Shunji Hasegawa

Clarithromycin Inhibits NF-κB Activation in Human Peripheral Blood Mononuclear Cells and Pulmonary Epithelial Cells

ABSTRACT Macrolide antibiotics modulate the production of proinflammatory cytokines in vivo and in vitro. Transcription of the genes for these proinflammatory cytokines is regulated by nuclear factor κB (NF-κB). We examined whether or not clarithromycin inhibits the activation of NF-κB induced by tumor necrosis factor alpha (TNF-α) or staphylococcal enterotoxin A (SEA) in human monocytic U-937 cells, a T-cell line (Jurkat), a pulmonary epithelial cell line (A549), and peripheral blood mononuclear cells (PBMC). Flow cytometry revealed that clarithromycin suppresses NF-κB activation induced by TNF-α in U-937 and Jurkat cells in a concentration-related manner. Western blot analysis also demonstrated that clarithromycin inhibits NF-κB activation induced by TNF-α in U-937, Jurkat, and A549 cells and PBMC and by SEA in PBMC. Western blot analysis of cytoplasmic extracts of A549 cells revealed that this inhibition is not linked to preservation of expression of the IκBα protein. The chloramphenicol acetyltransferase assay indicated that NF-κB-dependent reporter gene expression is suppressed in U-937 cells pretreated with clarithromycin. These findings are consistent with the idea that clarithromycin suppresses the production of proinflammatory cytokines via inhibition of NF-κB activation.

Inhibition of NK cell activity by IL-17 allows vaccinia virus to induce severe skin lesions in a mouse model of eczema vaccinatum

Threats of bioterrorism have renewed efforts to better understand poxvirus pathogenesis and to develop a safer vaccine against smallpox. Individuals with atopic dermatitis are excluded from smallpox vaccination because of their propensity to develop eczema vaccinatum, a disseminated vaccinia virus (VACV) infection. To study the underlying mechanism of the vulnerability of atopic dermatitis patients to VACV infection, we developed a mouse model of eczema vaccinatum. Virus infection of eczematous skin induced severe primary erosive skin lesions, but not in the skin of healthy mice. Eczematous mice exhibited lower natural killer (NK) cell activity but similar cytotoxic T lymphocyte activity and humoral immune responses. The role of NK cells in controlling VACV-induced skin lesions was demonstrated by experiments depleting or transferring NK cells. The proinflammatory cytokine interleukin (IL)-17 reduced NK cell activity in mice with preexisting dermatitis. Given low NK cell activities and increased IL-17 expression in atopic dermatitis patients, these results can explain the increased susceptibility of atopic dermatitis patients to eczema vaccinatum.

Anti-inflammatory effect of 1α,25-dihydroxyvitamin D3 in human coronary arterial endothelial cells: Implication for the treatment of Kawasaki disease

Kawasaki disease (KD) is an acute febrile vasculitis in childhood that is associated with inflammatory cytokines, in which the vascular inflammation results in damage to the coronary arteries. The active form of vitamin D, 1alpha,25-dihydroxyvitamin D(3) {1alpha,25-(OH)(2)D(3)} exhibits anti-inflammatory activities. In this study, we determined the mRNA and protein expression of the vitamin D receptor in human coronary arterial endothelial cells (HCAEC) by RT-PCR and Western blotting, respectively. We examined whether or not 1alpha,25-(OH)(2)D(3) inhibits the tumor necrosis factor-alpha (TNF-alpha)-induced activation of nuclear transcription factor-kappaB (NF-kappaB), which is essential for the expression of proinflammatory cytokines in HCAEC, by ELISA. In addition, we determined the inhibitory effect of 1alpha,25-(OH)(2)D(3) on E-selectin expression induced by TNF-alpha in HCAEC by flow cytometry. RT-PCR revealed mRNA for the vitamin D receptor in HCAEC. Western blotting demonstrated vitamin D receptor protein in HCAEC. ELISA showed that pretreatment with 1alpha,25-(OH)(2)D(3) significantly inhibited the TNF-alpha-induced NF-kappaB activation in HCAEC. Moreover, flow cytometry revealed that pretreatment with 1alpha,25-(OH)(2)D(3) significantly inhibited the TNF-alpha-induced expression of E-selectin on HCAEC. Our results suggest that adjunctive 1alpha,25-(OH)(2)D(3) may modulate the inflammatory response during KD vasculitis.

