长链非编码RNA H19促进脑缺血再灌注损伤的机制研究

Ischemic stroke is one of the leading causes of human death and disability around the world. Thrombolytic therapy is the only accepted treatment in clinic so far for ischemic stroke due to its complex pathogenesis and long duration. Thus fully understanding of the epigenetic molecular mechanism of ischemic stroke is of vital importance for the prevention and treatment of this disease. We first investigated the role of H19 gene polymorphism and the expression of lncRNA H19 in cerebral Ischemia/Reperfusion(I/R) injury, and found that the expression of lncRNA H19 was upregulated by cerebral I/R in rats, as well as by oxygen/glucose deprivation/reperfusion(OGD/R) in vitro. lncRNA H19 also takes part in the regulation of autophagy and microglia polarization in the process of I/R injury. However, the exact epigenetic mechanism remains unclear. In this project, we aim to investigate whether lncRNA H19 functions as a competing endogenous RNA (ceRNA) in the modulation of autophagy-associated gene 12 (Atg12) expression and TLR4 signaling through decreasing miR-200b activity in cerebral I/R and OGD/R cells. This study may elucidate the important role of lncRNA H19 in neuronal autophage and inflammation of ischemia reperfusion injury, which not only provides a new idea for the study of mechanism of ischemic stroke, but also provides a new target and strategy for the drug therapy of ischemic stroke.

缺血性脑卒中是人类致死和致残的主要神经系统疾病，目前除溶栓外尚无有效的治疗方法。缺血性脑卒中发病机制复杂，因此充分掌握缺血性脑卒中的病理过程及其表观遗传分子调控机制是预防及治疗该病的关键。我们在前期研究中首次发现长链非编码RNA H19（lncRNA H19）参与缺血性脑卒中的发病且lncRNA H19可通过激活神经元自噬和小胶质细胞M1型极化诱导缺血再灌注损伤。为深入探究lncRNA H19调控缺血再灌注损伤的表观遗传机制，本课题将在完善前期研究的基础上采用基因修饰法联合缺血再灌注细胞模型和动物模型，探讨lncRNA H19是否通过竞争性抑制miR-200b调控Atg12和TLR4信号通路，进而促进神经元自噬和小胶质细胞炎症反应，从而阐明lncRNA H19在脑缺血再灌注损伤中的重要调控作用。这不仅为缺血性脑卒中发病机制的研究提供了新的思路，更为该病的临床治疗提供新的靶点。

缺血性脑卒中具有高发病率、高复发率、高致残率和高死亡率的特点。我国每年死于脑卒中的患者高达 190 多万，约占全球卒中死亡人数的三分之一。缺血性脑卒中现有的临床治疗手段有限且效果不佳，即使目前临床普遍认可的血管再通手段也因其治疗窗窄、可导致出血及再灌注损伤等问题，在应用中受到极大限制。该项目前期研究基于临床样本，揭示了lncRNA H19参与缺血缺氧所致神经损伤的病理生理过程，通过进一步开展基础研究探究lncRNA H19通过调控自噬相关通路、诱导小胶质细胞表型极化、抑制神经血管新生及阿司匹林抵抗多种途径介导缺血性脑卒中的神经功能损伤。重要研究结果如下：1.明确缺血性脑卒中过程中lncRNA H19通过诱导自噬体的形成激活自噬过程并减低神经元细胞活力；2.证实lncRNA H19参与调控小胶质细胞细胞活力及增殖，促进其M1表型转化及炎性因子的释放；3.证实神经元通过释放外泌体将lncRNA H19运输至星形胶质细胞并通过lncRNA H19/miR-18a/VEGF信号通路参与血脑屏障的破坏；4.证实lncRNA H19通过激活lncRNA H19/p53/Notch轴抑制神经血管新生；5. 明确lncRNA H19参与调控缺血性脑卒中患者阿司匹林抵抗，并以外泌体为载体激活H19/let-7a/IGF1R轴诱导缺血性脑卒中的发病。科学意义一：揭示缺血性脑卒中进展的新机制，首次发现了lncRNA H19是促进卒中进展的关键因子，可用于缺血性脑卒中的诊断以及病情严重程度的预判，为卒中治疗提供了新的药物研发靶点。科学意义二：国内首次以自噬为中心针对多靶点开发缺血性脑卒中临床治疗新策略。通过抑制lncRNA H19调控自噬减轻缺血缺氧所致神经损伤、促进卒中后神经功能恢复，显著降低缺血性脑卒中的临床致残及致死率，为缺血性脑卒中的治疗提供了新选择。