| Objective:1. Explore the effect of RNA components from lactobacillus metabolites on Toll like receptor pathways.2. Study the effect of RNA components from lactobacillus metabolites on the recovery of intestinal flora in unbalanced model mice.Methods:1. The effect of RNA components to Toll like receptor pathways on people colonic cancer cell lines HT-29 cell(1) Adjusted the concentration of HT-29 cell and divided HT-29 cell into three groups, negative control group, experiment group and positive control group. Then RNA components were put in experiment group,and CPG-ODN2006 were put in positive control group, culturing 48h.(2) Detecting the change of TLR9, NF-κBp65 by RT-PCR and assaying the change of IL-6 in culture of HT-29 by ELISA.2. The effect of RNA components on epithelial cells of intestinal mucosa in mice(1) We established mice model for dysbiosis of Intestinal flora by ceftriaxone, and then, to detect targets every 7 days by the secretive RNA 5mg/kg/day,7.5mg/kg/day, lOmg/kg/day for 3 days.(2) TLR9, TLR4, TLR2 and NF-κBp65 were detected by RT-PCR; LR9 and NF-κBp65 were detected by immunohistochemistry.(3) IL-6 was. detected by ELISA assay in whole blood; detection of cytokines changes.3. Intestinal flora was detected by analysis with caecum of mice Results:1. The experimental results of the effect of RNA components to Toll like receptor pathways on people colonic cancer cell lines HT-29 cell(1) There is effect with the expression of TLR9 and NF-κBp65 in HT 29 cell by RNA components. Compared with the control group, the expression of TLR9 in experiment group is slightly higher than the negative control group,and lower than positive control group.The expression of NF-κBp65 in experiment group is slightly higher than the negative control group, and lower than positive control group in HT-29 cells too.(2) There is effect with the expression of IL-6 in Cell cultures of HT 29 cell by RNA components.Compared with the control group, the expression of IL-6 in experiment group is higher than the negative control group (P<0.05),and lower than positive control group (P<0.05).2. The experimental results of the effect of RNA components on epithelial cells of intestinal mucosa in mice(1) There is effect with the expression of TLR9 in intestinal epithelial cells of mice. TLR9 expression is extremely low in the normal mice,and there is a sharply rise in unbalanced model mice.The expression of TLR9 is clearly drop tendency with RNA components. The expression of TLR9 came back with RNA components rise,and the time longer,the drop better in IECs. 21d of RNA components (10mg/kg/d group) was the best.The result of expression of protein level of TLR9 is the same with the mRNA level before,RNA of lOmg/kg/d group can put the expression of protein level of TLR9 down.(2) Compared with the normal mice, the expression of TLR4 is slightly rise in IECs of unbalanced model mice,and the expression of TLR4 came back by RNA components. The expression of TLR2 is obviously rise in IECs of unbalanced model mice,and is back down trend though the expression of TLR2 is higher compared with the normal mice by RNA components.(3) The expression of protein level and mRNA level of NF-KBp65 is extremely low in the normal mice,and there is a sharply rise in unbalanced model mice,and then the expression of NF-KBp65 is drop by RNA components 21d. (4) There is effect with the expression of IL-6 in blood of mice by RNA components.IL-6 were increased significantly compared with normal mice in mice plasma then IL-6 levels resumed trends significantly by the RNA components and returned to normal statistically (P<0.05).(5) There is effect with the appendix index of unbalanced model mice by RNA components. Compared with the normal mice, the appendix index is obviously rise of unbalanced model mice,and the appendix index came back by RNA components.And the appendix index is clearly drop with RNA components rise, and the time longer, the drop better.(6) Actobacillus return to normal levels the first statistically, all of the bacteria have returned to normal levels statistically (P<0.05) by the RNA components,but the yeast and the peptococcus.(7) There is effect with the blood tests by RNA components. expression of WBC is rising trend in nature resume group and RNA components can rein excessive elevated of WBC expression. Percentage came back constantly by RNA components,and returned to normal level nearly, percentage is avoid to rise excessively. Compared with the normal mice, PLT is obviously rise of unbalanced model mice,and the change trend is same in nature resume group and experiment group by RNA components.Conclusion:1. RNA components of Lactobacillus metabolites can control the TLR pathways.2. RNA components of Lactobacillus metabolites can restore advantage bacterium group in unbalanced model mice.3. RNA components of Lactobacillus metabolites can control Blood tests and IL-6. |
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