Project description:Mast cells are known to be the key players in type I hypersensitivity reactions in humans and mice. They are critically involved in the development of allergic rhinitis, allergic asthma and systemic anaphylaxis. In this study we investigated the role of the transcriptional regulator MAZR in mast cells by comparing the expression profile of mast cells generated from wild-type (MazrF/F) and MAZR-deficient (MazrF/F x Vav-iCre) bone marrow cells. Our results from the array data demonstrate that MAZR acts preferentially as a transcriptional repressor in mast cells.
Project description:Objective:Cigarette smoke (CS) exposure reportedly enhances allergic airway inflammation. However, some studies have shown an association between current cigarette smoke exposure and a low risk for allergic rhinitis. Thus, the impact of CS exposure on allergic rhinitis remains poorly understood. The purpose of this study was to investigate the effects of CS on the respiratory mucosa (RM) and the olfactory epithelium (OE) of mice with allergic rhinitis, as the effects may differ depending on the nasal histological compartments. Methods:Eight-week-old male BALB/c mice were used for this study. We developed a mouse model of smoking by intranasally administering 10 doses of a CS solution (CSS), and a mouse model of allergic rhinitis by sensitization with intraperitoneal ovalbumin (OVA) injection and intranasal challenge with OVA. We examined the effects of CS on the nasal RM and OE in mice with or without allergic rhinitis using histological, serum, and genetic analyses. First, we examine whether CSS exposure induces allergic responses and then, examined allergic responses in the OVA-sensitized allergic rhinitis mice with or without CSS exposure. Results:Short-term CSS administration intensified allergic responses including increased infiltration of eosinophils and inflammatory cells and upregulation of interleukin-5 expression in the nasal RM of OVA-immunized mice, although only CSS induced neither allergic responses nor impairment of the RM and OE. Notably, repetitive OVA-immunization partially impaired the OE in the upper-lateral area, but CSS administration did not reinforce this impairment in OVA-induced allergic mice. Conclusion:Short-term CSS exposure strengthened allergic responses in the nasal RM and did not change the structure of the OE. These results suggest that patients with allergic rhinitis could experience exacerbation of allergic symptoms after CS exposure.
Project description:(1) Background: The atopic march is defined by the increased prevalence of allergic diseases after atopic dermatitis onset. In fact, atopic dermatitis is believed to play an important role in allergen sensitization via the damaged skin barrier, leading to allergic diseases such as allergic asthma and allergic rhinitis. The eosinophil, a pro-inflammatory cell that contributes to epithelial damage, is one of the various cells recruited in the inflammatory reactions characterizing these diseases. Few studies were conducted on the transcriptome of this cell type and even less on their specific microRNA (miRNA) profile, which could modulate pathogenesis of allergic diseases and clinical manifestations post-transcriptionally. Actually, their implication in allergic diseases is not fully understood, but they are believed to play a role in inflammation-related patterns and epithelial cell proliferation. (2) Methods: Next-generation sequencing was performed on RNA samples from eosinophils of individuals with atopic dermatitis, atopy, allergic rhinitis and asthma to obtain differential counts of primary miRNA (pri-miRNA); these were also analyzed for asthma-related phenotypes such as forced expiratory volume in one second (FEV<sub>1</sub>), immunoglobulin E (IgE) and provocative concentration of methacholine inducing a 20% fall in forced expiratory volume in 1 s (PC<sub>20</sub>) levels, as well as FEV<sub>1</sub> to forced vital capacity (FEV<sub>1</sub>/FVC) ratio. (3) Results: Eighteen miRNAs from eosinophils were identified to be significantly different between affected individuals and unaffected ones. Based on counts from these miRNAs, individuals were then clustered into groups using Ward's method on Euclidian distances. Groups were found to be explained by asthma diagnosis, familial history of respiratory diseases and allergic rhinitis as well as neutrophil counts. (4) Conclusions: The 18 differential miRNA counts for the studying phenotypes allow a better understanding of the epigenetic mechanisms underlying the development of the allergic diseases included in the atopic march.
