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Notes: Background Both prostaglandin (PG) D receptor (DP) and CRTH2 (chemoattractant receptor-homologous molecule expressed on Th2 cells)/DP2 are high-affinity receptors for PGD2. Previous studies have demonstrated that PGD2 enhances releasability and induces CRTH2/DP2-mediated migration in human basophils, but the precise effects of PGD2 on basophils as well as receptor usage have not been fully clarified.Objective We comprehensively explored the roles of DP and CRTH2/DP2 in basophil functions by using selective agonists and antagonists for each receptor.Methods DP and CRTH2/DP2 transcripts were quantified by real-time PCR. We studied the effects of selective agonists (DP: BW245C; CRTH2/DP2: 13,14-dihydro-15-keto (DK)-PGD2) and/or antagonists (DP: BWA868C; CRTH2/DP2: ramatroban) on Ca2+ mobilization, migration, degranulation, CD11b expression and survival of human basophils.Results Basophils expressed transcripts of both DP and CRTH2/DP2, but the levels of CRTH2/DP2 transcripts were ca. 100-fold higher compared with DP transcripts. Ca2+ influx was induced in basophils by either PGD2 or DK-PGD2/CRTH2 agonist but not by BW245C/DP agonist. Basophils treated with PGD2 were completely desensitized to subsequent stimulation with DK-PGD2, but not vice versa. DK-PGD2 as well as PGD2 up-regulated CD11b expression, induced migration and enhanced degranulation, and those effects were completely antagonized by ramatroban/CRTH2 antagonist. In contrast, BW245C/DP agonist exhibited an inhibitory effect on basophil migration and IgE-mediated degranulation, and the migration inhibitory effect was effectively antagonized by BWA868C/DP antagonist. On the other hand, while PGD2 significantly shortened the basophil life-span, neither DK-PGD2/CRTH2 agonist nor BW245C/DP agonist did.Conclusion CRTH2/DP2 is primarily responsible for the pro-inflammatory effects of PGD2 on human basophils, while DP introduces negative signals capable of antagonizing the effects of CRTH2/DP2 in these cells. The effects of PGD2 on longevity imply a mechanism(s) other than via DP or CRTH2/DP2. CRTH2/DP2 on basophils may afford opportunities for therapeutic targeting in allergic inflammation.

Notes: Background Th2 and Th1 cells have been suggested to express CCR3/CCR4 and CCR5/CXCR3, respectively.Objective We examined CCR3, CCR4, CCR5 and CXCR3 expression and cytokine production in peripheral blood CD4+ T cells from patients with atopic dermatitis (AD), which has been postulated to be a Th2-type cell-mediated disease, and then analysed the possible correlation between these values and the levels of several clinical parameters.Methods Intracellular cytokine production and chemokine receptor expression in peripheral blood CD4+ T cells from 40 AD patients and 20 sex- and age-matched healthy control subjects were studied by flow cytometry.Results The frequencies of IL-4- and IL-13-producing CD4+ T cells from patients with AD were significantly higher than those from healthy control subjects (IL-4:3.9 ± 2.1% vs. 1.6 ± 0.7%, P = 0.0005, IL-13:4.0 ± 2.1% vs. 1.8 ± 0.8%, P = 0.0023), whereas the frequencies of IL-2- and IFN-γ-producing CD4+ T cells were significantly decreased in AD patients (IL-2:38.1 ± 10.3% vs. 51.3 ± 6.3%, P = 0.0003, IFN-γ: 9.9 ± 3.5% vs. 26.4 ± 4.6%, P 〈 0.0001). The percentage of CCR4+ cells in CD4+ CD45RO+ T cells in AD patients was significantly higher than that in healthy control subjects (24.4 ± 8.0% vs. 10.9 ± 2.3%, P 〈 0.0001) and was correlated positively with the total serum IgE, serum lactic dehydrogenase (LDH) level, eosinophil number, eruption score, and IL-4 and IL-13 secretion in CD4+ T cells, and inversely with IL-2 and IFN-γ secretion in CD4+ T cells. In contrast, CCR3 was not detected on circulating CD4+ T cells even in AD patients. On the other hand, the percentage of CCR5+ or CXCR3+ cells in CD4+ CD45RO+ T cells in AD patients was significantly decreased (CCR5:23.2 ± 7.0% vs. 28.4 ± 5.4%, P = 0.023, CXCR3:29.9 ± 11.4% vs. 38.5 ± 6.7%, P = 0.028) and was positively correlated with eruption score (P 〈 0.05). Multiple regression analyses showed that the percentage of CCR4 expression highly correlated with serum IgE, LDH, eosinophil number and eruption in AD patients.Conclusion CCR4+ cells might be involved in the aetiopathogenesis of AD.