Cysteine, histidine and glycine exhibit anti-inflammatory effects in human coronary arterial endothelial cells

The activation of nuclear factor‐kappa B (NF‐κB) in vascular endothelial cells may be involved in vascular pathogeneses such as vasculitis or atherosclerosis. Recently, it has been reported that some amino acids exhibit anti‐inflammatory effects. We investigated the inhibitory effects of a panel of amino acids on cytokine production or expression of adhesion molecules that are involved in inflammatory diseases in various cell types. The activation of NF‐κB was determined in human coronary arterial endothelial cells (HCAECs) because NF‐κB modulates the production of many cytokines and the expression of adhesion molecules. We examined the inhibitory effects of the amino acids cysteine, histidine and glycine on the induction of NF‐κB activation, expression of CD62E (E‐selectin) and the production of interleukin (IL)‐6 in HCAECs stimulated with tumour necrosis factor (TNF)‐α. Cysteine, histidine and glycine significantly reduced NF‐κB activation and inhibitor κBα (IκBα) degradation in HCAECs stimulated with TNF‐α. Additionally, all the amino acids inhibited the expression of E‐selectin and the production of IL‐6 in HCAECs, and the effects of cysteine were the most significant. Our results show that glycine, histidine and cysteine can inhibit NF‐κB activation, IκBα degradation, CD62E expression and IL‐6 production in HCAECs, suggesting that these amino acids may exhibit anti‐inflammatory effects during endothelial inflammation.

Characteristics of atopic children with pandemic H1N1 influenza viral infection: pandemic H1N1 influenza reveals 'occult' asthma of childhood.

To cite this article: Hasegawa S, Hirano R, Hashimoto K, Haneda Y, Shirabe K, Ichiyama T. Characteristics of atopic children with pandemic H1N1 influenza viral infection: Pandemic H1N1 influenza reveals ‘occult’ asthma of childhood. Pediatr Allergy Immunol 2011; 22: e119–e123.

Serum and cerebrospinal fluid cytokine profile of patients with 2009 pandemic H1N1 influenza virus-associated encephalopathy

PURPOSE Since April 2009, the number of patients with 2009 pandemic H1N1 influenza virus infection has been increasing in Japan just as in the rest of the world. Patients with 2009 pandemic H1N1 influenza-associated encephalopathy (pIE) have also been reported. The common clinical symptoms of this condition are seizures and progressive coma with high-grade fever. We previously reported the possible association between seasonal influenza-associated encephalopathy (sIE) and proinflammatory cytokines. However, the pathogenesis of pIE remains to be elucidated. RESULTS In pIE patients with a poor outcome, the serum levels of interleukin (IL)-6, IL-10, and soluble tumor necrosis factor (TNF) receptor (sTNFR1) were significantly higher than those in pIE patients without neurological sequelae. Similarly, the cerebrospinal fluid (CSF) IL-6 levels in pIE patients with a poor outcome were significantly higher than those in pIE patients without neurological sequelae. CONCLUSION Our results suggest that IL-6, TNF-α, and IL-10 play important roles in pIE, and that the serum levels of IL-6, IL-10, and sTNFR1 and the CSF levels of IL-6 are related to neurological complications.

