目的·探究免疫抑制性受体白细胞免疫球蛋白样受体亚家族B成员2（leukocyte immunoglobulin-like receptor subfamily B member 2，LILRB2）在新型冠状（新冠）病毒感染所导致的免疫细胞炎症因子释放中的作用及机制，为新冠病毒肺炎治疗提供潜在靶点。方法·收集包含刺突蛋白胞外段（S-ECD）的细胞上清液，分别使用Western blotting及流式细胞术检测上清液中蛋白表达情况及活性；通过流式细胞术及免疫共沉淀技术检测该上清液中S-ECD与LILRB2的结合；用刺突蛋白处理人单核细胞系THP1或人外周血单个核细胞（peripheral blood mononuclear cell，PBMC），24 h后收集细胞，使用实时定量PCR技术检测相关炎症因子基因mRNA水平改变，同时使用酶联免疫吸附法检测细胞培养液中白细胞介素6（interleukin-6，IL-6）及IL-1β含量；将siLILRB2通过Lipofectamine 3000试剂转染至人外周血CD33⁺髓系细胞中，24 h后使用流式细胞术检测基因干扰效果；在对照组及LILRB2敲低的CD33⁺髓系细胞中加入刺突蛋白培养24 h，使用酶联免疫吸附试剂盒检测细胞培养液中IL-6的含量变化。结果·实验成功构建可分泌具有生物活性的新冠病毒S-ECD的293T细胞转染体系；免疫共沉淀实验显示刺突蛋白与LILRB2蛋白存在相互作用，且流式细胞术结果提示刺突蛋白胞外段能够与细胞表面的LILRB2结合；与对照组相比，经刺突蛋白处理24 h的THP1细胞中IL-6、IL-8、精氨酸酶（arginase 1）及IL-2基因表达水平均有明显上调（均P<0.05）；PBMC经刺突蛋白处理24 h后，IL-6、转化生长因子-β（transforming growth factor-β，TGF-β）、IL-8、IL-10及IL-1β的mRNA水平均显著上升（均P<0.05）；酶联免疫吸附实验结果显示，在PBMC培养液中加入刺突蛋白能够提高上清液中IL-6及IL-1β的浓度（均P<0.05）；使用S-ECD或刺突蛋白处理CD33⁺髓系细胞，IL-1β及IL-6含量随之升高（均P<0.05）；并且过表达LILRB2的THP1细胞经刺突蛋白刺激后展现了更强的IL-6分泌能力（P<0.05）；流式细胞术检测结果显示，2条siRNA均能敲低CD33⁺髓系细胞中的LILRB2，且siLILRB2-1敲除效果更好；LILRB2缺失后，刺突蛋白则不能促进CD33⁺髓系细胞的IL-6分泌。结论·新冠病毒刺突蛋白通过与细胞表面分子LILRB2结合，引起髓系细胞释放IL-6、IL-1β等炎症因子，导致患者出现细胞因子释放综合征。

Objective ·To explore the possible roles of immune inhibitory receptor leukocyte immunoglobulin-like receptor subfamily B member 2 (LILRB2) in the immune inflammation after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, and provide a potential therapeutic way for the coronavirus disease 2019 (COVID-19). Methods ·The supernatants containing the extracellular domain of spike protein (S-ECD) were collected, and the detection of the protein expression and activity in the conditional medium by Western blotting and flow cytometric analysis was followed by. The binding of S-ECD with LILRB2 was measured by co-immunoprecipitation and flow cytometric analysis. The mRNA expression levels of several inflammation genes in a human mononuclear cell line (THP1) or peripheral blood mononuclear cells (PBMC) were measured after spike protein stimulation for 24 h by quantitative RT-PCR. The protein levels of interleukin-6 (IL-6) and interleukin-1β (IL-1β) in the conditional medium were examined by enzyme-linked immunosorbent assay (ELISA). The siLILRB2 was transferred into CD33⁺ myeloid cells purified from human peripheral blood with Lipofectamine 3000 reagents. The knockdown efficiency was detected 24 h after transfection by flow cytometric analysis. The difference in the protein levels of IL-6 between the control cells and LILRB2-knocked-down cells after spike protein treatment was evaluated by ELISA. Results ·The study established a transfection system with 293T cells by which the SARS-CoV-2 S-ECD could be secreted to supernatants with normal biological activities. The interaction and the binding of spike protein with LILRB2 were evaluated by a co-immunoprecipitation assay and flow cytometric analysis, respectively. The mRNA expression levels of IL-6, IL-8, arginase 1 and IL-2 in THP1 cells were significantly up-regulated 24 h after spike protein treatment compared to the control cells (all P<0.05). Consistently, the mRNA levels of IL-6, transforming growth factor-β(TGF-β), IL-8, IL-10 and IL-1β in PBMC were notably increased after spike protein stimulation (all P<0.05). In addition, spike protein could also induce the release of IL-6 and IL-1β in PBMC as measured by ELISA (all P<0.05). More importantly, spike protein was able to increase the secretion of IL-1β and IL-6 by CD33⁺ myeloid cells 24 h after treatment (both P<0.05). LILRB2-overexpressing THP1 cells produced more IL-6 24 h after treatment with spike protein than the control cells (P<0.05). Two siRNAs could efficiently down-regulate the expression of LILRB2 in CD33⁺ cells as evaluated by flow cytometric analysis. Consistently, spike protein had no effect on the IL-6 secretion to supernatant from LILRB2-knockdown CD33⁺ myeloid cells. Conclusion ·SARS-CoV-2 can induce cytokine release syndrome by inflammatory factors, such as IL-6 and IL-1β, released by myeloid cells through spike protein binding to LILRB2.