PARs受体通路参与天然水蛭素改善大鼠超长随意皮瓣微循环的分子机制研究

Super-long random skin flaps, commonly used in plastic and reconstructive surgery，which are usually subjected to ischemia-induced necrosis due to the ratio of pedicle width to flap length is far less than traditional ratio and thus limited its clinical application. Our previous studies had shown that hirudin could not only improve skin flap survival via anticoagulation, but also have the functions of microvascular reconstruction and cytoprotection. However, the molecular mechanisms of those functions are not well understood. In this study, we intend to establish the rat super-long random skin flap model and investigate the effects of natural hirudin on flaps microvascular reconstruction. Meanwhile, the single protease-activated receptors (PARs) pathway activation model would be utilized in vascular endothelial cells (VECs). With this model, the impact of natural hirudin on the PARs signaling transduction and the regulation of node genes (COX-1/2, NF-kB) as well as downstream proteins would be studied to investigate the molecular mechanisms of how natural hirudin improves microcirculation of flap and its cellular protective effects. The above research methods were critical for demonstrating molecular mechanisms of microvascular reconstruction and cytoprotection by natural hirudin in super-long random skin flap and solved the blood circulation disorder. And it might also provide the theoretical basis for the clinical application of natural hirudin to treat the super-long random skin flap ischemia in orthopedic, plastic and reconstructive surgery.

整形修复重建外科常用的超长随意皮瓣因长宽比例超常而导致的远端淤血、缺血坏死一直未能解决，限制了其临床应用。我们的前期研究发现天然水蛭素在辅助构建超长随意皮瓣中不但具有抗凝血功能，还可能具有微循环功能重建及细胞保护的作用，但其作用机理尚待进一步研究证实。本课题拟构建大鼠背部超长随意皮瓣模型，研究天然水蛭素对皮瓣微循环重建的效果；同时利用血管内皮细胞PARs单一受体激活模型，研究天然水蛭素对PARs跨膜信号的影响和该通路中的节点基因（COX-1/2、NF-kB）及下游各功能蛋白表达的调控，从而分析天然水蛭素改善皮瓣微循环及细胞保护的分子机制。上述研究对明确天然水蛭素在构建超长随意皮瓣中的微循环改善与细胞保护作用具有重要意义，或将使其血运障碍问题得到解决，进而为天然水蛭素辅助构建超长随意皮瓣在创伤及整形修复外科领域的应用提供理论依据。

整形修复重建外科常用的超长随意皮瓣因长宽比例超常而导致的远端淤血、缺血坏死一直未能解决，限制了其临床应用。本课题构建大鼠背部超长随意皮瓣模型，研究天然水蛭素对皮瓣微循环重建的效果，并进一步深入探索其作用分子机制。我们的实验结果证明局部应用天然水蛭素能有效提高超长随意皮瓣的成活率。我们的研究结果发现，水蛭素能有效增加超长随意皮瓣术后微血管生成并改善皮瓣血运。在促血管生成作用机制方面，水蛭素通过拮抗凝血酶/PAR1通路来调控下游p38 MAPK-ERK1/2通路间的交汇作用，促使皮瓣组织中ERK1/2磷酸化提高而降低p38 MAPK的激活，进而调控血管生成因子(VEGF)和抑制因子(TSP-1、endostatin)的分泌平衡，促进皮瓣组织血管的生成。在细胞保护作用中，水蛭素通过p38 MAPK通路调控炎症因子IL-6、TNFα及ICAM-1的表达，减少皮瓣组织中炎性细胞的浸润。同时，水蛭素通过抑制凝血酶诱导的p38 MAPK/IKK/NF-κB通路和JAK2/STAT通路减少细胞凋亡，起到抗凋亡的作用。上述研究结果明确了天然水蛭素在构建超长随意皮瓣中的微循环改善与细胞保护作用，为天然水蛭素辅助构建超长随意皮瓣在创伤及整形修复外科领域的应用提供重要的理论依据。