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1.
Biosci Biotechnol Biochem ; 87(6): 649-652, 2023 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-36945060

RESUMO

The antiallergic properties of phlorotannins, algal polyphenols, have been widely reported. This study examined the soothing effect of phlorotannin concentrate (PTC) from Eisenia nipponica on cedar pollinosis in Cry j 1-stimulated mice. PTC reduced the mice's sneezing and nasal rubbing, which was attributed to decreased levels of immunoglobulin E and Th2-type cytokines [interleukin (IL)-4, IL-5, and IL-13].


Assuntos
Cryptomeria , Rinite Alérgica Sazonal , Camundongos , Animais , Alérgenos , Proteínas de Plantas , Antígenos de Plantas , Citocinas
2.
Biosci Biotechnol Biochem ; 85(10): 2089-2092, 2021 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-34410303

RESUMO

As a method of evaluating the effect of inactivators on allergens while suppressing the effect of inactivator on the assay, we developed new dot-blot method that combines immunostaining and protein detection methods. This method is useful for evaluating whether the inactivator can inactivate allergens rather than removing them from the assay.


Assuntos
Alérgenos , Animais , Cryptomeria , Ácaros
3.
Clin Immunol ; 210: 108310, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31743749

RESUMO

Sublingual immunotherapy (SLIT) with Japanese cedar (JCe) pollinosis was expected to be effective for Japanese cypress (JCy) pollinosis. However, only a half of JCy pollinosis patients clinically improved. Therefore, we examined the immunological effect of SLIT for JCy pollinosis. Peripheral blood mononuclear cells (PBMCs) from patients with JCe and JCy pollinosis who did and did not receive SLIT were incubated with Cry j 1, Cha o 1 and Cha o 3 antigens. Basophil activation test (BAT) were performed. Production of IL-5 and IL-17 induced by antigens was inhibited in the SLIT group. Cry j 1-specific production of IL-10 was increased, and serum Cry j 1-specific IgE and -IgG4 were elevated. However, Cha o 1- or Cha o 3-specific production of IL-10 and specific IgG4 was not increased. Antigens-specific BAT did not decrease after SLIT. New SLIT with JCe and JCy is needed for patients with combined JCe and JCy pollinosis.


Assuntos
Leucócitos Mononucleares/imunologia , Rinite Alérgica Sazonal/terapia , Imunoterapia Sublingual/métodos , Adulto , Antígenos de Plantas/imunologia , Teste de Degranulação de Basófilos , Células Cultivadas , Chamaecyparis/imunologia , Cryptomeria/imunologia , Citocinas/metabolismo , Feminino , Humanos , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Masculino , Pessoa de Meia-Idade , Extratos Vegetais/imunologia , Proteínas de Plantas/imunologia , Pólen/imunologia , Estudos Prospectivos , Rinite Alérgica Sazonal/imunologia
4.
Protein Expr Purif ; 109: 62-9, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25665505

RESUMO

Peptides containing T-cell epitopes from allergens, which are not reactive to allergen-specific IgE, are appropriate candidates as antigens for specific immunotherapy against allergies. To develop a vaccine that can be used in practical application to prevent and treat Japanese cedar pollen allergy, four major T-cell epitopes from the Cry j 1 antigen and six from the Cry j 2 antigen were selected to design cry j 1 epi and cry j 2 epi, DNA constructs encoding artificial polypeptides of the selected epitopes. To apply cholera toxin B subunit (CTB) as an adjuvant, cry j 1 epi and cry j 2 epi were linked and then fused to the CTB gene in tandem to construct a fusion gene, ctb-linker-cry j 1 epi- cry j 2 epi-flag. The fusion gene was introduced into a pET-28a(+) vector and expressed in Escherichia coli BL21(DE3). The expressed recombinant protein was purified by a His-tag affinity column and confirmed by western blot analysis using anti-CTB and anti-FLAG antibodies. The purified recombinant protein also proved to be antigenic against anti-Cry j 1 and anti-Cry j 2 antibodies. Expression of the recombinant protein induced with 1mM IPTG reached a maximum in 3-5h, and recovery of the affinity-purified recombinant protein was approximately 120mg/L of culture medium. The present study indicates that production of sufficient amounts of recombinant protein with antigenic epitopes may be possible by recombinant techniques using E. coli or other bacterial strains for protein expression.


