Gremlin调节TGF-ß/Smad信号通路在慢性血栓栓塞性肺动脉高压血管重构中的作用

Gremlin is an antagonist of the bone morphogenetic protein(BMP), which belongs to transforming growth factor-β(TGF-β) superfamily. Gremlin directly binds with high affinity to BMP and blocks the TGF-β/Smad signal pathway. The balance between matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) plays an important role in the regulation of the ECM degradation and EnMT, which are crucial mechanisms of pulmonary vascular remodeling. Based on our previous work, we hypothesis that Gremlin can modify the MMPs/TIMPs balance, regulate the ECM degradation and EnMT, and impact the process of pulmonary vascular remodeling, via the blocking of TGF-β/Smad pathway. The present research is designed to explore the role of Gremlin and TGF-β/Smad pathway in the regulation of MMPs/TIMPs balance and EnMT by the overexpression and RNA interference of Gremlin in cultured cells and CTEPH animal models. Moreover, we will use clinic samples to identify the application of Gremlin/MMPs/TIMPs as biomarkers in the diagnosis and therapeutic monitoring. With this project, we hope to provide new biomarkers for screening and diagnosis, as well as the new interventional targets in clinical practice.

Gremlin是TGF-β超家族成员骨形成蛋白（BMP）的拮抗剂，通过调节TGF-β/Smad信号通路参与血管重构。我们研究发现，Gremlin在肺动脉平滑肌上有较高表达，影响细胞外基质（ECM）沉积和内皮细胞间质转化（EnMT），但Gremlin在慢性血栓栓塞性肺动脉高压(CTEPH)发病中的作用尚属未知。本研究在前期工作基础上：(1)细胞水平探讨Gremlin过表达和RNA干扰调节TGF-β/Smad信号通路、基质金属蛋白酶（MMPs）与内源性组织抑制因子（TIMPs）平衡,参与调节ECM降解；(2) 构建动物模型，应用Gremlin诱导剂和抗体干预，探讨其对EnMT和肺血管重构的影响；(3) 利用临床标本，探讨Gremlin/MMPs/TIMPs作为标志物在CTEPH治疗和预后监测中的价值。本研究将进一步加深对CTEPH发病机制的认识，并为临床筛选新的治疗靶点提供理论依据。

Gremlin-1在特发性肺动脉高压及低氧性肺动脉高压中具有重要作用。但Gremlin-1对慢性血栓栓塞性肺动脉高压 (CTEPH)的作用尚不明确。.本研究在临床中收集CTEPH患者血标本及血栓内膜剥脱术后组织，完善了100例CTEPH临床信息库和50例手术标本库建设，发现CTEPH患者肺组织和血清中Gremlin-1表达增加，CTEPH患者肺血管组织中存在EndMT现象。.细胞层面分离培养肺动脉平滑肌细胞（PASMCs）及肺动脉内皮细胞（PAECs），对细胞进行表型鉴定及基本细胞学行为检测，Gremlin-1可以促进PAECs发生EndMT；并可促进PAECs迁移及血管生成。BMP 7可以抑制Gremlin-1诱导的EndMT，并有效缓解Gremlin-1对BMP/Smad通路的抑制作用及Gremlin-1对 PAECs迁移及血管生成能力的促进作用。Gremlin-1刺激正常人及CTEPH患者PASMCs，可显著促进其增殖及迁移能力。BMP-7可有效缓解Gremlin-1对BMP/Smad通路的抑制作用及Gremlin-1对PASMCs增殖能力的促进作用。.CTEPH大鼠模型的建立探索，栓塞模型制备采用Sugen-5416联合自体血栓反复注入法，进一步优化模型建立。模型组大鼠肺动脉平均压有升高趋势，肺小动脉中膜厚度增加。.本研究首次明确Gremlin-1可促进PASMCs的增殖及迁移能力并可诱导PAECs发生 EndMT。且发现Gremlin-1可通过调控BMP/Smad通路促进PASMCs增殖及迁移并促进PAECs的EndMT，从而参与CTEPH的血管重构。提示Gremlin-1在CTEPH形成中具有重要的作用，靶向抑制Gremlin-1将有可能成为CTEPH治疗的新靶点。动物模型构建模拟临床CTEPH发生发展过程，优化CTEPH小动物模型建立方法，为CTEPH小动物模型构建提供了参考。