Study On Immune Intervention In Systemic Lupus Erythematosus Mice With Anti-CD137L Monoclonal Antibody

Objective To study the lymphocyte costimulatory signal change in systemic lupuserythematosus (SLE) mouse model induced by pristane, and to investigate impact ofblocking CD137/CD137L costimulatory signal on disease of systemic lupuserythematosus (SLE) mouse model in vivo. Method Firstly, systemic lupuserythematosus (SLE) mouse model induced by pristane was created, which6-8weeksfemale BALB/c mice were single intraperitoneal injected by pristane0.5ml, whilecontrol group mice were injected by PBS0.5ml. Secondly, systemic lupuserythematosus (SLE) mouse model was identified, and respectively urinary protein wastested by bromophenol blue indicator, and peripheral blood autoantibody (such asanti-dsDNA, anti-Sm/RNP) content was tested by ELISA, and pathological injury wasassessed through HE staining. Thirdly, peripheral blood and splenic lymphocyte wereisolated at first, and then CD28, CD40L, CD137molecules in T cell surface and CD40,CD137L molecules in B cell were detected through flow cytometry (FCM). Fourthly, totest the expression of costimulatory molecules CD28, CD40L, CD137in lung tissue ofmodel and control mice via immunohistochemistry analysis.Fifthly, semi-quantitativeRT-PCR was used to detect expression of TNF-α, IFN-γ in the spleen of mice. Sixthly,anti-CD137L monoclonal antibody was used to immunologic intervention, after3months pristane injection, mice were randomly divided into four groups, each groupwas respectively injected anti-CD137L monoclonal antibody0,10,50,100ug, andcontrol group were treated with normal saline, which is once a week and totally3times.Seventhly, biology and pathology identification of different doses treatment groups andcontrol group mice were appraised, respectively urinary protein was tested by albustix,and peripheral blood autoantibody (such as anti-dsDNA, anti-Sm/RNP) content wastested by ELISA, and pathological injury was assessed through HE staining. Eighthly,Quantitative PCR was used to detect TNF-α and IFN-γ expression in spleen of differentdoses treatment groups and control group. Result:1.Pristane can successfully induce thesystemic lupus erythematosus (SLE) mouse model. there was proteinuria began toappear in a month in model group and gradually increased over time, in the other hand, proteinuria in control group mice turned out to be negative. Anti-Sm/RNP andanti-dsDNA in serum of model mice began to be higher than control group in the thirdand fourth month,and then would be remaining high level until those mice were put todeath, but control group mice almost kept as usual. Joint swelling and facial erythemawere found in model mice,but no abnormal change in control group.④Those lungs andlivers, and kidneys of model group displayed pathological changes in different extent.2.FCM results indicated that peripheral blood lymphocyte surface costimulatorymolecules CD28up-expressed after first month and then gradually down-expressedeven lower than normal control group (P<0.01), expression of CD40L in T cell had nostatistical significance(P>0.05),CD40in B cell up-expressed after the third month anddown-expressed lower than normal control group after the sixth month (P<0.05), andcostimulatory molecules CD137/CD137L up-expressed at the third month and kept highlevel until those mice were put to death (P<0.05).Expression of peripheral bloodlymphocyte surface costimulatory molecules of control group mice had no statisticalsignificance. At the same time, lymphocyte surface costimulatory molecules CD28,CD40L/CD40, CD137/CD137L in spleen expressed almost same as in peripheral blood.3.Immunohistochemistry showed lung organ costimulatory molecules CD28, CD40Land CD137in model group were positive, and control group were nearly allnegative.4.The results of semi-quantitative RT-PCR showed that TNF-α and IFN-γ inmodel group mice expressed higher than control group.5.After intervention in vivo withanti-CD137L monoclonal antibody, part of the clinical symptoms and signs of lupusmice was improved,so as the pathological injury of livers and lungs and kidneys.Compared with the control group,the expression levels of TNF-α and IFN-γ mRNA inthe spleen of the model groups which were injected with anti-CD137L monoclonalantibody is lower. Conclusion:1. The costimulatory molecules of lupus mice expressabnormally,and the expression level of TNF-α and IFN-γ mRNA in the model group ishigher,this may be associated with the development of SLE disease.2.Interventiontreatment with anti-CD137L monoclonal antibody may be helpful for SLE disease,andprovide a new way to cure SLE.