水蛭地龙提取液通过VEGF-Notch/DLL4信号通路调控随意皮瓣成活的机制研究

Random skin flap is very common used in the tissue repair, however, the limitation of its length/width ratio and its ischemic necrosis at distal end hinder its clinical application.It is valuable to enhance its length/width ratio and improve its blood supplement. Our previous studies found that traditional Chinese medicine of Shuxuetong injection significantly enhance the expression of VEGF and inhibit the expression of Nocth4 and DLL4 to improve the survivorship random skin flap. In this project, we use traditional Chinese medicine of Shuxuetong injection to treat random flap in SD rat model (modified McFarlane method), the VEGF-Notch/Dll4 signaling pathway will be investigated. The survival of flap will be observated；Immunohistochemical staining, Western Blot, Real-Time PCR, Protein and Genetic chip will be applied. the key protein and gene expression of Notch, DLL4,VEGF, NICD, Hes, Hey will aslo be investigated. We will establish related gene expression profiles and a number of differences in the molecular target of random flap,clarify the molecular mechanism of Shuxuetong injection to promote random flap survival, and provide theoretical and experimental evidence to support its further clinical application.

随意皮瓣常用于临床组织缺损修复，但其长宽比例受限制，远端容易出现缺血坏死，影响其临床应用。如何在增加随意型皮瓣长宽比例的同时，改善皮瓣微循环及血运，保证较高的成活率，对扩大其临床应用范围有重要意义。前期我们研究发现水蛭地龙提取液可以促进VEGF的表达，Notch4和DLL4表达下降，从而促进皮瓣存活，但VEGF-Notch/DLL4具体调控机制未明。本项目拟建立SD大鼠超长随意皮瓣模型，以水蛭地龙提取液作为干预手段，VEGF-Notch/Delta信号通路为研究切入点，通过组织病理、细胞分子生物、蛋白、基因芯片等技术深入探讨活血药物治疗下皮瓣的Notch、DLL4、VEGF、NICD、Hes、Hey等为主导的基因表达和调控的变化，确立此类药物治疗下随意皮瓣的相关基因差异表达谱和若干分子靶标，阐明其分子作用机制，为进一步临床应用提供理论支持和实验依据。

临床上，随意皮瓣已经成为修复组织缺损的常见技术之一。然而随意皮瓣因不含知名血管，当其长宽比超越1.5-2：1，其远端将部分或者完全坏死，从而影响随意皮瓣在组织创面修复等中的临床使用。本课题旨在研究探讨水蛭地龙提取液对大鼠缺血随意皮瓣成活的影响及可能机制。. 建立改良的McFarlane大鼠缺血随意皮瓣模型。将雄性成年Sprague- Dawley大鼠按随机对数表法分为水蛭地龙实验组和生理盐水对照组。术后观察随意皮瓣存活大体情况及测算皮瓣存活面积比；术后第7天苏木精-伊红（H＆E）染色评估皮瓣组织病理学情况；免疫组化法测定VEGF、Notch1、Dll4表达；激光多普勒血流测量仪检测皮瓣血流灌注情况；明胶-氧化铅血管造影观测皮瓣新生血管情况、Western Blot检测VEGF、Notch1、Dll4等蛋白表达。. 水蛭地龙提取液可以促进大鼠缺血随意皮瓣的成活，其机制可能是水蛭地龙提取液下调Nothch1、Dll4的表达，上调皮瓣VEGF的表达这VEGF-Notch/DLL4信号通路，刺激皮瓣远端血管新生，改善皮瓣血供，从而提高皮瓣的成活面积。本课题研究结果为后期药物对随意皮瓣存活的临床转化研究提供了新的思路。