| Objective and significanceSystemic lupus erythematosus (SLE) is a diffuse connective tissue disease, which is mediated by autoimmune and considered as the prototype autoimmune disease characterized by many autoantibodies and causing multiple organ damage. SLE is widely distributed around the world, and the prevalence rate is 0.017-0.048% worldwide. An investigation in textile workers of Shanghai demonstrated the prevalence rate is 0.07%, and the prevalence rate of women is up to 1.13%, which is somewhat higher than western countries. SLE has high incidence in the young female, and the general age is 15-45 years old with a female:male ratio of 7-9:1.Kidney damage affected by SLE is called lupus nephritis (LN), which is the most common and serious manifestation of SLE. According to the clinical manifestations, renal involvement is more than 70% of patients, while almost all the patients had kidney damage in SLE if based on renal biopsy. The severity of renal disease directly impacts prognosis in SLE. Its clinical manifestations are complex and diversified, and change with different severity. The clinical manifestation is proteinuria, hematuria, urinary tube, or with experimental evidence of kidney damage and renal dysfunction, such as hypertension and edema. Some patients can progress to end-stage renal disease (ESRD), which is one of the major causes of death.Although the exact pathogenesis and mechanisms of LN are not clear, the common opinion is that the environments and endocrine disorders act on body with certain heredity factors, which induce immune function disorder, including the abnormal activation of T and B cells, dysregulation of cytokine network, complement activation and apoptosis, and produce many autoantibodies, such as antinuclear antibodies (ANA), anti-double-stranded DNA (dsDNA) antibodies and anti-Sm antibodies and immune complexes. These autoantibodies mainly aim for nuclear antigen composition, especially for nuclear DNA antigen. Antigen-antibody immune complexes deposit in the glomerular, and then activate complement, lead to inflammation cell infiltration, coagulable factor activation and inflammatory mediators release, result in kidney damage.Glucocorticoid, cytotoxic drugs and mang new menthods (such as biological agents, plasma exchange, stem cell transplantation, large doses of immunoglobulin shock therapy treatment) in modern medicine have been applied in the treatment of LN and shown good therapeutic effects. But many patients are often difficult to insist on the long-term treatment because of high recurrence rate, side effects and expensive prices of these drugs. Furthermore therapeutic strategy of simple intervention a certain taches are often difficult to achieve good effects due to the complexity and heterogeneity of SLE. Traditional chinese medicine (TCM) is a valuable medical resources in china. Clinical evidences have proven that TCM has overall adjustment functions in multiple places, many link and many targets, and good effects in enhancing therapeutic effects, improving prognosis, reducing the recurrence, maintaining long-term stablility of illness, and helping to stop using or reduce maintaining doses and side-effects of corticosteroid, which have extremely significant meaning in treating patients with SLE, who have a multiple gene regulation, complex pathogenesis and changeful conditions.Total glucosides of paeony (TGP) is officially approved for production listed as the first anti-inflammatory immunomodulatory drug in treatment of rheumatoid arthritis. TGP is an effective composition, extraceted from the dry roots of herbaceous peony, which is mainly a group of glycoside substances including paeoniflorin, hydroxy-paeoniflorin, paeonin, albiflorin, benzoylpaeonfiflodn and so on. More than 90% of total glucosides is paeoniflorin, which is the main effective composition. Pharmacology and clinical study found that TGP has good effects in anti-inflammatory, analgesic, immune regulation. In addition, it has few side-effect and good tolerance. In early and remission stage of some autoimmune diseases, TGP is a satisfactory choice for patients who are unfavorable to use powerful immunosuppressive or glucocorticoid. TGP has gradually applied in treating many autoimmune diseases such as SLE, ankylosing spondylitis and sjogren syndrome and shown the unique application value and good prospect in treating autoimmune diseases. But there are less clinical reports and no experimental reports about LN.MRL/lpr mice are the most commonly used spontaneous animal models to study SLE, which were successfully cultivated by Murphy and Roths in Jackson laboratory of United States in 1978. The mice contain Fas gene recessive mutant which is related to cell apoptosis, and appear lymphocyte proliferation genes. Thus T cells escape thymus negative selection and release into the peripheral organs, which produce spleen and lymph node enlargement, many antibodies,γ-globulin and immune complex correlation nephritis. These are very similar with human LN.This study will adopt MRL/lpr mice as experimental subjects. To observe the effect of TGP on urinary protein content, anti-dsDNA antibodies, ANA, renal pathological changes, spleen and thymus indexes and general conditions in MRL/lpr mice with LN, investigate the immune regulation mechanism of TGP and provide