香烟烟雾诱导的巨噬细胞条件培养基通过mTORC1/p70S6K通路促进气道上皮细胞炎症反应OA
Cigarette smoke-induced macrophage-conditioned medium triggers airway epithelial cell inflammatory responses through the mTORC1/p70S6K pathway
目的 观察香烟烟雾提取物(CSE)诱导的巨噬细胞条件培养基对气道上皮细胞炎症反应的影响并探讨其机制.方法 用10％的CSE刺激佛波12-肉豆蔻酸酯13-乙酸酯(PMA)诱导分化的单核/巨噬细胞(THP-1)24 h,收集条件培养基,作为刺激物诱导人支气管上皮细胞(BEAS-2B).采用CCK-8法检测条件培养基对BEAS-2B细胞活力的影响;RT-qPCR法检测条件培养基对BEAS-2B细胞炎症因子mRNA表达的影响;ELISA检测条件培养基对BEAS-2B细胞炎症因子分泌的影响;免疫荧光染色法观察条件培养基对BEAS-2B细胞形态的影响;Western blot法检测条件培养基对BEAS-2B细胞哺乳动物雷帕霉素靶蛋白复合物1(mTOR)/p70核糖体蛋白S6激酶(p70S6K)通路相关蛋白表达的影响.结果 与0％组相比,6.25％、12.5％、25％条件培养基作用6、12、24、48 h对BEAS-2B细胞无明显毒性,50％、75％、100％条件培养基显著降低BEAS-2B细胞活力(P＜0.05,P＜0.01);6.25％、12.5％、25％的条件培养基显著升高BEAS-2B细胞的炎症因子IL-1β、IL-6、IL-8、TNF-α mRNA表达及分泌(P＜0.05,P＜0.01),对细胞形态无显著影响;12.5％、25％的条件培养基可显著升高 BEAS-2B 细胞 p-mTOR、p-p70S6K、p-GSK3β(Tyr216)、p-NF-κB p65 的表达(P＜0.05,P＜0.01);mTORC1抑制剂雷帕霉素可显著降低25％的条件培养基诱导BEAS-2B细胞中炎症因子mRNA表达.结论 CSE诱导的巨噬细胞条件培养基可通过激活mTORC1/p70S6K通路促进气道上皮细胞炎症反应.
Objective This study investigated the effects of cigarette smoke extract(CSE)-induced macrophage-conditioned medium on airway epithelial cells' inflammatory responses,and explored the underlying mechanisms involved.Methods Phorbol myristate acetate-induced differentiated monocytes and macrophages(THP-1)were cultured with 10％CSE for 24 h;the conditioned medium was collected to induce inflammatory responses in human bronchial epithelial cells(BEAS-2B).The effects of THP-1-conditioned medium on BEAS-2B cells viability were detected using the CCK-8 assay.The effects of conditioned medium on mRNA expression levels of inflammation-related factors in BEAS-2B cells were analyzed by RT-qPCR,and inflammatory cytokine secretion was measured with enzyme-linked immunosorbent assay.The effects of conditioned medium on BEAS-2B cells morphology were observed with immunofluorescence staining.The expression of proteins related to the mTOR/p70S6K pathway was examined by Western blot.Results BEAS-2B cells viability after treatment with 6.25％,12.5％,or 25％conditioned medium showed no significant changes after 6,12,24,or 48 h,while 50％,75％,and 100％induced significant reductions in cell viability(P＜0.05,P＜0.01).Conditioned medium at 6.25％,12.5％,and 25％significantly increased the mRNA expression and secretion of inflammatory factors interleukin(IL)-1β,IL-6,IL-8,and TNF-α in BEAS-2B cells(P＜0.05,P＜0.01),but had no significant effect on morphology.Levels of p-mTOR,p-p70S6K,p-GSK3β(Tyr216),and p-NF-KB p65 were significantly increased in conditioned medium-treated cells(P＜0.05,P＜0.01);the mTORC1 inhibitor rapamycin significantly reduced inflammatory factor mRNA expression in BEAS-2B cells induced by 25％conditioned medium.Conclusions CSE-induced macrophage-conditioned medium significantly induced airway epithelial cell inflammatory responses through activating the mTORC1/p70S6K pathway.
宋雨薇;王茜茜;杨博斐;秦燕勤
河南中医药大学,河南省中医药防治呼吸病重点实验室,呼吸疾病中医药防治省部共建协同创新中心,郑州 450046||河南中医药大学,中医药科学院,郑州 450046河南中医药大学,河南省中医药防治呼吸病重点实验室,呼吸疾病中医药防治省部共建协同创新中心,郑州 450046||河南中医药大学,中医药科学院,郑州 450046河南中医药大学,河南省中医药防治呼吸病重点实验室,呼吸疾病中医药防治省部共建协同创新中心,郑州 450046||河南中医药大学,中医药科学院,郑州 450046河南中医药大学,河南省中医药防治呼吸病重点实验室,呼吸疾病中医药防治省部共建协同创新中心,郑州 450046||河南中医药大学,中医药科学院,郑州 450046
医药卫生
慢性阻塞性肺疾病炎症反应气道上皮细胞巨噬细胞mTORC1/p70S6K通路
chronic obstructive pulmonary diseaseinflammatory responseairway epithelial cellsmacrophagesmTORC1/p70S6K pathway
《中国比较医学杂志》 2026 (8)
1-9,9
国家自然科学基金项目(82104662)河南省自然科学基金项目(252300420126)中华中医药学会青年人才托举工程项目(CACM-2024-QNRC2-B35)河南省科技攻关计划项目(222102310141).
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