Project description:Information regarding the prevalence of infectious agents in mice in pet shops in Japan is scarce. This information is particularly useful for minimizing the risk of potential transmission of infections to laboratory mice. Therefore, we surveyed infectious agents in mice from pet shops in Kanagawa and Tokyo, Japan. The survey was conducted in 28 mice from 5 pet shops to screen for 47 items (17 viruses, 22 bacteria and fungi, 10 parasites) using culture tests, serology, PCR, and microscopy. The most common viral agent detected was murine norovirus (17 mice; 60.7%), followed by Theiler's murine encephalomyelitis virus (13 mice; 46.4%), and mouse hepatitis virus (12 mice; 42.8%). The most common agent amongst the bacteria and fungi was Pasteurella pneumotropica (10 mice; 35.7%), followed by Helicobacter ganmani and Pneumocystis murina (8 mice; 28.5%, for both). Tritrichomonas muris was the most common parasite (19 mice; 67.8%), followed by Spironucleus muris (13 mice; 46.4%), Aspiculuris tetraptera, and Syphacia obvelata (8 mice each; 28.5%). Remarkably, a zoonotic agent, Hymenolepis nana, was found in 7 mice (25%). Given these results, we suggest that the workers in laboratory animal facilities should recognize again the potential risks of mice outside of the laboratory animal facilities as an infectious source, and avoid keeping mice as pets or as feed for carnivorous reptiles as much as possible for risk management.
Project description:MicroRNAs (miRNAs) are emerging as important regulators of allergic inflammation and potential therapeutic targets. We sought to identify which miRNAs are expressed in CD4+ T-cells and determine whether allergic stimuli or glucocorticoids alter their expression.After IRB approval, blood was collected from dust mite (DM) allergic rhinitis subjects (n=20), non-allergic controls (n=8), and asthmatics (n=16). Peripheral blood mononuclear cells were incubated with dust mite extract (DME), diluent control, or DME + dexamethasone (0.1 µM). CD4+ T-cells were collected by magnetic bead column, and RNA was isolated by guanidinium/phenol-chloroform extraction. MicroRNA expression was measured using Nanostring microarray and quantitative real time PCR (qPCR).We identified 196 miRNAs that were stably expressed in circulating CD4+ T-cells. Allergen stimulation of CD4+ T-cells with DME differentially induced miR-155 expression in cells of DM-allergic subjects as compared to non-allergic subjects. Induction of miR-155 expression was also observed with anti-CD3/anti-CD28 simulation and phorbol-12-Myristate-13-Acetate (PMA) treatment, and further augmented by calcium inophore and bromocyclic AMP in the latter treatment. The level of miR-155 expression was positively associated with expression of the TH2 cytokines IL-5 and IL-13. Inhibition of miR-155 in Jurkat T-cells inhibited the production of these cytokines. Glucocorticoids attenuated the effects of dust mite allergen, raising the possibility that inhibition of this miRNA could be a mechanism through which glucocorticoids exhibit their anti-inflammatory effects. The CD4+ T-cells had a higher level of miR-155 expression in asthma compared to in allergic rhinitis and non-asthmatics. The inhibitory effects of glucocorticoids on CD4+ T-cell miR-155 expression were lost in severe asthmatics.Mir-155 is differentially expressed in allergic T-cells exposed to DM extract compared to in non-allergic cells and it is inhibited by glucocorticoids. MiR-155 may play a role in mediating allergic inflammation in T-cells and could be an anti-inflammatory target of steroids. This pathway may be de-regulated in severe asthma.
Project description:INTRODUCTION:MicroRNAs (miRNAs) contribute to the regulation of dendritic cell (DC) polarization, thereby influencing the balance of adaptive immune responses. Herein, we studied the expression of miRNAs in polarized DCs and analyzed whether expression of these miRNAs could be associated with allergic rhinitis and allergen immunotherapy (AIT) outcome. METHOD:Using specific culture conditions, we differentiated immature human monocyte-derived DCs into DC1, DC2, and DCreg subsets (supporting the differentiation of TH 1, TH 2 or regulatory T cells, respectively). Profiling of miRNA expression was performed in these DC subpopulations using microarrays. Levels of miRNAs specific for polarized DCs were then evaluated in a cohort of 58 patients with allergic rhinitis and 25 non-allergic controls, as well as in samples from 30 subjects treated with sublingual grass pollen tablets or placebo for four months. RESULTS:We successfully identified 16 miRNAs differentially regulated between immature DCs, DC1, DC2, and DCreg cells. In allergic rhinoconjunctivitis patients, the expression of two of those miRNAs (miR-132 and miR-155), was down-regulated compared to non-allergic individuals. However, the levels of these miRNAs were not significantly modified following four months of grass pollen immunotherapy. CONCLUSIONS:Studying polarized DCs and clinical samples from subjects with or without allergic rhinoconjunctivitis, we demonstrated that the expression of two miRNAs linked to effector DCs (i.e., DC1 and/or DC2 cells), was reduced in the blood of patients with allergic rhinoconjunctivitis. Nevertheless, these miRNAs did not represent relevant biomarkers to predict or follow-up AIT efficacy.
