Ag85B DNA vaccine suppresses airway inflammation in a murine model of asthma

Jian Wu 1 2 Jun Xu 0 Chuang Cai 2 Xinglin Gao 1 Li Li 0 Nanshan Zhong 0 0 Guangzhou Institute of Respiratory Disease, First Affiliated Hospital of Guangzhou Medical College , Guangzhou 510120 , PR China 1 Department of Respiratory Disease, East District, Guangdong General Hospital, Guangdong Academy of Medical Science , Guangzhou 510080 , PR China 2 Department of Respiratory Disease, Peking University First Hospital , Beijing 100034 , PR China Background: In allergic asthma, Th2 lymphocytes are believed to play important roles in orchestrating airway eosinophilia and inflammation. Resetting the Th1/Th2 imbalance may have a therapeutic role in asthma. The mycobacterium tuberculosis 30-kilodalton major secretory protein (antigen 85B, Ag85B) can protect animals from M. tuberculosis infection by inducing a Th1-dominant response. Methods: In this study, the Ag85B gene was cloned into pMG plasmids to yield the pMG-Ag85B plasmid. The expression of Ag85B gene in murine bronchial epithelia cells was detected by Western blotting and immunohistochemical staining after intranasal immunization with reconstructed pMG-Ag85B plasmids. The protective effect of pMG-Ag85B plasmids immunization in airway inflammation was evaluated by histological examination and bronchoalveolar lavage (BAL). IL-4 and IFN-g levels in the BAL and supernatant from splenocyte culture were determined using ELISA kits. Results: The Ag85B gene was successfully expressed in murine bronchial epithelia cells by intranasal immunization with reconstructed pMG-Ag85B plasmids. Using a murine model of asthma induced by ovalbumin (OVA), pMG-Ag85B immunization significantly inhibited cellular infiltration across the airway epithelium with a 37% decrease in the total number of cells (9.6 2.6 105/ml vs. 15.2 3.0 105/ml, p < 0.05) and a 74% decrease in the number of eosinophils (1.4 0.2 105/ml vs. 5.4 1.1 105/ml, p < 0.01) compared with the OVA-sensitized control group. There was no difference in the number of neutrophils in BAL fluid between the pMG-Ag85B group, the OVA-sensitized control group and the empty pMG group. IL-4 production was significantly decreased in the BAL fluid (32.0 7.6 pg/ml vs. 130.8 32.6 pg/ml, p < 0.01) and in the splenocyte supernatant (5.1 1.6 pg/ml vs. 10.1 2.3 pg/ml, p < 0.05) in the pMG-Ag85B group compared with the OVA-sensitized control group, while IFN-g production was increased in the BAL fluid (137.9 25.6 pg/ml vs. 68.4 15.3 pg/ml, p < 0.05) and in the splenocyte supernatant (20.1 5.4 pg/ml vs. 11.3 3.2 pg/ml, p < 0.05). Conclusion: In a murine model of asthma induced by OVA, intranasal immunization with pMG-Ag85B significantly reduced allergic airway inflammation with less eosinophil infiltration. This protective effect was associated with decreased IL-4 and increased IFN-g production in the BAL fluid and in the supernatant of cultured splenocytes. - Background Allergic bronchial asthma is a complex syndrome characterized by airflow obstruction, bronchial hyper-responsiveness and airway inflammation [1]. Elevated levels of type 2 T cell cytokines such as IL-4, IL-5 and IL-13 are recognized as factors that initiate and accelerate allergic inflammation in asthma. These cytokines promote IgE synthesis, stimulate eosinophil growth and differentiation, and augment mucus production. In contrast, type 1 T cell cytokines such as IL-2, IFN-g and IL-12, initiate the clearance of viruses and other intracellular organisms by activating macrophages and cytotoxic T cells. The two subgroups of helper T cells are stimulated in response to different immunogenic stimuli and cytokines, and constitute an immune regulatory loop [2,3]. An imbalance between Th1 cells and Th2 cells plays an important role in the development of asthma [4]. Previous research revealed that Th2 cells could provoke airway inflammation with the restricted influence of IFN-g [5,6]. Therefore, a strategy of upregulating the Th1 immune response or downregulating the Th2 immune response may be valuable in the prophylaxis and management of bronchial asthma [7,8]. It has been hypothesized that the increased prevalence of atopy in developed countries may be associated with the declining prevalence of some infectious diseases such as tuberculosis [9]. Since Shirakawa [10] demonstrated an inverse association between exposure to mycobacteria and the subsequent development of atopy among Japanese school children, mycobacterium exposure and its relationship to asthma has gained increasing attention. Bacille Calmette-Gurin (BCG), a live attenuated Mycobacterium bovis, which is commonly used in many countries as a vaccine against human tuberculosis, has been shown to strongly induce a Th1-like response [11]. In a murine asthma model, intranasal administration of BCG suppressed airway eosinophilia, inflammation and airway hyper-responsiveness, and was accompanied by decreased Th2 cytokine levels in BAL fluid [6,12]. It has also been reported that the BCG vaccine had a (...truncated)