Immune Netw.  2013 Dec;13(6):295-300. 10.4110/in.2013.13.6.295.

Influence of the Adjuvants and Genetic Background on the Asthma Model Using Recombinant Der f 2 in Mice

• Chang YS
• Kim YK
• Jeon SG
• Kim SH
• Kim SS
• Park HW
• Min KU
• Kim YY
• Cho SH

• KMID: 2150791
• DOI: http://doi.org/10.4110/in.2013.13.6.295

Abstract

Der f 2 is the group 2 major allergen of a house dust mite (Dermatophagoides farinae) and its function has been recently suggested. To determine the optimal condition of sensitization to recombinant Der f 2 (rDer f 2) in murine model of asthma, we compared the effectiveness with different adjuvants in BALB/c and C57BL/6 mice. Mice from both strains sensitized with rDer f 2 by intraperitoneal injection or subcutaneous injection on days 1 and 14. The dosage was 20 microg. Freund's adjuvants with pertussis toxin (FP) or alum alone were used as adjuvants. On days 28, 29, and 30, mice were challenged intranasally with 0.1% rDer f 2. We evaluated airway hyperresponsivenss, eosinophil proportion in lung lavage, airway inflammation, and serum allergen specific antibody responses. Naive mice were used as controls. Airway hyperresponsiveness was increased in C57BL/6 with FP, and BALB/c with alum (PC200: 13.5+/-6.3, 13.2+/-6.7 vs. >50 mg/ml, p<0.05). The eosinophil proportion was increased in all groups; C57BL/6 with FP, BALB/c with FP, C57BL/6 with alum, BALB/c with alum (24.8+/-3.6, 20.3+/-10.3, 11.0+/-6.9, 5.7+/-2.8, vs. 0.0+/-0.0%, p<0.05). The serum allergen specific IgE levels were increased in C57BL/6 with FP or alum (OD: 0.8+/-1.4, 1.1+/-0.8, vs. 0.0+/-0.0). C57BL/6 mice were better responders to rDer f 2 and as for adjuvants, Freund's adjuvant with pertussis toxin was better.

Keyword

Allergy; Asthma; Rodent; House dust mite; Der f 2

MeSH Terms

Animals
Antibody Formation
Asthma*
Bronchoalveolar Lavage
Eosinophils
Freund's Adjuvant
Hypersensitivity
Immunoglobulin E
Inflammation
Injections, Intraperitoneal
Injections, Subcutaneous
Mice*
Pertussis Toxin
Pyroglyphidae
Rodentia
Freund's Adjuvant
Immunoglobulin E
Pertussis Toxin

Figure

• 

  Figure 1 Molecular cloning, functional expression and purification of recombinant Der f 2. (A) 12% SDS-PAGE analysis of the expressed Der f 2 allergen. M; molecular size marker, Lane 1, 2; homogenates from JM 109 transformed of pET-15b+Der f 2 cDNA in induced condions, Lane 3-5; purified recombinant Der f 2 allergens. (B) 12% SDS-PAGE and Western blot analysis of the expressed Der f 2 allergen. A; coomassie blue staining of SDS-PAGE gel. B; immunoblot with pooled patients sera sensitized against Dermatophagoides farinae.

• 

  Figure 2 Experimental design. Twenty µg of recombinant Der f 2 was injected for sensitization and boosting using Freund adjuvants with pertussis toxin (A) or alum (B) as adjuvants. On day 28, intranasal (IN) challenge was performed with 50 µl of 0.1% recombinant Der f 2 and for consecutive 3 days.

• 

  Figure 3 Evaluation of asthma phenotypes. (A) Airway hyperresponsiveness measured using whole body plethysmograph, (B) airway eosinophilia from bronchioalveolar lavage fluid, (C) Serum anti-recombinant Der f 2 (rDer f2) specific IgE measured by ELISA. Each bar indicates the mean (±SEM) value. *p<0.05, compared with the control groups. BALF, bronchoalveolar lavage fluid.

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