白血病抑制因子ceRNA调控网络在周围神经损伤再生中的作用机制研究

Peripheral nerve injury is a common clinical problem. However, its clinical effect is far from satisfactory. Gaining a better understanding of the molecular mechanisms underlying peripheral nerve injury will largely contribute to the development of clinical treatments. In the current study, we analyzed outcomes from deep sequencing by using bioinformatic tools and constructed the competitive endogenous RNA (ceRNA) network based on the regulatory characteristics of long non-coding RNAs (lncRNAs) and microRNAs (miRNAs). We then screened and verified ceRNA network that might be related with peripheral nerve repair and regeneration and found that lncRNA XLOC_174535 could bind to miR-494-3p and regulate LIF expression. We also demonstrated that LIF could regulate the migration and myelination of Schwann cells. LIF plays important roles in the central nervous system. But its effect on the peripheral nerve system remains unrevealed. We will perform both in vitro and in vivo experiments to identify the effect of LIF ceRNA network on Schwann cells as well as peripheral nerve repair and regeneration. Moreover, by using chemical activators, we aim to illustrate the downstream signaling pathways of LIF ceRNA network. Taken together, our study will deepen the understanding of the molecular mechanisms regulating peripheral nerve regeneration, offer exciting prospects for the identification of new therapeutic targets, and provide some guidance for relevant research works.

周围神经损伤是临床常见病，其疗效尚不十分理想。为了加深对其分子机制的理解以更好地服务于临床治疗，基于深度测序和生物信息分析，根据长链非编码RNA（lncRNA）和microRNA（miRNA）的调控特点，我们构建了竞争性内源RNA（ceRNA）调控网络，并对可能与神经再生过程相关的ceRNA调控网络进行实验验证和筛选。前期结果显示lncRNA XLOC_174535竞争性结合miR-494-3p调控白血病抑制因子（LIF）的表达，而且LIF影响了施旺细胞的迁移和成髓鞘。LIF在中枢神经系统具有重要的调控作用，但在周围神经系统的作用尚不明确。本项目将结合体内外实验，探讨LIF ceRNA调控网络对施旺细胞及周围神经损伤再生的调控作用，阐述该调控网络发挥作用的下游通路。本项目旨在从新的角度深入探索周围神经再生的分子机制，并可能为神经损伤修复的治疗提供新的靶点，也能为其它领域的相关研究提供借鉴。

周围神经损伤是临床常见疾病，给患者带来极大痛苦，也造成巨大的社会和经济负担。从细胞和分子层面深入探索周围神经损伤的内在机制，有利于寻求神经再生干预治疗的分子药物或靶标。.本研究通过分析大鼠坐骨神经损伤后的基因测序表达谱，筛选了显著差异表达的编码和非编码RNA，发现白血病抑制因子（LIF）富集于多个显著生物学过程，并根据LIF和其上游非编码RNA的表达相关性和结合位点构建了lncRNA XLOC_174535/miR-494-3p/LIF的ceRNA调控网络。基因和蛋白表达验证结果表明大鼠坐骨神经损伤后，LIF在神经损伤处显著上调，提示LIF可能影响施旺细胞这一坐骨神经处的主要细胞类型。细胞实验表明，施旺细胞转染LIF-siRNA后，其增殖迁移能力显著升高，而转染LIF过表达慢病毒后，增殖迁移能力降低，表明LIF影响施旺细胞表型。与细胞实验结果一致，体内注射LIF-siRNA后，施旺细胞的增殖迁移能力显著增加。形态学观察表明敲降LIF也促进了轴突的生长、髓鞘碎片的清除和新生髓鞘的包绕。足迹观察、电生理测量和痛觉检测表明敲降LIF加速了受损坐骨神经的运动和感觉功能的恢复。对于转染LIF-siRNA、LIF过表达慢病毒或相应对照的施旺细胞测序结果表明，LIF影响施旺细胞的代谢过程且抑制ERFE这一调控糖脂代谢基因的表达。进一步研究发现与敲降LIF相反，敲降ERFE抑制了施旺细胞的增殖和迁移能力，表明LIF可通过负向调控ERFE的表达影响施旺细胞表型。.综上，本研究挖掘了LIF这一周围神经损伤后表达发现显著改变的基因，检测了LIF对施旺细胞功能和周围神经损伤修复的影响，并阐明了LIF的上下游调控分子。本研究加深了对于周围神经损伤分子机制的理解，探讨了治疗周围神经损伤的潜在靶点，对优化周围神经损伤再生的治疗具有重要科学意义和临床应用价值。