Page 23 - 南京医科大学学报自然科学版

第43卷第7期 南京医科大学学报（自然科学版）
2023年7月 Journal of Nanjing Medical University（Natural Sciences） ·909 ·

·基础研究·

cGAS⁃STING通路介导酸性去氧胆酸诱导人正常食管上皮细胞

炎症的机制研究

张丹萍，苏 鑫，朱 宏 *

南京医科大学第一附属医院消化内科，江苏 南京 210029

［摘 要］ 目的：探讨酸性去氧胆酸（deoxycholic acid，DCA）诱导人正常食管上皮细胞（human esophageal epithelial cell，HEEC）
线粒体DNA（mitochondrial DNA，mtDNA）损伤及释放与环岛苷酸⁃腺苷酸合酶⁃干扰素基因刺激蛋白（cyclic GMP⁃AMP synthase⁃
stimulation of interferon gene，cGAS⁃STING）通路在食管上皮炎症发生发展中的关联。方法：将HEEC分为对照组和酸性DCA处
理组。CCK⁃8 法检测细胞存活率；荧光显微镜及流式细胞术检测活性氧（reactive oxygen species，ROS）、线粒体活性氧（mito⁃
chondrial reactive oxygen species，mtROS）及线粒体膜电位（mitochondrial membrane potential，MMP）的变化；化学发光法检测三
磷酸腺苷（adenosine triphosphate，ATP）水平；透射电镜观察线粒体超微结构改变；RT⁃qPCR 检测 mtDNA 拷贝数变化；Western
blot 检测γH2AX、cGAS、STING、p⁃NF⁃κB p65 及 NF⁃κB p65 的蛋白表达水平；RT⁃qPCR 检测炎症因子白细胞介素（interleukin，
IL）⁃6及IL⁃1β的mRNA 表达水平。结果：酸性DCA 处理后细胞存活率呈剂量⁃时间依赖性降低；细胞内ROS及mtROS 产生增
多，同时MMP降低，ATP含量减少；与对照组相比，酸性DCA处理后γH2AX的表达水平升高；mtDNA释放入胞质，mtDNA拷贝
数减少；cGAS、STING 和 p⁃NF⁃κB p65 表达升高；炎症因子 IL⁃6 及 IL⁃1β表达升高；cGAS 抑制剂 RU.521 预处理抑制了 cGAS、
STING的表达水平及部分抑制了p⁃NF⁃κB p65的表达水平，炎症因子IL⁃6及IL⁃1β水平降低。结论：体外实验表明，酸性DCA诱
导HEEC线粒体功能障碍，mtDNA损伤及释放，介导HEEC炎症反应，其机制可能与cGAS⁃STING通路的激活有关。
［关键词］ 反流性食管炎；线粒体DNA；cGAS⁃STING通路
［中图分类号］ R571 ［文献标志码］ A ［文章编号］ 1007⁃4368（2023）07⁃909⁃08
doi：10.7655/NYDXBNS20230703

Mechanism of cGAS ⁃ STING pathway mediating acidic deoxycholic acid inducing
inflammation of human esophageal epithelial cell

ZHANG Danping，SU Xin，ZHU Hong *
Department of Gastroenterology，the First Affiliated Hospital of Nanjing Medical University，Nanjing 210029，China

［Abstract］ Objective：To investigate the relationship between acidic deoxycholic acid induced mitochondrial DNA（mtDNA）
damage and release of human esophageal epithelial cells and cGAS⁃STING pathway in the development of esophageal epithelial cell
inflammation. Methods：HEECs were divided into control group and acidic deoxycholic acid treatment group. The viability of cells was
measured by CCK⁃8 assay. Changes of reactive oxygen species，mitochondrial reactive oxygen species and mitochondrial membrane
potential were detected by fluorescence microscope and flow cytometry. ATP was detected by the luminometer. The ultrastructure of
mitochondria was observed by transmission electron microscope. The mtDNA copy number was evaluated by qPCR. The expressions of
γH2AX，cGAS，STING，p⁃NF⁃κB p65 and NF⁃κB p65 were detected by Western blotting. The mRNA expressions of inflammatory
cytokines IL ⁃ 6 and IL ⁃ 1β were detected by qPCR. Results：CCK ⁃ 8 assay showed that the viability of cells treated with acidic
deoxycholic acid decreased in a dose⁃time dependent manner. The production of intracellular ROS and mtROS increased，while MMP
and ATP decreased. Compared with the control group，the expression of γH2AX increased after acidic deoxycholic acid，mtDNA
released into the cytoplasm，mtDNA copy number reduced，the expressions of cGAS，STING and p⁃NF⁃κB p65 were increased，and the
expressions of inflammatory cytokines IL⁃6 and IL⁃1β were elevated. After pretreatment with cGAS inhibitor RU.521，the expression

［基金项目］ 江苏省高层次人才“六个一”科研项目（LGY2016010）
∗
通信作者（Corresponding author），E⁃mail：zhuhong@1059@126.com