Effect of treatment with geraniol on ovalbumin-induced allergic asthma in mice.

BACKGROUND: Asthma, a complex highly prevalent airway disease, is a major public health problem for which current treatment options are inadequate. OBJECTIVE: To evaluate the antiasthma activity of geraniol and investigate its underlying molecular mechanisms. METHODS: In a standard experimental asthma model, Balb/c mice were sensitized with ovalbumin, treated with geraniol (100 or 200 mg/kg) or a vehicle control, during ovalbumin challenge. RESULTS: Treatment of ovalbumin-sensitized/challenged mice with geraniol significantly decreased airway hyperresponsiveness to inhaled methacholine. Geraniol treatment reduced eotaxin levels in bronchoalveolar lavage fluid and attenuated infiltration of eosinophils induced by ovalbumin. Geraniol treatment reduced TH2 cytokines (including interleukins 4, 5, and 13), increased TH1 cytokine interferon γ in bronchoalveolar lavage fluid, and reduced ovalbumin-specific IgE in serum. In addition, treatment of ovalbumin-sensitized/challenged mice with geraniol enhanced T-bet (TH1 response) messenger RNA expression and reduced GATA-3 (TH2 response) messenger RNA expression in lungs. Furthermore, treatment of ovalbumin -sensitized/challenged mice with geraniol further enhanced Nrf2 protein expression and activated Nrf2-directed antioxidant pathways, such as glutamate-cysteine ligase, superoxide dismutase, and glutathione S-transferase, and enhanced formation of reduced glutathione and reduced formation of malondialdehyde in lungs. CONCLUSION: Geraniol attenuated important features of allergic asthma in mice, possibly through the modulation of TH1/TH2 balance and activation the of Nrf2/antioxidant response element pathway.

Glutathione-S-transferase: a minor allergen in birch pollen due to limited release from hydrated pollen.

BACKGROUND: Recently, a protein homologous to glutathione-S-transferases (GST) was detected in prominent amounts in birch pollen by proteomic profiling. As members of the GST family are relevant allergens in mites, cockroach and fungi we investigated the allergenic relevance of GST from birch (bGST). METHODOLOGY: bGST was expressed in Escherichia coli, purified and characterized by mass spectrometry. Sera from 217 birch pollen-allergic patients were tested for IgE-reactivity to bGST by ELISA. The mediator-releasing activity of bGST was analysed with IgE-loaded rat basophil leukaemia cells (RBL) expressing human FcεRI. BALB/c mice were immunized with bGST or Bet v 1. Antibody and T cell responses to either protein were assessed. IgE-cross-reactivity between bGST with GST from house dust mite, Der p 8, was studied with murine and human sera in ELISA. The release kinetics of bGST and Bet v 1 from birch pollen were assessed in water, simulated lung fluid, 0.9% NaCl and PBS. Eluted proteins were quantified by ELISA and analysed by immunoblotting. PRINCIPLE FINDINGS: Only 13% of 217 birch pollen-allergic patients showed IgE-reactivity to bGST. In RBL assays bGST induced mediator release. Immunization of mice with bGST induced specific IgE and a Th2-dominated cellular immune response comparably to immunization with Bet v 1. bGST did not cross-react with Der p 8. In contrast to Bet v 1, only low amounts of bGST were released from pollen grains upon incubation in water and the different physiological solutions. CONCLUSION/SIGNIFICANCE: Although bGST is abundant in birch pollen, immunogenic in mice and able to induce mediator release from effector cells passively loaded with specific IgE, it is a minor allergen for birch pollen-allergic patients. We refer this discrepancy to its limited release from hydrated pollen. Hence, bGST is an example demonstrating that allergenicity depends mainly on rapid elution from airborne particles.

Vaccination of buffaloes with Fasciola gigantica recombinant glutathione S-transferase and fatty acid binding protein.

Fasciola gigantica, causative agent of tropical fasciolosis, inflicts substantial economic losses on the livestock industry, affecting severely buffalo productivity in the tropical countries. Very few vaccination trials with different target antigens against F. gigantica infection have been conducted in this host. Present study describes a vaccination trial in buffaloes with F. gigantica recombinant glutathione S-transferase and fatty acid binding protein. The two recombinant proteins were expressed in Escherichia coli and evaluated for their immunoprophylactic potential in buffalo calves, using montanide 70 M-VG, a mineral oil-based adjuvant, for delivering the antigens. Buffalo calves were distributed in three groups, with group I, II and III calves immunized with recombinant glutathione S-transferase, fatty acid binding protein and a cocktail of these two antigens, respectively. Immunization of the calves evoked a mixed IgG1 and IgG2 antibody response. Present vaccination trial in these animals achieved a maximum protection level of 35%, when the two antigens were used in combination. Eosinophils were measured in both immunized and non-immunized challenge control animals, which showed a steady increase in their count in response to immunization with both the antigens and infection with F. gigantica, respectively.

Recombinant glutathione-S-transferase a major allergen from Alternaria alternata for clinical use in allergy patients.

Recombinant proteins are used for vaccines, therapy and diagnosis of many diseases. Biological activity of these may differ from native counterpart and needs investigation. The present study aimed to compare recombinant (r) and native (n) glutathione-S-transferase (GST) from Alternaria alternata. Glutathione-S-transferase sequence showed an ORF of 696bp encoding 26-kDa protein with N-terminus conserved domain. Secondary structure of both forms was comparable with melting temperature of 57 and 59 degrees C, respectively. rGST and nGST showed similar enzymatic activity, allergenicity and potency by ELISA inhibition. Histamine release was comparable in 14/17 patients for both the GSTs. rGST and nGST induced proliferation in PBMC at different concentration. Cell supernatant revealed higher IL-4 and IL-5 levels with low levels of IFN-gamma. In summary, recombinant and native GST demonstrated similar physio-chemical, biological and immunological properties and induced comparable cell mediated and humoral response to be used for diagnosis and specific immunotherapy for the fungal allergy cases.

Immunogenicity of Borrelia burgdorferi arthritis-related protein.

Immunization against arthritis-related protein (Arp) elicits antibody in mice that resolves arthritis but is not protective against challenge with Borrelia burgdorferi. In mice immunized against Arp, an unrelated 37-kDa protein (P37-42), outer surface protein A (OspA), or glutathione S-transferase (GT) and then challenged by syringe or tick, only OspA conferred protection. Passive transfer of Arp antiserum into infected SCID mice induced arthritis resolution, but antisera to P37-42, OspA, GT, or six overlapping Arp peptide fragments did not. Results suggest that the arthritis-resolving immunogenicity is specific to Arp, but the relevant epitopes may be conformational.

[Expression and purification of p11 fusion protein in E. coli and preparation of antiserum against GST-p11].

OBJECTIVE: To obtain p11 fusion protein and prepare specific polyclonal antibody against p11. METHODS: A full-length human p11 gene was cloned into expression vectors, pGEX-4T-2 and pQE30, and transformed into E. coli. The expressed proteins were purified from lysates with Glutathione Sepharose 4B and the Ni-NTA agarose column, respectively. The purified GST-p11 was mixed with Freund's complete or incomplete adjuvant and immunized rabbits. RESULTS: A high level of expression of target proteins was detected after IPTG induction and purified proteins were obtained by affinity chromatography with Glutathione Sepharose 4B and the Ni-NTA agarose column, respectively. Western blotting analysis suggested that the polyclonal antibody can recognize 6xHis-p11 and GST protein. CONCLUSION: The antiserum against p11 prepared by prokaryotic expression of GST-p11 fusion protein has good specificity.