Project description:Liver fibrosis is a major pathological feature of chronic liver diseases, including liver cancer. MicroRNAs (miRNAs), small noncoding RNAs, regulate gene expression posttranscriptionally and play important roles in various kinds of diseases; however, miRNA-associated hepatic fibrogenesis and its acting mechanisms are poorly investigated. Therefore, we performed an miRNA microarray in the fibrotic livers of Mus musculus treated with carbon-tetrachloride (CCl₄) and analyzed the biological functions engaged by the target genes of differentially-expressed miRNAs through gene ontology (GO) and in-depth pathway enrichment analysis. Herein, we found that four miRNAs were upregulated and four miRNAs were downregulated more than two-fold in CCl₄-treated livers compared to a control liver. Eight miRNAs were predicted to target a total of 4079 genes. GO analysis revealed that those target genes were located in various cellular compartments, including cytoplasm, nucleolus and cell surface, and they were involved in protein-protein or protein-DNA bindings, which influence the signal transductions and gene transcription. Furthermore, pathway enrichment analysis demonstrated that the 72 subspecialized signaling pathways were associated with CCl₄-induced liver fibrosis and were mostly classified into metabolic function-related pathways. These results suggest that CCl₄ induces liver fibrosis by disrupting the metabolic pathways. In conclusion, we presented several miRNAs and their biological processes that might be important in the progression of liver fibrosis; these findings help increase the understanding of liver fibrogenesis and provide novel ideas for further studies of the role of miRNAs in liver fibrosis.
Project description:BACKGROUND:Identification of protein-protein interactions is an important first step to understand living systems. High-throughput experimental approaches have accumulated large amount of information on protein-protein interactions in human and other model organisms. Such interaction information has been successfully transferred to other species, in which the experimental data are limited. However, the annotation transfer method could yield false positive interologs due to the lack of conservation of interactions when applied to phylogenetically distant organisms. RESULTS:To address this issue, we used phylogenetic profile method to filter false positives in interologs based on the notion that evolutionary conserved interactions show similar patterns of occurrence along the genomes. The approach was applied to Mus musculus, in which the experimentally identified interactions are limited. We first inferred the protein-protein interactions in Mus musculus by using two approaches: i) identifying mouse orthologs of interacting proteins (interologs) based on the experimental protein-protein interaction data from other organisms; and ii) analyzing frequency of mouse ortholog co-occurrence in predicted operons of bacteria. We then filtered possible false-positives in the predicted interactions using the phylogenetic profiles. We found that this filtering method significantly increased the frequency of interacting protein-pairs coexpressed in the same cells/tissues in gene expression omnibus (GEO) database as well as the frequency of interacting protein-pairs shared the similar Gene Ontology (GO) terms for biological processes and cellular localizations. The data supports the notion that phylogenetic profile helps to reduce the number of false positives in interologs. CONCLUSION:We have developed protein-protein interaction database in mouse, which contains 41109 interologs. We have also developed a web interface to facilitate the use of database http://lgsun.grc.nia.nih.gov/mppi/.
Project description:Rodent betaherpesviruses vary considerably in genomic content, and these variations can result in a distinct pathogenicity. Therefore, the identification of unknown betaherpesviruses in house mice (Mus musculus), the most important rodent host species in basic research, is of importance. During a search for novel herpesviruses in house mice using herpesvirus consensus PCR and attempts to isolate viruses in tissue culture, we identified a previously unknown betaherpesvirus. The primary PCR search in mouse organs revealed the presence of known strains of murine cytomegalovirus (Murid herpesvirus 1) and of Mus musculus rhadinovirus 1 only. However, the novel virus was detected after incubation of organ pieces in fibroblast tissue culture and subsequent PCR analysis of the supernatants. Long-distance PCR amplification including the DNA polymerase and glycoprotein B genes revealed a 3.4 kb sequence that was similar to sequences of rodent cytomegaloviruses. Pairwise sequence comparisons and phylogenetic analyses showed that this newly identified murine virus is most similar to the English isolate of rat cytomegalovirus, thereby raising the possibility that two distinct CMV lineages have evolved in both Mus musculus and Rattus norvegicus.