important theoretical and experimental evidence for further exploring its mechanism and clinical application.Methods and contents1. Grouping:To detect urinary protein content, anti-dsDNA antibodies and ANA of MRL/lpr mice after quarantining for a week. When anti-dsDNA antibodies and ANA were positive and urinary protein content was greater than or equal to 100mg·dl-1, model was established a successful model with LN. LN mice were randomized into model group and TGP group according to the number random list, 10 rats per group. According to《between human and animal body surface area ratio of equivalent dose conversion table》, TGP group was given TGP 234mg·kg-1 to intragastric administrationd once a day, for 30 consecutive days.2. Detection method:(1) Observed general conditions of mice befor and after administration, including mental state, hair, diet and activities.(2) Weighed body weight and collected urine at a fixed time before and 15,30 days after administration. Urinary protein content was detected using the method of coomassie brilliant blue.(3) Collected blood samples from tail veins before and 15 days after administration, and by removing the eyes 30 days after administration. The levels of antibodies were measured by enzyme-linked immunosorbent assay (ELISA).(4) Killed mice and removed spleen, thymus and kidey. Spleen and thymus wet weight were weighed, and spleen and thymus indexes were calculated. Renal pathology changes were observed using light microscope.3. Statistical analysis:Analyzing data used statistical software SPSS 13.0, and measurement data was indicated in the format of mean±standard deviation (X±S). Comparison of spleen and thymus wet weight and indexes used independent-samples t test. Comparison body weight, urinary protein content and antibodies level used ANOVA of repeated measurement data. Multiple comparisons used least significant difference method (LSD) test. The significance level was a=0.05.Results(1) General conditions:The hair was smooth and activity was sensitive. As time went by, The model group showed reduction in mental, appetite and activities, the hair was few, scattered and dull, and some hair in the neck is significantly loss, while the TGP group was lighter than model group. There was no death in the two groups.(2) Body weight:The body weight 30 days after administration was significantly higher than that of 15 days after administration in the TGP group (P<0.05), while the difference in the model group significantly decreased (P<0.05).(3) Spleen and thymus wet weight and indexes:The weight and index of spleen in the TGP group were significantly lower than that of model group (P<0.05), while the difference of thymus was no statistically significance (P>0.05).(4) Urinary protein content:Urinary protein content in the TGP group was significantly lower than that of model group 15,30 days after administration (P<0.05, P<0.01). At 15,30 days after administration, as compared with before administration, urinary protein content in the TGP group decreased significantly (P<0.05), while the difference in the model group was no statistically significant (P>0.05).(5) The level of anti-dsDNA antibody:The level of anti-dsDNA antibody in the TGP group was significantly lower than that of model group 30 days after administration (P<0.05). At 30 days after administration, as compared with before and 15 days after administration, the level of anti-dsDNA antibody in the TGP group decreased significantly(P<0.05). At 15,30 days after administration, as compared with before administration, the level of anti-dsDNA antibody in the model group increased significantly (P<0.05, P<0.01). (6) The level of ANA:The level of ANA in the TGP group was significantly lower than that of model group 30 days after administration (P<0.05). At 15,30 days after administration, as compared with before administration, the level of ANA in the TGP group was significantly decreased (P<0.05), while the difference in the model group was no statistically significant (P>0.05).(7) Renal pathology:It appeared that glomerular mesangial cells and endothelial cell proliferation, accompanied by inflammatory cell infiltration, basement membrane thickening slightly, some glomerular volume increasing, renal tubular epithelial cells swelling and cast were observed in the model group, while the renal pathology damage was eased obviously in the TGP group 30 days after administration.Conclusions1. TGP can improve the general conditions, increase the body weight and lighten the swollen spleen of MRL/lpr mice with LN, which show that TGP can improve the quality of life and has a protective effect on immune organ.2. TGP can reduce the levels of anti-dsDNA antibodies and ANA, which indicate that TGP has moderating effect on immune system. TGP may be through an access of immune network to inhibit the production of autoantibodies.3. TGP can reduce glomerular mesangial cells and endothelial cell proliferation, the glomerular inflammation and renal tubular injury, improve renal pathology damage and reduce urinary protein content. This shows that TGP has the role of delaying progress of the disease and protecting kidney.4. TGP has a certain effect on MRL/lpr mice with LN. As to underlying mechanism of its role, we will explore in future research. |
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