonvulsants and the HLA locus on chromosome 6. Another potentially important area is assessing risk factors to prevent allergic reactions to antibiotics, especially with regard to the effective treatment of MRSA.

The adaptor molecule CARD9 is essential for tuberculosis control.

The cross talk between host and pathogen starts with recognition of bacterial signatures through pattern recognition receptors (PRRs), which mobilize downstream signaling cascades. We investigated the role of the cytosolic adaptor caspase recruitment domain family, member 9 (CARD9) in tuberculosis. This adaptor was critical for full activation of innate immunity by converging signals downstream of multiple PRRs. Card9(-/-) mice succumbed early after aerosol infection, with higher mycobacterial burden, pyogranulomatous pneumonia, accelerated granulocyte recruitment, and higher abundance of proinflammatory cytokines and granulocyte colony-stimulating factor (G-CSF) in serum and lung. Neutralization of G-CSF and neutrophil depletion significantly prolonged survival, indicating that an exacerbated systemic inflammatory disease triggered lethality of Card9(-/-) mice. CARD9 deficiency had no apparent effect on T cell responses, but a marked impact on the hematopoietic compartment. Card9(-/-) granulocytes failed to produce IL-10 after Mycobacterium tuberculosis infection, suggesting that an absent antiinflammatory feedback loop accounted for granulocyte-dominated pathology, uncontrolled bacterial replication, and, ultimately, death of infected Card9(-/-) mice. Our data provide evidence that deregulated innate responses trigger excessive lung inflammation and demonstrate a pivotal role of CARD9 signaling in autonomous innate host defense against tuberculosis.

Ran overexpression leads to diminished T cell responses and selectively modulates nuclear levels of c-Jun and c-Fos.

Ras-related nuclear protein (Ran) is a Ras family GTPase, and its documented functions are the regulation of DNA replication, cell cycle progression, nuclear structure formation, RNA processing and exportation, and nuclear protein importation. In this study, we performed detailed mapping of Ran expression during mouse ontogeny using in situ hybridization. High Ran expression was found in various organs and tissues including the thymus cortex and spleen white pulp. Ran was induced in T cells 24 h after their activation. The function of Ran in the immune system was investigated using Ran transgenic (Tg) mice. In Ran Tg T cells, there was compromised activation marker expression, lymphokine secretion, and proliferation upon T cell receptor activation in vitro when compared with wild type T cells. Tg mice also manifested defective delayed type hypersensitivity in vivo. Upon PMA and ionomycin stimulation, Tg T cells were defective in nuclear accumulation of AP-1 factors (c-Jun and c-Fos) but not NF-kappaB family members. Our experiments showed that Ran had important regulatory function in T cell activation. One of the possible mechanisms is that intracellular Ran protein levels control the nuclear retention for selective transcription factors such as c-Jun and c-Fos of AP-1, which is known to be critical in T cell activation and proliferation and lymphokine secretion.