| Atopic dermatitis is a chronic, recurring, inflammatory dermatosis, which is T-cell mediated, characterized by intense pruritis and somnipathy, and accompanied by asthma,allergic rhinitis and conjunctivitis. Its pathogenesis is still unclear, but the immune anomaly of T-cell has been held to play a significant role in atopic dermatitis breakout,which, along with continuing skin lesion, has to do with T-cell overactivity and abnormal cytokine function.Costimulatory molecules are critical in T-cell overactivity. In the research, the cure by combining the receptors and ligands of costimulatory molecules can both prevent acute transplant immunity rejection response and maintain autoimmune inhibition. Herpesvirus entry mediator ligand, namely LIGHT or HVEML, is a newly-found member of tumor necrosis factor(TNF) receptor super-family, which can be detected in many inflammatory diseases and whose receptors are herpesvirus entry mediators or HVEM mainly exhibiting on activated T-cells.It has been proved by a study that LIGHT takes part in rheumatoid arthritis,ankylosing spondylitis and inflammatory bowel diseases, and the expressive level of AD patients’ peripheral blood serum, measurably related to the level of Ig E in serum and eczema area and severity index(EASI), is appreciably higher than that of the average.Accordingly, the thesis aims at analyzing the effects from LIGHT-HVEM blockage antibody or LTβR-Fc on the mouse model suffering ovalbumin-induced dermatitis in order to further explore how LIGHT-HVEM works on AD mechanism.Part I The expression of IFN-r, IL-4 and IL-5 on the mouse model of dermatitisObjective: To evaluate the expressions of IL-4,IL-5 and INF-γ in the skin lesionsand the splenocyte supernate of the mouse model with atopic dermatitis established by using the blockage antibody of LIGHT-HVEM or LTβR-Fc, and to analyze the effects of LIGHT-HVEM on the atopic dermatitis.Methods:Thirty BALB/c mice were randomly and evenly divided into 3 groups: the blank control group treated with 100 μl of sodium chloride physiological solution,the model group sensitized with 100 μl of sodium chloride physiological solution containing100 μg ovalbumin,and the blocker group firstly blocked with 100 μl of sodium chloride physiological solution containing 100 μg LTβR-Fc followed by sensitization with 100 μl of sodium chloride physiological solution containing 100 μg ovalbumin at 24 hours after the blocking. All the mice were sacrificed after the extraction of serum from their orbital cavities, tissue specimens obtained from skin lesions, and single cell suspensions of the spleen prepared. At first, RT-PCR was performed to detect m RNA expressions of interferon γ(IFN-γ), interleukin 4(IL-4) and IL-5 in murine lesions, ELISA to measure IFN-γ, IL-4 and IL-5 levels in culture supernate of murine splenocytes, as well as the total and ovalbumin-specific Ig E and Ig G1 levels in murine serum.Results : LTβR-Fc significantly inhibited the inflammatory response in the mouse model of dermatitis induced by ovalbumin. A significant fall was observed in the m RNA expressions of IL-4(0.88 ± 0.25 vs. 1.81 ± 0.25, P < 0.05), IL-5(0.75 ± 0.15 vs. 1.24 ±0.26, P < 0.05) and IFN-γ(0.62 ± 0.09 vs. 1.11 ± 0.19, P < 0.05) in murine lesions, and in supernate levels of IL-4(9.58 ± 1.44 ng/L vs. 20.12 ± 5.39 ng/L, P < 0.05), IL-5(11.37 ±2.02 ng/L vs. 22.77 ± 4.07 ng/L, P < 0.05) and IFN-γ(16 167 ± 950.40 ng/L vs. 23 930 ±44.20 ng/L, P < 0.05) in the blocker group compared with the model group.Conclusion:LTβR-Fc can measurably decrease the levels of IL-4,IL-5 and IFN-γ of the mouse model with ovalbumin-induced atopic dermatitis,which suggests that LIGHT-HVEM may serve as a potential target for the treatment of dermatitis(such as atopic dermatitis).Part II The expression of the total Ig E and Ig G1 and of ovalbumin-specific Ig Eand Ig G1 in the serum of the mouse model of dermatitisObjective: To evaluate the eczema area and severity index or EAST of the mouse model established by using the blockage antibody of LIGHT-HVEM or LTβR-Fc, and to identify the expressions of the total Ig E and Ig G1 and of ovalbumin-specific IgE and Ig G1 in the serum of the model with atopic dermatitis to analyze how LIGHT-HVEM works on atopic dermatitis mechanism.Methods:Thirty BALB/c mice were randomly and evenly divided into 3 groups: the blank control group treated with 100 μl of sodium chloride physiological solution, the model group sensitized with 100 μl of sodium chloride physiological solution containing100 μg ovalbumin,and the blocker group firstly blocked with 100 μl of sodium chloride physiological solution containing 100 μg LTβR-Fc followed by sensitization with 100 μl of sodium chloride physiological solution containing 100 μg ovalbumin at 24 hours after the blocking. Disease severity was evaluated by eczema area and severity index(EASI) score,and lesion size measured on days 0, 4, 8, 12, 15, 20, 23, 27, 31 and 34 after the first sensitization. A total of three sessions of sensitization were carried out. All the mice were sacrificed after the extraction of serum from their orbital cavities, tissue specimens obtained from skin lesions, and single cell suspensions of the spleen prepared. Finally,ELISA was performed to measure the expressions of ovalbumin-specific Ig E and of the total Ig E and Ig G1 in the serum of the mouse model with dermatitis.Results:LTβR-Fc considerably inhibited inflammatory response in the mouse model of dermatitis induced by ovalbumin. Compared with the model group, the blocker group showed significantly decreased lesion areas and EASI score(both P < 0.05). Although there being statistical difference of the total Ig E(32277.00 ± 407.53 vs.2744367.00 ±2025.64)and Ig G1(2323.33 ± 520.43 vs.1296.33 ± 32.72.64) in the serum between the control group with dermatitis and that with blocking agent, the ovalbumin-specific Ig E(0.84±0.11 vs.0.46±0.10) and Ig G1(0.87± 0.0.07 vs.0.62±0.10)in serum were still statistically meaningful.Conclusion:The EASI of the dermatitis model can be reduced by LTβR-Fc, so can the total both of Ig E and of Ig GI,and the ovalbumin-specific Ig E and Ig G1 in serum,suggesting that this signaling pathway may serve as a potential target for the treatment of dermatitis(such as atopic dermatitis). |
PDF Full Text Request