人类白细胞抗原-B27相关性前葡萄膜炎患者差异基因的表达特征

Screening of key genes and inflammatory signalling pathway involved in the pathogenesis of HLA-B27-associated acute anterior uveitis by gene expression microarray

目的研究人类白细胞抗原（HLA）-B27相关性前葡萄膜炎患者外周血单核细胞炎症通路差异基因的表达特征。方法实验研究。抽取3例HLA-B27阳性前葡萄膜炎患者及2例健康对照者外周血，分离后获得的单核细胞经含霍乱弧菌的脂多糖刺激后提取RNA，使用基因表达谱芯片进行检测，实时荧光定量PCR验证芯片结果，并通过基因功能分析、KEGG数据库等生物信息分析技术对筛选出的差异表达的基因进行分析。结果HLA．B27阳性前葡萄膜炎患者的外周血单核细胞经脂多糖刺激后出现了1105个表达水平上调超过2倍的基因；而健康对照组只有25个。Geneontology聚类分析及信号转导通路分析结果显示，上调的基因主要参与蛋白转运、蛋白折叠，在炎症反应中起重要作用。脂多糖与Toll样受体4结合后激活了Toll样受体转导通路及霍乱弧菌感染相关通路，是位于整个炎症反应网络中较上游的转导通路。其中PIK3CA、PIK3CB、AKT3及MAPKl基因可能是炎症反应中较关键的基因。结论HLA-B27阳性前葡萄膜炎患者及健康对照者的外周血单核细胞经脂多糖刺激后差异表达的基因有显著的差异。Toll样受体转导通路在致病中起重要作用。

Objective To investigate the genes and signalling pathways located upstream of the inflammatory processes in human leukocyte antigen （HLA）-B27-associated acute anterior uveitis by gene expression microarray. Methods Experimental study. HLA-B27-positive and-negative monocytes isolated from human peripheral blood were stimulated with Vibrio cholera lipopolysaccharide （LPS）. Gene expression microarrays were used to identify the differentially expressed genes. Differentially expressed （DE） genes were testified by real-time PCR and analyzed by a series of bioinformatics-based techniques such as Gene Ontology, Kyoto Encyclopedia of Genes and Genomes. Results Gene expression microarray analysis revealed marked differences between HLA-B27-positive acute anterior uveitis（AAU） and HLA-B27-negative healthy control peripheral monocytes in the genes that were upregulated in response to LPS stimulation with 1105 genes and 25 genes respectively. Gene Ontology enrichment and pathway analysis indicated that genes participating in protein transport and folding were essential to the inflammatory process. The LPS receptor- Toll-like reeeptor（TLR）4 induced TLR signalling pathway and pathway related to Vihrio cholerae infection were located upstream of the network and contribute to the overall response. Among the DE genes ,PIK3CA, PIK3CB,AKT3, and MAPK1 might play critical roles in inflammation. Conclusions Equivalent LPS stimulation induces a different response in HLA-B27-positive peripheral monocytes compared to normal control,, suggesting that the TLR pathway is involved in the pathogenesis of HLA-B27-associated AAU.