Yoshinobu Fukui

Genetic polymorphisms at FCER1B and PAI-1 and asthma susceptibility

Background We previously detected a promoter polymorphism (−109C/T) in the gene for the β‐chain of the high‐affinity receptor for IgE (FCER1B), which was associated with total serum IgE levels but not with asthma in a Japanese population. A genetic interaction is biologically plausible between FcεRI‐β and the plasminogen activator inhibitor 1 (PAI‐1), which is highly expressed in mast cells in asthmatics and plays an essential role in airway remodelling. We hypothesized that FCER1B promoter polymorphisms, by modifying the intensity of mast cell activation signals, modulate the genetic effects of a functional 4G/5G polymorphism in the PAI‐1 gene on asthma.

Genetic Impact of Functional Single Nucleotide Polymorphisms in the 3′-UTR Region of the Chemoattractant Receptor Expressed on Th2 Cells (CRTH2) Gene on Asthma and Atopy in a Japanese Population

Background: The human chemoattractant receptor expressed on Th2 cells (CRTH2), the receptor for prostaglandin D2, induces cell migration in eosinophils, basophils, and Th2 cells. The gene encoding CRTH2 is located on chromosome 11q13. Several groups, including ours, have reported significant associations between this region and various traits associated with allergic diseases such as asthma and atopy. Two single nucleotide polymorphisms in the 3′-UTR of the CRTH2 gene (1544G→C and 1651G→A) are associated with the mRNA stability of the gene; they have also been associated with asthma in both African American and Chinese populations. Methods: Because CRTH2 is a biologically important candidate gene on chromosome 11q13, we conducted a case-control analysis using 787 Japanese subjects (384 asthmatics and 403 controls) to evaluate the genetic impact of the CRTH2 gene on asthma and asthma-related traits. Four polymorphisms [1544G→C (rs11571288), 1651G→A (rs545659), 11336T→C (rs2074422), and 12375G→T (rs561285)] were studied. Results: The allele, genotype, or haplotype frequencies for 2 functional polymorphisms in our Japanese population were significantly different from those in the Chinese or African American populations. No association was found between any polymorphisms or haplotypes in the CRTH2 gene and asthma, atopy, or total serum IgE levels in a Japanese population. Conclusions: Our data failed to support previous associations of functional polymorphisms at the 3′-UTR of the CRTH2 gene implicated in asthma. We did show a significant difference in the allele and genotype frequencies as well as different haplotype frequencies among African American, Chinese, and Japanese populations, suggesting that the genetic impacts of these functional polymorphisms on asthma and asthma-related phenotypes may vary in different populations.

Sequence variants of the secreted phosphoprotein 1 gene are associated with total serum immunoglobulin E levels in a Japanese population.

Background Secreted phosphoprotein 1 (SPP1) is a cytokine with pleiotrophic immunological activities, including activation of macrophage chemotaxis and T‐helper type 1 (Th1) immune responses. SPP1 gene polymorphisms have been shown to be associated with several immune inflammatory diseases including multiple sclerosis (MS), which is characterized by fewer allergic symptoms and lower numbers of allergen sensitizations.

Enhanced Expression of Interleukin-18 Receptor α Chain by CD4+ T Cells in Sarcoidosis

Study objectives To investigate the expression of interleukin-18 receptor α chain (IL-18Rα) in BAL and peripheral blood (PB) T cells in patients with sarcoidosis compared with control subjects, to evaluate the relationship between the expression and clinical manifestations, and to clarify the mechanisms of altered expression. Subjects and methods The study subjects consisted of 21 patients with sarcoidosis and 8 normal control subjects. The expression of IL-18Rα was examined by flow cytometry. Results The proportions of BAL CD4+ and PB CD4+ T cells expressing IL-18Rα were significantly increased in patients with sarcoidosis compared to control subjects. BAL CD4+ T cells expressed IL-18Rα in a higher proportion than did paired CD8+ T cells in patients with sarcoidosis but not in control subjects. Greater proportions of BAL CD4+ T cells and BAL CD8+ T cells than of their PB counterparts expressed IL-18Rα in both patients and control subjects. CD4+ T cells were more sensitive to the induction of IL-18Rα by cytokines in vitro, such as interleukin (IL)-2, IL-12, and tumor necrosis factor-α than were CD8+ T cells. Increased expression of IL-18Rα by BAL T cells commonly observed in patients and control subjects was associated with the expansion of CD45RO+ cells in BAL T cells. However, there were no significant correlations between the expression of IL-18Rα by any cell populations and BAL findings, serum angiotensin-converting enzyme activities, radiograph stages, or clinical courses. Conclusion The overexpression of IL-18Rα predominantly by CD4+ T cells in sarcoidosis emphasizes crucial roles played by T-helper type 1 cells in the IL-18/IL-18Rα system in sarcoidosis.

