Preliminary study on the effect of acyl-homoserine lactones on macrophage model
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摘要: 酰化高丝氨酸内酯类（Acyl-homoserine lactones,AHLs）化合物是革兰氏阴性菌群体感应系统中最重要的一类信号分子,本文模拟淡水鱼革兰氏阴性腐败菌产生的5种主要AHLs（C4-HSL、C6-HSL、C8-HSL、C10-HSL和3OC12-HSL）。通过构建Ana-1和RAW 264.7两种巨噬细胞模型,筛选敏感识别特定AHLs的巨噬细胞株,为构建基于细胞传感的淡水鱼腐败菌的检测方法进行前期初步研究。结果表明,5种AHLs均能造成Ana-1和RAW 264.7细胞的活性降低;胞内Ca2+荧光检测发现,在C10-HSL的刺激作用下,Ana-1细胞的活性与胞内Ca2+水平变化呈现一定的相关性,由此推测C10-HSL具有诱导Ana-1细胞胞内Ca2+上升,进而引发细胞内钙离子探针的荧光反应。同时采用流式细胞术和电镜分析对Ana-1细胞识别C10-HSL的敏感性进行了验证,为构建基于Ana-1细胞的淡水鱼腐败菌信号分子C10-HSL的荧光传感检测方法提供了前期研究基础。Abstract: Acyl-homoserine lactones ( AHLs) compound is a class of the most important signaling molecule in quorum sensing system of gram-negative bacterium, this paper simulates 5 dominant AHLs ( C4-HSL, C6-HSL, C8-HSL, C10-HSL, and 3OC12-HSL) generated from gram negative bacteria in freshwater fish, and screens the macrophage strain with sensitive identification of specific AHLs by constructing two macrophage models of Ana-1 and RAW 264.7.The results showed that each of the 5 AHLs could cause the decrease of Ana-1 and RAW 264.7 cell activeness.Through the intracellular Ca (2 +) fluorescence detection, under C10-HSL stimulation, the activity of Ana-1 cells showed a certain correlation with intracellular Ca (2 +) , it was speculated that C10-HSL could induce the intracellular Ca (2 +) increase in Ana-1 cells, thus trigger the fluorescence reaction of intracellular Ca (2 +) probe. Meanwhile the sensitivity of Ana-1 cells C10-HSL was verified by flow cytometry and electron microscopy, which provided the basis and guidance for the construction of ANA-1 cell based fluorescent sensing method for detection of signaling molecule C10-HSL in freshwater fish spoilage organisms.
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