Abstract:

Objective To investigate the effect of Der f 1 mRNA molecules for specific immunotherapy on murine model of asthma. Methods Fifty BALB/c mice were randomly divided into 5 groups： PBS group， Der f 1 sensitization group， Der f 1 specific immunotherapy（SIT） group， β-actin mRNA SIT group， and Der f 1 mRNA SIT group. On days 0， 7 and 14， mice in PBS group received PBS injection； mice in the other groups were intraperitoneally injected with 10 μg Der f 1. At day 21， the mice in the 4 experimental groups were challenged with a 30-min inhaled dose of Der f 1（100 μg/ml） for 7 successive days. Two weeks after the final sensitization， the mice in the above five groups were immunized by intradermal injection with PBS， 1 μg Der f 1， 10 μg Der f 1， 2 μg β-actin mRNA， and 2 μg Der f 1 mRNA， respectively for 3 times at one-week intervals. Two weeks after the last intradermal injection， all mice were sacrificed and bronchoalveolar lavage fluid （BALF） was collected. ELISA was performed to detect the levels of IFN-γ and IL-13 in BALF， the number of eosinophils in the BALF was recorded. Splenocytes were prepared， and cultured with Der f 1 allergen （10 μg/ml） for 72 h. Splenocytes of PBS group was cultured without Der f 1 allergen. The levels of IFN-γ and IL-13 in splenocyte culture supernatant were measured by ELISA， as well as serum antibody levels of total IgE， allergen-specific IgE （sIgE）， sIgG1， and sIgG2a. Lung sections were stained in hematoxylin and eosin, and observed under the microsope. Results Except for PBS group， mice in the other 4 group showed symptoms of acute asthma attack. Compared with Der f 1 sensitization group ［（897.56±105.73） pg/ml］ and β-actin mRNA SIT group ［（219.47±64.72） pg/ml］， the level of IFN-γ in BALF from Der f 1 mRNA SIT group ［（897.56±105.73） pg/ml］ and Der f 1 SIT group ［（864.48±70.62）pg/ml］ significantly increased（P<0.01）. However， the level of IL-13 in BALF from Der f 1 mRNA SIT group ［（241.64±31.41） pg/ml］ and Der f 1 SIT group ［（321.94±41.07）pg/ml］ was significantly lower than that of Der f 1 sensitization group ［（520.62±43.77） pg/ml］ and β-actin mRNA SIT group ［（507.22±42.26） pg/ml］（P<0.01）. The number of eosinophils in Der f 1 mRNA SIT group ［（1.33±0.44）×105/ml］ and Der f 1 SIT group ［（1.48±0.39）×105/ml］ was also lower than that of Der f 1 sensitization group ［（3.54±0.52）×105/ml］ and β-actin mRNA SIT group ［（2.98±0.53）×105/ml］（P<0.01）. The levels of IFN-γ and IL-13 in splenocyte culture supernatant showed that IFN-γ level in Der f 1 mRNA SIT group ［（420.91±69.92） pg/ml］ and Der f 1 SIT group ［（334.92±43.72） pg/ml］ was significantly higher than that of Der f 1 sensitization group［（123.75±15.48） pg/ml］ and β-actin mRNA SIT group［（128.84±59.00） pg/ml］ （P<0.01）. However， IL-13 level of Der f 1 mRNA SIT group ［（268.51±40.42） pg/ml］ and Der f 1 SIT group ［（285.26±62.21）pg/ml］ was significantly lower than that of Der f 1 sensitization group ［（613.89±51.54） pg/ml］ and β-actin mRNA SIT group ［（524.05±39.12） pg/ml］（P<0.01）. Compared with Der f 1 sensitization group ［total IgE： （94.34±11.66） ng/ml， sIgE： （65.67±9.47） ng/ml， sIgG1： （75.18±9.52） ng/ml， sIgG2a： （2.81±1.17） ng/ml］ and β-actin mRNA SIT group［total IgE： （86.48±10.26） ng/ml， sIgE： （62.36±8.35） ng/ml， sIgG1： （69.51±8.98） ng/ml， IgG2a： （1.06±0.11） ng/ml］， the serum antibody levels of total IgE ［（33.72±9.78） ng/ml］， sIgE ［（22.76±8.09） ng/ml］， sIgG1 ［（17.87±7.59） ng/ml］ of Der f 1 mRNA SIT group decreased significantly（P<0.01）， whereas the level of IgG2a ［（7.74±0.88） ng/ml］ increased（P<0.01）. Compared with Der f 1 sensitization group， the asthmatic symptoms were relieved after immunization with Der f 1 mRNA for specific immunotherapy， including intact structure of respiratory and alveolar epithelial cells， decreased inflammatory cell infiltration， and similar to those in Der f 1 SIT group. However， the breakage and detachment of bronchial epithelial cells occurred in β-actin mRNA SIT group. Conclusion Der f 1 mRNA vaccine can correct Th1 and Th2 imbalance.

Key words:

Dermatophagoides farinaemso-ascii-font-family: Calibri, mso-ascii-theme-font: minor-latin, mso-fareast-font-family: 宋体, mso-fareast-theme-font: minor-fareast, mso-hansi-font-family: Calibri, mso-hansi-theme-font: minor-latin">；Allergenmso-ascii-font-family: Calibri, mso-ascii-theme-font: minor-latin, mso-fareast-font-family: 宋体, mso-fareast-theme-font: minor-fareast, mso-hansi-font-family: Calibri, mso-hansi-theme-font: minor-latin">；mRNA vaccinemso-ascii-font-family: Calibri, mso-ascii-theme-font: minor-latin, mso-fareast-font-family: 宋体, mso-fareast-theme-font: minor-fareast, mso-hansi-font-family: Calibri, mso-hansi-theme-font: minor-latin">；Specific immunotherapy