Effects Of STAT6 Decoy Oligonucleotides On Airway Inflammation And Th1/Th2 Balance In Mice Models Of Asthma

Bronchial asthma is a chronic airway inflammatory disease,which is infiltrated with eosinophil (EOS),involved in many kinds of inflammatory mediators (including related cytokine, chemotactic factor and so on) causing the airway inflammation and the airway hyperreactivity (AHR).Among these,Th1/Th2 imbalance, especially in Th2 differentiation advantage is thought to lead to airway inflammation and airway hyperreactivity (AHR),which plays an important factor in occurring and development in asthma.The signal transduction and activator of transcription 6 (STAT6) is one of the initiating factors in Th1/Th2 imbalance and Th2 differentiation advantage,which causes high expressing interleukin -4 (Interleukin-4, IL-4), while decreases y-interferon (IFN-γ)'s expression,and causes many inflammatory cell inducing EOS and other cells to aggregate in airway,which leading to airway inflammation and airway hyperreactivity. This research aimed to observe the effect of STAT6 decoy Oligonucleotides on the EOS, WBC, LYM, and inflammatory cytokines IL-4, INF-γexpression in bronchoalveolar lavage fluid of asthmatic mice (BALF) and the lung tissue pathology, and to investigate the effect of STAT6 decoy ODN on bronchial airway inflammation. Methods:32 BALB-C female mice were randomly divided into four groups:control group (C), asthmatic group(A), STAT6 decoy asthmatic (D) groupand scrambled ODN asthma (S)group, there were eight mice in each group; The asthmatic models of BALB/C mice were established by immunized with ovalbumin(OVA), in which group A, D, S was sensitized by intraperitoneal injection in the same day,7th,14th days,and inhalation excited 21st to 28th days, group C with saline instead of OVA; STAT6 decoy ODN and scrambled decoy ODN were designed and synthesized, some of STAT6 decoy ODN and scrambled decoy ODN were marked with FITC(fluorescein isothiocyanate, FITC-ODN). Before the last challenge, some of mice in group D and S used FITC (fluorescein isothiocyanate, FITC) labeled decoy ODN and random ODN intranasal administrations. Then the STAT6 decoy ODN and scrambled ODN were respectively transfered to the group D and S by local intranasal administrations. Then mice were sacrificed to obtain specimens of the next day.The eosinophil, neutrophil, lymphocyte counts of bronchoalveolar lavage fluid (BALF)were made with Wright'S staining.The levels of IL-4 and IFN-γin BALF were detected by ELISA; The lungs were used for frozen section, the distribution of FITC-ODN in lung tissue Frozen sections was observed by fluorescence microscope; Pathological changes were observed by HE staining.The tissues can be assessed for general morphology and cellular infiltration,and the standard of this method discriminates around brochioles,blood vessels and the number of patchy cellular infiltration (ScoreO-3),then statistics.Results:1. The frozen sections of lung tissue of the two groups with FITC-ODN administration directly deserved in the fluorescence microscope, found that decoy FITC-ODN in mice lung tissue showed green fluorescence, a clear outline of the organizational structure especially more pronounced in the epithelial cells in the airway and inflammatory cells infiltrating under the airway, while the group S, although lung tissue of mice have the green fluorescence, but it was clutter and no organizational structure, as the performance of spontaneous fluorescence;2. The white blood cells, lymphocytes, eosinophils in the branehoalveolar lavage fluid of asthmatic group were significantly higher than those in the group C (P<0.01),and these cells in group D is lower than those in asthmatic group(P<0.01),while there was no significant differences (P>0.05)between the S group and asthmatic group; 3. IL-4 in BALF of asthmatic group was significantly higher than the control group (P<0.01), group D was significantly lower (P<0.01)than that in the asthma group,while there was no significant differences (P>0.05)between the group S and asthmatic group, IFN-y in asthmatic group was significantly lower than that in control group (P<0.01), between group D and asthma group, there are differences, group D was higher than that in asthmatic group (P<0.01), while the group S and asthmatic group were no significant difference (P> 0.05), the rate of INF-y/IL-4 in group D is higher than that in asthmatic group,there was significantly increased(P<0.01), while there was no significant differences (P>0.05)between the group S and asthmatic group;4. HE staining sections of lung tissue were used to observed:①Lung tissue around the bronchioles, blood vessels, and pulmonary interstitial tissue showed very small amount of inflammatory cells in group C.②Contrast to group C,the asthmatic group histological results showed extensive infiltration of inflammatory cells around bronchioles, blood vessels and pulmonary interstitial tissue, especially eosinophils.③Decoy ODN treated group:the scope of airway inflammation and cell infiltration was reduced comparing with asthmatic group, statistics was significant(P<0.01).④But the group S showed the extensive of inflammatory cells infiltration around bronchioles, blood vessels andpulmonary interstitial tissue was like the asthma group(P>0.05). Conclusion:STAT6 decoy ODN can transfer into lung tissues of mice by local intranasal administration;STAT6 decoy ODN can inhibit the asthmatic airway production of IL-4, rectify the Thl/Th2 cytokine imbalance in asthmatic airway, cause the immune response from the Th2 type to Thl-type reverse,therefore regulate the inflammation of airway. STAT6 decoy ODN has led to specificity rather than nucleic acid on the impact of asthma...