Therapeutic Effects And Possible Mechanisms Of Artesunate On Lupus-prone MRL/lpr Mice

ObjectiveSystemic lupus erythematosus(SLE)is a serious chronic recurrent autoimmune disease with abnormal immune function.Autoimmune responses in lupus patients can attack multiple systems and organs.The most common desease is lupus nephritis(LN),which is a major risk factor for death in SLE patients.At present,the most classical SLE animal model in the world is MRL/lpr mice,which are used as the research object.Follicular helper T cells(Tfh)is a subpopulation closely related to CD4+ lymphocytes.Tfn can assist B cells to induce an effective humoral immune response.Studies have confirmed that Tfh cells are involved in the pathogenesis of various autoimmune diseases,including SLE.Several studies have also found that the Jak/Stat signaling pathway contributes to the differentiation of Tfh and cytokine response.Artesunate(ART)is one of the most effective and safe drugs for clinical treatment of falciparum malaria.Recent studies have found that ART can alleviate the lupus phenotype of lupus mice by regulating the body’s immune system,which means that ART has anti-inflammatory immunosuppressive effects and it may be beneficial for the treatment of autoimmune diseases.Based on the above reseach background,this study further explored the therapeutic effects of ART on MRL/lpr mice,and the effect of ART on Tfh cells and Tfr cells in the spleen of MRL/lpr mice.It may provide a theoretical basis for the application of ART in the adjuvant therapy of SLE in the future.Research methodsFemale BALB/c mice,which did not carry the lpr gene,were selected as normal controls.Female MRL/lpr mice were randomly divided into the blank vehicle solution-treated model group,the prednisolone-treated positive control group and 2.5 mg/kg and 5 mg/kg ART treatment group according to body weight and urine protein levels.Mice were treated at eight weeks of age,twice a day for eight weeks.After eight weeks,mice were anesthetized to collect the serum.The renal tissue was fixed at 4%paraformaldehyde and embedded in paraffin,and sections were prepared for hematoxylin and eosin(HE)staining and immunohistochemistry.The images were collected by a microscope.Urine protein concentration,urea nitrogen and creatinine levels in the serum of the mice were determined according to the corresponding instructions.Spleen cell suspension was obtained,then stained with Alexa Fluor 488 anti-mouse CD4,APC anti-mouse CD 185(CXCR5)and brilliant violet 421 anti-mouse FOXP3 antibody.The number of Tfh and Tfr cells were analyzed by flow cytometry.The expression levels of IL-6,IFN-y,IL-10 and IL-21 were detected by ELISA and qPCR.The expression levels of Statl,p-Statl,Stat3,p-Stat3,Jak2 and p-Jak2 in the kidneys of the mice were detected by Western blotting.ResultsART not only significantly prolonged the survival of MRL/lpr mice,but also delayed the occurrence of urinary protein,decreased the levels of serum urea nitrogen and creatinine.ART significantly attenuated renal damage and decreased the levels of anti-dsDNA 1gG antibodies deposited in the kidney.ART could inhibit the expression of MCP-1 in the kidney,and suppress the expresion of SLE-related inflammation factors IL-6,IFN-y,IL-21.Upon oral administration,ART significantly inhibited Tfh cell differentiation in the spleen of MRL/lpr mice and restored the radio of Tfr/Tfh.In addition,ART significantly inhibited the phosphorylation levels of Jak2 and Stat3 in the MRL/lpr mice.ConclusionsART significantly attenuated the lupus phenotype of MRL/lpr mice.ART might restore the normal ratio of Tfr/Tfh and inhibit the production of inflammatory cytokines by inhibiting the differentiation of Tfh cells and the Jak2-Stat3 signaling pathway in the MRL/lpr mice.Thus,the pathogenesis of MRL/Ipr mice was delayed.This study may provide a theoretical basis for ART to become an adjuvant therapy for SLE in the future.