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研究烟曲霉菌(AF)、脂多糖(LPS)能否引起角膜细胞的耐受反应,增强角膜对病原体刺激的防御功能。小剂量AF、LPS分别预处理永生化人角膜上皮细胞(THCE)和永生化人角膜基质细胞(THSF),20 h后给予大剂量AF或LPS再次刺激。于再次刺激后4 h收集细胞上清,酶联免疫吸附试验(ELISA)检测细胞因子IL-8的分泌,趋化实验检测LPS预处理引起THSF对PMN趋化的改变。于再次刺激后1 h收集细胞,实时RT-PCR检测细胞因子IL-8 mRNA和抗菌肽CCL20和LL-37 mRNA的表达。结果显示AF、LPS分别预处理THCE和THSF均可显著抑制IL-8的分泌。LPS预处理THSF抑制IL-8 mRNA的表达,增加CCL20和LL-37 mRNA的表达,并且抑制中性粒细胞(PMN)向角膜细胞的趋化。因此角膜细胞的耐受反应可避免过度的炎症反应,减轻角膜感染,在角膜炎症中起保护性作用。
To study whether Aspergillus fumigatus (AF) and lipopolysaccharide (LPS) can induce corneal cell tolerance and enhance the corneal defense against pathogen stimuli. Small doses of AF, LPS were pretreated immortalized human corneal epithelial cells (THCE) and immortalized human corneal stromal cells (THSF), 20 h after high-dose AF or LPS re-stimulation. Supernatants were harvested at 4 h after re-stimulation, and the secretion of IL-8 was detected by enzyme-linked immunosorbent assay (ELISA). Chemotaxis assay was used to detect the chemotactic changes of PMN induced by LPS pretreatment. Cells were collected 1 h after re-stimulation, and the expression of cytokines IL-8 mRNA and antimicrobial peptides CCL20 and LL-37 mRNA were detected by real-time RT-PCR. The results showed that AF, LPS pretreatment THCE and THSF can significantly inhibit IL-8 secretion. LPS preconditioning THSF inhibited the expression of IL-8 mRNA, increased the expression of CCL20 and LL-37 mRNA, and inhibited chemotaxis of neutrophils (PMNs) to corneal cells. Therefore, corneal tolerance reaction can avoid excessive inflammatory response, reduce corneal infection, play a protective role in corneal inflammation.