Masaki Hayama

Effects of Probiotics on Allergic Rhinitis Induced by Japanese Cedar Pollen: Randomized Double-Blind, Placebo-Controlled Clinical Trial

Background:Lactobacillus casei strain Shirota (LcS) has been found to exert antiallergic effects in animal experiments, but there is little information about its clinical effects in human patients with allergy. Methods: We performed a randomized double-blind, placebo-controlled study to investigate the effects of LcS in patients with allergic rhinitis triggered by Japanese cedar pollen (JCP). Participants were asked to drink fermented milk containing LcS (LcS group) or placebo (control group) for 8 weeks. Clinical symptoms and immunological parameters were compared between the two groups. Results: Symptom-medication scores (SMS) worsened in accordance with the increase in the amount of scattered JCP. In terms of the nasal and ocular SMS, there was no significant difference between the LcS group and the placebo group during the ingestion period. In the subgroup of patients with moderate-to-severe nasal symptom scores before starting the ingestion of test samples, supplementation with LcS tended to reduce nasal SMS. Conclusion: These results indicate that fermented milk containing LcS does not prevent allergic symptoms in patients sensitive to JCP, but may delay the occurrence of allergic symptoms in patients with moderate-to-severe nasal symptom scores.

Serase-1B, a new splice variant of polyserase-1/TMPRSS9, activates urokinase-type plasminogen activator and the proteolytic activation is negatively regulated by glycosaminoglycans

Polyserase-1 (polyserine protease-1)/TMPRSS9 (transmembrane serine protease 9) is a type II transmembrane serine protease (TTSP) that possesses unique three tandem serine protease domains. However, the physiological function of each protease domain remains poorly understood. We discovered a new splice variant of polyserase-1, termed Serase-1B, which contains 34 extra amino acids consisting a SEA module (a domain found in sea urchin sperm protein, enterokinase and agrin) adjacent to the transmembrane domain and the first protease domain with a mucin-like box at the C-terminus. The tissue distribution of this enzyme by RT (reverse transcription)-PCR analysis revealed high expression in the liver, small intestine, pancreas, testis and peripheral blood CD14+ and CD8+ cells. To investigate the role of Serase-1B, a full-length form recombinant protein was produced. Interestingly, recombinant Serase-1B was partly secreted as a soluble inactive precursor and it was also activated by trypsin. This activated enzyme selectively cleaved synthetic peptides for trypsin and activated protein C, and it was inhibited by several natural serine protease inhibitors, such as aprotinin, alpha2-antiplasmin and plasminogen activator inhibitor 1. In addition, Serase-1B efficiently converted pro-uPA (urokinase-type plasminogen activator) into active uPA and this activation was strongly inhibited by these natural inhibitors. Furthermore, this activation was also negatively regulated by glycosaminoglycans. Our results indicate that Serase-1B is a novel member of TTSPs that might be involved in uPA/plasmin-mediated proteolysis and possibly implicated in biological events such as fibrinolysis and tumour progression.

Identification and analysis of the promoter region of the type II transmembrane serine protease polyserase-1 and its transcript variants

Abstract Polyserase-1/TMPRSS9 and its alternative transcripts, serase-1B and serase-2B, are novel type II transmembrane serine proteases that may regulate physiological and pathological phenomena on the cell surface. To understand the mechanisms of gene expression and regulation of these transcripts, we cloned and characterized the 5′ promoter region of the mouse polyserase-1 (mpolyserase-1) gene. Using 5′-rapid amplification of cDNA ends, we located the transcription initiation site 272 nucleotides upstream of the translation initiation site. Luciferase reporter gene analysis revealed that the region from +186 to +272 bp in the 5′-untranslated region (UTR), containing the GATA motif (AGATAA), glucocorticoid responsible element (TGTTCT), and E-box sequence (CAGGTG), is required for maximal promoter activity. Mutations introduced into the E-box sequence but not elsewhere in the promoter region caused a selective decrease in transcriptional activity. Furthermore, a DNA probe (+229 to +255 bp) containing the E-box sequence formed a single nuclear protein complex in a sequence-specific manner. These data suggest that the expression of mpolyserase-1 and its transcript variants is positively regulated by the E-box in its 5′-UTR, which might be responsible for the binding of basic helix-loop-helix transcription factors involved in the development of various organelles.