A comparison of FcεRI-mediated RANTES release from human platelets between allergic patients and healthy individuals

Background: Recently some studies have suggested that human platelets may play an important role in allergic inflammation through the high affinity IgE receptor (FcΕRI), the low affinity IgE receptor (FcΕRII/CD23) and the low affinity IgG receptor (FcγRIIA/CD32) expressed on the cell surface. We reported that human platelets via the FcΕRI induced the release of the chemical mediator serotonin and the chemokine RANTES (regulated upon activation, normal T expressed and presumably secreted), but the biological implication of human platelets in type I allergy has not yet been understood clearly. Methods: We compared the levels of RANTES release from platelets obtained from allergic patients and healthy individuals, stimulated with monoclonal antibody (Ab) to human FcΕRI α-chain, or human myeloma IgE and anti-human IgE Ab. Results: We confirmed that the level of RANTES release from platelets of allergic patients stimulated with human IgE and anti-human IgE was significantly higher than that of healthy individuals. Conclusions: We demonstrated that the surface expression levels of FcΕRI on the platelets from allergic patients and healthy individuals were not significantly different, but that the platelets of allergic patients were more activated by the IgE-FcΕRI pathway than those of healthy individuals. Taken together, these results suggest a novel and important role for human platelets in perpetuating allergic inflammation through the IgE and FcΕRI.

Pranlukast inhibits NF‐κB activation in human monocytes/macrophages and T cells

Background Pranlukast is a leukotriene 1 (LT1) receptor antagonist and is effective against bronchial asthma. Pranlukast inhibits contraction of the tracheal muscle, and thereby antagonizes the binding of LTC4, LTD4 and LTE4. However, the action of pranlukast on monocytes/macrophages and T cells is unknown.

Molecular epidemiological study of human rhinovirus species A, B and C from patients with acute respiratory illnesses in Japan

Recent studies suggest that human rhinovirus species A, B and C (HRV-ABCs) may be associated with both the common cold and severe acute respiratory illnesses (ARIs) such as bronchiolitis, wheezy bronchiolitis and pneumonia. However, the state and molecular epidemiology of these viruses in Japan is not fully understood. This study detected the genomes of HRV-ABCs from Japanese patients (92 cases, 0-36 years old, mean±sd 3.5±5.0 years) with various ARIs including upper respiratory infection, bronchiolitis, wheezy bronchiolitis, croup and pneumonia between January and December 2010. HRV-ABCs were provisionally type assigned from the pairwise distances among the strains. On phylogenetic trees based on the nucleotide sequences of the VP4/VP2 coding region, HRV-A, -B and -C were provisionally assigned to 14, 2 and 12 types, respectively. The present HRV-A and -C strains had a wide genetic diversity (>30 % divergence). The interspecies distances were 0.230±0.063 (mean±sd, HRV-A), 0.218±0.048 (HRV-B) and 0.281±0.105 (HRV-C), based on nucleotide sequences, and 0.075±0.036 (HRV-A), 0.049±0.022 (HRV-B) and 0.141±0.064 (HRV-C) at the deduced amino acid level. Furthermore, HRV-A and -C were the predominant species and were detected throughout the seasons. The results suggested that HRV-A and -C strains have a wide genetic divergence and are associated with various ARIs in Japan.

Polyclonal IgE Induces Mast Cell Survival and Cytokine Production

BACKGROUND Ag-dependent activation of IgE-bearing mast cells is a critical first step in immediate hypersensitivity and other allergic responses. Recent studies have revealed Ag-independent effects of monoclonal mouse IgE molecules on mast cell survival and activation. However, no studies have been performed on the effects of polyclonal IgE molecules. Here, we tested whether polyclonal mouse and human IgE molecules affect survival and cytokine production in mast cells. METHODS Mast cells were cultured in the presence of polyclonal mouse and human IgE molecules, and cell survival and cytokine production were analyzed. RESULTS Polyclonal mouse IgE molecules in sera from mice with atopic dermatitis-like allergic skin inflammation, enhanced survival and cytokine production in mast cell cultures. Similar to the effects of monoclonal IgE, the polyclonal IgE effects were mediated by the high-affinity IgE receptor, FcepsilonRI. Human polyclonal IgE molecules present in sera from atopic dermatitis patients were also capable of activating mast cells, and inducing IL-8 production in human cord blood-derived mast cells. CONCLUSIONS These results imply that polyclonal IgE in atopic dermatitis and other atopic conditions might modulate mast cell number and function, thus amplifying the allergic response.