Assuntos
Alérgenos/imunologia , Bioquímica/métodos , Toxina da Cólera/metabolismo , Cryptomeria/metabolismo , Epitopos de Linfócito T/metabolismo , Escherichia coli/metabolismo , Pólen/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Alérgenos/química , Sequência de Aminoácidos , Antígenos de Plantas , Western Blotting , DNA/metabolismo , Eletroforese em Gel de Poliacrilamida , Epitopos de Linfócito T/química , Epitopos de Linfócito T/isolamento & purificação , Dados de Sequência Molecular , Peptídeos/química , Plasmídeos/metabolismo , Reação em Cadeia da Polimerase , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/isolamento & purificação
5.
Allergol Int ; 64(2): 161-8, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25838092

RESUMO

BACKGROUND: Short-term oral immunotherapy (OIT) using the Cry j1-galactomannan conjugate for Japanese cedar pollinosis may be effective and relatively safe. However, a treatment regimen has not been established. In the present study, we examined a new OIT regimen with a build-up phase and extended the maintenance phase of OIT to the peak period of the pollen season to enhance the therapeutic effect and safety of OIT. METHODS: A prospective, randomized, open-label trial was conducted over a period of 4 months. Participants were randomly divided into two groups. The OIT group comprised 23 subjects. The build-up phase was initiated 1 month before the expected pollen season. The maintenance phase was continued for 51 days during the peak pollen season. The control group comprised 24 subjects. The symptoms and medication score, levels of allergen-specific serum antibodies throughout the pollen season, and adverse effects with OIT were evaluated. RESULTS: Participants receiving OIT showed significant improvements in total symptom scores, medication score, and total symptom-medication scores throughout the pollen season compared with the control group. The levels of allergen-specific serum IgG4 were significantly increased in the OIT group but not in the control group throughout the cedar pollen season. Importantly, no severe adverse effects were observed with OIT. CONCLUSIONS: The new regimen of short-term OIT using the Cry j1-galactomannan conjugate for Japanese cedar pollinosis is effective, relatively safe and induces immune tolerance. Thus, OIT using allergen-galactomannan conjugates may provide a rapid, effective, and thus convenient immunotherapy for pollinosis instead of SLIT or SCIT.


Assuntos
Antígenos de Plantas/imunologia , Dessensibilização Imunológica , Mananas/imunologia , Proteínas de Plantas/imunologia , Rinite Alérgica Sazonal/terapia , Administração Oral , Adulto , Antígenos de Plantas/química , Contagem de Células , Cryptomeria/imunologia , Dessensibilização Imunológica/efeitos adversos , Feminino , Galactose/análogos & derivados , Humanos , Masculino , Mananas/química , Pessoa de Meia-Idade , Proteínas de Plantas/química , Pólen , Resultado do Tratamento , Adulto Jovem
6.
Nutrients ; 14(13)2022 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-35807837

RESUMO

The association between pollen food allergy syndrome (PFAS) and allergic march remains unclear. In this prospective cohort study of the general population in Tokyo (T-Child Study), we found that sensitization to Cry j 1 and Fel d 1 at ages 5 and 9 years was associated with an increased risk of PFAS at 13 years old (at 5 years, Cry j 1: adjusted odds ratio aOR, 2.74; 95% confidence interval CI, 1.53-4.91; Fel d 1: aOR, 2.61; 95% CI, 1.31-5.19; at 9 years, Cry j 1: adjusted odds ratio aOR, 4.28; 95% confidence interval CI, 1.98-9.25; Fel d 1: aOR, 2.40; 95% CI, 1.33-4.32). In particular, sensitization to Bet v 1 at ages 5 and 9 years was associated with a strong risk of PFAS at the age of 13 years (at 5 years: aOR, 10.6; 95% CI, 2.64-42.5; at 9 years: aOR, 9.1; 95% CI, 4.71-17.6). PFAS risk by age 13 years was increased by any allergic symptom at 5 or 9 years, a combination of wheezing, eczema, and rhinitis, and Bet v 1 sensitization. Our findings suggest that PFAS may be associated with allergic march.


Assuntos
Fluorocarbonos , Hipersensibilidade Alimentar , Adolescente , Alérgenos , Criança , Pré-Escolar , Hipersensibilidade Alimentar/diagnóstico , Humanos , Pólen , Estudos Prospectivos , Síndrome
7.
Asia Pac Allergy ; 11(3): e26, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34386402