Cytokines and Th2 cells in AEP of smoking.

. A MAJOR CAUSE of IgE-mediated allergy in bakers is the inhalation of wheat flour. After 2D electrophoresis more than 100 of spots of a water/salt-soluble fraction of wheat proteins are recognized by serum IgE antibodies from wheat-allergic patients (1). UsingDNAtechnology,wheatallergens can be cloned, biotechnologically produced as pure recombinant proteins, and characterized. An aliquot (1310 pfu) of a phagemid preparation, derived from in vivo mass excision (2) of a wheat endosperm cDNA library (3) (CSIRO, Canberra, Australia) was digested with the restriction enzymes EcoRI and XhoI. The cDNA inserts in the size region between 500 and 3000 base pairs (bp) were ligated into EcoRI/XhoI restricted pJuFo2 vector (4). Electrocompetent E. coli XL1-blue cells were electroporated with the ligation mixture, according to the manufacturers instructions (Bio-Rad, Munich, Germany). Thereafter, the biopanning procedure was applied to enrich IgEbinding phagemids as described (4) using a pooled serum from eight allergic bakers. The wheat-specific IgE antibody concentration of the serum pool was 54.7 kU/l, determined with the Pharmacia CAP method. Five rounds of affinity selections resulted in a library with 6310 cfu potentially coding for wheat allergens. In our previous study (1) triosephosphate isomerase (TPIS) was found to be detected by five out of 10 sera. TPIS could be amplified from the pJuFo cDNA expression library with the following specific primer pair allowing site-directed cloning into EcoRI/HindIII-cut pMal c2 expression vector (New England Biolabs, Frankfurt, Germany): TPIS_5’: 5’-CCGAATTCATGGGCCGCAAGTTCTTCGTC-3’ TPIS_3’: 5’-GAAAGCTTTTAGGCGGACTTCACGGCGGC-3’ PCR conditions were 968C, 5 min; (948C, 1 min; 548C, 1 min; 728C, 2 min) 32 cycles; 728C, 10 min. The resulting construct generates a fusion protein with a maltose binding protein (MBP) coded by pMal c2 suitable for affinity purification with an amylose resin (New England Biolabs). This is the first detection of TPIS in wheat (EMBL accession number: AJ278243). Sequence comparison of the deduced amino-acid sequence of thewheat TPIS gene, against other cereal triosephosphate isomerases, demonstrated a high level of sequence identity (88.1–96.4%). Comparison with data of Posch et al. (5) showed that seven of eight amino acids of a latex allergen, which was identified as a TPIS, are identical to the wheat TPIS. Sera obtained from 60 allergic bakers withawheat-specific IgEantibodyrangeof 3.65–24.9 kU/l, including four of the sera used for biopanning, were tested by EAST (Allergopharma, Reinbek,Germany) with self-prepared allergen disks with rTPISMBP according to the method described byCeska et al. (6).MBP served as negative control. Only one of the sera tested reacted with rTPIS-MBP (2.3 kU/l), without reaction toMBP (,0.35 kU/l). This serum belonged to the serum pool used for biopanning. Preincubation of this serum with native wheat-flour extract (80 mg) inhibited binding to rTPIS-MBP completely, indicating identity of allergenic epitopes in native and recombinant TPIS. We are grateful to Prof. K. Blaser for his continuous support. Work at SIAF was supported by the Swiss National Science Foundation Grants no. 31–50515.97 and 31–63381.00. This work is part of the BGFA-project All/Zen8, supported by theHauptverband der gewerblichen Berufsgenossenschaften (HVBG), Sankt Augustin, Germany.