【摘要】 该文旨在研究复合菌培养物及其有效成分对脂多糖(LPS)诱导的小鼠RAW264.7细胞炎症反应的影响。实验分为7组,分别为对照组、LPS组、LPS+N-乙酰谷氨酸(NAG)组、LPS+花生四烯酸(ARA)组、LPS+牛磺酸(TAU)组、LPS+有效成分复合物(COMP)(NAG+ARA+TAU)组、LPS+复合菌培养物提取物(CME)组,利用色谱法测定NAG、ARA和TAU在复合菌培养物中的含量, ELISA、Griess、RT-q PCR法检测炎症因子的蛋白及mRNA的表达情况, Western blot法检测TLR4/NF-κB通路中关键蛋白的表达情况,细胞免疫荧光法检测转录因子NF-κB亚基P65入核情况。结果显示,NAG、ARA和TAU含量分别为193.60 mg/kg、4.28 mg/kg和129.70 mg/kg;与LPS组比较, LPS+NAG组、LPS+ARA组、LPS+TAU组、LPS+COMP组和LPS+CME组均不同程度地抑制了炎症因子一氧化氮(NO)、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白细胞介素-12(IL-12)以及趋化因子CXCL1的蛋白和mRNA的表达, COMP的效果优于CME;且各给药组均不同程度地降低了TLR4的表达水平以及P65和IκBα的磷酸化水平,同时抑制了P65的入核,其中LPS+COMP组的效果优于其他各组。结果表明, CME具有抗炎作用,而COMP的抗炎作用优于三种有效成分和CME;且COMP抑制LPS-Toll样受体4(TLR4)/核转录因子-κB(NF-κB)信号通路的效果优于CME。更多还原

【Abstract】 This study was conducted to investigate the effects of complex microbial culture and its active ingredients on LPS(lipopolysaccharide)-induced inflammatory response of RAW264.7 cells in mice. Seven groups were established: control group, LPS group, LPS+NAG(N-acetylglutamate) group, LPS+ARA(arachidonic acid)group, LPS+TAU(taurine) group, LPS+COMP(active ingredient complex)(NAG+ARA+TAU) group, and LPS+ CME(complex microbial extract) group. The content of NAG, ARA and TAU in the complex microbial culture was determined by chromatography. The levels of inflammatory factors were determined using the ELISA method and Griess method, and their mRNA expression was tested with the RT-qPCR method. The expression of key proteins in the TLR4(Toll-like receptor 4)/NF-κB(nuclear transcription factor-κB) pathway was tested by the Western blot assay. The cell immunofluorescence method was used to detect the nuclear translocation of the subunit P65 of the transcription factor NF-κB. The results showed that the contents of NAG, ARA and TAU were 193.60 mg/kg, 4.28 mg/kg and 129.70 mg/kg, respectively. Compared with the LPS group, the protein and mRNA expression levels of inflammatory factors, including NO(nitric oxide), IL-1β(interleukin-1β), IL-6, TNF-α(tumor necrosis factor-α), IL-12, and CXCL1 [chemokine(C-X-C motif) ligand 1], were inhibited in the LPS+NAG group, the LPS+ARA group, the LPS+TAU group, the LPS+COMP group and the LPS+CME group to different degrees. The effect of COMP was better than that of CME. In these groups, the expression of TLR4 and the phosphorylation levels of P65 and IκBα were reduced to varying degrees, and the nuclear translocation of P65 was also inhibited. The LPS+COMP group reported better effects than the other groups in respect of these indicators. The results revealed that CME had an anti-inflammatory effect, and COMP provided a superior anti-inflammatory effect to the three active ingredients and CME. COMP also inhibited the LPS-TLR4/NF-κB signaling pathway more effectively than CME.更多还原