RESUMO

BACKGROUND: Symptoms of rhinitis and asthma can be exacerbated during Japanese cedar pollen (JCP)-scattering season, even in subjects who are not sensitized to JCP, suggesting that innate immune responses may contribute to this process. We previously reported that house dust mite directly activates the effector functions of eosinophils. Similar mechanisms may play roles in the JCP-related aggravation of allergic diseases. OBJECTIVE: To investigate whether JCP or Cry j 1, a major allergen of JCP, can modify the effector functions of eosinophils. METHODS: Eosinophils isolated from the peripheral blood of healthy donors were stimulated with either JCP or Cry j 1, and their adhesion to human intercellular adhesion molecule-1 was measured using eosinophil peroxidase assays. The generation of eosinophil superoxide anion (O2 -) was measured based on the superoxide dismutase-inhibitable reduction of cytochrome C. Concentrations of eosinophil-derived neurotoxin in the cell media were measured by enzyme-linked immunosorbent assay as a marker of degranulation. RESULTS: Both JCP and Cry j 1 directly induced eosinophil adhesiveness, generation of O2 -, and release of eosinophil-derived neurotoxin. Both anti-αM and anti-ß2 integrin antibodies blocked all of these eosinophil functions induced by JCP and Cry j 1. Similarly, PAR-2 antagonists also partially suppressed all of these effector functions induced by JCP and Cry j 1. CONCLUSION: JCP and Cry j 1 directly activate the functions of eosinophils, and both αMß2 integrin and partly PAR-2 are contributed to this activation. Therefore, JCP-induced eosinophil activation may play a role in the aggravation of allergic airway diseases in nonsensitized patients as well as in JCP-sensitized patients.

8.
Vet Sci ; 5(3)2018 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-30208581

RESUMO

Levels of Japanese cedar pollen (Cryptomeria japonica) have increased in Japan and cedar pollinosis caused by Japanese cedar pollen has been reported in dogs. Serum levels of immunoglobulin E (IgE) against Cry j 1 and Cry j 2 in dogs raised in institutes and treated at veterinary hospitals in Japan were thus investigated. A total of 71 sera obtained from two institutes and 87 sera obtained from veterinary hospitals in the Hyogo and Kanagawa Prefectures were analyzed in this study. Serum levels of IgE were measured using the enzyme-linked immunosorbent assay with commercial purified Cry j 1 and Cry j 2. IgE against Cry j 1 and Cry j 2 in sera obtained from the two institutes were detected, despite the dogs being bred in enclosed areas. Moreover, significant differences were noted in the serum levels of IgE against Cry j 1 and Cry j 2 between the two institutes. The number of samples showing Cry j 1 or Cry j 2 levels above the cut-off values was greater in the Kanagawa Prefecture than in the Hyogo Prefecture. In total, 14 dogs showed Cry j 1 and Cry j 2 levels greater than the cut-off values in the Hyogo Prefecture, and only three such dogs were seen in the Kanagawa Prefecture. A significant correlation between serum levels against both allergens was observed (r² = 0.6931, p < 0.0001).

9.
Eur J Pharmacol ; 798: 105-112, 2017 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-28119075

RESUMO

Fab fragments (Fabs) of antibodies having the ability only to bind to specific allergens lack effector functions due to the absence of the Fc portion. In the present study, we examined whether IgG1 monoclonal antibody (mAb) Fabs targeting Japanese cedar pollen (JCP) Cry j 1 were able to regulate JCP-induced allergic conjunctivitis in mice. BALB/c mice actively sensitized with JCP were repeatedly challenged by topical administration of JCP eye drops. Fabs prepared by the digestion of anti-JCP IgG1 mAbs (P1-3 and P1-8) with papain were applied to the eye 15min before the JCP challenges followed by measurement of the clinical conjunctivitis score. In the in vitro experiments, P1-3 and P1-8 showed specific binding to JCP Cry j 1. Furthermore, intact P1-3 binding to Cry j 1 was inhibited by P1-3 Fabs, but not P1-8 Fabs; additionally, P1-8 Fabs, but not P1-3 Fabs, suppressed the intact P1-8 binding, suggesting that the epitopes of Cry j 1 recognized by P1-3 and P1-8 were different. Topical ocular treatment with P1-3 Fabs or P1-8 Fabs was followed by marked suppression of JCP-induced conjunctivitis (P<0.01). In histological evaluation, P1-8 Fabs showed a reduction in eosinophil infiltration in the conjunctiva (P<0.01). These results demonstrated that topical ocular treatment with IgG1 mAb Fabs to Cry j 1 was effective in suppressing JCP-induced allergic conjunctivitis in mice. Furthermore, it suggests the possibility that some epitopes recognized by Fabs could be used as a tool to regulate allergic conjunctivitis.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Antígenos de Plantas/imunologia , Conjuntivite Alérgica/tratamento farmacológico , Conjuntivite Alérgica/imunologia , Fragmentos Fab das Imunoglobulinas/administração & dosagem , Fragmentos Fab das Imunoglobulinas/imunologia , Proteínas de Plantas/imunologia , Administração Tópica , Animais , Anticorpos Monoclonais/uso terapêutico , Especificidade de Anticorpos , Conjuntivite Alérgica/sangue , Epitopos/imunologia , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Fragmentos Fab das Imunoglobulinas/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pólen/imunologia
10.
Arch Dermatol Res ; 308(1): 49-54, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26498292