Notes: Background: Cytokines liberated by TH2 cells play crucial roles in the pathogenesis of bronchial asthma. Recent studies have demonstrated that CC chemokine receptor (CCR)4 is preferentially expressed by TH2 cells. These facts suggest possible involvement of two CCR4-specific ligands i.e., thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC), in the pathogenesis of bronchial asthma via recruitment of TH2 cells to inflammatory sites. We investigated the levels of TARC and MDC in the serum and induced sputum of asthmatics. Methods: The levels of TARC in the serum (46 asthmatics and 26 healthy subjects) and induced sputum (30 asthmatics and 6 healthy subjects) were measured using a highly sensitive ELISA system. The levels of eotaxin and MDC were also measured by ELISA. Results: TARC, but not MDC, was significantly increased in asthmatic sera (P〈0.001). Although MDC was undetectable in the sputum of most cases by our assay system, sputum TARC was significantly increased (P=0.027). Conclusions: The elevated TARC levels in asthmatics might be involved in the pathophysiology of asthma.

Notes: Background: Th2 cells are thought to be involved in eosinophilic inflammation of the lung. CC chemokine receptor 4 (CCR4) has been identified as a specific receptor for both thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC), and is preferentially expressed on Th2 cells.Objective: Our aim was to evaluate the role of Th2 cells in the lung of patients with eosinophilic pneumonia (EP).Methods: The concentrations of TARC, MDC, and interleukin (IL)-5 were measured in bronchoalveolar lavage fluid (BALF) by ELISA. Proportion of CCR4-expressing CD4+ T cells (CCR4+ CD4+ T cells) was determined by flow cytometry.Results: TARC and MDC concentrations in BALF were higher in patients with EP than in normal subjects. The proportion of CCR4-expressing cells among CD4+ T cells was higher in BALF than in peripheral blood of patients with EP. There was a significant correlation between the number of CCR4+ CD4+ T cells and the levels of TARC, MDC, and IL-5 in BALF of patients with EP.Conclusions: Our data suggest that Th2 cells, which express CCR4 and its ligands (TARC and MDC), contribute to the pathogenesis of EP in the lung.

Notes: Summary Background Bullous pemphigoid (BP) is an autoimmune blistering disease characterized by eosinophilia and high serum IgE levels. The accumulated evidence suggests that various cytokines are involved in the lesional skin of patients with BP. Recently, thymus and activation-regulated chemokine (TARC/CCL17), a CC chemokine, was identified as a selective chemoattractant for CC chemokine receptor 4 (CCR4)-expressing cells. Objective In this study, we examined the involvement of TARC in patients with BP. Methods We determined the fluid and serum TARC levels in patients with BP by enzyme-linked immunosorbent assay and compared the serum TARC levels with the eosinophil numbers in peripheral blood. We also compared the serum TARC levels in five patients with BP before and after they were treated. Moreover, we examined TARC, CCR4 and CXC chemokine receptor 3 (CXCR3) expression in the lesional skin of patients with BP by immunohistochemical procedures. Furthermore, we measured CCR4 positivity in CD4+ CD45RO+ cells of peripheral blood mononuclear cells (PBMCs) in patients with BP and healthy control subjects. Results The fluid TARC levels in patients with BP were significantly higher than those in blisters from burn patients or suction blisters of healthy control subjects. The serum TARC levels in patients with BP were also significantly higher than those in pemphigus vulgaris (PV) patients and healthy control subjects, and decreased after the treatment. The serum TARC levels in patients with BP significantly correlated with the eosinophil numbers in peripheral blood (r = 0·72, P 〈 0·002). Immunohistochemistry showed a strong reactivity of TARC in the epidermal keratinocytes (KCs) of BP. Moreover, both CCR4 and CXCR3 were expressed on the dermal infiltrating CD4+ T cells mainly beneath the bullae of patients with BP. Fluorescence-activated cell sorting analysis showed a higher percentage of CCR4 positivity in CD4+ CD45RO+ cells of PBMCs in patients with BP than that in healthy control subjects, while there was no significant difference of CXCR3 positivity in CD4+ CD45RO+ cells of PBMCs between patients with BP and healthy control subjects. Conclusions These findings strongly suggest that TARC may be one of the important chemokines that are involved in the pathogenesis of BP.