树突状细胞内RNA编辑酶ADAR1调节移植免疫反应及其机制研究

The major problem of allotransplantation is immune rejection, and an important idea to solve this problem is immune tolerance induced by immature dendritic cell (imDC). But in vivo, imDC easily loses the ability to inducing tolerance and blows up immune rejection because of converting into mature dendritic cell (mDC). We have tried to maintain the immature state of imDC with IL-10 or GM-CSF and induce immune tolerance with exosome secreted by imDC, but all these efforts have limited effects on suppressing immune rejection after organ transplantation. So, the key scientific issues are clarifying the molecular mechanism of imDC conversion mDC and searching effective interventional targets. We found that the mRNA and protein expression level of ADAR1 increased significantly when imDC converted into mDC, which was induced by stimulation of the foreign antigens. ADAR1 expression was significantly higher in the receptor spleen and transplant liver in acute rejection in rat liver transplantation models, and this effect can be blocked by FK506. This suggests that ADAR1 may affect allograft rejection by mediating maturation of imDC. But there aren’t any reports about relationship between ADAR1 and DC. This subject is intended to regard ADAR1 as a research core, and carry out in vitro and in vivo studies to clarify the biological function of ADAR1 in the process of transformation from imDC to mDC and explore its possible mechanism of mediating immune rejection. Furthermore, siRNA technology will be used to cut ADAR1 expression in imDC, and treated transplant recipients with the exosome which is secreted by imDC. The effect of the combined treatment on suppressing allograft immune rejection will be observed and provide valuable intervention targets and intervention programs for clinical transplantation immunotherapy.

免疫排斥是器官移植的难题，利用imDC诱导免疫耐受是重要解决思路。但imDC易转化为mDC而失去诱导免疫耐受的能力，并放大免疫排斥。课题组曾用IL-10、GM-CSF等维持DC的未成熟状态，并以其分泌的exosome诱导免疫耐受，但效果有限。明确imDC向mDC转化的机制，寻找干预靶点是亟需解决的关键科学问题。前期研究发现体外imDC向mDC转化中，ADAR1的mRNA和蛋白表达升高；大鼠肝移植急性排斥模型中，ADAR1在受体脾脏和移植肝脏内表达显著高于对照组，且应用他克莫司后ADAR1表达和排斥反应均下调。这提示ADAR1可能介导imDC的成熟而影响排斥反应，但两者的关系及作用机制尚无报道。因此，本课题拟通过体内外研究，阐明ADAR1在imDC向mDC转化中的功能，探讨其调节免疫排斥的可能机制；调控其表达以阻断imDC成熟，再以分泌的exosome诱导免疫耐受，为防治免疫排斥提供新方法。

研究背景：免疫排斥是器官移植的难题，巨噬细胞是介导移植排斥的主要细胞，ADAR1分子在先天免疫应答中起着不可或缺的调控作用，其在巨噬细胞中的表达明显高于其他先天免疫细胞。探索ADAR1调控巨噬细胞功能的机制，可能为解决免疫排斥提供新的思路和靶点。研究内容：1.调控ADAR1表达对巨噬细胞极化的影响。2.调控ADAR1表达对参与巨噬细胞极化的相关miRNA的作用。3.ADAR1对miRNA-21前体的编辑作用。4.调控miRNA-21表达对FOXO1的作用和对巨噬细胞极化的影响。5.调控ADAR1表达对同种异体皮片移植免疫排斥的作用。重要结果：1.ADAR1促进RAW-264.7细胞向M2极化。2.ADAR1促进巨噬细胞向M2极化过程中主要通过调控miR-21功能体的生成产生作用。3.ADAR1通过对PRI-miR-21的加工，导致Pre-miR-21及miR-21成熟体减少。4.miR-21通过抑制FOXO1转录，抑制IL-10的产生，促进RAW-264.7细胞向M1极化。5..ADAR1通过调控巨噬细胞向M2极化促进小鼠皮片移植存活，减轻移植排斥反应。科学意义：首次发现并报道ADAR1对miRNA-21前体的编辑作用，首次发现并报道ADAR1通过调控miRNA成熟干预巨噬细胞极化。进一步明确了调控ADAR1表达减轻移植排斥反应。