CpG Oligodeoxynucleotides Attenuate OVA-Induced Allergic Airway Inflammation Via Suppressing JNK-Mediated Endoplasmic Reticulum Stress

Background Bronchial asthma is a chronic and heterogeneous disease of the lower respiratory tract involving various cells and cellular components such as eosinophils,mast cells,neutrophils and airway epithelial cells.Hyperresponsiveness and airway remodeling are typical features.As one of the serious public health problems,the incidence of asthma is increasing year by year.Although glucocorticoids and bronchodilators are the main treatment for asthma,there are still some patients who are insensitive to these treatments and their symptoms are not well controlled.In recent years,many studies have shown that allergen-induced endoplasmic reticulum（ER）stress plays an important role in the pathophysiological process of asthma.Among them,the increased expression of ER stress-related protein CHOP leads to apoptosis,XBP1 induces the production of inflammatory factors and mucus secretion-related factors,GRP78 and ATF6α further increase the transcription of ER stress-related proteins.In our previous study,it has been confirmed that CpG-ODN can reduce the occurrence of allergic airway inflammation.But does CpG-ODN play a protective role in the treatment of allergic asthma by regulating ER stress?ObjectiveOVA was used to induce allergic airway inflammation animal and cell models.We aimed to investigate the role of CpG-ODN in the treatment of allergic airway inflammation and its underlying molecular mechanism.MethodsFive-week-old C57BL/6J female mice were randomly grouped into two parts.Part 1comprised four groups（Control,CpG-ODN,OVA,OVA + CpG-ODN）;Part 2 comprised four groups（OVA,OVA + SP600125,OVA + CpG-ODN,OVA + CpG-ODN +SP600125）.In vitro experiments,the cultured RAW264.7 cells were grouped into two parts,Part 1 comprised four groups（Control,OVA,OVA + 1.5 μM CpG-ODN,OVA + 5μM CpG-ODN）;Part 2 comprised four groups（OVA,OVA + 10 μM SP600125、OVA +1.5 μM CpG-ODN、OVA + 10 μM SP600125 + 1.5 μM CpG-ODN）.The total number of cells in the bronchoalveolar lavage fluid were counted with a hemocytometer.The cellularity of BALF was classified and counted after Wright Giemsa staining.HE and PAS staining methods were applied to analyze airway inflammation,goblet cell hyperplasia and mucus secretion.Immunohistochemistry was used to analyze the infiltration of macrophages in lung tissues.The protein levels of IL-4,IL-5,IL-13,p-JNK,JNK,CHOP,XBP1,ATF6α and GRP78 in lung tissues were detected by Western blotting.Correspondingly,the ER stress markers were detected by Western blotting and immunofluorescence in RAW264.7 cells.ResultsIn OVA-induced allergic airway inflammation,CpG-ODN significantly suppressed inflammatory cells infiltration,goblet cell hyperplasia and the protein expression of Th2 cytokines.Moreover,OVA exposure strongly increased the activation of ER stress with higher protein expressions of CHOP,XBP1,ATF6α and GRP78.However,these OVAinduced increase of ER stress markers were markedly suppressed by CpG-ODN treatment.In addition,exposure to OVA significantly increased the phosphorylation of JNK,which was significantly reduced by CpG-ODN treatment.Remarkably,single treatment of SP600125,an antagonist of JNK,functioned similarly as CpG-ODN in mitigating allergic airway inflammation and suppressing OVA-induced activation of ER stress,however,no significant synergistic effect was evidenced by combined treatment of SP600125 and CpG-ODN.Furthermore,in OVA-stimulated RAW264.7 cells,we also found that OVA stimulation increased the expressions of ER stress markers,and CpG-ODN significantly reduced their expression levels via suppressing the phosphorylation of JNK.ConclusionThese results indicated that CpG-ODN mitigates allergic airway inflammation by inhibiting the activation of JNK-medicated ER stress.Our study further provides theoretical basis for CpG-ODN as an adjuvant therapy in allergic airway inflammation.