RESUMO

Cry j1 is the major peptide allergen of Japanese cedar (Sugi), Cryptomeria japonica. Since some allergens disrupt epidermal permeability barrier homeostasis, we hypothesized that Cry j1 might have a similar effect. Intracellular calcium level in cultured human keratinocytes was measured with a ratiometric fluorescent probe, Fura-2 AM. Application of Cry j1 significantly increased the intracellular calcium level of keratinocytes, and this increase was inhibited by trypsin inhibitor or a protease-activated receptor 2 (PAR-2) antagonist. We found that Cry j1 itself did not show protease activity, but application of Cry j1 to cultured keratinocytes induced a rapid (within 30 s) and transient increase of protease activity in the medium. This transient increase was blocked by trypsin inhibitor or PAR-2 antagonist. The effect of Cry j1 on transepidermal water loss (TEWL) of cultured human skin was measured in the presence and absence of a trypsin inhibitor and PAR-2 antagonist. Cry j1 significantly impaired the barrier function of human skin ex vivo, and this action was blocked by co-application of trypsin inhibitor or PAR-2 antagonist. Our results suggested that interaction of Cry j1 with epidermal keratinocytes leads to the activation of PAR-2, which induces elevation of intracellular calcium and disruption of barrier function. Blocking the interaction of Cry j1 with epidermal keratinocytes might ameliorate allergic reaction and prevent disruption of epidermal permeability barrier homeostasis.


Assuntos
Alérgenos/imunologia , Antígenos de Plantas/imunologia , Cálcio/metabolismo , Epiderme/patologia , Queratinócitos/metabolismo , Proteínas de Plantas/imunologia , Junções Íntimas/patologia , Células Cultivadas , Cryptomeria/imunologia , Epiderme/imunologia , Humanos , Hipersensibilidade/imunologia , Técnicas de Cultura de Órgãos , Peptídeo Hidrolases/metabolismo , Proteínas de Plantas/metabolismo , Pólen/imunologia , Inibidores de Proteases/metabolismo , Receptor PAR-2/metabolismo , Junções Íntimas/imunologia
11.
Auris Nasus Larynx ; 43(1): 50-5, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26197940

RESUMO

OBJECTIVE: We have recently reported that a new regimen of short-term oral immunotherapy (OIT) with the Cry j1-galactomannan conjugate for Japanese cedar pollinosis (JCP) is effective to the improvement in the symptoms and medication use during the pollen season and relatively safe. The effect of OIT on quality of life (QOL) of JCP patients has not been assessed. Therefore, we evaluated for the first time the effect of OIT on QOL during the Japanese cedar/cypress pollen season. METHODS: A prospective, randomized, open-label trial was conducted over a period of 4 months. Participants were randomly divided into two groups. The OIT and control groups comprised 23 and 24 subjects, respectively. The build-up phase was initiated 1 month before the expected pollen season. The maintenance phase was continued for 51 days during the peak of the cedar pollen season. The QOL score in the Japan Rhinoconjunctivitis Quality of Life Questionnaire (JRQLQ) No. 1 and visual analog scale (VAS) throughout the pollen season were evaluated. RESULTS: Participants receiving OIT showed significant improvements in the total QOL score and VAS throughout the pollen season compared with the control group. In addition, the mean total QOL score and VAS correlated in both groups during the pollen season. CONCLUSION: The new regimen of short-term OIT using the Cry j1-galactomannan conjugate results in meaningful improvements in QOL of JCP patients. Our findings suggest that short-term OIT using allergen-galactomannan conjugates, as well as sublingual and subcutaneous immunotherapy, improves QOL of patients with pollinosis. The study was registered in UMIN-CTR (UMIN000013408) as the name of "a prospective, randomized, open study of oral Cry j1-galactomannan conjugate immunotherapy for Japanese cedar pollen allergy".


