老龄反向调节血管内皮衰老和平滑肌增殖的分子协同和交互作用机制

Generally the proliferative potential of somatic cells decreases along with aging. The vascular wall, however, displays over-activity of smooth muscle cells (SMC) proliferation. Our previous findings showed the bidirectional regulation of Jag1 on endothelial cells (EC) senescence and SMC proliferation during aging process, which was possibly mediated by the soluble proteins including IGFBP1/ADAM19/CXCL1. We therefore hypothesized that the decreased Jag1 expression with aging results in the imbalanced paracrine of EC, and the soluble proteins which targeted to Jag1 exert synergetic and reciprocal regulations, and further mediated the abnormal vascular repair by regulating the classical senescence and proliferative pathways. By using the carotid artery injury model from the endothelial Jag1 conditional knockout mice and wild type aging mice, this study was designed to observe the dynamic expression pattern of the target paracrine proteins as well as its relation with the vascular lesions. In the EC-SMC co-culture system, the gene synergetic and molecular reciprocal regulations were studied by using methods involving loss or gain of gene function, GST-pull down and mass spectrometric analysis. Finally, the molecular mechanisms will be investigated to clarify its relationship with the classical vascular senescence and proliferative pathways. This study was hopeful to address the key point of vascular EC regeneration and SMC proliferation in vascular repair.

随年龄增长体细胞增殖能力下降，血管系统却出现平滑肌反常增殖。我们前期研究发现内皮Jag1降低是老龄反向调节血管内皮衰老和平滑肌增殖的重要原因，可能与其调节内皮分泌IGFBP1/ADAM19/CXCL1蛋白有关。以此为基础进一步提出科学假设：老龄时Jag1表达减少导致血管内皮分泌功能失衡，以Jag1为靶标的分泌蛋白IGFBP1/ADAM19/CXCL1间发挥协同和交互作用，进一步通过调节细胞衰老和增殖途径导致血管不良修复。分别建立血管内皮Jag1条件敲除和野生型老年小鼠颈动脉损伤模型，阐明筛选分泌蛋白水平与血管病变和老龄的关系；建立内皮-平滑肌共培养体系，通过基因获得或缺失、GST-pull down、质谱分析等方法分析分泌蛋白在调节内皮衰老和平滑肌增殖中的协同及交互机制，并从与经典细胞衰老增殖机制的关系探讨其分子机制。本研究旨在阐明血管修复过程中内皮再生和平滑肌增殖矛盾的关键问题。

随年龄增长体细胞增殖能力下降，血管系统却出现平滑肌反常增殖。我们在前期研究发现内皮Jag1降低反向调节血管内皮衰老和平滑肌增殖的现象，可能与其调节内皮分泌IGFBP1/ADAM19/CXCL1蛋白有关的基础上，进一步通过建立内皮Jag1条件敲除小鼠和不同年龄野生型小鼠颈动脉损伤模型，观察IGFBP1、CXCL1和ADAM19在老龄促进血管重构过程中的动态表达，发现内皮Jag1条件敲除小鼠循环IGFBP1水平降低，CXCL1增高，ADAM19变化不大；而野生型老年和幼年小鼠颈动脉结扎损伤模型中，外周血IGFBP1和CXCL1蛋白水平在老龄小鼠均较幼年小鼠表达增高，ADAM19水平无明显变化；补充外源IGFBP1能减轻老年和内皮Jag1条件敲除小鼠血管内膜和中膜增殖。内皮-平滑肌共培养研究发现老龄时内皮细胞Jag1减少破坏了内皮细胞稳态，导致内皮细胞由静止表型直接转换为衰老表型，分泌蛋白IGFBP1可能通过Akt途径保护传代和H2O2引起的血管内皮细胞衰老，CXCL1不影响内皮细胞衰老，但能促进共培养的平滑肌细胞增殖和迁移。进一步分析IGFBP1的影响因素和调节机制，发现IGFBP1不仅受年龄和内皮Jag1表达影响，还受静脉表型基因COUP-TFII调节，外源性IGFBP1能增强COUP-TFII保护血管内皮抵抗衰老的作用。在临床转化研究部分，以112例临床患者为研究对象，发现IGFBP1与老龄加重冠脉病变有关，循环 IGFBP1水平是冠脉病变严重程度的独立预测因素。研究结果提示在老龄相关血管病变形成中，Jag1调节的分泌蛋白IGFBP1主要影响血管内皮衰老、CXCL1主要影响平滑肌增殖；其中IGFBP1除受Jag1调节外，还受多重调节机制影响，可能是老龄相关血管重构的内源性保护因子，不仅可能作为老龄患者估测冠脉病变程度的循环标志物，也是血管衰老防治的一个潜在靶点。