基于ERK/PI3K-Hif1α-PDGF通路调控周细胞募集探讨白芷促进糖尿病创面愈合的机制

Impaired neovascularization and related regulatory factors disorder is one of the important mechanisms of refractory diabetic ulcer. Angelica dahurica is a representative drug of the Tou-method in the traditional Chinese medicine formula Tuolixiaodusan. Our previous study (No:81273916) discovered that Angelica dahurica extract has profound angiogenic action. We propose a hypothesis of "neovascularization disorder in diabetic wounds - Angelica dahurica promotes the healing of diabetic wounds through recruiting pericytes and increasing neovascularization by regulation of ERK, PI3K, Hif-1α, PDGFβ signaling pathways". We apply the skin ulcer model of Akita diabetic mice, three-dimensional culture neovascularization model and endothelial cells-pericytes co-culture model to investigate the neovascularization disorder in diabetic wounds and use immunohistochemistry, double-labeled immunofluorescence, real-time fluorescence quantitative PCR, western blot and RNA interference to explore the effects of Angelica dahurica in the treatment of refractory diabetic wounds and to clarify the mechanisms of action. The study promises to explore active ingredients of traditional Chinese medicine and to provide new methods for the treatment of diabetic refractory ulcer.

新生血管化形成障碍和相关调控因子的紊乱是糖尿病溃疡难愈的重要机制之一，白芷是外科名方托里透毒散中透法的代表药物，我们前期的研究(No:81273916)发现白芷提取物具有明显的促血管生成作用。本课题围绕“糖尿病创面新生血管化障碍-白芷通过调控ERK、PI3K、Hif-1α、PDGFβ信号通路募集周细胞促进治疗性新生血管成熟加速创面愈合”工作假说，通过Akita糖尿病小鼠皮肤溃疡、动脉环三维培养的新生血管模型、内皮细胞/周细胞共培养模型，以创面新生血管化障碍为切入点，应用免疫组化、免疫荧光双标、实时荧光定量PCR、western blot、RNA干扰技术等手段，探讨白芷治疗糖尿病难愈创面的作用机制、作用环节，旨在阐明中药有效成分发挥作用的机制，为治疗糖尿病难愈性溃疡提供新的思路和手段。

糖尿病难愈性创面致病因素复杂，其愈合过程中伴随的一个潜在因素是血管发育不良，这种情况阻碍了创面的愈合。血管新生是创面愈合过程的关键阶段，新生血管成熟亦是慢性创面有效愈合的重要保证，而目前尚缺乏有效的治疗手段。白芷是中医外科临床治疗痈疽疮疡的常用药且效果显著，然而，白芷促进糖尿病创面愈合的机制目前仍不清楚。本研究旨在探讨白芷治疗糖尿病难愈性创面的效果，并从实验动物和细胞水平探索白芷的作用靶点和作用机制，为糖尿病创面愈合的治疗扩展新的治疗手段，也为促进传统中医理念与现代医学结合以及促进中医药现代化的发展提供科学的理论支持.研究结论：1.白芷具有明显改善糖尿病创面愈合的作用。2.白芷可促进糖尿病创面的血管新生和血管成熟，其机制与白芷增加HIF-1α的表达和转录功能，上调促血管生成因子PDGF-β和PDGFR-β的表达有关。3.白芷促进糖尿病创面血管新生的作用机制与激活PI3K/AKT、HIF-1α、PDGF-β/PDGFR-β信号通路有关。