基于新型circ-PRKCB靶向miR-339-5p调控p66Shc在肠缺血再灌注多器官损伤中的作用及机制研究

Massive intestinal epithelial cell apoptosis is the key mechanism of intestinal ischemia/reperfusion (II/R) injury, which contributes to intestinal barrier dysfunction and multiple organ dysfunction syndrome (MODS) induced by II/R. Our previous studies found that p66Shc acts as a crucial mediator of intestinal epithelial cell apoptosis. Our recent findings indicated: 1) p66Shc can be post-transcriptionally inhibited by miR-339-5p, which is significantly down-regulated after II/R; 2) Bioinformatics algorithms reveal that a novel circRNA circ-PRKCB may compete with miR-339-5p endogenously and the level of circ-PRKCB is dramatically increased after II/R, which is negatively correlated with miR-339-5p. This ncRNA network’s coordinated regulation on p66shc may be involved in II/R-induced intestinal epithelial cell apoptosis. Therefore, we hypothesize that modulation of circ-PRKCB/miR-339-5p axis may inhibit the expression level of p66Shc, decrease the p66Shc-mediated intestinal epithelial cell apoptosis after II/R, reduce intestinal mucosal barrier damage, and as a result, alleviate remote organ injury. In this study, by utilizing genetic animal model and RNA intervention, we investigate the role and mechanism of circ-PRKCB/miR-339-5p-p66Shc signaling in II/R damage and intend to provide a novel therapeutic strategy for II/R-induced MODS

肠上皮细胞过度凋亡是肠缺血再灌注（II/R）导致肠屏障损伤、进而发生多器官衰竭的关键机制。我们研究发现，p66Shc是介导肠上皮细胞凋亡的重要枢纽；进一步发现，1）miR-339-5p可靶向抑制p66Shc，且在II/R后显著降低；2）生物信息学预测新型circ-PRKCB与miR-339-5p具备竞争结合关系，实验显示其在II/R后表达升高，与miR-339-5p负相关。肠上皮细胞凋亡可能与此非编码RNA网络协同调控p66Shc有关。据此提出假说：通过circ-PRKCB/miR-339-5p信号轴抑制p66Shc表达及活性，可减轻p66Shc介导的II/R肠上皮细胞凋亡，减轻肠黏膜屏障损伤，进而改善多器官损伤。本研究利用基因模式动物、RNA干预等技术，探讨circ-PRKCB靶向miR-339-5p调控p66Shc在II/R损伤中的作用及机制，为II/R多器官损伤提供新的防治策略。

肠上皮细胞过度凋亡是肠缺血再灌注（II/R）导致肠屏障损伤、进而发生多器官衰竭的关键机制。我们研究发现，p66Shc是介导肠上皮细胞凋亡的重要枢纽；前期研究发现，1）miR-339-5p可靶向抑制p66Shc，且在II/R后显著降低；2）生物信息学预测新型circ-PRKCB与miR-339-5p具备竞争结合关系，实验显示其在II/R后表达升高，与miR-339-5p负相关。肠上皮细胞凋亡可能与此非编码RNA网络协同调控p66Shc有关。据此提出假说：通过circ-PRKCB/miR-339-5p信号轴抑制p66Shc表达及活性，可减轻p66Shc介导的II/R肠上皮细胞凋亡，减轻肠黏膜屏障损伤，进而改善多器官损伤。本研究利用基因模式动物、RNA干预等技术，探讨circ-PRKCB靶向miR-339-5p调控p66Shc在II/R损伤中的作用及机制，为II/R多器官损伤提供新的防治策略。结果发现：（1） p66Shc在介导肠I/R损伤后的ROS积累中起关键作用。（2）miR-339-5p参与调节p66Shc的表达。（3）miR-339-5p通过靶向p66Shc体外抑制H/ R诱导的氧化应激。（4）miR-339-5p在体内改善肠道I/R损伤和氧化应激（5）circ-PRKCB能够与miR-339-5p结合。（6）circ-PRKCB通过靶向miR-339-5p调控p66Shc表达和H/ R诱导的氧化应激。（7）抑制circ-PRKCB可减轻I/R引起的肠道氧化损伤。（8） circ-PRKCB/miR-339-5p/p66Shc信号通路在缺血性肠道中发挥一定作用。该研究以 ncRNA 调控网络作为切入点研究 p66Shc 在 II/R 后的表达监管机制，不仅有助于完善 II/R 多器官损伤的发病机制，且对进一步临床干预和治疗 II/R 多器官损伤具有重要意义。新型环状 RNA circ-PRKCB/miR-339-5p 信号轴可能是临床干预 II/R 的重要靶点，为重症病人 II/R 多器官损伤靶向治疗提供重要的理论依据。