Foodborne and Waterborne Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Department of Cell and Molecular Biology, Pharmaceutical Sciences Branch, Islamic Azad University, Tehran, Iran.
Probiotics Antimicrob Proteins. 2017 Mar;9(1):56-63. doi: 10.1007/s12602-016-9234-x.
Probiotics are used as a treatment for different intestinal disorders. They confer health benefits by different ways. This study was aimed to investigate immunomodulatory effect of Bacillus probiotic spores on the production of lipopolysaccharide (LPS)-induced interleukin 8 (IL-8) in HT-29 intestinal epithelial cells. Differentiated intestinal epithelial cell line was used as a model for the study of colonization of purified spores (Bacillus subtilis (natto) and B. coagulans) and their anti-inflammatory effects. MTT assay and trypan blue staining were used for the detection of optimal concentration of the purified spores and LPS. Pre-treatment assay was done by treatment of the cells with the purified spores for 2 h, followed by challenges with LPS for 3 and 18 h. Post-treatment assay was done by initial treatment of the cells with LPS for 18 h, followed by the spores for 3 and 6 h. Levels of IL-8 secretion and its mRNA expression were measured by ELISA and relative Q real-time PCR. Our results showed similar rates of adherence to intestinal epithelial cells by the spore probiotics, while displaying no cytotoxic effect. In the pre-treatment assay, a significant decrease in IL-8, at both protein and mRNA levels, was measured for B. coagulans spores after the addition of LPS, which was higher than those observed for Bacillus subtilis (natto) spores. In the post-treatment assay, while Bacillus subtilis (but not B. coagulans) diminished the LPS-stimulated IL-8 levels after 3 h of incubation, the inhibitory effect was not constant. In conclusion, ability of Bacillus spore probiotics for adherence to intestinal epithelial cell and their anti-inflammatory effects, through interference with LPS/IL-8 signaling, was shown in this study. Further studies are needed to characterize responsible bacterial compounds associated with these effects.
益生菌被用作治疗不同肠道疾病的方法。它们通过不同的方式对健康有益。本研究旨在研究芽孢杆菌益生菌孢子对脂多糖（LPS）诱导的 HT-29 肠上皮细胞白细胞介素 8（IL-8）产生的免疫调节作用。分化的肠上皮细胞系被用作研究纯化孢子（枯草芽孢杆菌（纳豆）和凝结芽孢杆菌）定植及其抗炎作用的模型。MTT 测定法和台盼蓝染色法用于检测纯化孢子和 LPS 的最佳浓度。通过用纯化孢子处理细胞 2 小时来进行预处理实验，然后用 LPS 处理 3 和 18 小时。通过先用 LPS 处理细胞 18 小时，然后再用孢子处理 3 和 6 小时来进行后处理实验。通过 ELISA 和相对 Q 实时 PCR 测量 IL-8 分泌及其 mRNA 表达水平。我们的结果表明，两种孢子益生菌对肠上皮细胞的黏附率相似，同时没有细胞毒性作用。在预处理实验中，在添加 LPS 后，B. coagulans 孢子的 IL-8 蛋白和 mRNA 水平均显著降低，这比枯草芽孢杆菌（纳豆）孢子观察到的效果更高。在后处理实验中，虽然枯草芽孢杆菌（但不是凝结芽孢杆菌）在孵育 3 小时后降低了 LPS 刺激的 IL-8 水平，但抑制作用并不稳定。总之，本研究表明，芽孢杆菌孢子益生菌通过干扰 LPS/IL-8 信号通路具有黏附到肠上皮细胞的能力和抗炎作用。需要进一步的研究来表征与这些作用相关的负责细菌化合物。