Induction and maintenance of anti-influenza antigen-specific nasal secretory IgA levels and serum IgG levels after influenza infection in adults

Please cite this paper as: Fujimoto et al. (2012) Induction and maintenance of anti‐influenza antigen‐specific nasal secretory IgA levels and serum IgG levels after influenza infection in adults. Influenza and Other Respiratory Viruses 6(6), 396–403.

Evaluation of nasal IgA secretion in normal subjects by nasal spray and aspiration

OBJECTIVE Nasal washing (NW) is a popular method for collecting human nasal lavage fluid. However, for NW the subject must be trained, and the method is unsuitable for field studies on untrained subjects. To overcome this problem, we have developed an easy and painless method, a nasal spray and aspiration (NSA) method. METHODS This method is different from NW in that the nasal cavity is misted over with saline, and the nasal lavage fluid is aspirated from the nostrils through a silicon tube. First, nasal lavage fluid was obtained twice by NSA with an interval of a week between lavages to evaluate intraindividual variability, and the IgA and protein levels in the nasal lavage fluid were measured by enzyme-linked immunosorbent assay and bicinchoninic acid assay, respectively. Next, the IgA value determined by NSA was compared with that by NW in another 12 normal subjects 2 days after NSA. RESULTS In 10 normal subjects, mean volume of saline sprayed into the nose was 0.46+/-0.15 ml (mean+/-S.D.). Mean volume of aspirated nasal lavage fluid containing both sprayed saline and nasal secretion was 0.44+/-0.37 ml. The mean IgA level/mg protein in the nasal lavage fluid determined by NSA was 112+/-18 microg/mg protein at the first and 99+/-20 at the second times of measurement, being highly reproducible. The mean value by NSA was 114+/-19 microg/mg protein, being almost the same as that by NW of 99+/-27. CONCLUSION These findings suggest that the IgA level/mg protein in nasal lavage fluid determined by NSA instead of NW might be useful for assessing the variability of nasal IgA secretion.

Allergic conversion of protective mucosal immunity against nasal bacteria in patients with chronic rhinosinusitis with nasal polyposis

Background Chronic rhinosinusitis with nasal polyposis (CRSwNP) is characterized by eosinophilic inflammation and polyposis at the nose and paranasal sinus and a high concentration of IgE in nasal polyps (NPs). The causative antigen and pathogenesis of CRSwNP remain unknown. Objective We aimed to identify reactive allergens of IgE antibodies produced locally in NPs of patients with CRSwNP. We also attempted to unravel the differentiation pathway of IgE‐producing B cells in NPs. Methods IgE reactivity of patients with CRSwNP was investigated by characterizing single cell–derived mAbs. T‐cell response against identified allergens was investigated in vitro. NP‐infiltrating lymphocytes were characterized by using flow cytometry. Immunoglobulins expressed in NPs were analyzed by using high‐throughput DNA sequencing for immunoglobulin. Results About 20% of isolated IgE antibodies derived from NP‐residing plasmablasts specifically recognized surface determinants of nasal bacteria, such as Staphylococcus aureus, Streptococcus pyogenes, and Haemophilus influenzae. A TH2 response against S pyogenes was observed in patients with CRSwNP. Flow cytometric analysis revealed sizable germinal center B–like cell and plasmablast subsets expressing IgE on the cell surface in NPs. High‐throughput DNA sequencing immunoglobulin analysis highlighted the clonal connectivity of IgE with IgG and IgA1. The I&egr;‐C&agr;1 circle transcript was detected in NPs. Conclusions In patients with CRSwNP, nasal bacteria–reactive B cells differentiate into IgE‐producing B cells through IgG/IgA1‐IgE class switching, suggesting that allergic conversion of the mucosal response against nasal bacteria underlies disease pathogenesis. Graphical abstract Figure. No Caption available.

Eosinophil-derived neurotoxin enhances airway remodeling in eosinophilic chronic rhinosinusitis and correlates with disease severity

The role of eosinophil-derived neurotoxin in chronic rhinosinusitis