海洋链球菌次级代谢产物Streptochlorin对金黄色葡萄球菌诱发乳腺细胞氧化应激的保护作用  

The protective effects of Streptochlorin,marine Streptomyces sp.secondary metabolite,on oxidative stress of mammary cells induced by Staphylococcus aureus

作　　者：扶绍东 王子菁 王正磊 黄代钏 石皖荣 苗晋锋[1] 张明智 FU Shaodong;WANG Zijing;WANG Zhenglei;HUANG Daichuan;SHI Wanrong;MIAO Jinfeng;ZHANG Mingzhi(College of Veterinary Medicine,Nanjing Agricultural University,Nanjing 210095,China;College of Sciences,Nanjing Agricultural University,Nanjing 210095,China;The First Affiliated Hospital of Anhui Medical University,Hefei 230011,China)

机构地区：[1]南京农业大学动物医学院,江苏南京210095 [2]南京农业大学理学院,江苏南京210095 [3]安徽医科大学第一附属医院,安徽合肥230011

出　　处：《南京农业大学学报》2024年第3期522-529,共8页Journal of Nanjing Agricultural University

基　　金：国家自然科学基金项目(22177051,32061143045);中央高校基本业务费专项资金(KYCYXT2022010);四川省科技计划项目(2022YFN0068)。

摘　　要：[目的]本试验旨在探讨Streptochlorin对金黄色葡萄球菌(Staphylococcus aureus)感染引起小鼠乳腺上皮细胞氧化应激的影响及机制。[方法]采用海洋链球菌次级代谢产物Streptochlorin预处理小鼠乳腺上皮细胞指标检测(EpH4-Ev)2 h,适时加入S.aureus(MOI=10)共孵育4 h,收集样品;通过细胞活性检测、倒置荧光显微镜观察、生化指标检测、Western blot、RT-PCR等方法检测相关指标,并辅以计算机分子对接技术,明确Streptochlorin对S.aureus诱发乳腺上皮细胞氧化应激的影响及潜在机制。[结果]20μmol·L-1 Streptochlorin预处理能够明显改善S.aureus侵袭导致的乳腺上皮细胞排列紊乱、皱缩等形态改变,降低细胞损伤标志物乳酸脱氢酶(LDH)、N-乙酰-β-D-氨基葡萄糖苷酶(NAGase)的分泌以及肿瘤坏死因子(TNF-α)、白细胞介素1β(IL-1β)和白细胞介素6(IL-6)等炎性细胞因子的基因表达(P<0.05)。Streptochlorin预处理能显著降低乳腺上皮细胞胞内活性氧(ROS)水平,提高细胞总抗氧化能力(T-AOC)水平,减少丙二醛(MDA)水平(P<0.05)。Western blot检测以及分子对接结果显示,Streptochlorin可能通过Keap1-Nrf2信号通路促进下游分子血红素加氧酶1(HO-1)、NAD(P)H∶醌氧化还原酶1(NQO1)、过氧化氢酶(CAT)、谷胱甘肽-S-转移酶(GST)的基因表达发挥抗氧化作用(P<0.05)。[结论]Streptochlorin对S.aureus感染小鼠乳腺上皮细胞诱发的氧化应激和炎症反应具有显著的保护作用,这可能与Keap1-Nrf2信号通路的激活有关。[Objectives]The study aimed to explore the effect and mechanism of Streptochlorin on oxidative stress of mice mammary epithelial cells induced by Staphylococcus aureus infection.[Methods]Mice mammary epithelial cells(EpH4-Ev)were pretreated with Streptochlorin,then the S.aureus(MOI=10)was added for 4 h to collect samples to clarify the effect and underlying mechanism of Streptochlorin on oxidative stress induced by S.aureus in EpH4-Fv cells,the levels of related indicators were detected by cell viability test,inverted fluorescence microscope,biochemical test,Western blot,RT-qPCR,complemented by computer molecular docking technology.[Results]20μmol·L-1 Streptochlorin pretreatment was found to significantly ameliorate the morphological changes such as disorganization and wrinkling of EpH4-Ev cells caused by S.aureus invasion,and reduce the secretion of the cell supernatant damage markers lactate dehydrogenase(LDH)and N-acetyl-β-D-aminoglucosidase(NAGase)as well as the gene expression of classical inflammatory cytokines,such as tumor necrosis factor(TNF-α),interleukin 1β(IL-1β)and interleukin 6(IL-6)(P<0.05).Streptochlorin pretreatment significantly reduced intracellular reactive oxygen species(ROS)levels,increased total antioxidant capacity of cells(T-AOC)capacity,and decreased malondialdehyde(MDA)level in EpH4-Ev cells(P<0.05).Western blot and molecular docking results tentatively demonstrated that Streptochlorin might exert antioxidant effects through Keap1-Nrf2 signaling pathway and promote the gene expression of downstream molecules heme oxygenase 1(HO-1),NAD(P)H quinone oxidoreductase 1(NQO1),catalase(CAT),and glutathione-S-transferase(GST)(P<0.05).[Conclusions]Streptochlorin had significant protective effects on oxidative stress and inflammatory responses of EpH4-Ev cells induced by S.aureus infection,which might be associated with Keap1-Nrf2 pathway activation.

关 键 词：海洋链球菌 Streptochlorin 金黄色葡萄球菌 氧化应激 Keap1-Nrf2 

分 类 号：S852.2[农业科学—基础兽医学]