HLA-DQ在SAA系统免疫激活中作用机制的研究

In recent years, a large number of studies have shown that the severe aplastic anemia (SAA) is an immunological disease related with the hyperfunction of T lymphocyte, which destroyed the bone marrow hematopoietic cells of the patients. It is unclear why SAA patients can't identify their own hematopoietic cells and lead to self-attacks. MHCII molecules are expressed on the surface of the myeloid dendritic cell (mDC), which is an iconic molecule that identifies itself. In the study of the mDC proteomics of SAA patients, the expression of HLA-DQ on the surface of mDC increased. This topic proposed on the basis of previous work, used SAA mouse model and SAA patients as the research object, analyzed the expression of HLA-DQ of the SAA patients and its correlation with patients' immune status. At the same time, identified the gene mutation sites by the second generation sequencing technology. In vitro cell and mouse model of SAA, elucidated the effect and the mechanism of HLA-DQ on abnormal antigen presenting and activating T cell. And then we discussed the effects of immunosuppressive agents and MHCII monoclonal antibodies on HLA-DQ function. Through the above studies, we furtherly clarified the immunological pathogenesis of SAA and helped to open up new targeted therapy.

近年来大量研究显示重型再生障碍性贫血（SAA）为一种T淋巴细胞功能亢进损伤自身骨髓造血细胞的免疫性疾病。至于SAA患者为何不能识别自身造血细胞，而导致自我攻击，机制尚不明了。MHCII类分子表达于髓系树突细胞（mDC）表面，是机体识别自我和非我的标志性分子。申请人既往主要参与SAA免疫发病机制的研究，在对SAA患者mDC蛋白质组学的研究中发现，患者mDC细胞内HLA-DQ表达增高。本次课题拟在前期工作的基础上，以SAA患者及其小鼠模型为研究对象，对SAA患者体内HLA-DQ水平及其与患者免疫状态之间的关联性进行分析，同时进行二代测序，明确该基因发生突变的位点；建立体外细胞及小鼠模型，阐明HLA-DQ对mDC细胞异常抗原呈递并激活T细胞的影响及其分子机制；探讨免疫抑制剂及MHCII单抗对HLA-DQ功能的影响。通过上述研究进一步明晰SAA的免疫发病机制，为开辟新的靶向治疗提供帮助。

近年来大量研究显示重型再生障碍性贫血（SAA）为一种T淋巴细胞功能亢进损伤自身骨髓造血细胞的免疫性疾病。至于SAA患者为何不能识别自身造血细胞，而导致自我攻击，机制尚不明了。MHCII类分子表达于髓系树突细胞（mDC）表面，是机体识别自我和非我的标志性分子。研究结果提示SAA初治组和缓解组患者外周血mDC HLA-DQ表达率高于健康对照组；HLA-DQB1 mRNA相对表达量自然对数高于缓解组和健康人；WB检测显示SAA患者mDC内HLA-DQ表达水平高于健康人。SAA初治患者外周血mDC HLA-DQ表达率与外周血白细胞计数、中性粒细胞绝对值、网织红细胞百分比、CD4+T/CD8+T比值、Treg比例和骨髓粒、红系百分比均呈负相关，与骨髓淋巴系百分比和mDC上CD86表达率呈正相关，与外周血mDC比例呈正相关。HLA-DQB1*06:01:01在SAA频率低于健康人。HLA-DQ敲低组mDC增殖水平低于对照组；HLA-DQ敲低组mDC吞噬率低于对照组；HLA-DQ敲低组mDC表面CD86表达率低于对照组；HLA-DQ敲低组mDC总凋亡细胞比例高于对照组。HLA-DQ敲低组共培养上清IL-2浓度为低于对照组，IFN-γ浓度低于对照组，TNF-α浓度低于空白对照组。AA小鼠外周血mDC比例高于正常对照组，mDC表面CD40表达率高于正常对照组，mDC表面CD86表达率高于正常对照组。环孢素治疗组小鼠mDC表面CD40、CD86表达率下降、血常规逐渐恢复、生存期延长；左旋咪唑干预组mDC共刺激分子表达变化无统计学意义，左旋咪唑干预组小鼠血常规在第17天时，明显低于其他各组，但经左旋咪唑处理后存活下来的小鼠，血常规各系细胞随时间推移存在缓慢恢复。研究结果提示HLA-DQ在SAA免疫发病机制中的作用有很大潜在研究价值，可能成为探索新免疫治疗方法的突破口。