Assuntos
Antígenos de Plantas/uso terapêutico , Dessensibilização Imunológica/métodos , Mananas/uso terapêutico , Proteínas de Plantas/uso terapêutico , Qualidade de Vida , Rinite Alérgica Sazonal/tratamento farmacológico , Administração Oral , Adulto , Antialérgicos/uso terapêutico , Cryptomeria/imunologia , Feminino , Galactose/análogos & derivados , Humanos , Imunoglobulina E/imunologia , Japão , Masculino , Pessoa de Meia-Idade , Rinite Alérgica Sazonal/imunologia , Inquéritos e Questionários , Escala Visual Analógica , Adulto Jovem
12.
J Mol Biol ; 426(17): 3016-27, 2014 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-25020231

RESUMO

The major allergen, Cry j 1, was isolated from Japanese cedar Cryptomeria japonica (Cry j) pollen and was shown to react with immunoglobulin E antibodies in the sera from pollinosis patients. We previously reported that the frequency of HLA-DP5 was significantly higher in pollinosis patients and the immunodominant peptides from Cry j 1 bound to HLA-DP5 to activate Th2 cells. In the present study, we determined the crystal structure of the HLA-DP5 heterodimer in complex with a Cry j 1-derived nine-residue peptide, at 2.4Å resolution. The peptide-binding groove recognizes the minimal peptide with 10 hydrogen bonds, including those between the negatively charged P1 pocket and the Lys side chain at the first position in the peptide sequence. We confirmed that HLA-DP5 exhibits the same Cry j 1-binding mode in solution, through pull-down experiments using structure-based mutations of Cry j 1. We also identified the characteristic residues of HLA-DP5 that are responsible for the distinct properties of the groove, by comparing the structure of HLA-DP5 and the previously reported structures of HLA-DP2 in complexes with pDRA of the self-antigen. The comparison revealed that the HLA-DP5·pCry j 1 complex forms several hydrogen bond/salt bridge networks between the receptor and the antigen that were not observed in the HLA-DP2·pDRA complex. Evolutionary considerations have led us to conclude that HLA-DP5 and HLA-DP2 represent two major groups of the HLA-DP family, in which the properties of the P1 and P4 pockets have evolved and acquired the present ranges of epitope peptide-binding specificities.


Assuntos
Antígenos de Plantas/química , Cadeias alfa de HLA-DP/química , Cadeias beta de HLA-DP/química , Fragmentos de Peptídeos/química , Proteínas de Plantas/química , Animais , Sítios de Ligação , Cryptomeria/química , Cristalografia por Raios X , Cadeias alfa de HLA-DP/genética , Cadeias beta de HLA-DP/genética , Humanos , Ligação de Hidrogênio , Modelos Moleculares , Filogenia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Células Sf9 , Spodoptera
13.
Auris Nasus Larynx ; 41(4): 350-8, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24698163

RESUMO

OBJECTIVE: Among many immunotherapeutic approaches, oral immunotherapy (OIT) is thought to be an effective route for desensitization against a variety of allergens. However, there is little evidence that OIT is effective for airway allergic diseases such as pollen allergy. Thus, in the present study, we assessed the safety, efficacy and immune response of OIT using the Cry j1-galactomannan conjugate for Japanese cedar pollen allergy. METHODS: An open trial was conducted over a period of 4 months. The OIT group comprised of 23 subjects. Treatment was initiated 1 month before the estimated pollen season and continued for 1 month. The control group (the pharmacological treatment group without OIT) comprised of 11 subjects. The symptoms and medication score, levels of allergen-specific serum antibodies, cellular components of lymphocytes and cytokine production from peripheral blood mononuclear cells (PBMCs) were evaluated throughout the pollen season. RESULTS: The participants receiving OIT treatment showed significant improvements in total symptom scores and symptom-medication scores during the pollen season compared with the control group. The levels of allergen-specific serum IgG4 and IL-10 production in PBMCs were significantly increased in the OIT group compared with that in the control group. Importantly, no severe adverse effects were observed in the participants receiving OIT treatment. CONCLUSION: Short-term OIT using the Cry j1-galactomannan conjugate is effective, relatively safe and induces tolerant immune responses such as increased allergen-specific serum IgG4 and IL-10 production in PBMCs. These results suggest that OIT using allergen-galactomannan conjugates may provide a rapid, effective, and safe immunotherapy regimen for cedar pollen allergy.


Assuntos
Antígenos de Plantas/imunologia , Dessensibilização Imunológica/métodos , Mananas/imunologia , Proteínas de Plantas/imunologia , Rinite Alérgica Sazonal/terapia , Administração Oral , Adulto , Anticorpos/análise , Antígenos de Plantas/administração & dosagem , Antígenos de Plantas/efeitos adversos , Cryptomeria/imunologia , Citocinas/biossíntese , Feminino , Galactose/análogos & derivados , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Mananas/administração & dosagem , Mananas/efeitos adversos , Pessoa de Meia-Idade , Proteínas de Plantas/administração & dosagem , Proteínas de Plantas/efeitos adversos
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