摘要
目的:探讨寿胎丸调控miR-374c-5p/ATG12信号轴减轻滋养细胞自噬治疗原因不明复发性自然流产(URSA的作用与机制。方法:收集正常早孕妇女(NP)和URSA患者绒毛组织各30例,Western blot检测LC3Ⅱ/Ⅰ、P62及ATG12蛋白表达;qRT-PCR检测ATG12 mRNA及miR-374c-5p表达。调控HTR-8/SVneo细胞miR-374c-5p表达,观察其对ATG12表达及细胞自噬的影响。体外观察寿胎丸含药血清对HTR-8/SVneo细胞miR-374c-5p、ATG12表达及自噬的影响;建立URSA小鼠模型,随机分为对照组、寿胎丸组、寿胎丸+INC组及寿胎丸+miR-374c-5p inhibitor组,观察各组胚胎吸收率,检测胎盘组织miR-374c-5p、ATG12及自噬相关蛋白表达。结果:与NP组相比,URSA患者绒毛组织LC3Ⅱ/Ⅰ比值及ATG12表达显著升高,P62表达显著降低,miR-374c-5p表达显著降低且与ATG12呈显著负相关。上调miR-374c-5p表达可显著降低ATG12表达及LC3Ⅱ/Ⅰ比值,提高P62蛋白表达;下调miR-374c-5p表达作用相反。双荧光素酶报告基因证实miR-374c-5p可与ATG12 mRNA 3'UTR直接结合。寿胎丸含药血清作用后,HTR-8/SVneo细胞miR-374c-5p及P62表达显著升高,ATG12及LC3Ⅱ/Ⅰ比值显著下降。与对照组相比,寿胎丸组小鼠胚胎吸收率明显降低,胎盘组织miR-374c-5p及P62表达显著升高,ATG12表达及LC3Ⅱ/Ⅰ比值显著下降;miR-374c-5p inhibitor可拮抗寿胎丸的妊娠保护作用。结论:miR-374c-5p表达降低可促进ATG12介导的滋养细胞自噬参与URSA发生过程,寿胎丸可靶向调节miR-374c-5p/ATG12信号轴减轻滋养细胞自噬进而治疗URSA。
Objective:To investigate effect and mechanism of Shoutai Pill on regulating miR-374 c-5 p/ATG 12 signaling axis and reducing trophoblast autophagy in treatment of unexplained recurrent spontaneous abortion(URSA).Methods:30 villous tissues were collected respectively from normal early pregnant women(NP)and URSA patients.LC 3Ⅱ/Ⅰ,P 62 and ATG 12 protein expres⁃sions were detected by Western blot.ATG 12 mRNA and miR-374 c-5 p expressions were detected by qRT-PCR.miR-374 c-5 p expres⁃sion in HTR-8/SVneo cells was regulated,and its effects on ATG 12 expression and autophagy were observed.Regulatory effects of Shoutai Pill containing serum on expressions of miR-374 c-5 p and ATG 12,and autophagy of HTR-8/SVneo cells were observed in vitro.URSA mice model was constructed and randomly divided into control group,Shoutai Pill group,Shoutai Pill+INC group and Shoutai Pill+miR-374 c-5 p inhibitor group.Embryo absorption rate of each group was observed,and expressions of miR-374 c-5 p,ATG 12 and autophagy-related proteins in placenta were detected.Results:Compared with NP group,ratio of LC 3Ⅱ/Ⅰand ATG 12 expressions in villi of URSA patients was increased significantly,while expression of P 62 were decreased significantly,expression of miR-374 c-5 p was decreased markedly and negatively correlated with ATG 12.miR-374 c-5 p overexpression could significantly down-regulate ATG 12 expression and ratio of LC 3Ⅱ/Ⅰ,but increase expression of P 62 protein;while miR-374 c-5 p inhibition had opposite effect.Double luciferase reporter gene experiment confirmed that miR-374 c-5 p could directly bind to ATG 12 mRNA 3'UTR.After treatment of Shoutai Pill containing serum,expressions of miR-374 c-5 p and P 62 in HTR-8/SVneo cells were increased significantly,while ATG 12 expression and ratio of LC 3Ⅱ/Ⅰwere decreased significantly.Compared with control group,embryo absorption rate of Shoutai Pill group was decreased significantly,expressions of miR-374 c-5 p and P 62 were increased while ATG 12 and LC 3Ⅱ/Ⅰratio were de
作者
付笑笑
褚楚
黑国真
魏然
朱肖肖
张振
赵霖
郭强
许珂
李霞(指导)
FU Xiaoxiao;CHU Chu;HEI Guozhen;WEI Ran;ZHU Xiaoxiao;ZHANG Zhen;ZHAO Lin;GUO Qiang;XU Ke;LI Xia(Innovative Institute of Chinese Medicine and Pharmacy,Shandong University of Traditional Chinese Medicine,Jinan 250355,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2022年第3期358-364,共7页
Chinese Journal of Immunology
基金
国家自然科学基金面上项目(81873337)
山东省自然科学基金面上项目(ZR2019MH039)
山东省中央引导地方科技发展资金项目(YDZX20203700001407)
泰山学者青年专家项目(Tsqn201812125)
山东第一医科大学“学术提升计划”(2019RC005)
济南市科技局科研带头人工作室项目(2020GXRC050)
山东省医药卫生科技发展计划项目(202005021490)资助。
