基于CD169表达水平的小鼠脾红髓巨噬细胞分型研究

Objective: Based on the expression level of sialoadhesin (Sn), namely CD169 molecule, in mouse splenic red pulp macrophages, to study the differences in gene expression profiles between CD169+ and CD169- red pulp macrophage subsets. Methods: Flow cytometry and immunofluorescence were used to detect the expression of CD169 in splenic red pulp macrophages of C57BL/6J wild-type (WT) mice, with CD169 knockout (CD169KO) mice as a negative control. The F4/80+ splenic red pulp macrophages were enriched and two subsets of CD169+ and CD169- were sorted out for RNA sequencing. The DESeq2 software was used to screen out differentially expressed genes with the condition of P < 0.05 and |log2FC| ≥ 1. Through the Kyoto Encyclopedia of Genes and Genomes (KEGG) functional analysis, the differential genes were classified according to the pathways or functions they participated in. Some of the differential genes were verified by quantitative real-time PCR (qPCR). Results: Flow cytometry and immunofluorescence confirmed that some red pulp macrophages expressed CD169. There were 485 differentially expressed genes in the CD169+ and CD169- subsets. Some genes related to mediating inflammation were highly expressed in the CD169- subset. Conclusion: CD169+ and CD169- red pulp macrophages have different transcriptional profiles, and the CD169- red pulp macrophage subset has more characteristics of M1-type macrophages.

目的·基于小鼠脾红髓巨噬细胞唾液酸黏附蛋白(sialoadhesin,Sn)即CD169分子的表达水平,研究CD169~+ 和CD169~- 红髓巨噬细胞亚群的基因表达谱差异。方法·采用流式细胞术和免疫荧光法检测C57BL/6J野生型(WT)小鼠脾红髓巨噬细胞中CD169的表达,以CD169敲除(CD169KO)小鼠为阴性对照。富集F4/80~+ 的脾红髓巨噬细胞并分选出CD169~+ 和CD169~- 2个亚群,进行RNA测序。利用DESeq2软件以P<0.05且|log_2FC| ≥ 1为条件筛选出差异表达基因,通过京都基因与基因组百科全书(Kyoto Encyclopedia of Genes,KEGG)功能分析,将差异基因按照参与的通路或功能进行分类。通过实时荧光定量PCR(quantitative realtime PCR,qPCR)验证部分差异基因。结果·流式细胞术和免疫荧光法证实部分红髓巨噬细胞表达CD169。CD169~+ 和CD169~- 亚群拥有485个差异表达基因。一些与介导炎症相关的基因在CD169~- 亚群中高表达。结论·CD169~+ 和CD169~- 红髓巨噬细胞有不同的转录谱,CD169~- 红髓巨噬细胞亚群具有更多M1型巨噬细胞的特征。