| Toxoplasma gondii is an obligate, intracellular, parasitic protozoan that causes the disease Toxoplasmosis. Found worldwide, T. gondii is capable of infecting virtually all warm-blooded animals. Previous studies estimate that about5-20%of the Chinese population has been exposed to and may be chronically infected with T. gondii. Most infection generally produces no symptoms in healthy human adults. But the infection can cause serious and occasionally fatal illness in infants. Thus the infection of T. gondii has been taken as a part of ToRCH screen. IgG/IgM ELIS A for T. gondii is the most widely used test in China, and the result of IgM test has great value for clinical diagnosis and prognosis. In order to keep the accuracy of tests and run the external quality assessment program, QC material is essential. However, the traditional QC material for T. gondii ELIS A test, which usually derived from infected patients, has many flaws:the limited source, lot-to-lot deviation, ethical issue and potential infectiousness. Thus we prepared the anti-T. gondii human-mice chimeric IgM antibody by genetic engineering, and used this antibody as QC material in ELISA test, evaluating its stability and commonality. As expected, this kind of chimeric antibody had been prepared successfully, and had following characters:ⅰ. stability which kept stable for one month in37℃or room temperature, or for at least two months in4℃or-20℃; ⅱ. commonality that all four ELISA kits tested in this study could identify the chimeric IgM antibody; ⅲ. without infectiousness; ⅳ. ease processings. without ethical problems. In conclusion, the preparation of human-mice chimeric antibody described in this part not only shows a new way in T. gondii ELISA QC material production, but also has great value in research for QC material of other pathogens.Systemic lupus erythematosus (SLE) is a chronic, complex, and debilitating systemic autoimmune disease characterized by variable involvement of different organ systems. Aberrant activation of T and B lymphocytes and their subsequent production of inflammatory cytokines and autoantibodies have been shown to be one of the main disease signs in SLE patients. MicroRNA (miRNA) is a type of20～24nucleotide non-coding RNA, which can bind to the3’-untranslated region of target mRNA resulting in its degradation and/or translational suppression. The powerful gene-regulatory role of miRNAs is now well recognized, and some miRNA based therapeutic strategies have been recently introduced. A major obstacle in miRNA-related therapy is the availability of an effective delivery system, owing to the instability and anionic charge of miRNA. In this part, we first constructed a MS2viral like particles (VLPs)-based miRNAs delivery approach, and then utilized this method to transfer miR-146a into lupus-prone mice to explore the possibility that miR-146a may act therapeutically, suppressing the production of autoantibodies. It showed that the concentration of miR-146a kept a4-5-fold increase in vitro or rose about2folds in vivo (with a half-live of43hours) after the MS2-miR146a VLPs treatment. What’s more, the VLPs used in this study is easy to prepare, stable and with relative low toxicity. Treatment with MS2-miR146a VLPs also showed profound effects on lupus-prone mice, including an increased level of mature miR-146a, which led to a significant reduction in the expression of autoantibodies and total IgG. Remarkably, these mice also exhibited reduced levels of proinflammatory cytokines, including IFN-a, IL-1β, and IL-6. In addition, we proved that the TLR pathway was involved in this regulation. The innovativeness of this study is based on:ⅰ. our MS2VLPs-based delivery system is an effective, stable, hypotoxic and versatile approach in RNA interference; ⅱ. restoring the loss of miR-146a is effective in eliminating the production of autoantibodies and ameliorating SLE progression in lupus-prone mice. Thus, the induction of dysregulated miRNAs by an MS2VLP-based delivery system may lead to novel therapies. |
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