为探讨高核苷酸酵母水解物对RAW264.7小鼠巨噬细胞免疫活性调节及其相关作用机制,采用脂多糖(1 μg/mL)刺激RAW264.7细胞来建立体外细胞炎症模型,以不同质量浓度的高核苷酸酵母水解物干预来明确其抗炎效果。ELISA法检测细胞培养上清中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)的含量,RT-PCR法检测相关mRNA表达水平,Western blot法检测细胞iNOS及胞核内核转录因子NF-κBp65蛋白水平。结果表明:高核苷酸酵母水解物作用RAW264.7细胞的安全范围≤150 μg/mL。与LPS模型组相比,质量浓度60~150 μg/mL的高核苷酸酵母水解物能显著增强RAW264.7吞噬能力,高核苷酸酵母水解物(60~150 μg/mL)能有效抑制脂多糖(lipopolysaccharide,LPS)诱导RAW264.7细胞释放NO、TNF-α、IL-1β和IL-6炎症因子及相关mRNA表达,降低iNOS及NF-κB蛋白水平。研究结果证实了高核苷酸酵母水解物对LPS刺激RAW264.7细胞炎症的保护作用,作用机制与NF-κB通路有关,为食疗干预慢性病提供一定的理论依据。
This study aimed to investigate the effects of high-nucleotide yeast hydrolysates on the immune activities of macrophages in RAW264.7 cells and the related mechanisms. In vitro cell inflammation model was created by stimulating RAW264.7 cells with 1 μg/mL lipopolysaccharide (LPS). Different concentrations of high-nucleotide yeast hydrolysates were used as interventions to determine their anti-inflammatory effects. ELISA was used to detect the contents of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β) in cell culture supernatant. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect related mRNAs, and Western blot was used to detect the levels of iNOS and nuclear transcription factor NF-κBp65 at protein levels in cells. The results showed that ≤150 μg/mL high nucleotide yeast hydrolysates were safe for RAW264.7 cells. Compared with the LPS model with no intervention, 60-150 μg/mL high-nucleotide yeast hydrolysates could significantly enhance the phagocytic ability of RAW264.7 cells, inhibit the releases of NO, TNF-α, IL-1β, and IL-6 inflammatory factors and their related mRNAs in LPS-induced RAW264.7 cells. Besides, iNOS and NF-κB decreased at protein levels. The results confirmed the protective effects of high nucleotide yeast hydrolysates on LPS-induced inflammation in RAW264.7 cells, and the mechanism was related to the NF-κB pathway, which provides a theoretical basis for therapeutic intervention by foods for chronic diseases.
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