纳豆芽孢杆菌B4对Caco-2细胞活性、黏附与细胞因子分泌的影响

Effects of Bacillus subtilis var. natto B4 on Activity,Adhesion and Cytokine Secretion of Caco-2 Cells

本试验以Caco-2细胞为体外模型研究纳豆芽孢杆菌B4对其活性、黏附与细胞因子分泌的影响。采用台盼蓝排斥试验和乳酸脱氢酶细胞毒性检测研究纳豆芽孢杆菌B4对Caco-2细胞活性的影响;采用荧光标记法评价大肠杆菌K88和霍乱沙门氏菌的黏附性能,并通过竞争、排斥和置换试验检测纳豆芽孢杆菌B4对这2株病原菌黏附的阻断作用;采用酶联免疫试剂盒测定Caco-2细胞分泌的细胞因子含量。结果表明:1)纳豆芽孢杆菌B4对Caco-2细胞存活率无显著影响(P>0.05),且对其无毒害作用。2)纳豆芽孢杆菌B4对大肠杆菌K88和霍乱沙门氏菌黏附Caco-2细胞的黏附率无显著影响(P>0.05),且对这2株病原菌黏附Caco-2细胞无阻断作用。3)纳豆芽孢杆菌B4极显著提高了Caco-2细胞分泌促炎细胞因子增殖诱导配体含量(P<0.01),对白细胞介素-6和白细胞介素-8含量无显著影响(P>0.05),极显著降低了Caco-2细胞分泌抗炎细胞因子白细胞介素-10含量(P<0.01)。综上所述,纳豆芽孢杆菌B4对Caco-2细胞无毒性,对大肠杆菌K88和霍乱沙门氏菌黏附Caco-2细胞无阻断作用,可通过调节Caco-2细胞的细胞因子分泌发挥一定的免疫功能。

Using Caco-2 cells as an in vitro model to investigate the effects of Bacillus subtilis var. natto B4 on the activity, adhesion and cytokine secretion of Caco-2 cells. The trypan blue dye exclusion test and extracellu- lar lactate dehydrogenase activity determination were conducted to research the influence of Bacillus subtilis var. natto B4 on the activity of Caco-2 cells; the fluorescence labeling technology was used to investigate the adhesive properties of Escherichia coli K88 and Salmonella choleraesuis to Caco-2 cells. The competition, re- jection and displacement experiments were carried out to determine the blocking action of Bacillus subtilis var. natto B4 to the adhesion of the two pathogenic bacterial strains; the cytokines secreted by Caco-2 cells were measured by ELISA. The results showed as follows ： 1 ） Bacillus subtilis var. natto B4 did not significantly af- fect survival rate of Caco-2 cells （ P 〉 0.05 ） and had no cytotoxicity on Caco-2 cells. 2 ） Bacillus subtilis var. natto B4 had no significant effects on the adhesion rate of Escherichia coli I（88 and Salmonella choleraesuis to Caco-2 cells （ P 〉 0.05 ）, and had no blocking action on the adhesion of the two pathogenic bacterial strains to Caco-2 cells. 3 ） Bacillus subtilis var. natto B4 significantly increased the content of pro-inflammatory cytokine inducing ligand （ P 〈 0.01 ）, and had no significant influence on the contents of interleukin-6 and interleukin-8 （ P 〉 0.05 ）, whereas Bacillus subtilis var. natto B4 significantly reduced the content of anti-inflammatory cy- tokine interleukin-10 （P 〈0.01 ）. In conclusion, Bacillus subtilis var. natto B4 has no cytotoxicity against Caco-2 cells, and can modulate the cytokine secretion function of Caco-2 cells to induce some immune respon- ses rather than block the adhesion of Escherichia coli K88 and Salmonella choleraesuis to Caco-2 cells.